Abstracts of the 11th Annual Meeting of the Israel Society for Neuroscience

Directed convergence of dendrites causes synaptic clusterin and strengthenin Abi-Jaoue E. 1, Blinder P.2’3.’Lafrenirre-Roula M. 1, Cove j.2,3, Devroye L. andD. JDept. ofAnatomv and Cell Bioloer; McGill Universit); Montreal, Canada; 2Dept. ofLife’8iences, Ben Gurion University ofthe Negev, Beer heva; 3Zlowtovski Centerfor Neuroscience, Be ’er Sheva; 4Dept. ofComputer Science, McGill University, Montreal, Canada Synaptic clustering influences synaptic efficacy by increasing cooperative interactions among neighboring synaptic inputs, however, the mechanism of synaptm clustrring is unknown. We found that in hippocampal neurons in culture synaptic clustering as well as synaptic strength increase at the point of intersection between dendrites compared to other areas along the dendrites. The clustering in the intersection increases linearly and the strengthening exponentially with an increase in the number of dendrites crossing the intersection. Intersections containing multiple dendrites are frequent (4 times more than in computer simulations ofrandomly distributed dendrites) and are formed by direct growth of dendrites toward pre-existing dendritic intersections. This process is disrupted by inhibitors of synaptic activity and of glutamate receptors. Thus, an activity-dependent mechanism exists that causes dendrites to converge to a single intersection, resulting in an increase in synaptm clustering and synaptie activity at the intersection. This may se.rve as a mechanism for activity-dependent pre-.synaptie plasticity. In addition to converging, dendrites in the culture tend to grow in parallel, even when separated over large distances, leiding to formation of areas where specific dendritic orientations are dominant. This order restricts the location of the dendritic intersections and their synaptie dusters. Thus, directed convergence combined with parallel growth of dendrites shapes the synaptie map in cultured neuronal networks.

mammals. This hypothesis is based on the demonstration tha.t melanopsin is expressed in retinal ganglion cells that respond directly to light and that innervate the suprachiasmatic nucleus.
In the course of studying a potential role for melanopsin in photic entrainment, we found a fiber plexus immunoreactive to melanopsin in the retinorecipient region of the suprachiasmatic nucleus of the rat. To explore this further, we assessed the expression of melanopsin in the retina and suprachiasmatic nucleus of rats that were treated with the neurotoxin, monosodium glutamate, during the neonatal period. This treatment strongly reduced the number of retinal ganglion cells expressing mdanopsin and abolished the expression of melanopsm in the suprachiasmatic nucleus. Because photic entrainment is spared in spite of the massive damage to retinal ganglion cells bought about by neonatal treatment with glutamate, these findings suggest that retinal ganglion cells that project to the suprachiasmatic nucleus and that contain melanopsin in their axon terminals do not play a critical role in photic entrainment in rats. A possible role for these melanopsin containing ganglion cells in mediating the masking effect of light on behavior will be discussed. Certain CNS disorders such as Alzheimer's disease (AD), and traumatic brain injury (TBI) are accompanied by significant increase in inflammatory markers. Furthermore, AD and TBI in humans include reduction in certain cholinergic markers.
Based on cholinergic hypofunction observed in AD several cholinesterase inhibitors (ChEI) (e.g. Aricept, Exeleon and Reminyl) have been developed and approved for human use.
We have synthesized a series of bifunctional compounds containing both non-steroidal anti-inflammatory drug (NSAID) and cholinergic up-regulation (CURE) moiety. The treatment The lipophilic transition metal chelator DP-109 attenuates asymmetric rotations in the 6-hydroxydopamine partial lesion model of Parkinson's disease in adult rats Aran A., Friedman J.E., Shapiro I., Angel I. and Kozak A.
D-Pharm, Ltd., Kiryat Weizmann Science Park, Bldg. 7, Rehovot 76123 Perturbations in the homeostasis of transition metals such as copper, iron and zinc can cause oxidative stress in neurons. These metals are considered to be involved in neurodegenerative disorders such as Parkinson's and Alzheimer's disease. Antioxidants and metal chelators have been found to be beneficial in various models of these diseases, but are problematic due to a poor safe_ty profile and low rate of penetration across the blood brain bairier. To address this problem, we have developed a family of lipophilic chelators selectively activated in the vicinity of cell membranes. The lead compound, DP-109, was designed to chelate bivalent ions such as Zn, Cu, Fe, and Ca. DP-109 was tested in the unilateral 6-OHDA substantia nigra lesion model using adult male Wistar rats. Five days post-lesion animals were screened for apomohine-induced rotations over a 5 min period. Responding animals subsequently received either DP-109 (lO-500g/kg) or vehicle daily, p.o., for a period of up to 45 days. Animals were tested weekly for apomorphine-lnduced rotations. Results were standardized and the increase in asymmetric rotations, indicating progressive neurodegeneration, was calculated. Vehicle treated animals increased their rotations approximately 15-fold within 30 days. DP-109 treated animals demonstrated significantly fewer rotations in a dose-dependant manner. DP-109 (100g/kg) significantly attenuated (p<0.05) both the rate of increase and number of rotations by 70%. We suggest that DP-109 Pe s t . of molecular microbiology and biotechnology, faculty of cience, Tel-Aviv university, Tel Aviv 69978 The transmissible spongiform encephalopathies (TSE) or better known as prions, are rare neurodegenerative diseases which include scrapie in sheep, BSE in cattle and Kuru, Creutzfeldt-Jaob disease, Gerstmann-Strussler-Schienker syndrome (GSS) and fatal familial insomnia (FFI) in human. It is believed that prion propagation is caused by refolding of a normal endogenous glycoprotein callec], prion protein (PrPc): PrPc is expressed mostly n the central nervous system and lymphoid tissue. In the course of the disease PrPc changes its structure into an abnormal (-sheet rich structure termed PrPsc which is the only known particle in the prion infectious age.nt.
The abnormal protein is neurotoxic and-leads to death wthin few month. As for today there is no effective therapeutic agent to prion diseases. Antibodies are known to act as chaperones and are able to stabilize protein structure and/or induce conformational changes. Thus, our working hypothesis is that site directed antibbdies towards PrP may interfere with aggregation processes and/or inhibit prion replication.
We developed in our lab an active immunization procedure towards human Helix of the whole prion protein. Helix was chosen since it appears to be a putative key position in the protein induced conformational changes. In order to overcome the poor mmunogenicity of short peptides in general and the high sequence smilanty between the human and mouse peptides in particular, we chose the MAP (Multiple Antigen Peptides) as the antigen of choice. T.he MAP is a branching structure of lysine residues coupled to the desired peptide. The immune response that our peptide elicits was unexpectedly high. Antibodies raised were oflgG isotype. The antibodies that were produced recogmze the whole protein and are now being tested for their ability to inhibit the PrPc to PrPsc conversion in cell culture model. Keywords: PrP (prion protein), chaperons, MAP Sub-millisecond precision of the input-output transformation function mediated by fast sodium dendritic spikes in basal dendrites of CAI pyramidal neurons Ariav G..., Polsky A. and Schiller J.

Dept. of Physiology, Technion Medical School, Haifa
The ability of cortical neurons to perform sub-millisecond scale computations has been shown to be important for encoding of information in the cortex (1-2). Tiffs ability necessitates involvement of special mechanisms in order to overcome the relative long membrane time-constant and the significant dendritic filtering of excitatory postsynaptic 9.otentials (EPSP) in cortical neurons. Here we show that coincident activation of closely sp.aced basal inputs in CA1 pyramidal neurons resulted in significant sharpening (872+375%) and supra-linear amplification (156+37%) of the summed potential, as compared to the individual EPSPs. These phenomena are medmted by initiation of a local dendritic spike composed of an early fast sodium spike followed bv a slower NMDA spike.
When paired with apical EPSPs fast'local basal sodium spikes significantly improved the temporal precision of output action potentials by markedly decreasing the temporal jitter of action potentials (from 2.174:1.12 to 0.28+0.17 msec).
We examined the behavioral consequences of earl), exposure to stress, and in particular, on the abdity to cope with stress in adulthood. Post-weaning and Pre-puberty, at the age of 4 weeks, rats were placed on a platform in the middle of a water pool, for 30 minutes IPlatform stress).
In the experiment, we used as late stress the Host-intruder interaction at the age of 8 weeks. All rats were then tested in an open-field, the Morris water-maze and the startle-reflex test. In the 2 experiment adult rats were trained under stress condition (two-way avoidance task) in a shuttle box, followed affective examination.
report here that an early exposure to stress led to elevated levels of anxiety, as measured in the open-field and startle response tests. However, the rats that were exposed to the combinati,on of early and late stress exhibited the highest level of anxiet). In addition, and as was previously reported, the early + late stress group acquired a stressful spatial learning task faster. Furthermore, rats that were exposed to stress early in life did not learn the two-way avoidance task in adulthood. These findings suggest that an early exposure to stress may have lasting effects on both emotional and cognitive abilitis in adulthood, as is indicated by its effects on anxiety levels and on the performance in a spatial learning and two-way-avoidance tasks.
Supported by a grant 52/2000 from the Israel Foundation Trusties to G. Keywords: early stress, anxiety, learning Post-weaning pre-puberyexposure to stress effects on DHEA and DHEA-S in the rat hypothalamus and entorhinal cortex Avital A. , Jacobson $.x, Maayan 1z, Ram E. , Weizman A. d Richter-Levin G." Dept.  Recent evidance support the hypothesis that exposure to stress or trauma during eaxly childhood may disturbe the formation of functional brain pathways, in particular of the limbic circuits. Thus, we examined the effects of an early exposure to stress on behavior and neurosteroids levels in adulthood. Post-weaning and Pre-puberty, at the age of 4 weeks rats were placed on a platform in the middle of a water pool, for 30 minutes. As late stress, we used the Host-intruder interaction at the age of 8 weeks. All rats were then tested in an open-field, the Morris water-maze and the startle-reflex test. Immediately after the last behavioral test, rats were decapitated and the hyothalamus and entorhinal cortex (EC) were removed for measuring the neurosteroids DHEA and DHEA-S. We report here that an early exposure to stress led to elevated levels of anxiety, as measured in an open-field and startle response tests. In a spatial learmng task, the early stress group didn't reach criterion. In the hypothalamus, early exposure to stress did not alter either DHEA or DHEA-S levels, whereas exposure to stress in adulthood increased their levels. On the other hand, in the EC, early exposure to stress increased both DHEA and DHEA-S levels, whereas exposure to stress in adulthood decreased their levels comparedto the control and early stress groups. These findings indicate that an early exposure to stress has a differential effect on DHEA and DHEA-S levels in the hypothalamus versus EC. The relevance of which to the elevated anxiety is yet to established.
Supported bya grant 52/2000 from the Israel Foundation whisking field at different radial positions. We recorded single-unit responses and captured video-images of the whisker at ms resolution. This allowed us to accurately correlate the moment when the whisker touched the object with neural events.
We report here preliminary results from 'object detectors', i. e. single units that responded selectively to the contact between whisker and object (see accompanying abstract by Szwed et al.). We examined the responses of these neurons to objects positioned at three radial distances: 30%, 60%, and 90% of whisker length. We analyzed four response variables: spike-count (number of response spikes per whisking cycle), amplitude (peak fu'ing-rate), duration (of response burst), and latency to 1/2 of peak response. These data show that radial object position is encoded most consistently by spike-counts, less by amplitude, and much less by duration or latency. As radial distance increased, spike-count and amplitude decreased.
In contrast, as reported in the accompanying abstract, horizontal position was most consistently encoded by latency. These preliminary results suggest that horizontal and radial coordinates of object position are encoded differently: primarily by spike-count m the radial direction and primarily by latency in the horizontal direction. Supported by ISFgrant 3 77/02-1. Keywords: object localization, neural code, sensory encoding Multi-unit recordings from the gustatory cortex of.the freely behaving rat reveal differential response to familiar and unfamiliar taste during a distinct phase of the response Bahar A.,_Dudai Y. and Ahissar E. Dept. of Neurobiology, Weizmann Institute of Science, Rehovot 76100 The ability to distinguish an, unfamiliar tastant from a familiar one is crucial to the animal s survival, as the unfamiliar may poison. Ample molecular and cellular data support the hypothesis that the gustatory cortex (GC) plays an important role in the neural processing underlying detection of unfamiliar tastants, and transforming them to familiar stimuli. We investigate the neural correlates of gustatory unfamiliarity/familiarity encoding in the GC of the freely behaving rat. To this end, we employ chronically implanted multi-wre electrodes to record the extracellular activity of multiple units in the GC of rats, while they lick unfamiliar or famihar solutions. We calculate the ratio of responses to taste vs. water and compare them between exposures to unfamiliar and familiar tastants. We find that most recorded units demonstrate a typical lingering response (7 sec) to lsec of licking. The ratio of response to taste vs. water, in the initial 2 sec following drinking onset, is independent of whether the taste is unfamiliar or familiar. This ratio, however, increases significantly during the following 5 sec. Thus, our data imply that the GC responds differentially to familiar vs. unfamiliar tastants during a distinct phase of the neural response. Our data corroborate the notion that multiple taste attributes, including familiarity, are encoded in the GC and that at such attributes undergo experience dependent modifications. Supported by the Minerva  Recently it was shown in our laboratory that immune intervention by passive or active vaccination directed to tissue specific abundant antigens protect neurons from consequences of insult wthin the same tissue. Thus myelin associated antigens protect from consequences of acute axonal injury being a crush of the optic nerve or spinal cord contusion (Moalem et al. Nat. Med., 1999 Hauben et.al., J Neurosci 20,6421-6430 [20001. In contrast, retinal-derived abundant peptides protect from glutamate toxicity inflicted directly to the retinal ganglion cells (Mizrahi et al., J. Immunol. 2002).
Vaccination with the random copolymer, Copaxone (Cop-l), circumvented the tissue specificity barrier of protective vaccination. In this work we use an animal model for chronic neurodegenerative disease characterized by high intra ocular pressure (IOP) and progressive optic nerve de[eneration resulting in gradual visual field loss. We show that autoimmune neuroprotection is effective in chronic conditions.
We show that rats with high lOP arc amenable to protection by vaccination with antigens residing in the retina and not in the optic nerve. We show that the efficacy of Cop-I is dose and 69 regimen dependent and that its effect is superior to pharmacological intervention such as that achieved with ot2-adrenoreceptor agonist. It is suggestive that vaccination help homing of T cells to the site of damage. Such T cells are locally activated by the relevant antigen presenting cells and in turn amplify and regulate the tissue ability to fight against the local threat. Monoelonal antibody (mAb) 5.5 raised against Torpedo nicotinic acetylcholine receptor (AChR) is specific for the binding site t;f AChR and competes with the binding of t-bungarotoxin (ot-BTX) and other cholinergic ligands to the receptor. Here we show that mAb5.5 binds to snail acetylcholine binding-protein (AChBP), the structure of which has recently been deciphered. As the binding affinity of mAb5.5 to AChBP is considerably hig.h...er than that of ot-BTX to AChBP, we focused on the capability of this antibody to bind either AChR and or AChBP. We thias employed pepfides corresponding to the binding site loops of Torpedo and human AChR, and of snail AChBP. We tested the potency of the peptides to compete with the binding of mAb5.5 to either Torpedo AChR or snail AChBP. A peptide corresponding to amino acid residues 187-200 of Torpedo AChR ot-subunit (WVYYTCCPDTPYLD, inhibits the binding of mAb5.5 to bgth Torpedo AChR and to AChBP (IC50 values of 10 and 10"M, respectively). A peptide corresponding to the homologous residues in AChBP (SVTYSCCPEAYED, inhibits only the binding of mAb5.5 to its parental molecule, AChBP (IC5o of 10" M). Likewise, the peptide corresponding to amino acid residues 187-200 of human AChR tx-subunit (SVTYSCCPDTPYLD,, that has a high sequence similarity to the AChBP peptide , ,also inhibits only the bnding of mAb5.5 to AChBP (IC5o of 10M). Our findings demonstrate that mAb5.5 interacts directly with the ligand-binding site of AChBP, and makes this antibody a good candidate for analyzing the ligand-binding site of snail AChBP as well as of proteins that bind acetylcholine, in eyv_ords: nicotinic acetylcholine receptor; acetylcholine binding site; acetylcholine binding proten Pitch discrimination as a marker of reading and reading related cognitive difficulties Banai K.1 and Ahissar M. ' lnterdisciplinary Center for Neural Computation, Hebrew University of Jerusalem, Jerusalem; Dept. of Psychology, Hebrew University of Jerusalem, Jerusalem Previous studies documented strong correlations between auditory frequency discrimination thresholds (JND) and "high-level" cognitive abilities among reading disabled and normally reading adults. In this study we screened a regular class and a 'learning-disabled' (LD)class from two junior high-schools, respectively. In addition to reading, phonological awareness and memory we measured frequency discrimination. Two methods were used: pitch judgment (hi-lo) and similarity judgment (same/different-SD). In both, tones were 50ms long with Is inter-stimulus-interval. In each trial one tone was of 1000Hz. In the hi-lo method the other tone was always higher. In the SD method, the other tone was higher in halfthe trials and same in rest. We asked whether frequency JNDs are a good marker of reading related abilities. Repficating the adult findings, using the hi-lo paradigm, we found that the disabled readers in both classes (10% and 50% in the regular and LD class, respectively) were also those with the highest frequency JNDs (>30%) and with particularly impaired verbal memory. Hi-lo Frequency JNDs were correlated with verbal memory scores, but not with non-verbal memory. Surprisingly, JNDs on the SD task did not correlate with any of the reading or reading related measures, nor with the hi-lo JNDs. The finding that behavioral paradigm is an important factor in the correlation to memory and reading scores suggests that these relations do not simply result from variability in low-level auditory processing.__ Although the causal direction is n, ot yet clear, we suggest that hi-lo frequency discrimination ma) serve as a useful screening tool for memory related learning difficulties in schools.
Keywords: auditory processing, reading disability, frequency discrimination 70 Plasma membrane targetting of syntaxin la is enhanced following expression of M2 muscarinic acetylcholine receptors in PC12 cells Baram D., Shilkrot R. and Linial M.
In mammalian brain, the m2 subtype of the presynaptic muscarinic acetylcholine receptors (mAChR) are predicted as autoreceptors for ACh transmission. Preliminary results in our laboratory suggest a functional and molecular link between syntaxin, a crucial member of the exocytotic apparatus, and the rn2 mAChR. This link is mediated by the alpha subunit of the Go protein.
A key component in the functionality of G protein-coupled receptors is in their life cycle: intracellular trafficking, targeting to the membrane and internalization. Herein, we show that in the neuroendocrine secretory cell line PC 12, expression of m2 mAChRs results in efficient targeting of the receptors to the cell surface while altering the subcellular profile of syntaxin. In control PC 12 cells, that do not express endogenous m2 mAChR, syntaxin expression is partitioned between the Golgi and the plasma membrane. However, in cells expressing exogenous rn2 receptors, syntaxin is primarily localized to the plasma membrane. This phenomenon could be observed by both immunofluorescence staining and following subcellular fractionation of the cells. Furthermore, m2 mAChRs expression in these neuroendocrine cells causes a dramatic decrease of synaptophysin from the secretory granule fraction, whereas the protein remains located exclusively to a lighter membrane fraction that represents the small synaptic-like vesicles. The data suggest that m2 mAChR receptors are directly involved in the efficient trafficking of synaptic proteins, including syntaxin. The involvement of the G-proteins in the process and the physiological consequence of the observed phenomena are currently being investigated.
Keyw.ords: muscarinic ACh receptor, syntaxin, targeting, secretion Dynamic changes in mitochondriai function after closed head injury in mice Barash .1 Barnea D. 1, Alexandrovich A.2, Beni SM 2 and Dept. of Clinical Biochemistry, Hadassah Hospital; 2Dept. of Pharrnacology,'Hebrew University, Jerusalem Traumatic brain injury is associated with a rapid burst of reactive oxygen species (ROS), leading to oxidative stress.
Additionally, ischemia and energy failure occur within minutes to hours after trauma. Mitoehondria dysfunction is a major common pathway to these pathological events. The present study examines the fuctional activity of the mitochondria by measurment of ATPproducion at various time intervals after closed head injury (CHI) in mice. CHI was induced by a weight-drop device on 8-10 week old male Sabra mice, as described earlier. Mitochondrial fractions were isolated at 1, 4 or 24 h after CHI, and ATP production was followed by addition of glutamate (10mM) and malate (0.5raM) for 5 mins. The accumulating ATP was measured spectrophotometrieally. In addition, enzymatic activities of complex I-III and of complex IV were determined in the same preparations. A significant decrease in ATP production was noticed already at hpost CHI (-22% decrease form basal activity, p<0.05). A transient return towards baseline activity was found at 4 h (-8% decrease, not significant), which was later declined again, and a,t 24 h a decrease of-20% was recorded (p<0.05).
Interestingl), a similar bi-phasic pattern was reportedfor the endogenous low-molecular weight antioxidants, in the same model. The en,zyme activity assays revealed no changes in NADH-eytochrome-e oxidoreduetase and in cytochrome-c oxidase at any time tested. Citrate synthase activity, a marker for mitoehondrial content, was also unaffected by CHI. We propose that CHI-indueed effects which interfere with ATP-producing pathways by the mitochondria, contributes to the aggravation of cellular damage. Keywords: traumatic brain injury, mitochondria, oxidative phosphorylation High expression of apolipoprotein -E and Cathepsin B/D in spinal cords of amyotrophic lateral sclerosis patients and mice expressing human SOD mutation Barhum y.l, Ransmayr G. , Melamed E. and Offen D.
1Dept. of Neurolo_ gy, Felsenstein Medical Resear--nter, Rabin Medical Center, Petah-Tiqva 49100 and Sackler School of Medicine, Tel Aviv University, Tel Aviv 69978; : Dept. of Neurology and Psychiatry, General Hospital, 4020 Linz. Austria gerhard ransmayr@akh, linz. at Amyo_trophic lateral sclerosis (ALS) is a progressive, neurodegenerative disease characterized by loss of motor neurons in the cerebral cortex, brain stem, and spinal cord. A minori of ALS patients (5-10%) have a familial form (FALS) and 20 of these demonstrate mutations in the Cu/Zn SOD1 gene. Mice expressin.g, the human mutant SOD1 gene develop age-dependent ALS-like neurological symptoms. We studies the mRNA expression profile in post-mortem spinal cord sections from sporadic ALS patients and human controls. Using cDNA microarray gene expression we found significant increases in mRNA ot" cathepsin B (484%) and cathepsin D (206%), which are cysteine proteases that mediate intracellular protein turnover in the lysosme. MRNA of apolipoprotein E which is closely associated with the pathogenesis of neurodegenerative diseases, was also markedly increased (290%). Further analysis with specific probes revealed that the expression of these genes also increases in hSOD1-G93A transgenie mice. They show enhancement with disease progression and peak at the end stage of the illness. Our data from ALS patients supported by trmdings using the transgenic The nonpsychotropic dextrocannabinoid, PRS-211,092, a novel analog of Dexanabinol, does not bind to the NMDA receptor, but fias powerful anti-inflammatory properties (in-vtro). PRS-211,092 inhibits the ischemia-[nduced increased gene expression of cyclooxygenase-2 (COX-2) and proinflammatory cytokines and chemokines in brain of transient middle cerebral artery occluded (MCAo) animals. The aim of the present study was to determine whether PRS-211,092 had long-term beneficial effects on functional following focal brain iscnemia.
The MCA of Sprague Dawley male rats was occluded for 120 minutes by intraluminal suture under halothane anesthesia. PRS-211,092 5 mg/kg iv or PEG/ethanol vehicle alone were administered hour after the end of the ischemic insult. The neuroprotectiv,e efficacy of the compound was evaluated by the "staircase test' ( PRS-211,096 demonstrated dose-dependent (2 to 10 mg/kg) analgesia, which at 10mg/kg, was smilar to that of morphine (5mg/kg) 30 min after di'ug injection. In animals treated with PRS-211,096, significant analgesia was still evident at 90 min (by which time morphine analgesia was no longer detectable), and at 150 min after injection. PRS-211,335 also induced significant analgesia at dosages of 2 and 4 mg/kg at 30 min .after !njection, a trend that was still evident at 90 min after mjectlon.
PRS-211,096 was also tested for possible efficacy as an analgesic for neuropathic pain using the model of Bennett and Xie in the rat. Administered IP at doses of 0.5, 1.0 and 5.0 mg/kg, PRS-211,096 reduced tactile hyperalgesia, allodynia and thermal hyperalgesis in a dose-response manner. The efficacy of PRS-211,096 at 5.0 mg/kg was similar to that produced by morphine at a dosage of 5.0mg/kg. These cannabinoids may be novel and potent analgesics for the treatment of noxious and neuropathic pain in man.
Dept. of Behavioral Sciences and Zlotowski Center for Neuroscience, Ben-Gurion University of the Negev Acetylcholinesterase inhibitors (AChE-I) are widely used as domestic and a_gricultural organophosphate pesticides, exposing a substantial population to the risk of neurological damage. ACh plays a major role in eneoding attention and regulation of cortical development. Sexual dimorphism has been found in cholinergic development and in behavioral deficits after treatment with AChE-I's. A single exposure to (AChE-I) in 3-10 day old mice caused down-regulation of muscarinic receptors, impaired spatial learning and altered motor behavior in females only (Dam et al. Developmental Brain Research 121:179-187 [2000]). The present study examined the effect of chronic developmental exposure to AChE-I on avoidance learning. Neonatal C57BL/65 mice of both sexes w,ere injected with lmg/kg sc DFP or saline on postnatal da)s 4-10 and tested at age 4 months on the step-down passive avoidance paradigm. On Day 1, mice were placed individually on a vibrating platform above a metal grid floor. Upon stepping down, the mouse received a 0.5 mA footshock for 5 see. The procedure was repeated until the mouse made 2 escape responses up to a maximum of 10 trials.
The test phase, 24 hr later, consisted of a single trial without footshock. Females administered DFP had shorter step down latencies than controls on the test day, whereas no difference was found between DFP and control males (sex x treatment x trial interaction, F, 04=4.30, p<0.02). No difference in pain thresholds was found between DFP and controls. The results suggest a greater sensitivity of females to deficits in fear-motivated learning, following early exposure to AChE-I's. G-protein coupled receptors are not considered to exhibit voltage sensitivity. Here, using Xenopus ooc)etes, we showed that tlae muscarimc M (m2R) and M (mlR) receptors, known to modulate neurotransmitter release, are voltage sensors. The m2R-mediated G-protein gated potassium channel currents were used to assay the activity of the m2R. We found that the apparent affinity of the m2R toward acetylcholine (ACh) was redqced upon depolarization. The endogenous m lRmediated Ca dependent chloride currents were used to assay the activ.i.ty, of the m lR. The m lR also shown exhibits voltage sensavty but its apparent affinity toward ACh was increased upon depo|arization. Direct binding experiments of ['H]-ACh to individual oocytes expressing either m2R or mlR conf'trmed the electrophysiological findings. The cumulative results indicate that membrane potentialaffects either the G-proteins coupled to the m2R and the mlR or the receptors themselves.
Based on the fact that G-proteins are peripheral membrane proteins and considering the large similarity between the different G-proteins coupled to the mlR and the m2R, we favor 71 the possibility that the mlR and the m2R are by themselves voltage sensors. Keywords: G-protein coupled receptors, muscarinic receptor, voltage sensitivity, Xenopus oocytes CuZnSOD deficient mice show late-phase improvement of neurobehaviorai recovery after CHI and altered cortical reductiv capacity Beni SM Krool-Gal-ron N.. Kohen R., Tremtover v. E. and Barzillai A. 2 Depts. oflpharmacology and pharmaceutics, the Hebrew University, Jerusalem;-Dept. Of Neurobiochemislry, Tel-Aviv University Impaired redox homeostasis after brain trauma mediates cell-death pathways. CuZnSOD catalyzes O2 to H202 conversion, and along with other antioxidants constitute endogenous defense mechanisms. SOD administration was not beneficial in clinical trials, yet, SOD-transgenic mice display decreased lesion and apoptosis after brain trauma/ischemia. We assessed the effect of CuZnSOD deficiency on on outcome and reductive capacity_ after closed head injury (CHI). CHI was induced in CuZnSOD knockout (KO) and wild type (WT) mice, using a weight-drop devise. Neurobehavioral function was evaluated up to 14d after CHI, using Neurological Severity Score (NSS). The difference between NSS at lhr post CHI and at later times (ANSS) reflects recovery. Brain water content was measured 24hr after CHI. Reductive capacity was evaluated in sham (non-injured) and 5min post-CHImice using cyclic voltammetry. Sham KO mice displayed greater failure to exit a circle (p<0.0001) and to balance over a beam (p<0.005).
After CHI water content increased in the injured cortex (p<0.05) but to a similar extent in KO and WT. Yet, unexpectedly, at 14d after CHI, KO mice showed greater recovery (ANSS 3 vs 1; p<0.02). Cortical reductive capacity was higher in sham KO compared to WT (p<0.05) and decreased at 5 minpost CHI (p<0.05), whereas no changes occured in the WT. Hepatic reductive capacity was also higher in sham KO (p<0.0001) but was not affected by CHI, in contrast to WT m which it increased (p<0.01). We propose that KO mice displayed late-phase better recovery after CHI which is not accounted for by the early post CHI alterations in reductive capacity. Keywords Noladin ether (Nol) was isolated from porcine brain and identified as an endocannabinoid binding to CB cannabinoid receptor.
Aims: To evaluate the effect of noladin ether in mice under diet restriction (DR) as possible treatment for Anorexia Nervosa. Methods: Female Sabra mice assigned to 4 groups were fed on 2.5 hours a day diet for 14 days. Correspondingly were administrated i.p. different Nol treatment (Vehicle, nol 0.001, 0.01, 0.1 mg/kg). Food consumption (FC) and weight were measured. Cognitive function was evaluated using 8-arm maze and activity by x/y beam apparatus. After 14 davsDR all groups were fed ad-libitum.
Results: FC of mice administered 0.001 mg/kg/day Nol was significantly higher than those received vehicle (p<0.0001).
However, 0.01 and 0.1 caused significantly lower consumption cl?o<0.0001, <0.02 respectively). gnitive function: 0.1 Nol showed a significant decline in maze performance vs vehicle 0.001 and 0.01 nol. (p<0.05). Activity: 0.001 and 0.1 nol. attained significantly higher beam aaplparatussp,erformance than control.(p<0.00,) Weight: No sgmficant weight differences were obtained after one month of trial. Nevertheless, all mice groups lost an average of 10% in their weight.
Mice on diet restriction under noladin trea, tment (0.001mg/kg) increased their food intake while their actis itv was significantly higher than control and their weight didn't differ from vehicle. FC of mice under noladin (0.1mg/kg) decreased while their activity increased whereas their weight didn't differ probably due to decrease in BMR through IP3 signaling system(Leptin and endocannabinoids). A bi-phasic dose response in the FC (nol 0.001>vehicle>0.1 mg/kg) was noted. Cognitive function of vehicle, nol 0.001. nol 0.01 was improved, whereas on nol 0.1 it declined.
Noladin has significant effect on food consumption and activity. Thus, low doses of noladin may possibly enhance appetite in patients with Anorexia Nervosa and weight loss such as cancer and Aids cachexia. No such treatment is yet available. Keywords: Noladin, endocannabinoids, diet restriction, anorexia, food consumption, activity Visual and auditory "retain-and-compare" deficits in dyslexia Ben-Yehudah G. and Ahissar M. L3 Depts. of NeurobiologyI, Psychology: and3Center for Neural Computation, Hebrew UniversityofJerusalem, Jerusalem Previously, we found that dyslexics' contrast detection of temporally modulated gratings was impaired only when they were required to "retain-and-compare" between sequentially presented stimuli (Ben-Yehudah et al., Brain 124:1381-95 [2001). We now ask whether similar deficits characterize the performance of dyslexic adults when the stimuli are stationary. Using both auditory (tones) and visual (sinusoidal gratings) stimuli, we found that dyslexics' sequential frequency discrimination is significantly impaired in both modalities. We further compared visual frequency discrimination under two conditions. In the simultaneous condition, observers were asked whether the stripes were denser in the upper or lower half of the screen. In the sequential condition, observers compared stripe density between two successive intervals.
Consistent with our ,revious findings, the majority of the dyslexic group (74%) was significantly impaired on the sequential condition. A significantly smaller group effect was found for the simultaneous condition. Classifying dyslexic participants according to their auditory frequenc,, Under physiological conditions, T-cells may encounter dopamine in brain, blood, and various peripheral-organs.
Moreover, dopamine is elevated in the plasma in certain conditions of uncoping-stress, and injected iv in several emergency conditions. Can dopamine directly trigger T-cell function?
We recently discovered that dopamine interacts directly with its D3 and D2 receptors on normal human T-cells, activates 131-integrins, and induces subsequent T-cell adhesion Can dopamine also drive T-cells into cytokine secretion? To challenge this question we applied dopamine or dopaminergic D1-D4 receptor agonists to normalperipheral, cancer, andantigen-specific human T-cell clones (ofTh0, Thl, and Th2 phenotype), and analyzed the secretion of IFNT, TNFc, IL-4 and IL-10. We found that dopamine, in the absence of any additional molecule, can induce significant cytokine secretion from different T-cell types. The most prominent effect was the triggering of IL-10 secretion, a Th2characteristic anti-inflammatorv and suppress.ive cytokine. Dopamine also triggered a rharked secretion of TNFt, a potent pro-inflammatory Thl-associated cytokine, playing a crucial role in various inflammatory-diseases. Dopamine's effects were mimicked by specific dopaminergic-agonists, and blocked by the respective antagonists. Interestingly, the results suggested that dopamine may trigger IL-10 and TNFt secretion via different receptor subtypes. Taken together, we speculate an active role for dopamine in various IL-10 and TNFt-dependent T-cell activities. Dopamine's effects may be either beneficial or detrimental, depending on the context. Tel Aviv Sourasky Medical Center, Tel Aviv; :Tel Aviv University, Tel-Aviv, The Wohl Institute for Advanced Imaging, Tel-Aviv Sourasky Medical Center Introduction: What bridges the gap between stimulus and feelina? Originally James claimed that this gap is filled merely by a "feedback from an arousal response, while later others introduced the concept of cognitive assessment and appraisal of that arousal (1-3). These processes could well be complementary and may depend on the brain region and presentation mode of the stimulus. In this study we examined the interactions between the presentation mode of stimuli and their negative emotional appraisal in the brain by fMRI. Discussion." Stimuli affected activation level in sensory, limbic and language regions depending on the mode or presentation.
In contrary, emotional appraisal effect was found only in sensory and language regions. These results suggest that the distinction between processes that link stimulus and feeling is reflected in differential activation of sensory, language and limbic regions. This difference depends on either mode of presentation and/or appraisal ofnegative emotional stimuli. References: Slow twitch muscles are readily recruited and are active continuously while fast-twitch muscles are recruited when increased motor effort is needed. The synaptic activity of muscles involves acetylcholinesterase (ACHE) which is concentrated at the neuromuscular junctions and determines the duration of the neurotransmissin activating the muscles. Because synaptic AChE depends on neuromuscular activity we examined the regulation of AChE in fast and slow-twitch rat muscles following strenuous exercise. Rats were trained by walking on motor-driven treadmill (10d, lh/d at a speed of 9m/min with 2 min sprints of 17m/min, every 10min), so as to activate the fast-twitch leg muscles, not usually recruited in sedentary animals. Fast and slow-twitch leg muscles were isolated, AChE was extracted and its levels and isoform composition in trained and control-untrained muscles were analyzed. The results show increase in AChE content (per protein) in all examined fast-twitch muscles but not in the slow-twitch soleus muscle. The most significant increase was measured in trained gracilis anterior muscles (24%). All muscles contained globular G I+G2, G4 and asymmetric A12-AChE isoforms (4-6S, 10S and 16S sedimentation values, respectively). While GI+G2 and A12-AChE activities in both fast and slow-twitch muscles remained unchanged after training, the tetramer, G4-AChE, increased significantly in the exercised fast-twitch muscles (25-60%). These observations suggest that the G4-tetramer might become significant at the endplates of fast-twitch fibers following exercise. body-weight in infant OLETF vs. their controls, Long Evans Tokushima (LETO) rats, on postnatal days 2-4 and 9-11, during a 30-min independent ingestion test. In the current study we compared feeding suppression by 50% BW gastricpreloads of corn oil and mineral oil in 18-20 days old OLETF and LETO pups, and in Long-Evans pups on postnatal days 9-11. C-fos immunoreactivi.ty was examined in area postrema (AP), the nucleus of the sohtary tract (NTS), and in hothalamic areas implicated in feeding regulation. LETO rats ingested significantly less after corn oil than after mineral oil preloads. Their c-fos immunoreactivity in AP and NTS was ncreased compared to sham-preload controls. The effect of corn oil was not evident in OLETF pups; their mean number of c-fos expressing cells was increased in the caudal, subpostremal, and intermediate NTS, and in the AP, but not as much as in LETO pups. Intake of Long-Evans pups was not lower after corn oil compared to mineral oil preloads, replicating previous ontogenetic findings in Sprague-Dawley rats (Weller et al. Phvsiol Behav, 62:871-874[1997]). Their activation patterns in th AP and NTS corresponded to the intake results. These findings provide support for the role of CCKA receptors in mediating intake-reduction early in ontogeny. Supported by the US-Israel Binatonal Science Foundation K.e),words: CCK, c-fos, feeding, area postrema, nucleus of soltary tract Differentiation and neurotransmitter phenotype acquisition in developing neurons-differential expression of genes and roteins ogoc.h Y., TaTar S., Bledi Y. and Linial M. Det. of Biological Sciences, Silberman Institute ofLife sciences, The Hebrew University, Jerusalem, Israel P 19 cells are embryonal carcinoma cells that serve as a model for studying differentiation processes including commitment to 75 cell lineage. We have studied P 19 cells following activation of neuronal differentiation. The potential of these cells to mature and efficiently release neurotransmitter (NT) was established in our lab. We discovered that several variables, most notably, cell density and various neurotrophic factors affects neuronal maturation, survival, and most surprisingly, the choice of NT phenotype. We have observed that NT phenotype acquisition in P19 cells is mediated by cell-cell contact and not by soluble factors. We have shown that changes in cell density are associated with changes in gene expression. A large-scale holistic view on gene expression was obtained by DNA chip technology. The data obtained from this screen was extensively analyzed with various bioinformatical methods. Among the genes found in that screen are members of the Wnt and Cullin families which function in transducing a signal from the cell surface.
Mass Spectroscopy comparative proteomics approach was applied on sparse and dense P 19 induced cultures. Membranes were collected at different time windows following neuronal induction. Differentially expressing proteins were excised and sent to Mass-spectroscopy analysis. Currently, over 30 proteins were successfully analyzed by such methodology. Using 2D gels we were able to improve detection level and could identify relatively low expressing proteins. Most intriguing proteins are a variant of Drebrin a putative dentritic-shaping molecule and several cytoskeletal-signaling molecules. Those and other proteins reflect the morphological reorganization that takes place while establishing PI 9 neuronal fate. Recent studies indicate that the endogenous circadian system is sensitive to much lower levels of light than initially suspected (Boivin DB et al Nature 1996;379:540-542.). It was also recently reported that 70% of the resetting effect of a bright light stimulus can be preserved even though it is interrupted by episodes of complete darkness for 63% of the time (Rimme'r DW et al. Am J Physiol Regulatory Integrative Comp Physiol 2000;279 :R1574-R1579). The usefulness of a judicious schedule of exposure to room light (Ira-st study) or to intermittent bright light (second study) to treat circadian maladaptation to jet lag or shift work was tested. In the first study, 15 healthy young men participated to a laboratory simulation of a Montreal to London voyage (Boivin DB and James Ft. J Biol Rhythms 2002;17(3):266-276 ). They were exposed to 380 lux for 6 hours each day either before bedtime or n the morning. This study demonstrated that the schedule of exposure to room light can substantially affect circadian adaptation to a shifted sleep/wake schedule. In the second stud.y, 15 nurses working permanent night schedules were studed for 3 consecutive weeks (in press). They were either exposed to bright light intermittently for 6 hours in their workplace (treatment group) or worked under their usual lighting conditions (control group). Circadian reentrainment was complete in the treatment group only. Results of these two studies underline the importance of the control of the overall pattern of light exposure in circadian adaptation to shifted sleep/wake schedules. We have previously showed that this phenomenon is unlikely to reflect retinal suppression, sensory, masking or adaptation, but the mechanisms involved in MIB are largely unknown. Recent evidence suggest that parietal mechaniss representing space in different reference frames may be involved in the control of awareness, and possibly in MIB.
Here we ask what is the frame of reference'in which disappearance during MIB occur. To answer this question we measured the magnitude of disappearance at different spatial locations in three different conditions: (1) retinotopic .mapping in which a single target dot was presented at different locations relative to fixation, (2) a head-centered condition in which the direction of the head was displaced by 20 deg. to the right or left keeping direct fixation and (3) object-centered condition in which a target dot in fixed retinal location was surrounded by an elliptic contour with different relative displacement along it's main axis. Results show the effect of all three manipulations on disappearance. Anisometropic disappearance was found for all observers with more disappearance in the upper and upper-left visual fields. Disappearance map changed for different angles of head rotation even though the retinal locations were identical. The elliptic contour induced more disappearance around its focal points and less in its center.
These results suggest that disappearance in MIB involves or is affected by head-centered and object-centered mechanisms. Keywords: visual disappearance, visual awareness, visual space Evaluating ion channel modulating activities in a Tarantula venom Brass D., Dan P., Magnushevsky N., Marom A., Meidan O. and Meir A. Alomone Labs Ltd. P.O. Box 4287, Jerusalem 91042 The flow of ions through membrane channels is an essential component of cellular function. Recently, ion channels have emerged as potential targets for diagnostic and therapeutic agents due to their membrane localization and their contribution to the physiology and pathophysiology in many biological systems. The venomous animal world offers a large variety of peptide toxins, with some exerting their biological activity on various ion channel activities, by means of high affinity interaction and selective modulation.
Alomone Labs R & D focus on the discovery of new effective and specific ion channel peptide toxins. These toxins may serve as potential leader compounds for diagnostic and therapeutic agents, which affect various diseases, such as cancer, cardiovascular diseases and neurological disorders. Alomone Labs has developed a technological platform for finding, characterizing, producing and modifying peptide-toxin ion channel modulators. It includes: Separation and purification of peptide toxins from whole native venom.
Assay.s for fmding molecularly def'med ion channels as possible targets for the purified toxins, and for evaluating the characteristics of the toxin pharmacology.
Rec.ombinant production and purification of a desired peptide toxin.
Here we focus on the separation and dissection of ion channel modulating activities found in the venom of one spider species. The spider Grarnmostola spatulata (Brown Morph Grammostola-BMG) venom effects on various cloned voltage dependent ion channels, was examined using two-electrode voltage clamped Xenopus oocytes.
The BMG whole venom strongly inhibited the following channels: Kv2.1, Nay 1.5, Cav 1.2, Cav2.2 and Cav3.1. BMG venom was subjected to SP-sepharose fractionation, yielding 8 separate peaks and the blocking activity, within the venom was narrowed to specified peaks for each of the channels examined.
A peak containing single peptide that was identified by mass spectroscopy and amino-acid analysis as the previously described o-Grammotoxin SIA specifically blocked Cav2.2 channels.
Strong blocking activity towards Kv2.1 channels was found in two separate peaks that contain several peptides each.
However, further analysis showed that this peak contained several different peptides.
In conclusion, we show wide range and specific ion channel blocking activities, carried out by separated fractions of Tarantula spider venom. Some of these activities affect channels with poor specific pharmacology such as the Car3.1 channel.
Keywords: channels, toxin, Tarantula Maternal deprivation increases alcohol consumption in adulthood in the rat Brave __M., Berger B.D. and Richter-Levin G.
Dep. of Psychology & Brain and Behavior Research Center, Haifa University,-Haifa Early maternal deprivation (MD) is considered an adverse stressful experience, associated with long-lasting effects in adulthood. It is suggested that this kind of stress may be a risk factor for the development of high alcohol consumption in adulthood. The aim of the present study was to examine, in a rat model, the relationship between early maternal deprivation, and subsequent development of high alcohol consumption. We further examined the influence of long-term isolation on MD modulation of alcohol consumption in adulthood. Method: Male Wistar rat pups were exposed to one of 4 different rearing, conditions: 1) Maternal separation from PND 7 for 7 consecutive days, for one hour every day. 2) Long-term social isolation immediately post weaning. 3) Maternal deprivation and social isolation. 4) Regular rearing conditions. Pups were weaned at PND 28. At the age of 3 months all animals were examined in a free choice paradigm for alcohol and water consumption.
Results: We observed a significantly high rate of alcohol ption only in rats that were maternally deprived 3 months before.
These finding indicate that MD has long lasting effects on emotional development.
In contrast to previous reports studying social isolation as a stressful experience, social isolation m this study was found to attenuate the effect the MD treatment. The main difference between the current and previous studies was that in the present study social isolation began immediately after weaning while in previous studies rats were exposed to social isolation in adulthood. Further examination is required to clarify these differences.
Supported by a grant from the Israel Foundation Trusties (2000) to G.R-L. Keywords play an important role in the modulation of behavour and are involved in a series of neuropsychiatric disorders. Despite the importance of these cells, little is known about the molecular mechanisms governing their development. During embogenesis midbrain dopaminergic neurons are specified rostral to the mid-hindbrain organizer (MHO) and hmdbrain serotonergic neurons caudal to it. We report that in transgenic mice in which the MHO is shifted caudally, the midbrain dopaminergic neuronal population expands to the ectop.ic positioned MHO and s enlarged. Complementarily, the extension of the hindbrain serotonergic cell group is decreased. In adulthood these changes are preserved and the additional, ectopic dopaminergic neurons project to the striaturn, which is a proper dopaminergic target area. Also in mutants in which the MHO is shifted rostrally, dopaminergic and serotonergic neurons are relocated at the newly positioned MHO. However, in these mice the size ratio between these two cell populations is changed in favour of the serotonergic cell population. To investigate whether the position of the MHO during embryogenesis is also of functional relevance for adult behaviour we tested mice with a caudally shifted MHO and report that these mutants show a higher locomotor activity. Taken together, we provide evidence that the position of the MHO determines the location and size of midbrain dopaminergic and hindbrain serotonergic cell populations tn vivo. In addition our data suggest that the position of the MHO during embryogenesis can modulate adult locomotor activity. Keywords: mid-hindbrain organizer, dopamine, serotonin, attention-deficit hyperactivity, disorder Olfactory-learning induced reduction in post-burst AHP in piriform, cortex neurops is due to a decrease in slp Brosh I.' and Barkai E.
Zlotowski Center for Neuroscience, Ben-Gurion University. Beer Sheva; :Center for Brain and Behavior, Haifa University We have previously shown that olfactory learning results with reduced post-burst afterhyperpolarization (AHP) in layer II pyramidal neurons of the priform cortex. In the present study we examine which of the currents that generate the post-burst AHP ,I and IAm, is reduced after learning Rats were trained in an olfactory dscnrmnatlon task to distinguish between positive and negative odor cues until they demonstrated rule learning. Three days after completion of training, intracellular recordings from pyramidal neurons were performed in piriform cortex brain slices. As previously shown, AHP amplitude was significantly smaller (P<0.02, one way ANOVA) in neurons from trained rats, compared with neurons from pseudo trained and naive rats (-4.98_+0.30 mV, n=32 in trained, -6.79+0.38 mV, n=23 in pseudo trained and -6.76_+0.37 mV, n=4 in naive). In the presence of 101M NE, AHP amplitude in neurons from trained rats did not differ from those of pseudo trained or naive rats (-6.32_q: 0.71 mV, n=14 in trained, -5.20 +0.54 mV, n=12 in pseudo trained and -5.64+0.40 mV, n=24 in naive). In the presence of 50nM of the specific IAHP blocker Apamin, AHP amplitude remained significantly smaller (P<0.03, one way ANOVA) in neurons from trained rats, compared with controls (-2.76+ 0.41 mV, n=17 in trained, -4.67 :k-0.59 mV, n=16 in pseudo trained and -4.73+0.31 mV, n=9 in naive). Furthermore, the reduction in AHP amplitude was stron er in neurons from trained rats (45%) then that in O O pseudo trained (31Yo) or naive rats (30 %). These data indicate that learning induced post-burst AHP reduction is accompanied by reduction in ratao of SlAHP /IAHP conducnatnes. We Suggest that the differential effect of noradrenaline is due to this change.
Keywords: olfactory-learning, piriform cortex, AHP, Our in vitro studies showed that co-culturing of microglia with autoimmune T cells resulted in up-regulation of the expression of MHC-II and B7-2 (both reminiscent of the activity of antigen-presenting cells) as well as increased expression of TNF-(, TNF-RI and TNF-RII in early interaction arid later on, decreased expression of TNF-RII (possibly associated with termination of the microglial activity). We suggest that autoimmune T cells exert their protective effect through dialog with local microglia and invading macrophages enabling these cells to express their beneficial activity, while at the same time imposing strict regtdation so as to avoid their potentially detrimental long-term effects. Keywords: CNS ,spinal cord injury, secondary degeneration.,, autoimmune neuroprotection, microglia/astrocytes, TNF-, The ischemic synapse and the role of mitochondria Carlen P.

University of TOronto, Canada
Mitochondria are increasingly implicated in neurodegenerative mechanisms from several perspectives, including intracellular calcium regulation, free radical production, energy utilization, and release of apoptotic factors. Synaptic transmission could be an early and sensitive target forcerebral ischemia and other neurodegenerative processes.
We have shown that an eight min ischemic (hypoxia/hypoglycemia) insult to an organotypic hippocampal slice culture, promoted glutamate-mediated generation of free radicals with concomitant elevation of intracellular calcium (Perez Velazquez et al., 1997). In this same model, the mitochondrial complex inhibitor, rotenone, the mitochondrial permeability transition blocker, cyclosporin A (CsA), and a blocker of NAD+, nicotinamide, decreased ischemia-induced free radical generation and increased mitochondrial calcium (Frantseva et al., 2001). Interestingly, CsA did not diminish the increase in the cytoplasmic calcium, but did reduce the increased mitochondrial calcium, sugsesting that mitochondrial calcium could be the most important mediator of neurodegenerative processes. Preliminary experiments, in acutely prepared rat hippocampal slices, have shown that during and following 8 min. of hypoxia/hypoglycemia, strattm radiatum evoked synaptic transmission in the CA1 subfield is depressed along with an increase in presynaptic c)oplasmic and mitochondrial calcium, which persists for up to one hour following the ischaemic insult. The presynaptic increase in calcium is reduced by BAPTA-AM. We hypothesize that the neuroprotective actions of intracellular calcium chelation could be by limiting rises in intramitochondrial calcium and that the maintenance of 'healthy' mitochondria will be neuroprotective ainst ischaemic insults. orted by the CIHR Keywords: ischemia, s)naptic transmission, mitochondria, presynaptic calcium Can T-cells be driven to de novo gene-expression under the exclusive command of a neuropeptide?
In this stiady, we investigated whether T-cells can be directly activated by gonadotropm-releasing-hormone-II (GnRH-II), a unique neuropeptide conserved through 500 million years of evolution, and recently identified in the brain of'various mammals, as well as in various peripheral tissues. We found that exposure of normal human or mouse T-cells to either GnRH-II or GnRH-I (the two neuropeptides sharing 70% homology but encoded by different genes) leads to de novo gene-transcription of various genes, among them the cell-surface-expressed 67-kDa non-integrin laminin-receptor, that plays a key role in cellular adhesion and migration, tumor invasion/metastasis, and specific infections.
GnRH-II or GnRH-I also directly induced cell-surface-expression of the 67-kDa-laminin-receptor, adhesion to laminin, chemotactic migration towards SDF-lot, and augmented entry in vivo of metastatic T-lymphoma into specific organs. Further, homing of normal T-cells oecmT"ed to a lesser extent in GnRH-I-knockout mice than in normal syngeneic mice. A ,.sjaecific GnRH-I receptor-antagonist blocked the induced effects of GnRH-I but not of GnRH-II, suggesting signaling through distinct putative binding sites.
Finally, we found that normal human and leukemia T-cells produce GnRH-II and GnRH-1. Taken together, we suggest that GnRH-II and GnRH-I, secreted from nerves or autocrine/paracrine sources, interact directly with T-cells, and trigger specific gene-transcription, adhesion, migration and homing of T cells across blood and tissue barriers. We further speculate that these GnRH-T-cell interactions may be beneficial in conditions of normal T-cell behavior, but detrimental when cancer or autoimmune T-cells are involved. coupling ratio of the neuron signal of about 0.5 together with relatively large transistor trans-conductance. The combination of the FG structure with depletion .t)pe device, leads to the following advantages: a) no DC bias between the biological solution and the transistor with direct consequences to the neuron as well as the silicon die durabili.ty. b) The sensing area of the neuron activity is separated from the active area ofthe transistor. Thus, it is possible to design the sensing area and the channel area separately, c) The diannel area, which is the most sensitive part of the transistor, can be insulated and shielded from the iofiic solution, d) The option to add a switchi.ng transistor to the FG and use the FG also for the neuron stimulation.

Keywords: MOS transistor, action potential
The neural activity in rat motor cortex changes during the learning of a conditional sensory discrimination task Cohen D. and Nicolelis M.A.L.
of Neurobiology, Duke UniversityMedical Ctr., Durham,770.5 Neural activity in motor cortex has been intensively studied for many years. Yet, little is known about the changes that occur in these neurons' activity, as animals learn a conditional auditory discrimination task. In this study, arrays of microwires were chronically implanted in motor cortex (MI) of adult rats to allow the recording of single unit activity as the rats learn to associate an arbitrary tone with a movement. This approach allowed us to characterize changes in cortical activity due to learning at the level of single neurons as well as the ensemble. Adult rats were trained to nose poke a center hole and wait for a go cue which was either a high or a low tone. Depending on the tone, the rat was required to nose poke a hole located either to the right or to the left of the center hole. Following correct trials, the rat received water reward. In this task, the rats learn to associate each tone with a specific movement in addition to improving the movement performance via repetitions. Our results show that during the learning process, the behavioral attributes of reaction time, movement time and error frequency decreased with practice. The improvement in task performance was accompanied by changes in the firing patterns of neurons associated with the task. These changes included sharpening of the response to the task related events and modification of the average firing rate. Keywords: chronic recording, motor learning ERK activation is correlated with s_ynaptic modifications after odor rule learninl in the Diriform cortex 2" ohen S. Shiboleth A. Rosenllum K. and Barkai E. Zlotowsk Center for Neuroscience Ben-Cmrion University Beer Sheva; "Center for Brain and Behavior, Haifa Unlverstty, Haifa We have shown previously that post-burst afterhyperpolarization (AHP) and paired pulse facilitation (PPF) is reduced in piriform coaex ofpyramidal neurons, days after olfactory-trainning. We assume that this long term cellular modifications subserve at least part of the odor rule learning and are looking into possible molecular mechanisms for these long term modifications (See also Shiboleth et al.). Extra cellular regulated kinas (ERKIIII) were found by others and us to play major role in learning and memory as well as svnaptic plasticity in cortical areas. We began to study the possible involverhent of ERKI/II in long-term modulation of intrinsic and s3rnaptic properties correlated with odor rule 79 learnin^in,e piriform cortex. The specific MEK inhibitor PD9809 (4o IsM) caused a significant increase in PPF litude in neurons from trained rats only. Consequently, the rence in PPF amplitude between trained and pseudo rats was diminished (from 1.22_+0.25, n=21 to 1.56+ 0.29, n=7 in trained and from 1.34+0.20, n=6 to 1.44+0.18, n=5 in pseudotrained). PD98059 did not have an effect onpost-burst AHP amplitude in any group on neurons (from 6.25+2.29, n=44 to 6.53+ 2.72, n=19 in trained and from 7.82_+1.74, n=20 to 7.31_+1.74, n=8 in pseudotrained). Using sub-cellular fractionation and quantitative immunoblot analysis of anti phospho-specific antibody and protein antibodies we are examining ERKI/II activation in the piriform cortex. In preliminary research we detected 20% increase (n=4) in ERKI/II activation in the cytosolic fraction of trained (n=4) compare to control rats. We conclude that ERKI/II is involved with long-term modulation of synaptic transmission in the piriform cortex of trained rats but not in post burst AHP 3 days following training. Keywords: olfactory-learning, piriform cortex, paired-pulse facilitation, ERK PorOSapsin induces dendritic clustering of GluR1 and spine rmation Cove J.', Morales C. and Baranes D. ' Dept. ofLife Sciences Ben-(htrion University of the Nee,ev. We found that in hippocampal neurons prosaposin is localized in vesicles within dendrites, but is absent from axons and synaptic connections. Depletion of prosaposin in knockout mice is associated with an approximately 3-fold lower expression level of dendritic Glutamate Receltor (GIuRI) in the hippocampal dentate gyrus and CA! regions compared to the wild type. Supportingthis, application ofan anti-prosaposin antibody to the growth medium of hippocampal cultures reduces dendritic GIuRI cluster density b 44% and GluRI cluster size bv 56%. These effects are specifically on GluRI since the treatment with anti-prosaposm has n) effect on NMDAR1 clusterin.g. Furthermore, ant-prosaposin reduces the density of GluRl-containing spines by 88%, whereas application of exogenous prosaposin increased this density by 91%. Our results show that extra-cellular ac(ivilv of prospaosin clusters GluR1, but not NMDARI in dendrites, andpromotes ine formation. ywords: dendritic spines, glutamate receptor, prosaposin However, intracellular accumulation of AI3 also occurs at early stages of AD. In vitro studies have shown that intracellular AI3. (AI3i) can disrupt cellular organelles and MAP kinases. We have continued investigations regarding the consequences of elevated levels of AI3i in a rat transgenic model, expressing human transgene genes related to familial forms of AD, whose CNS pathology is limited to intraneuronal accumulation of in the hippocampus and neocortex. In this transgenic rat model we have observed that the presence of intraneuronal AI3, in the absence of amyloid plaques, is sufficient to provoke up-regulation of the MAP kinase ERK2 in the hippocampus at 9 months. This is a dvsregulation, which appears specific, since no changes were c;bserved in other putative tau kinases.
such as GSK3 or-13 and CDK5. These rats displayed a concomitant increase in the level of tau phosphorvla/ion at ERK2 target sit,es. Confocal microscopy studies revealed that neurons displaxing AI3i, also manifest changes in the Golgi complex and l\,sosomes. Interestingly, these morphological and biochemital alterations vere 'llowed by behavioral cognitive impairments which became apparent at 16 months of 8O age, thus suggesting that in the absence of amyloid plaques, accumulation of intracellular AI3 peptides is sufficient to initiate steps in the tau-phosphorylatlon cascade as well as to provoke late impairments ofhigher CNS functions. We are currently investagating other potential protein dysregulations within our transgenic rat model of early AD pathology, in an effort to identify the molecular pathology preceding plaque formation. Dendritic spines, the site of synaptic connections between central neurons, play an important role in information pressing and memory formation. Corticosterone, one of the major stress-induced hormones in mammals, may have profound effects on the morphology and functions of specific brain areas. One of the brain areas most affected by stress hormones is the hippocampus, a limbic structure, known for its involvement in learning and short-term memory. To examine the effects of corticosterone on the structure and functions of hippocampal neurons in relative isolation, we exposed dissociated hippocampal neurons cultured for two weeks in-vitro to 0.11aM corticosterone for 24 hrs, and measured the effect of the hormone on several morphological indices. The most striking and consistent effect of corticosterone was a decrease of about 26% in the density of dendritic spines without a concomitant change in other morphological parameters, including soma size, and number of primary dendrites per neuron. Also, there was no difference in the size of the spines as a result of exposure to corticosterone. Interestingly, the presence of the steroid antagonist RU486 by itself caused a significant increase in spine density abovb control level, and blocked the effects of corticosterone.
Acute (30 min) corticosterone treatment caused a significant increase in NMDA receptor-independent rise in [Ca]i reactivity of the cultured neurons to topical application of glutamate.
These results indicate that cortcosterone causes pruning of dendritic spines by increasing calcium load through an enhanced voltage gdted calcium channel activity, as suggested elsewhere. Keywords: corticosterone, hippocampus, dendritic spines Intermittent pentobarbitai microinfusion into the rat MPTA induces long lasting anesthesia Del Canho S. and Devor M.
Dept. of Cell and Animal Biology, Institute ofLife Sciences. Hebrew University of .lerusalem, Jerusalem Aims of!nvestigation: We have identified an area in the rat brainstem mesopont!n.e t.egmentum (MPTA) where minute pentobarbital mlcromlectons induce flaccidity, analgesia, slow-wave EEG and d.pparently loss of consciousness (Devor & Zalkind Pain, 2001). Following single pentobarbital bolus microinjection the anesthetic state typically lasts 20-30 min. In many types of neurobiologicalresearch it would be advdntageous to anesthetize animals without flooding the neural tissue under investigation with anesthetic agents. We therefore asked whether more prolonged anesthesia can be generated by pentobarbital microinfusion into MPTA.
_M_ethods. WJstar-derived Sabra strain rats (280-300g) were nesthetized and implanted with bilateral stainless steel guide-cannulae aimed at MPTA. After full recovery (one week) a 30g microinjection needle was inserted through each guide cannula bringing its tip within MPTA. We selected rats in which pentobarbitalmicronjection induced moderate to deep anesthesia 11-16 on the previously used 16-point scale). lesu/ts. After inducing anesthesia with a bolus microinjection (100lag in 0.5tl per side), microinfusion was initiated (10g in 0.05 pl per minute). Animals tended to lighten, awakening after 40min. However, intermittent microinfusions ot" 20tg in 0. ltl per minute or 40tg in 0.2tal per minute for periods of 5-10 rain every 30 min induced stable anesthesia for periods of 2-3 hours (6 experiments in 4 rats). Conclusion: Prolonged anesthesia can be obtained with repeate b---her microinfusions, although similar doses infused continuously at lower rate, appear to be less effective. MPTA microinfusion is a potentially useful tool for inducing stable, prolonged anesthesia without delivering drugs to the systemic circulation.
Keywords: microinjection, general anesthesia, microinfusion Differential pattern of CREB activation in the rat brain  [1998]). This implies that activated CREB could be used as a metabolic marker to delineate neuronal circuits that subserve specific items in memory. However, the question how CREB activation in identified neurons contributes to the specificity of the internal representation encoded b.' the relevant circuit, is still unexplored. As a fir.st step m this direction, we describe here the differential activation of CREB in the rat brain as a consequence of two related yet distinct forms of aversive conditioning, conditioned taste aversion (CTA) and conditioned context aversion. Using immunohistochemistry, we found that CTA induces strong CREB activation in the insular cortex (IC) and the lateral septum (LS), but not in the parietal cortex (PC) and the medial septtun (MS). In contrast, contextual conditioning resulted in strong activation in the PC and MS, but not in the IC and LS. are not yet known. Recent studies revealed that AI3 deposits occur in mice transgenic to amyloid precursor protein (APP). The occurrence of these deposits is age dependent and it is not yet known whether they can be cleared or degraded in vivo.
The brain neutral endopeptidase, Neprilysi-n, is a major All degrading protease. In the present study we inhibited brain neprilysin activity by prolonged i.c.v injection of Thiorphan, a selectwe inhibitor of Neprilysin, and examined the resulting effects on the rate and extent of AI3 deposition in the brain as well as on the extent to which these deposits can be cleared and degraded. Accordingly, i.c.v administration into the lateral ventricle of Thiorphan by Alzet osmotic pumps (0.5mM at a flow rate of 0.25.d/hour) revealed a time dependent increase in the number of immunohistochemicaly detectable AI3 deposits.
The level of AI deposits increased as a function of time for at least month after initiation of treatment. Examination of the spatial distribution of the AI3 deposits in the brain revealed maximal levels in the cortical areas whose ventral surface was the largest. The extent to which AI3 deposition is affected by AD genetic risk factors such as the apoE genotypes and the degree to which these deposits can be cleared from the brain will be discussed. that demonstrated a more rapid onset of clinical effects than classical antidepressants. However, the mechanism that enables some drugs to have a faster onset of action than others is poorly understood. The aim of the current study was to determine dynamic changes in the brain in response to a fast-acting antidepressant to elucidate possible neurochemical parameters that correspond to the improvement of depressive behavior. In our previous studies, we found that Flinder sensitive line (FSL) rats, an animal model of depression, are characterized by: a) increased immobility during the forced swim test, b) absence of serotonin-induced dopamine release in the nucleus accumbens, and c) abnormal expression and increased inhibitory-like activity of 5-HT2c receptor. All of these parameters were normalized after chronic (14 day) treatment with antidepressants. In the current study, we found that subchronic (7 day) treatment with nefazodone (a fast-onset antidepressant), but not with despiramine (a classical antidepressant), normalized immobihty time in the swim test in FSL rats. Nefazodone treatment of FSL rats also restored normal activity of the 5-HT2c receptor and accumbal serotonin-dopamine interaction after a subchronic treatment. We conclude that the restoration of accumbal serotonin-dopamine interaction via the 5-HT2c receptor seems to be a critical parameter for the fast Results from two neglect patients showed significantly longer reaction times (RT) and movement times (MT), compai'ed with control subjects. Moreover, a contralesional to ipsilesional gradient was fomd in both RT and MT.
The initial direction of motion in both 'oung and elderly healthy controls depended on the modification time (RT Inter-Stimulus-Interval), and included averaged/non-averaged and direct trajectories. In contrast, both patients showed no such dependence and no averaged modified trajectories. These findings suggest the patients' inability to amend their ongoing movements in response to a sudden change in the target location. Furthermore, a higher proportion of direct trajectories was observed in DS movements, for movements that were initially directed toward the left side of space.
In addition, whereas in elderly subiects and patients, some of the DS movements containeda pause before the direction of the trajectory was modified, (15% and 35% of the DS movements respectively), the patients displayed significantly longer pauses. MoreoVer, pauses were significantly longer when the first target appeared ipsilesionally, with respect to the trunk-midline. These findings reflect the existence of a competitive bias in favor of ipsilesional stimuli. Results from a perceptual control experiment demonstrated the existence of a spatial perception deficit in both patients. Taken together, our findings suggest that the impairment seen in the two patients could not be related to either pure perceptual or ure motor deficits.
eywords: neglect, reaching, competition 82 A new member of the cyclin family may play a role in long-term memory Edelheit S. Yahav S. and Meiri N.

Institute of Animal Science, Agriculture Research
Organization, Volcani Center, Bet-Dagan 50250 Identifying the molecular correlates of long term-memory has been a focus of research, nevertheless little progress has been made and only few molecules have been directly correlated with the neuronal processes underlying consolidation. In this project we were interested in findings gene products that are altered hours after learning. Since the prevailing hypothesis emphasizes the role of neuronal plasticity in memory consolidation we used a behavioral paradigm that occur during early developmental stages and hens amplify changes in memory-related gene expression. We used the one-day old chick passive-avoidance learning paradigm in which a chick learns to avoid eating a bead coated with an unpleasant tasting substrate. The intermediate medial h.yperstriatum ventrale (IMHV) is centrally involved in storing this memory. A screen using differential display for changes in mRNA expression in the IMHV between 2 and 24 hours after training chicks on the passive-avoidance memory task, revealed a new gene sequence with an homology of 80% with the cyclin family that was induced after training with a peak at 12 hours after training. Our results coincide with earlier findings suggesting selective induction of a different member of the cyclin family ania-6 by different types of neurotransmitters in the striatum. It is also known that certain cyclins can interact with RNA polymerase II. This might be a novel mechanism for regulation of neuronal gene expression which convergent with multiple lines of evidence suggesting that acute regulation of pre-mRNA splicing is important in neuronal plastici,. Keywords: passive avoidance, cyclin, chicks, differential display The extinction of fear conditioning in Medaka fish (Oryzias latipes) Eisenberg M. and Dudai Y.
Deptl of Neurobiology, Weizmann Institute of Science, Rehovot 76100 Ample evidence indicates that experimental extinction is learning rather than unlearning. However, it is yet unclear how different is the mechanism of extinction from that of acquisition. In the present study we used a developmental approach in order to address this question. Medaka fish of three different ages (lweek, 4 week and 7 months) were subjected to fear conditioning. They were trained by pairing light (conditioned stimulus, CS) with electric shock (unconditioned stimulus, US) for 20 trials over 2 days. Fear response was evaluated by computerized comparison of locomotion before and during CS presentation. Subjects were tested on memory retention starting 24 hours after the last training trial. Afterwards, the fish were subjected to 20-30 successive trials over 2-3 days in the absence of the US. Significant age-dependen( differences were unveiled in retention and extinction of conditioned response. Whereas all age groups readily acquired the task, the youngest fish (I week old) failed to retrieve memory of learned fear 24 hours later. Adult fish (7 month old) successfully extinguished the fear response in 20 trials, while the 4 week-old fish continued to exhibit a startle response to the CS even after 30 trials. These results suggest that extinction demands a more mature nervous system than the acquisition of conditioned fear. In addition, the results validate the ability to use Medaka fish, which are a convenient neurogenetic model, for the study of the ontogeny of learning and memory. Keywords: fear conditioning, extinction, acquisition, ontogenesis A novel amphiphilic motif in the N-terminal helix of heterotrimeric G-proteins Elia N., Kosloff M. and Selinger Z. Dept. ofBiological Chemistry, The Hebrew University, Jerusalem Heterotrimeric G-proteins relay signals between membrane-bound receptors and downstream effectors. The ot subunits of this super-family are anchored to the membrane by one or more lipid modification at their N-termini. These modifications can be palmitoylation, myristoylation or both. As no sequence determinant for palmitoylation is apparent, we used systematic homology modeling of all different human G, proteins to look for a three-dimensional structural determinant of palmitoylation, rather then a linear sequence motif. Comparison of the N-termini of this super-family revealed that all ot subunits modified only by palmitoylation contain a similar structural motif at their N-terminal helix. This motif is characterized by a prominent basic patch that exten a positive potential well beyond the molecular surface of the protein. Furthermore, this structural motif is oriented opposite to the face that interacts with the 157 subunits. Hence, these positive patches are free to interact with the negatively charged inner surface of the plasma membrane. Based on previous results, we suggest that that efficient palmitoylation of G proteins reqmr_" es prior targeting to the plasma membrane. The signal for this membrane localization can therefore be either .myristoylation or the novel motif that we identified. This signal s cooperative with the interaction of the tx subunit with the 13' complex. The N-terminus of a Go protein can therefore be described as amphiphilie, containing dual signals attracting it to the membrane and enabling it to undergo palmitoylation. As palmitoylation has been shown to modify a plethora of proteins extending beyond G-proteins, this motif could be widely applicable. Keywords: structure, G-protein, membrane-attachment Proteomics approach to study the rule of ERK in learning and olasticity Elkobi A, 1, Admon A.2. and Rosenblum K. Faculty ofScience and Science Education, University ofHaifa; :Dept. of fSiology Protein Center, Technion, Haifa We are interested in the role of the extracellular regulated kinase I/II (ERKI/II) in the formation of long-term memory and plasticity. ERK I/II is involved in both early and late phases LTP (Rosenblum et al. J. Neuroscience [2002]). In addition, ERKI/I activity is correlated and necessary for the formation of long-term memories (e.g. Berman et al., 1998).
Using subcellular fractionation, phospho-specific antibodies and proteomics (two Dimensional Electrophoresis, mass spectrometry, and bioinformatics) we are trying to identify ERKI/II substrates in the brain that are involved in memory formation. In order to identify ERKI/II substrates we are using antibody that recognizes phospho_rylated-Threonine only when followed by the armno acid Proline (the favorite phosphorylated site by ERKI/II). In addition, we test the mod/dation in synaptosomal protein expression and post translation modifications using different fractionation methods. The functionality of the identified spots or bands will be determined in the insular cortex both following learning (novel taste learning) and LTP. Keywords: consolidation, ERK, proteomics Oxidative stress induced by 6-hydroxydopamine affects ubiquitin-conjugates, protein degradation and proteasome activity in PC12 cells: Implications for the pathogenesis of Parkinson's disease _ElkonH., Melamed E. and Often D.
Dept. of Neurology, Felsenstein Medical Research Center, Rabin Medical Center, Petah-Tikva. 49100, and Sackler School Od/oMedicine, Tel Aviv University, Tel -Aviv ffen::&post.tau,ac.il Mutations in familial Parkinson's disease (PD) have been associated with the failure of protein degradation through the ubiquitin-proteasome system. Impairment of proteasome function has also been suggested to play a role in the pathogenesis of sporadic PD. Protein damage seen in the substantia nigra in PD was postulated to be nduced by the local oxidative metabolism of dopamine. We examined the proteasome activity in PC 12 cells treated .with 6-h;cdroxydopamine (6-OHDA), the dopamine synthetic denvate used in models of PD. We found that the treatment increased protein degradation, increased the levels of free ubiquitin and ubiquitm-conjugated proteins, in a dose dependent manner. In addition, there was an increase in proteasome trypsin-, chymotrypsin-and post acidic protease-like activities in cells treated with 10-100uM of 6-OHDA, whereas higher doses caused a dramatic decline.
Similarly, the presence of 0.3 mM 6-OHDA for up to 10 hours increased proteasome activities while further incubation (10-24 hrs) reduced them markedly. We demonstrated that 6-OHDA treatment increased the protein degradation, accumulation of carbonyls groups and caspase-3 activity, while addition of antioxidant, N-acetylcystene, prevented these effects. In conclusion, our data indicate that mild oxidative stress elevates proteasome activities in response to the increase in protein damage. Severe oxidative insult may lead to failure of the ubiqmtin system to clear defective proteins and cause protein aggregation and cell death. Control of protein clearance might offer a new strategy for therapy in neurodegenerative diseases in general and particularly for PD. Keywords: Parkinson's disease, 6-hydroxydopamine, ubiquitin-conjugates, protein degradation, proteasome activity Axotomy induced reversed microtubules polarity leads to the formation of a vesicles trap and the extension of a ezH. wth cone's lamellioodium , Shapira E. , lI0ogenraad C.C.2, de  Erasmus MC, the Netherlands.
The transformation of a stable axonal segment into a motile growth cone is a critical step in the regeneration of amputated axons. A strikinjg observation made in earlier studies from our laboratory, is nat down stream, in the cascade leading to growth cone formation, anterogradely transported vesicles accumulate at a defined compartment some 100-150rum proximal to the ti" p of the cut axonal segment. This compartment forms tlie GC's center. Here we began to explore the mechanisms by which vesicles are trapped at this specific location. To that end, we expressed EGFP-EB3 in cultured Aplysia neurons. EGFP-EB3 binds in stretches to the plus end of microtubules, moves with the growing MT's tips forming a comet tail like structure and thereby allows to Image the dynamics and polarity of MTs.
We found that axotomy leads to restructunng and re,orientation of the MTs polarity at the cut axonal end. 100-150pro from the cut axonal tip where the GC center is formed and vesicles are trapped, the MTs reorient such that plus ends point towards a common center-the trap. Proximally the trap is formed by MTs that maintain their original polarity. The distal boundary of the trap is formed by MTs with reversed polari_ty. Distal to that, following a short axoplasmic gap the MTs point their plus ends anterogradely. We propose that the vesicles trap is formed by the reorientation of the MTs polarity thus directing molecular motors to deliver vesicles into the trap. Keywords: axotomy, growth cone, microtubules, regeneration, EB3 ErbB4 receptor expression elevates following closed head inju_ry Er-lich S. Shohami E. and Pinkas-Kramarski R. tDept, ofNeurobiochemistry. Tel-Avv University, Tel-Aviv 69978; Dept. of Pharmacology, The Hebrew University, Jerusalem 91120 The ErbB-4 receptor tyrosine kinase and its ligand neuregulin are widely expressed in the nervous systems. To investigate their possible role in neurodegeneration we used the closed head injury (CHI) model. We demonstrate that levels of ErbB-4 are dramatically increased at the site of inj.ury. ErbB-4 levels in the cortical region at the site of injury were significantly increased starting at day 3 post trauma, lasted two weeks and then declined to basal levels. Notably, the highest levels of ErbB-4 expression at the site of injury were observed one week after CHI. The staining patterns indicate that activated microglia/macrophages and neurons but not astrocytes, constitute the major population of cells that highly express the receptor at the injury site. Confocal microscopy analysis suggests that the high levels of ErbB-4 protein in activated microglia/macrophages is probably due to phagocytosis of neuronal cells. These findings indicate that ErbB-4 receptors may play a role in brain responses to head trauma.
Keywords: tyrosine kinase, ErbB-4, brain, closed-head injury, microglia/macrophages NRG mediates neuromd differentiation and survival in PC12-ErbB4 cells Erlich S., Goldshmit Y. and Pinkas-Kramarski R. Dept. ofNeurobiochemistry, Tel-Aviv University. Tel-Aviv 69978 Neuregulins (NRGs), a large family of transmembrane polypeptide growth factors, mediate various cellular responses depending on the cell type and receptor expression. To examine the possible role of ErbB-4 in neurons we have used the PC12 cell. Expression of ErbB-4 in PC12 cells demonstrated that NRG induced signals and neurite outgrowth, which were indistinguishable of those mediated by 1VGF. In PC12-ErbB-4 cells, NRG induced an initial weak mitogenic signal and subsequently neurite outgrowth. The differentiation induced by_ NRG involves generation of reactive oxygen species (ROS). In PC12-ErbB-4 cells NRG can rescue from apoptotic cell death induced by serum deprivation or TNFt treatment. We also show that NRG induces a significant protective effect from H202 induced death. This effect of NRG is mediated by the PI3K signaling pathway, since NRG failed to rescue cell from H202 insult in the presence of the PI3K inhibitor, LY294002. Furthermore, the downstream effector of 83 PI3K, PKB/AKT, is activated by NRG in thepresence of H202 and PKB/AKT activation is inhibited by LY294002. In addition, our results demonstrate that the high levels of ROS induced by HzO2, is inhibited by NRG. LY294002, which blocks NRG mediated rescue, increases ROS levels. Moreover, both H202-induced ROS elevation and cell death are reduced bp3expression of activated PI3K. These results suggest that K dependent pathways may regulate toxic levels of ROS generated by oxidative stress. In conclusion, our results demonstrate that neurite outgrowth induced by ErbB-4 in PC 12 cells, requires MAPK and PKC signaling networks and NRG induced survival from apoptotic cell death requires PI3K signaling. Keywords: phosphatidylinositol 3-kinase (PI3K), ErbB-4, neuregulin (NRG), tyrosine kinase, reactive oxygen species ( and by other pro-apoptotic factors. In mouse brain, ARTS was seen m a number of brain areas, including several parts of the cortex, hippocampus, septal nuclei and bed nuclei of stria terminalis. In this work we explored the expression of ARTS in cerebellar granule neurons in primary culture by immunohistochemistry. Content of ARTS increased rapidly after plating. After 24 h m vitro, more than 90% of the neurons were strongly ARTS positive. The number of ARTS positive neurons was similar in cultures grown in medimn containing a .physiological concentration of potassium, and in 25 mM K* medium (in which the cells are viable for a long.er period in vitro). ARTS protein was seen to a similar extent m mitotic and non-mitotic neurons, as detected by BrDU incorporation. ARTS was detected by confocal microscopy in mitochondria and nuclei both in young (24 h) and mature (10 day-s in vitro) cultures. Since o_ur previous work in COS ceils has shown that migration of ARTS from mitochondrion to nucleus occurs at the onset of apoptosis, this finding may indicate which neurons in the culture are undergoing apoptosis. The finding of a high content of ARTS in cerebellar granule cells in culture contrasts with the near absence of ARTS in cerebellum of the post-natal rat as seen in sections of whole brain, and also correlates with the high degree of apoptotic cells (10% by TUNEL stain) in the neuronal culture in vitro.
Keywords: apoptosis, ARTS, cerebellar granule cells Transgenic excess AChE exerts non-catalytic effects on hippocampal LTP via PKC Farchi N., Hochner B. and Soreq H. empSCiences lnsamte and Interdisciplinary Center [br Neural utation, Hebrew Universi, JetTsalem AChE is known for its catalytic ACh hydrolyzing featm'es, but most interestingly acts as a sensitive marker for acute and chronic stress, as a specific splice variant of AChE is highly elevated under such insults. Transgenic models for chroniC: expression of either the stress-related AChE (AChE-R) or its primary synaptic variant (AChE-S) express both altered LTP patterns in hippoeampal CA1 svnapses and memory and learning impairments. Specificall,, LTP is enhancedunder transgenic excess of AChE-R (172 s. 138 in WT) whereas excess AChE-S leads to normal levels of potentiation that dramatically decays to 37% within 3h post-induction (vs. no decay in WT). lhtriguinglv, the decay phenotype was not rescued bv either phvsostigine (1 taM) or carbachol (0.5 tM) administration. Mtreover, it was also evident in the hippoeampus of transgenics expressing catal,1icallv inactive AChE-S, suggesting that non-catalytic features of AChE are involved in LTP maintenance. We examined the possible involvement of PKC in the distinct LTP responses of AChE-R or-S transgenic slices. Phorbol di-butvrate (5nM). a PKC activator, highly-facilitated svnaptic field "potentials in AChE-R slices (101% vs. 65% in WT) contrasted by poor (30%) facilitation in AChE-S slices. Furthermore, a tetanus following PKC activation induced stable late-phase LTP in AChE-S slices, thus rescuing the LTP decay. To conclude, we report the contribution of AChE non-catal'tic features involving PKC activity, and/or levels in LTP priming and maintenance. This study may demonstrate stress-related modulation of neuronal plasiicity.

84
The physiological relevance of the physical interaction of voltage gated K channels with protel'n components of the exocytotic machineryin PC12 cells Fill O., Singer-Lahat D., Chikvasgvili D. and Lotan I.
Dept. of Physiology & Pharmacology, Sackler Faculty of Medicine, Tel Aviv University, Tel Aviv 69978 Pmsvnaptic voltage-gated K (Kv) channels play a phygiological role m the regulation of transmitter release by virtue of their ability to shape presynaptic action potentials.
However, in a previous work (Fill et al, 2001) we showed that in brain synaptosomes and Xenopus oocytes Kvl.1 channels and SNAREs proteins display a physical interaction. Also, our preliminary results show that another Kv channel, Kv2.1, also interacts with the SNARE proteins in oocytes. In this study we aimed to attribute a physiological role to the physical interactions of these Kv channels with the exocytotic proteins using the cracked PC12 cell assay (Hay and Martin, 1992). Both channels, Kv 1.1 and Kv2.1, are expressed in PC 12 cells. In co-immunoprecipitation experiments we could observe a physical interaction between the SNARE proteins and the Kv2.1 channel in PC12 cells. We introduce GST-proteins corresponding to parts of the Kv 1.1 and Kv2.1 fragments of the two channels that were shown in in vitro binding assays to either bind or not SNAP-25 or syntaxm 1A and assayed their effects on the Ca regulated neurotransmitter secretion m ths system. Our results are consistent with the notion that the physical binding of Kv channels to the SNARE proteins modulates Ca + regulated neurotransmitter secretion. Keywords: Kvl.1, Kv2.1, SNARE proteins, PC 12 The endogenous cannabis system: role in the stress response and critical function in milk ingestion and survival in newborn mice Fride E. 1, Foox A. 1, Weidenfeld j.2. and Mechoulam R.

Dept. ofBehavioral Sciences College ofJudea and Samaria
Artel: "[-Iadassah Medical School and lept. of Medwmal Chemistry and Natural Products of the Hebrew University of Jerusalem The ability of cannabis to stimulate appetite has been used to benefit patients suffering from malhutrition. Endogenous cannabinoids ("endocannabinoids") are found in brain, peripheral organs, and maternal milk; they activate cannabinoid (CB 1-CB2) receptors. We showed recently that CB receptors are critical for mlk ingestion and survival in newborn mice.
Here, we further investigate the requirement for CB receptors in newborn feedin and development in CB receptor-knockout (CB'/ mice. Mil intake, weight gain, and development were recorded in CB and in wild type pups. Further, pups were injected with a CBI receptor antagonist (SR,141716A) on day of life. CB" pups did not ingest milk on the first day of life, and survival ,'as significantly lower compared to wild types. SR141716A completely inhibited milk intake in wild type pups and resulted in almost 100% mortality within 7 days. The antagonist partially affected milk ingestion, growth and survival in CB pups.
Throughout this study, we noticed a greater vulnerabilitv and mortality in CB mice, suggesting a role for" the endocannabinoid system in stress. Indeed, acute stress (4 min noise) induced a rise in corticosterone and in ACTH in the sera of CB "/ mice, which was respectively, twice and 4 times higher in knockout compared to wild type rice.
We conclude that 1) neonatal milk ingestion is entirely dependent on the endocannabinoid-receptor system. A major role is played by CBI receptors; however our data support evidence "for an additional ("CB3") receptor. 2) The cannabinoid system plays an important role in the response to stress.
Keywords: cannabinoids, stress, development, feeding T-cells express a functional ionotropic glutamate receptor GiuR3, and glutamate by itself directly activates T-ceil function. 12 Ganor Y., Besser M. and Levite M.' Dept. of Neurobiology Wei-]stitute of&'ience Rehovot 76/00. "7'he 3ackler laculty of Meaqclne, 7el-Arty University, Tel-Avv 69978 T-cells may encounter glutamate, the major excitatory neurotransmitter in the nervous system, when patrolling the brain, and in glutamate-rich peripheral organs. Moreover, CNS glutamate levels increase in various pathological conditions in which T-cells may play a beneficial or detrimental role.
Do T-cells express ionotropic glutamate receptors'? Can glutamate by itself trigger T-cell function? We found, for the first time, that normal human T-cells, human T-leukemia line; and mouse autoimmune anti myelin basic protein T-cells express high levels of specific ionotropic glutamate receptor ofthe AMPA subtle 3 (GluR3).

The evidence for GIuR3 expressmn in T-cells include
GluR3-specific RT-PCR and sequencing, showing identity to brain GluR3, western-blotting, and GluR3_ cell-surface expression revealed by immunocytochemical fluorescence staining and flow cytometry. Glutamate (10nM), n the absence of any additional molecule, activated specific T-cell functions, among them integrin dependent adhesion to laminin and fibronectin, and chemotactic migration towards the chemokine SDF-1.
AMPA receptor-agonists mimicked glutamate-induced effects, and the specific antagonists CNQX and NBQX blocked it. Taken together, we suggest that GluR3 expression in T-cells and glutamate-triggered T cell function could be .lmportant to numerous physiological and pathological conditmns, and especially relevant to: a) T-cell transmigration to the CNS, across laminin-containing blood-brain-barrier; b) T-cell mediated multiple sclerosis, c) Some human epilepsies in which specific anti-GluR3antibodies are found and suspected to play a neurotoxic role. Based on our fmding we speculate that granzyme-B-producing T-cells can cleave the GluR3B autoantigen in an autocrine (from their own surface-expressed-GluR3) or paracrine (from neighboring T-cells) fashion, not only from neuronal cells, as suggested thus far.
Laboratory of Neuroendocrinoloev of aging, CHUMResearch Center, Notre-Dame Hospital and'departments of2Medicine and 3Nutrition, University of Montreal, Montreal,Canada.
In aging mammals, a decreased responsiveness of somatotrophs to growth hormone-releasing hormone (GHRH) leads to low GHand IGF-I serum levels. This event was proposed to be responsible for a diminution of muscle mass, ncrease adiposity and deterioration of several tissues and organs. In the anterior pituitary, it has been related to changes in the level of GHRH-receptor (R) mRNA transcripts and immunoreactivity and to modifications of GHRH binding parameters. In long-term moderate calorie-restricted rats, a model of successful aging, youthful levels of pituitary GHRH-R mRNA transcripts and GHRH binding sites are maintained. Among regulatory factors positively impacting on GHRH-R, corticosterone and testosterone could be proposed as candidates. In 18-month-old calorie-restricted rats, serum corticosterone and testosterone are increased when compared to 18-month-old ad libitum-fed rats. An optimal control of such as in caloric restriction, could also protect the I-IRH-. In streptozotocin-diabetic rats, levels of pituitary GHRH-R mRNA transcripts exhibit several disturbances according to the lensth andseverity of diabetes. Moreover, a glucotoxicity state mimicking that found in aged or diabetic rat serum down-regulates the human GHRH-R, stably expressed in BHK cells, indicating that it may represent a common mechanism of GHRH-R aging. Altogether, these results suggest that circulating steroids and glucose may be critical to regulate the expression of the pituitary GHRH-R in aging. As seniors represent the fastest growing sgment of the population in industrialized countries, identific.ation of cellular an.d molecular mechanisms involved in somatotroph axis dysfunction may allow to design new interventions to delay somatopause.
Dept. of Molecular Genetics, Weizmann Institute of cience,

Rehovot 76100
Lissencephaly type is a severe human disease that affects 1/30,000 live births. It is characterized by a relatively smooth cerebral surface, and an abnormal organization of the cortical layers. This is the result of an abnormal embryonic neuronal migration. The clinical manifestations are short life span, epilepsy and mental retardation. Mutations in LISI and doublecortin (DCX) have been shown to cause Lissencephaly type I. A third gene, doublecortin-like kinase (DCLK) is thought to participate in the DCX pathway. Known protein interactions of LIS1 and DCX suggest their involvement in intracellular transport. Intracellular transport is a complicated problem for a highly polar cell like a neuron. A recent idea suggests that it would be advantageous for a neuron to assemble the components of a signaling pathway by loading them together on a scaffold protein. This scaffold protein s bound to a motor protein, and thus delivers the entire complex to the right place. This can ensure that the components of a pathway will be in the right ratio, all together, and promise spatial regulation on the pathway. According to this model, proteins of the same pathway are predicted to be transported together.
Here we show that both DCXand DCLK interact with a scaffold protein that assembles the mitogen activated protein kinase (MAPK) pathway in neurons.
This observation links for the first time neuronal migration with the MAPK pathway. Our hypothesis is that these interactions function to regulate the localization of MAPK module.
Beer Sheva Mental Health Center, Psychiatry Research Unit, Ben Gurion University of the Negev, Beer-Sheva It was previously shown that the dopamine D4 exon III repeat (D4DR) and the serotonin-transporter promoter region (5-HTTLPR) polymorphisrns are not only associated with adult personality traits but also with temperament in 2-week old neonates. We now report the results of a third study of these infants and their temperament at 12 months. This study examined the assocmton between two common polymorphisms, the DRD4 gene and the 5-HTTLPR and temperament m 12-month-old iants. 22 infants had a least one copy' of the 6-8 repeat DRD4 alleles (L-DRD4) and 39 had two copies of the 2-5 repeat allele (S-DRD4). 20 infants were homozygous for the short form (s/s) of 5-HTTLPR while 41 were ether heterozygous for the short and the long form (I/s) or were homozvgous for the long form (I/l). The infants were observed in a series of standard temperament episodes that elicited fear, anger, pleasure, interest, and activity. L-DRD4 infants showed less interest in a structured block play situation and more activity in a free play situation. They also displayed less anger in an episode of mild physical restraint. Infants -ith s/s 5-HTTLPR showed less fearful distress to stranger approach and less pleasure in a structured play situation than infants with 1/1 or I/s 5-HTTLPR. Duration of looking during block play was affected by a significant interaction between DRD4 and 5-HTTLPR. Shortest duration of looking was associated with the L-DRD4 and s/s 5-HTTLPR genotypes. These results support previous data showing that the origins of the molecular control of human temperament lie in infancy. Keywords: dopamine D4 receptor gene, serotonin transporter promoter genes, infant temperament DCX and development Ghosh I., Gdalyahu A., Levy T., Caspi M. and Reiner O.
Dept. of Molecular Genetics, Weizmann bstitute of Science,

Rehovot 76100
The Lissencephaly syndromes in humans involve abnormal cortical lamination and are medically categorized as neuronal migration defects. Two genes involved in Lissencephaly type have been identified: LISI and DCX. Doublecortin (DCX), a microtubule binding protein, maps to the X chromosome and mutations in this gene results in Lissencephaly in males or subcortical heterotopia in females. This strongly suggests a role for this gene product during neuronal migration. The protein consists of two 80 residue evolutionary conserved repeats and a serine proline rich C-terminal region. The repeats bind microtubules and most mutations that are tbund in Lissencephalv patients cluster in this domain The Cterminal portion has ben shown to interact with clathrin adapter proteins, AP-I and /or AP-2, thus suggesting a potential role of DCX in protein sorting or vesicular trafflcking.
Scaffold proteins laave been shown to regulate MAP kinase related pathways. We have identified a specific MAP kinase which phosphorylates DCX which maybe important during neuronal migration.
We have identified the phosphorylation sites on DCX and have also identified the regions through which it interacts with the MAP kinase complex. Using different biochemical approaches we have established that DCX is a key molecule in neuronal migration.
Dept. of Physiology and Pharmacology, SacMer School of Medicine, Tel Aviv University, Tel Aviv 69978 The KCNQ1 p0re-forming subunit belongs to a newly characterized K channel family, KCNO, which forms voltage-gated K channels. Here we used Ba 2 ions to probe the permeation pathway of homomeric KCNQ1 channels. In addition to its voltage-dependent block of the pore, Ba z/ also alters the gating of_homomeric KCNQ channels by favoring the closed state. Ba produces a marked rightward shift of the voltage-dependence ofactivation (+33 mV) and an acceleration of the deactivation kinetics. However, Ba z/ also reduces channel inactivation as revealed by' the suppression of the hook of the tail current, an additional effect that opposes KC_NQ current inhibition. To further investigate the effect of Ba / on KCNQ inactivation, we used the L273F KCNQ mutant. This naturally occurring mutation, located in the trans_rnembrane segment $5 produces macroscopic inactivation. Ba potently inhibits the maximum conductance and the macroscooic inactivation of the_ L273F KCNQ mutant. In high external the impact of Ba z/ on channel gating is relieved. At 50 mM external K/, Ba / neither shifts the voltage-dependence of activation nor accelerates deactivation kinetics of both WT and L273F KCNQ1 channels. Similarly, Ba2+ loses its ability to orevent chanoel inactivation in 50 mM K*. Interestingly, pore block by Ba was poorly aff+ected by 50 mM external K+, suggesting that Ba like K ions, may interact with the channel pore at more than one site. Though the mechanisms underlying the gating effects of Ba 2/ are unknown, we suggest that they are coupledto the permeation process. Keywords: permeation, gating, K+ channels Characterization of the antiproliferative activity of phenothiazines in neuroblastoma and in glioma cell-lines. Relevance to brain cancer therapy (1-20mcM) of perphenazine revealed a dose dependent increase in the G1, and a decrease in the S phase, suggesting inhibition of DNA synthesis. Determination of the Caspase-3 activity in the neuroblastoma cells treated with thioridazine showed a marked dose dependent increase in the activity 4hr after drug exposure. Ghoma (C6) cells co-stained with propidium iodide and hoechst reagents showed the characteristic reddish apoptotic bodies in the nuclei of the phenothiazine treated cells. bnclusions: Phenothiazines possess a dramatic cytotoxic activity in brain derived tumors. The effect of phenothiazines is associated with DNA fragmentation, arrest of cell cycle, and caspase 3 activation. We suggest that some cytotoxic neuroleptics could play a role in the therapy of brain derived tumors.
Keywords: phenothiazines, brain tumors, apoptosis Lithium modulates neuron survival and glial growth in cultureinvolvment of polyamines Gilad G.M. and Gilad V.H.

Research and Development. Laboralory of Neuroscience, Assaf
Haro.[bh Medical ('enter, Zr/in 70300 Lithium, the most prevalent treatment for manic-depressive illness, may have neuroprotective effects after brain injury. In youn 8 primary cultures, lithium can exert neurotoxic effects associated with reduction in polyamine synthesis, but neuroprotective effects as cultured neurons ma(ttre. Evidence indicating that lithium can affect glial cells directly, leads us to postulate that lithium may exert some of its effects on neurons indirectly, via glial cells. Here we used rat cerebellar cultures to ascertain the effects of lithium (LiCI) on the activity of ornithine decarboxylase (ODC, the e.nzyme catalyzing the first limiting step in polyamine synthesis) and on neuron survival, and to compare the effects of lithium with those of the ODC inhibitor, c-difluoromethylornithine (DFMO). Additionally, we examined the effects of lithium and DFMO on cultured cerebellar glial cells (astroglia and micro,g, lia). Switching cultures from high (25 mM) to low (5 mM) K (KC1) medium concentrations served as the traumatic insult. The results indicate that: 1) While high, depolarizing K concentration enhances neuron survival, il inhibits astroglial growth. 2) LiCI (1 8 mM) enhances neuronal survival, but inhibits astroglial growth in cultures switched from high to low K*. 3) LiC1 treatment leads to reduced ODC activity. 4) DFMO enhances neuron survival, but inhibits astroglial rowth in cultures switched to low K medium. 5) LiC'] or IFMO treatment of confluent glial cultures lead to transformation of astroglia from epithelioid (flat) to stellate morphology and to increased numbers of microglia. 6) Combined LiC1 and DFMO treatment is highly c0toxtc to both neurons and glia in culture. The findings indicate that lithium can exert direct effects on astrocytes, as reflected by growth retardation and altered cell morphology of cultured astroglia, and suggest that these effects may be associated with inhibition ofpolyamine synthesis.
Moreover, the results imply that direct effects on astrocytes and microglia may contribute to the neuroprotective effects of lithium.
Keywords: astroc)etes, alpha-difluoromethylornithine, microglia, neuroprotection, omithine decarboxylase, rat Therefore, inhibition of LO may be conducive for regeneration and recovery of function. Previously, we found evidence that active extracellular LO molecules appear in a spatiotemporal manner in sites of CNS injury and that treating adult mice with the LO inhibitor I-aminopropionitrile (BAPN) can result in accelerated and more complete functional recovery after spinal cord injury. In the present study, adult rats received daily treatment of BAPN (100 mg/kg, intraperitoneally) for 20 days after complete unilateral spinal cord transection. This treatment led to an accelerated and more complete functional recovery as compared to controls. Additionally, in contrast to controls, which showed relatively minor functional deficit after a second transection, produced at 90 days at the site of the first spinal cord injury_, BAPN rats demonstrated functional deficits and a functional recovery time course similar to those observed after the first injury. The mode of functional recovery of BAPN-treated rats suggests that axonal regeneration of long descending tracts may have occurred. These and previous fmdings indicate that BAPN, by  Oxidative stress is believed to play a crucial role in the initiation and enhancement of the inflammation process that lead to demyelination in multiple sclerosis. We examined whether treatment with N-acetylcysteine-amid (AD4), a novel thiol-antioxidant that readily crosses the blood brain barrier, might provide a beneficial effect in the chronic, experinaental autoimmune encephalomyelitis (EAE) model induced_ by myelin oligodendrocyte glcoprotein (MOG). Mice (C3H.SW, n=15) were injected s.c. twice with pMOG 35-55 (3 mg/kg on day and 8) to induce EAE. AD4 treatment (0.2g/kgx2 .p. and 0.3g/kg in drinking water daily) started in three groups on days 1, 8 and at day 14 after the appearance of the symptoms. We found that while most of the control MOG-treated mice developed significant clinical manifestations with complete hind limb paralysis, the AD4-treated mice were resistant when treated from day and showed only very mild and delayed clinical signs when treatment started at day 8 or 14.
Histological examination of spinal cord sections showed, in the MOG-injected group (in the absence of AD4), multifocal areas of pervascular lymphohistocytic inflammation containing activated macr0phages associated with loss of melin. In contrast, in the AD4-treated animals, only mild focal inflammatory ctmnges were observed.
Similarly, immunostaining and silver staining of the neurons indicated a massive axonal damage in the EAE control group, and only a minimal damage was seen in the AD4-treated groups. In conclusion, our study demonstrates that in a chronic EAE model, administration of a BBB penetrating thiol-antio.,ddant, prior to or even after the appearance of symptoms, significantly educes the induced axonal damage, motor dysfunction and aimals, there were only mild fo?zal inflammation changes. In parallel, silver staining (Bielshowsky) of the neurons indicated massive axonal damage in the EAE control group, while there was only minimal damage in GA-treated mice. Antibodies against amyloid precursor protein used to measure axonal damage demonstrated lower injury response, as measured bv the Image-j software (65% p<0.001). In conclusion, our stud' demonstrates that in a chronic EAE model, GA immunization attenuates the induced axonal damage. Keywords The neurotrophic factors, NGF and BDNF are involved in neuronal survival, morphoenesis and modulation of synaptic strength in the adult bramo Both factors are differentially regulated and their levels vary between different brain regions. Activation of interleukin-1 (IL-1) was implicated in the regulation of NGF and BDNF. Using the IL-1 knock-out (KO) balb/c mice we have examined the possibility that basal levels of IL-1 may control NGF and BDNF protein expression. Brain tissue of IL-la-KO, IL-lb-KO, IL-lab-KO, IL-1Ra-KO and wild-type (WT) were analyzed. BDNF levels were higher and more variable than NGF in all examined brain areas; cerebral cortex, thalamus, cerebellum and hippocampus. No significant difference was observed in both NGF and BDNFlevels between WT mad KO mice in all examined brain regions.
Nevertheless, BDNF/NGF ratio was constant in each tested region in balb/c mice; 5.2+/-2 in the cerebral cortex, 4.5+/-3 in the cerebellum and 3.7+/-1 in the hippocampus. All IL-1-KO mice to the same background preserve the ratio observed in the comparable brain region of the WT mice. In contrast, no relation between BDNF and NGF levels was detected in JBC-57 WT mice. However, ILlb-KO on JBC-57 background exhibit in the cerebral cortex a constant ratio of BDNF/NGF similar to that observed in this area in the balb/c KO and WT, which is significantly different from its control. Our data suggest that the involvement of IL in BDNF/NGF regulation is distinct between mice strains. In the JBC-57 mice the basal levels oflL-Ib may be implicated in the regulation of NGF and BDNF. Moreover, the neurotoxicity of the peptide requires the expression of endogenous PrP, and the peptide induces hypertrophy and proliferation of astroc)tes and activation of microglial cells in-vitro. These data suggest that the region includin.g residues 106-126 might be an essential site in the conversion of PrPc to PrPsc.
Antibodies are known to act as chaperons and are able to stabilize protein structure and/or induce conformational changes. Thus, site directed antibodies might interfere with aggregation processes and/or inhibit prion replication. In order to nvestigate antibodies ab.ility to interfer.e wi aggre.gat!on processes we used active and passive lmmumzaton approaches. Polyclonal sera generated by administration of fflhmentous phage displaying peptide 106-126 as the antigen of choice, was shown to prevent 106-126 peptide aggregation as well as dissolve already formed fibrils and protect PC 12 cells from death induced by 106-126 fibrils. Two monoclonal antibodies generated usmg KLH as the protein carrier were shown to share the sera characteristics towards 106-126 peptide and where further analyzed for their ability to induce those changes with the whole protein. or a novel brain-selective MAO inhitor, TV-3326." The female offspring of rats stressed bv daily restraint during the last week of pregnancy were housed in "a 0700-1900 hr light Cle and gven water, amitriptvline (4.5 mg/kg/dav) or -3326 (17 mg/kg/day) from ae of 6 -13 weeks [n the drinking water to avoid the stress of injection. When aged 16 weeks, blood (501xl) was taken from the tail at 0800, 1200, 1500 and 1800 hrs for determination of circadian rhthm of plasma COR. In other rats, blood was taken before (at 0800-0900) and 30 & 90 min after exposure a novel environment. Plasma COR peaked earlier in PS (at 1500 hr) than in C rats (1800 hr). In both groups, novelt, stress increased plasma COR 2.5-fold at 30 mn. This leclined significantly by 90 min in C but not in PS rats. Neither antidepressant normalized the circadian rh3hm in PS rats, but selectively reduced the plasma COR at 90 min in PS rats. In conclusion, antidepressants appear to be able to correct the abnormality, in feedback regulation of the HPA axis in PS rats in keeping with their antiartxiety effect, but do not affect the control of its circadian rhythm by the suprachiasmatic nucleus. (IL-lrKO). When exposed to mild 88 psychological (4-minutes loud noise) or metabolic (7.5mg/mouse 2-deoxyglucose) stressors, IL-lrKO mice displayed signific.antly lower increase in corticosterone secretion, compared to wild-type (WT) controls. However, when exposed to stronger stressors (60-minutes restraint or 15mg/mouse 2-deoxygl-ucose), IL-lrKO and WT controls showed a similar increase in corticosterone secretion. These results suggest that IL-1 plays an important role in the regulation of the HPA axis response to mild psychological and metabolic stressors. However, stronger stressors may induce other mediators that can activate the HPA axis in the absence of IL-1. Brain IL-1 was shown, by us and others, to be increased following adrenalectomy (ADX), suggesting a role for IL-1 in the regulation of the HPA axis feedback. To examine this hypothesis, mice were adrenalectomised, and post operational serum ACTH levels were measured. Five days following AD.X, both IL-lrKO and WT mice demonstrated a mild increase n ACTH levels compared to their sham-operated controls. Eight days following ADX, ACTH levels in IL-lrKO remained mildly increased, while WT mice displayed an 80-fold increase in ACTH compared to sham-operated mice.
These results suggest that the increase in IL-1 levels following stress and adrenalectomy plays a critical role in HPA axis regulation under these conditions.
Keywords: interleukin-(IL-1), stress, corticosterone, ACTH, In this work we propose that a similar effect can be achieved by manipulating the post traumatic immune response with the use of bone marrow derived dendritic cellspulsed with myelin peptides or with altered myelin peptides (A91) that lack the risk of causing experimental autoimmune encephalitis (EAE). We have found in a model of contusive spinal-cord injury in rats that local administration of dendritic cells prepulsed with myelin basic protein (MBP) or with A91, directly nto the site of injury, significantly improves recovery as measured by functional outcome of hind limb movement and confirmed by morphological criteria. Dendritic cells injected into the site of injury as far as two weeks after the injury had a beneficial effect on recovery as well. Nonpulsed dendritic cells or those pulsed with a nonrelevant antigen had no effect on recovery.. Otherpossible routs of administration of dendritic cells to the injured animal were also examined. The fact that a positive effect of the dendritic treatment could not be achieved in injured thymectomized rats, which are deprived of mature T cells, suggests that .the mechanism whereby dendritic cells display their effect is T cell dependant, similar to that of vaccina.tion involving antigen specific activation of the immune system. Vaccination with dendritic cells may be an effective way of achieving beneficial immunomodulation following spinal cord injury.
Keywords: .CNS trauma, dendritic cells, autoimmunity, MAM animals ex]aibited similar baseline startle reflex as controls, but displayed non significant decreased PPI in classical conditions (prepulse-pulse interval 80 ms; PD 20 ms). When prepulse-pulse interval was set at 20 ms, the difference between treated and control rats appeared to depend on PD: MAM animals had significantly impaired PPI responses when PD was 160 but not 5 or 20 ms; there was a tendency for impairment in 80 and 1280 ms PD conditions. In both groups, the maximum inhibition was observed in the 20 ms PD condition. There was no overall treatment effect, but a significant interaction between treatment and parametric conditions, suggesting that prepulse temporal conditions have a specific influence in MAM rats. This narrow "temporal window" for PPI efficiency in MAM treated animals might be somewhat comparable to the PPI modulation seen with dopaminergic and serotoninergic agonists. These findings have to be discussed in the light of developmental and pharmacological theories of schizophrenia. gestation: detected at E7.5 and increased on E9.5. Expression was augmented in the brain (El2) and was sustained throughout embryogenesis. A second step measured ADNP subcellular localization and potential processing in glial cells.
Results showed intact ADNP-like immunoreactivity (-120 kD) in the rat astrocytic nucleus, the cytoplasm and the extracellular milieu and increased ADNP content following. VIP treatment. Lower molecular weight ADNP-immunoreactave bands were also observed. Subcloning of ADNP into a vector containing the Herpes VP22 protein, that is able to penetrate through cell membranes, and assessments of VP22-ADNP neuroprotective activities indicated protection of rat pheochromocytoma cells against oxidative stress. Finally, establishment of ADNP gene Current studies show intact ADNP-like immunoreactivity (-120 kD) in the rat astrocytic nucleus, the cytoplasm and the extracellular milieu and increased ADNP content following VIP treatment. Subcloning of ADNP into a vector containing the Herpes VP22 protein that is able to penetrate through cell membranes and assessments of VP22-ADNP activities indicated protection of rat pheochromocytoma cells against oxidative stress. Part of the ADNP neuroprotection may be attributed to decreases in the pro-apoptotic protein p53. Peptide scanning analysis identified an eight amino acid peptide (NAP) within the ADNP sequence that mimics the ADNP neuroprotection (Leker et al, Stroke. 33: 1085-1092[2002). To further assess ADNP's roles in the intact organism, ADNP knockout mice were established. Results showed that homozygous ADNP-knockouts suffer cranial neural tube closure fadure and death on E8.5-9.0.
Expression of OCT4, a gene associated with germline maintenance, was markedly augmented, while expression of PAX6, a gene crucial for cerebral cortex formation, was abolished in the embryos and the brain primordial tissue of the knockout embryos, respectively. Incubation of E8.  (GDNF) have been implicated in many forms ofplasticity within the brain, including drug addiction. Previous!y we described that activation of the dopamine-1 (D0 receptor n a human astroc_yte cell line, SVG, increases GDNF levels. Therefore, we examined the effect of cocaine on GDNF and D receptor expression as compared to amphetamine and morphine in SVG cells. Twenty-four hr incubation of SVG cells with cocaine significantly lowered D receptor and GDNF mRNA levels (detected using Reverse Transcriptase Polymerase Chain Reaction [RT-PCR]) and increased levels of signal transducers and activators of transcription (STAT) (via scanning and analysis of cDNA arrays). Neither amphetamine nor morphine altered these parameters. Furthermore, in vivo studies demonstrate that rats that have been trained to self-administer cocaine have decreased levels of striatal GDNF, confirming and extending the in vitro findings. In conclusion, cocaine in vivo and in vitro has a direct and specific effect on extraneuronal cells, in addition to its known effect on the neuronal dopamine transporter. Decrease in GDNF neurotrophic support may increase local neuronal vulnerability.
Keywords: cocaine, astrocytes, GDNF, D receptor, STATs Apoptosis and alternative cell deaths in the retinal tissue Guimarae C.A., Benchimol M., Amarante-Mendes G. and Liriden R. r'' 1Institute of Biophysicsm UFRdl, Rio de Janeiro, Brazil: Univesdade Santa Ursula, Rio de Janeiro, BraziI; lnst#ute of Immunology-USP, Rio de daneiro. Brazil Distinct types of cell death (CD) occur either during development and pathologies. Apoptosis and autophagy share some lallmarks as cell shrinka.e and membrane integrity maintenance. Our aim was to identify the types of CD induced by anisomicyn in the retina and their mechanisms of execution. Ultrastructural analysis of explants treated with anisomycin showed of both autophagic and apoptotic features. Bongkrekic acid, a mitochondrial pore inhibitor, completely inhibited CD induced by anisomycm in retinal e_xplants. Ac-LEHD-CHO, Z-DEVD-FMK and Ac-VEID-CHO, caspase-9,-3 and-6 inhibitors respectively, partially inhibited CD induced by ANI.
Caspase-9 and caspase-6 inhibitors had synergic effect upon CD induced by amsomycin. Co-incubation of retinal explants with 3MA and caspase-9 inhibitors did not have synergic effect upon CD induced by anisomycin and TUNEL, activated 89 caspases-9 and activated caspase-3 staining decreased in explants treated with 3MA. Furthermore, co-incubating of explants with caspase-3 and caspase-6 inhibitors did not have synergic effect upon CD induced by anisomycin.
These results suggest that anisomycin induces distract types of CD, all dependent on mitochondrial commitment and caspase activation. The first type of CD involves autophagy, caspase-9 and-3 activation and is TUNEL+, suggesting an overlap between apoptosis and autophagy. The other CD type involves caspase-6 and is TUNEL-. A third CD pathway s dependent on caspase-9 activation and occurs under caspases-3 and -6 inhibition.
Keywords: cell death, caspases, autophagy, retina Orienta,on selectiv,ity in V1 of the ale monkey Gur M."'., Kagan I. and Snodderly DM." tBibmedical Engineering, Technion, Haifa; :Schepens Eye Research Institute, Harvard Medical School Area V1 is unusual among all primate cortical areas in its neural cell density, cortical thickness, and intricate histology. Physiological recordings, however, emphasize overall integrating features such as ocular dominance and orientation selectivity. We have previously shown that in the alert monkey physiological properties of single cells reflect alternating anatomical layers structure. Here we study the relation of single cells orientation selectivity to the cells spontaneous activity and receptive field (R.F) size, and how these properties are related to the anatomical location of the cell. Single cells were recorded from area V of an alert monkey perfOrming a simple fixating task. The cells spatial organization was studied by drifting increment and decrement bars and by sinusoidal gratings. Orientation selectivity was defined by the orientation tuning curve bandwidth and circular variance. Orientation selectivity was clearly correlated with RF size and spontaneous activity. These 3 measures were strongly predicted by the layer of origin such that small RFs, low spontaneous activity, and a high degree of orientation selectivity were found in the output layers 2/3, 4B and 5 while the reverse was true for the input layers 4A, 4C and 6. We conclude that the conjunction of these ph,vsiological measures and their anatomical characterization rellect mteractions between excitatory and inhibitor), mechanisms. When excitation is strong, large RFs, high spontaneous activity and a low degree of orientation tuning are found while when significant inhibition is present, RFs shrink, spontaneous activity almost disappears and orientation selectivity is high. Keywords: visual cortex, orientation selectivity Assesment of vasomotor reactivity for prediction of syncope in patients with orthostafic hypotension. Gurewch T. Yu. Gur A. Yu, Korczyn A.D., Giladi N. and Bornstein N. Neurology Dept., Tel-Aviv Sourasky Medical Center. Background: Orthostatic hypotension (OH) is a common neurological symptom. It is not clear why somepatients can tolerate OH while others faint. Syncope s a result of severe cerebral hypoperfusion and cerebral autoregulation failure. Objective." To assess cerebral autoregulataon in patients with OH and history of syncope. Methods." 29 patients with OH, age 72.9+9.8 years, mean orthostatic decrease of systolic blood pressure of 33.7+12.9 mmHg after three minutes of standing, were assessed for VMR of middle cerebral arteries (MCA) and vertebral arteries (VA), using TCD (Rimed) after injection of lg acetazolamide i.v. 13 patients had experienced at least 2 documented syncope episodes over the preceding year. Patients with carotid stenosis greater than 75% were excluded from the study. The percent difference between blood flow velocities before and after acetolamide injection was defined as VMR% and the results were compared by Mann 1Dept. of Human Genetics andMolecular Medicine, Sackler School Of Medicine, Tel Aviv University, Tel Aviv, Israel; 2Faculty _of Life Sciences, Bar-Ilan University, Ramat-Gan, Israel;-Genetica Medico, Dip. Patologia Generale, II Universite di Napoli and Tigem, Italy The past few years have provided an explosion in our understanding ofhow the inner ear functions. This dramatic increase is due in large part to the genes found to be associated with nonsyndromic hearing loss. Since 1997, 29 genes have been found, and these have provided clues about auditory transduction, ion homeostasis, and inner ear development. In particular, mouse models for human deafness, with mutations m orthologous genes, have revealed essential information about the pathophysiology caused by these mutations. One group of proteins frequently associated with hearing loss are the myosins, three of which were discovered thanks to their corresponding mouse mutants. Myosins are molecular motors that move along actin filaments and have been implicated in various cellular functions such as cell movement, membrane traffic, and signal transduction. An intraenic deletion in myosin VI (Myo6) leads to deafness and vestabular dysfunction in Snell's waltzer mice. A missense mutation in an Italian family is associated with autosomal dominant progressive hearing loss (C442Y) (Melchionda et al, AJHG 69:635-640 [200 I]). The following suggests that the deafness in this family is due to the MY06 missense mutation: the segregation of this mutation with the affected individuals in the family, the previous association of myosin VI with deafness, and the conservation of the mutated residue. We have now confnTned that this mutation is the cause of deafness in humans, since reproduction of this mutation in transgenic mice has revealed a similar progressive hearing loss. A morphological analysis of the tr.ansgenic mice demonstrates the pathophysiology of this mutation.
K.eywords: sensory system, hearing loss, myosin, transgenic mlce Global-local processing among elderly with and without insomnia: a comparison with young adults Hadad B. Shurkin D. and Haimov I.
Dept. of Behavioral Science, Emek Yezreel Academic College Disturbed sleep, an important cause of human miserv at any_ stage of life, exacerbates with advancing age. Complaints cf difficulty in initiating and maintaining sleep and-daytime drowsiness are more prevalent among the elderly than in any other group. Likewise, aging is associated with a decline in many aspects of visual processing including changes in global-local processing. However, little is known about the interacti6n between sleep disturbances and visual processing of global and local aspects among elderly people. Thus, we examined whether insomnia is indeed associated with age-related changes in this global-local processing. Results showed that the global and the local structures were equally manifested in young adults, along with a tendency toward a global advantage. Elderly subjects without insomnia, on the other hand, showed a reversed tendency of faster response time for the local aspects, indicating the processing dominance of the local level of visual stimuli n elderly. Furthermore, elderly subjects with insomnia showed a significant advantage for the local structure and an asymmetric local-to-global interference, suggesting that the ability to integrate individual elements into a coherent pattern deteriorates with age, and becomes substantially Impaired among elderly insomniacs. Overall, the fmdings imply that, at least in some aspects, sleep disturbances may account for perceptual and cognitive decline in the elderly. In contrast to other venomous animals that paralyze their prey, the venomous parasitoid wasp Ampulex Compressa subdues its prey by the induction of a bout of grooming followed by a five-week long lethargic state. During the lethargic state the cockroach is docile enough for the wasp to pull it into a burrow where the wasp larva eats it alive. The cockroach is always stung by the wasp twice, first in the thorax and then in the head.
To localize the site of stinging we injected wasps with a mixture of C 14 radiolabeled amino acids. After the amino acids were incorporated into the venom, we allowed the wasps to freely sting several cockroaches.
First, the amount of radioactivity in the ganglia of stung cockroaches was assessed using liquid scintillation of the different ganglia and tissue. Signific_antly higher levels of radioactivity were detected in the head ganglia and the first thoracic ganglion. In contrast, radioactivity levels in the third thoracic ganglion and non-neuronal tissue were much lower and comparable to control values. Second, microscopic emulsion autoradiography was carried out to determine the precise location of injection. Radioactivity was detected in the central area of the brain ganglion, the subesophageal ganglion and the first thoracic ganglion. No radioactivity cofild be detected in the second thoracic ganglion or in ganglia stung by non-radiolabeled wasps.
To our knowledge, this is the first direct evidence of a yenomous animal stinging into the central nervous system of its prey. Keywords Object images activate a large and complex array of high order visual areas in the human cortex, ranging from dorsal occipito-parietal cortex to ventral occipito-temporal cortex. To search for consistent patterns within this array we performed a detailed analysis of fMRI activation in 12 experienced subjects along four basic dimensions (object selectivity, eccentricity, motion selectivity and visual meridians). Our results reveal seven consistently distinct category-related entities situated in the cortex adjoining early visual areas. These include: two face-related regions, three object-related regions and two building-related regions. Interestingly, we found that the complex pattern is organized in a dorso-ventral, large-scale object-based mirror symmetry. Furthermore, correlating this pattern to the map of visual field eccentricity, we find that the entire network of object areas can be related to a single eccentricity map. We hypothesize that this large-scale organization points to a possible development of high order object areas through extension and specialization of a single proto-representataon. Funded by Zrael Academy 8009 and 644/99 grants. Keywords: visual cortex, fMRI, object recognition, topography Lack of paternal care reduces spine density in the limbic cortex Helmeke C., Ovtscharoffjun. W. and Braun K.
Dept. Zoology/Developmental Neurobiology, Otto von Cntericke University, 39008 Magdeburg, Germany. Emai katharina.b raun@nat,uni-magdeburg. de Previous studies in trumpet-tailed rat Octodon degus have demonstrated that, quite analogous to the functional maturation of sensory and motor cortices, which strongly depend on sensory or motor stimulation, the development of synaptic connections in limbic brain regions are modulated by early postnatal emotional experiences. Periodic and chronic interruptions of parental care during the first weeks of life induce a significant increase of dendritic spines in the limbic dorsal anterior cingulate cortex (ACd) (Helmeke C, Ovtscharoff jr et al, 2001, Cerebral Cortex, 11: 717). In the present study the contribution of paternal care on the development of synaptic composition in the ACd was analyzed light and electron microscopically. The comparison of pups, which were either raised with or without their father revealed o in the fatherless animals a 20, decreased spine density on apical and basal dendrites of layer II/III pyramidal neurons in the ACd. Such experience-induced modulations of synaptic inputs in higher associative and limbic cortical areas appear to shape the limbic synaptic networks and thereby may determine cognitive and psychosocial capacities during early and later stages of life. Supported by SFB 426. Keywords: limbic system, development, synoptic plasticity, dendritic spines Assessment of tissue functionality in vicinity of brain lesions using DTI and fMRI Hendler T.1, Pianka P., Sigal M. Ben-Bashat D. , Fried I.2, Graif M. land Assaf Y. 1,3 groh lnsatute for Advanced Imaging, Tel Aviv Sourasky Medical Center, Tel-Aviv; 2Functional Neurosumerv Unit, Tel Aviv Sourasky Medical Center, Tel-Aviv; 3Schoo'o/Chemistry, Tel Aviv University, Tel-Aviv 69978 In the last decade functional MRI (fMRI) gradually evolved to become a clinical tool for pre-surgical evaluation of critical functionality in vicinity of brain lesions, fMRI signals relate to activated gray matter only. However the connectivity of white matter is crucial for functionality of a region and should not be damaged during surgery. Diffusion tensor imaging (DTI) is a well-established method for in-vivo mapping of the white matter directionality and organization, but its clinical application has not been fully explored. Our aim in this study was to explore the added value of the combined methodologies in brain surgery. DTI is based on the anisotrppic motion of water molecules in white matter. While the difftision of water parallel to long the long axis of the neuronal fibers is free, perpendicular to the fibers it is disturbed. Recently, the concept of three-dimensional fiber tracking based on DTI data was introduced. With this methodology it ispossible to produces fiber tracts that corresponds with large fiber bundles such as the pyramidal tract, corpus callosum, optic radiation, corona radiata and thalamic radiation. In relation to brain tumors, one can think of three ways brain lesion and tumors can affect white matter fibers: cutting, pressing or infiltrating. Indeed, three-dimensional fiber tracking helped to visualize critical white matter bundles and their relation to the tumor. White matter bundles that were cut, pressed or infiltrated by the lesion could be identified and relation of these bundles to task related aY matter activation could be visualized. Axotomy induces profound changes in primary sensory neurons and often produces neuropathic pain states, but very little is known about its effects on satellite glial cells (SCs) ih dorsal root ganglia (DRG). We have shown previously that dye coupling among SCs was increased after axotomy. In this research we studied the ultrastructural basis of this change by correlating intracellular dye injections and serial section electron microscopy (EM). Lucifer yellow injections showed that in control ganglia 76.2% injected SCs were not coupled to other SCs, and 20.7% were coupled within a glial envelope around a given neuron, and very., few (3.1%) were coupled between glial envelopes around dfferent neurons. However, in axotomized ganglia the incidence of coupling between glial envelopes increased by 7-fold (p<0.0001) and those of coupling within an envelope increased to 30.5% (p<0.001).
Some g!ial processes from glial sheath extended into adjacent connectwe tssue. Neurons were not coupled to other ce|ls in control or axotomized ganglia. focused on one of its receptors, the D3 dopamine receptor (D3R), which is expressed on T cells. D3R is known to be involved in neuropsychiatric disorders and serves as a target for therapeutic treatment. We demonstrate that D3R on T cells is functionally active, and its activation affects immunological properties of the cells. Interestingly, the immunological changes observed, are confined to only a subclass of highly activated T cells, blasts. Unlike non-activated T cells, blasts can cross the BBB and bind NTs due to high expression of adhesion molecules. In addition, they also express higher levels of D3R than T cells. Dopaminergic hctivation of blasts changes their cytokine secretion profile and the expression of several surface markers. Since these effects do not occur in non-activated T cells it seems that the underlyingmechanism is meant to activate only those cells that can cross the BBB. Finally, we show that peripheral T lymphocytes of schizophrenic patients behave like blasts in several features.
They exhibit higher levels of both D3R mRNA and protein.
These findings suggest that a central disorder like schizophrenia leads to changes in peripheral immune cells, demonstrating a control pathway of the brain on immune cells. We are developing a new approach for blocking beta secretase activity, based on immunization with a small p.eptide representing the cleavage site of beta secretase. Antibodies raised recognize the cleavage site of APP and will hopefully interfere with AP formation. Limiting of beta amyloid production by this approach may become an important therapeutic target in Alzheimer disease (AD of PDL-I (3.4%, 9.3% and 12.2% for SP, RR and HC resp., 19<0.05). DCs incubated with serum from SP-MS (n=15) had higher IL-12 (3.53%) compared to RR-MS (1.27%, p=0.009, n=28) or HC (0.78%, p=0.007, n=17). Post-MLR T-cell stimulation showed T-cells polarized by SP-MS DCs secreted more TNF-a (225.9 pg/ml) vs. those polarized by HC DCs (22.7 pg/ml, p=0.049). The secreted TNF-x/IL-4 ratio from T-cells polarized by SP-MS DCs was higher than by HC DCs vs. 1.45, p=0.038)_.
nclusions." Myeloid l)tgs in SP-MS are activated compared to RR-MS and HC. They polarize naive T-cells into proinflammatory T-cells. Serum contents in SP-MS patients may play a role in DC activation, manifested by its effect on DC IL-12 production. These results demonstrate that the activated DCs may determine the type of immune activity in SP-MS. Keywords: dendritic cells, multiple sclerosis, co-stimulation, cytokines, T cells polarization The expression of mitochondrial complex I subunits, 24-and 51-kDa, is reduced in the frontal and elevated in the arietal cortices of schizophrenic patients arty R,, Klein E. and Ben Shachar D.
Lab.-Psychobiology, Dept. of Psychiatry, Rambam Medical center, B. Rappaportfaculty of MedWine, Technion Impairment of the mitochondria, which play a major role in cell function and survival, could provide an explanation for the tremendous heterogeneity of clinical and pathological manifestations in schizophrenia. Previous studies have shown abnormalities in brain mltochondrial morphology as well as in the respiratory chain enzymatic activity in schizophrenia. We have recently reported a disease stage dependent alteration in complex activity in platelets of schizophrenic patients, which was accompanied by alterations in the expression of the 24kDa and 5 lkDa, but not in the 75"ld:)a subunits of complex I. In the present study we compared the mRNA and protein levels of the three subunits in frontal and parietal post-mortem brain cortices of 14 schizophrenic patients, 15 unipolar patients (UP), 15 bipolar patient (BP) and 15 normal subjects, obtained from the Stanley Foundation Brain Bank. Both mRNA and protein levels of 24-and 51-kDa subunits were significantly reduced in the schizophrenic frontal cortex compared to the controls, while UP patients showed a reduction m 24kDa protein level, which was less significant than that of the schizophrenic group.
No such changes were observed for the BP. In contrast, a significantly increase was observed in mRNA and protein levels of the 24kDa subunit in the parietal cortex of the schizophrenic patients. Protein level of both subunits was also increased in the BP patients. The 75kDa subunit did not differ between groups. The present stu.dy further demonstrates the malfunctioning of complex in schizophrenia and supports the relevance of our peripheral findings to the CNS pathol6gy.
Keywords: schizophrenia, mitochondrial complex I, Yet, newborns previously exposed to hypoxia delayed in developing the ability to climb on inclining slope and hold themselves to rotating rod. On P8-P9,hypoxia exposed newborn were ble Jo climb on 10"-20 (mean) slope as compared to 40"-50 in the control group (p<0.0029,0.013, n=6-11, 9-10, respectively). Significant difference was also observed between the groups in the rotarod holding at P 10, P 11 (P00.006, 0.04, n=6-10,6-4 Psychophysical discriminations can be viewed as a comparison between two internal responses, each characterized by its mean value and noise amplitude. Performance, measured as percent correct discrimination, is effectively determined by signal to noise ratio. To separate these two response components, available methods impo_se additional constraints on the decision process or on the stimulus-response transformation.
Here we propose a novel approach that only assumes normally distributed one-dimensional internal responses. It is based on minimizing the residual error between measured and modeled psychophysical response levels. This procedure requires a number of stimuli pairs m order to derive an over-complete system of equations for mean and noise response amplitudes.
The method was ,applied to a contrast discrimination task.
Gabor patches xith three contrast levels in txo spatial configurations with flankers and without themwere used as antiphospholipid antibodies by immunization with 132-glycoprotein I. This immunization has previously been found to induce behavioral and cognitive dysfunction in a number of normal mouse strains, but not in C57BL, the background for the transgenic mice used in the present study. The mice were immunized at the age of 4-5 months and 4.5 months later were tested for hyperactivity and anxiety on a staircase apparatus and for cognitive function in a swim T-maze. There were significant differences in behavior between the APP transgemc mice compared to the controls in both tests. Induction of antiphospholipid antibodies impaired performance in the co_gitive test only in the APP transgenic mice and had no significant effect in the wild pe controls.
The immunization had no significant effect in the behavioral assay. These results indicate the potential role of immune mediated mechanisms in the pathogenesis of neurodegenerative processes and point to the potential use of immunomodulatory iherapies in such diseases. Serotonin2c receptors (5HT2cR) have been implicated in some psychiatric disorders, including depression. 5HT2cR undergoes editing whereby one to four specific adenosines are converted to inosines. Editing modulates receptor-G protein interactions, thereby altering receptor-mediated signaling. We tested 5HT2cR editing m the nucleus accumbens (N. Acc) of Flinder sensitive line (FSL, an animal model of depression) and control rats using the sequencing method. FSL rats exhibit 50% higher levels of the fully editedisoform (VSV) of 5HTzc than control rats. Only control rats exhibited the non-edited isoform (INI). Treatment with desipramine had no effect on RNA editing, although it improved swim behavior in FSL rats. Western blot analyss detected a decrease in protein levels of 5HT2cR in FSL rats after despiramine treatment although no significant changes were detected in mRNA and pre-mRNA levels of 5HT2cR in FSL rats or desipramine treated rats compared to controls. Acute exposure of the accumbal 5HT.cR to its antagonist, RS 102221, resulted in dopamine release and this was doubled in FSL as compared to control rats. Therefore, we suggest that the 5HT2cR in the N.Acc has an inhibitory-like role in tonic DA release. Furthermore, 5HTc receptor RNA editing and pharmacology are altered in association with depressive behavior. These data indicate that 5HT2c receptor could be a potential target for future treatment of depressive behavior.
Keywords: RNA editing, serotonin-2C receptor, microdialysis Autoinununity is the body's own mechanism against self-destructive compounds emerging as a consequence ofCNS insult. In this work we show that induction of tolerance to self myelin-specific antigens, what was believed for years to be a preferential state of autoreactive cells, is against 'individual's fitness' in coping with sta-essful conditions after CNS injury.
Neonatal immunization of rats with whole spinal cord homogenate, diminished the ability of adult rats to respond to myelin immunization and those animals show worse neuronal survival after optic nerve crush injury or spinal cord injury. As a corollary, immunization of adult animals with myelin-derived self-antigen or delgletion of endogenous SUlapressor T cells (e.g. CD4+CD25 reguqatory T cells) increased" fhe animals' ability to resist injurious conditions. These findings call for the redefinition of tolerance to self-antigens, and for the role of CNS In our study of adolescents with congenital adrenal hyperplasia (CAH) we find correlations between possible dehydrational events pre-and post-natal and the liking for salt taste expressed by adolescents. These events include scores of maternal vomiting and nausea when pregnant with the adolescent, and scores of the adolescent's levels of vomiting, diarrhea and infusions during infancy. We find positive mad significant relationships between these events and how much the adolescent likes salt, years later. Since these data are based on recall by the mother of events occurring years previously, we attemptl to control for the unreliabilit of the recall data by making comparisons within the family, i.e. comparing the recall data for siblings and its relataonship to adolescent salt-liking. The relationship between perinatal dehydrational events and adolescent Finally, we f'md a strong relationship between maternal nausea and vomiting and the severest from of CAH. However, here the causal relationship seems to be reversed-apparently carrying a fetus with CAH induces higher levels of nausea and vomiting in the mother. The covalent lipid modification of proteins plays a major role in targeting heterotrimeric (oq3y) G proteihs to cellular membranes. In the case of the Drosophila visual system, palmitoylation ofthe cysteine residues atposition 3 and 4 of an eye specific DGq-alpha is the sole lipid modification of the alpha subunit. Shannon-eickert C. , Kleinman JE. and Again G.
Stanlev Research Center, Ben-Gurion University of the Negev, Beer S[eva; 2Clinical Brain Disorders Branch, NIMH, Bethesda MD, USA Glycogen synthase kinase-3 (GSK-3) is a protein kinase highly abundant in brain and involved in signal transduction cascades, articularly neurodevelopment. Its activity and protein levels ave recently been found over 40% lower in postmortem frontal cortex of schizophrenic patients. To elucidate whether the low levels are a genetic marker or a consequence of a perinatal neurodevelopmental insult the following experiments were carried out: GSK-313 protein levels in the frontal cortex of rats treated with neuroleptics or exposed to cold restraint stress were assayed. In the schizophrenia-related neonatal ventral hippocampal lesion rat model we measured GSK-313 protein levels and activi_ty in the frontal cortex. To confirm our original finding in another brain area we studied mRNA levels of GSK-313 in postmortem dorsolateral prefrontal cortex (DLPFC) from schizophrenic patients. Chrome treatments of rats with neuroleptics or exposure to cold restraint stress did not alter GSK-313 protein levels, supporting the concept that low GSK-313 in schizophrenia is not secondary to drug treatment or stress. However, GSK-31 protein levels in lesioned rats were significantly, lower than in sham rats, favoring the perinatal insult possibility. GSK-313 mRNA levels were 36% Iower in postmortem DLPFC of schizophrenic patients, supporting our previous findings. An additional intriguing recent preliminary finding is that GSK-313 protein levels in CSF samples from schizophrenic patients were found 28% lower than n control subjects. Further studies will be aimed at determining whether nonspecific neonatal damage or only specific factors cause low GSK-3 as a late effect.

Keywords: schizophrenia, GSK-313, neurodevelopment
The cellular mechanisms underlying learning and stress Kogan I. and Richter-Levin G.

Det. of Psychology, University ofHaifa
Stress has multiple modes of influence o/a learning and memory, improving or impairing it under different conditions. The hippocampus is held accountable for explicit memory. The CA1 area of the hippocampus is reported to be invoh,ed in spatial memory. The amygdala is critically involved in m,diating stress-related effects on behavior.
Memory storage has a late phase that requires protein synthesis. The activation of the MAPK (ERK) cascade is related to the establishment of the late, protein-synthesis dependent phase of memory formation. Thus, the activation of ERK and of its down-stream substrates may serve as a biochemical indicator for the activation of long-term memory ost learning protocols involve also a component of stress. In the present study we attempt to separate between learning and stress induced haechanisms by examining the activation of members of MAPK cascade (ERK1/2, CREB and Elk-1) in the hippocampus and amygdala. Three groups of rats were tested: I. Learning subjected to a spatial learning protocol. 2.
Stressedsubjected to the water maze as the Learning group, but without an escape platform. 3. Naive. Ten minutes after the last training session, tissue was collected for analysis of ERK, CREB and Elkactivation. Only in the Learning group an activation of ERK was found in the CA1 area of the hippocampus but not in the amygdala. Other molecules are still under the examination in both brain areas. The findings are expected to contribute to the dissociation between learning-and stress-induced mechanisms of signal transduction. We have recently found that maternal immune activation during pregnancy by means of systemic administration of the synthetic cytokine rleaser poly I:C in rats, led in the offspring to neurochemical and histological brain aberrations, as well as long term behavioral deficits (Zuckerman & Weiner, submitted). Moreover, these deficits were not present before puberty but emerged at adulthood, implicating abnormal brain maturation processes. The present stud-y used this model for the analysis ofmultiple gene expression profiles in two brain regions, the striatum and hippocampus, from the offspring of poly I:C treated and control dams. To this end we employed Clontech eDNA macroarray technology containing 1200 rat genes relevant to neuroresearch and housekeeping genes. RNA preparations were pooled from four matched brain sections obtained from adult female offspring. RNA hybridization procedure was repeated in two independent trials. Our data indicated that genes associated with synaptic trafficking and neuronal growth, synapsins (1A, 13, 2A), syntaxin (B) and myelin basic proteins, were up-regulated in both striatum and hippocampus of the poly I:Coffspring. In addition, several genes of the signal transduction pathway, G-coupled proteins and kinases, were elevated. Few other genes of the excitatory glutamate-NMDA pathway were up-regulated specifically in the poly I:C offspring striatum, while the poly I:C offspring hippocampus was distinct by elevated neuro-endocrine proteins secretogranin and somatostatin. Although these findings need further validation by other molecular methodologies, they may suggest that prenatal administration of poly I:C interferes with neurodevelopmental processes involving neuronal remodeling, activity and proliferation, that might be associated with brain disorders such as schizophrenia. The mechanisms underlying short term facilitation in Aplysia sensory-motor synapse can be subdivided into two processes: (a) a spike duration dependent process (sensitization) that results from 5HT induced PKA activation, and (b) a spike duration independent process (dishabituation) that results from PKC activation. As a result of PKA activation the potassium conductance is reduced, leading to spike broadening and enhanced calcium influx. The cellular mechanisms underlying PKC dependent synaptic dishabituation are not entarely understood. It was suggested that vesicles mobilization, alterations in the release mechanism or local activation of specialized calcium channels might be involved. In contrast to earlier reports, we found that calpains (calcium activated cystein neutral proteinases), are involved in the cascade of events leading to synaptic habituation and dishabituation.

Supported by lhe lsrael Science Foundation
Application of the membrane permeable calpain inhibitors calaeptin or the nonspecific proteasome inhibitor MG132 increases the rate of s_ynaptic habituation and inhibits 5HT-induced synaptic dishabituation. On the other hand, sensitization is not affected by the inhibitors. The results are consistent with the hlgothesis that inhibition of constitutive proteolytic activity_ slow down or prevent the translocation of synaptic vesicles to the release sites. As a consequence the rate of synaptic habituation is accelerated in thepresence of the inhibltors, and after massive habituation, 5HTapplication does not lead to dishabituation. These results demonstrate that constitutive proteolytic activity is necessary for the induction of short-term neuronal plasticity. products that are altered during thermal conditioning by using molecular techniques. After thermal conditioning the AH/PO of young chicks was dissected in a time course ranging from minutes to 24 hours, and mRNA changes were "monitored. We applied two strategies: 1. General mRNA amplification changes screening using differential display technique 2. Since conditioning cause change in hypothalamic plasticity, we checked for changes in genes that are expected to be involved in growth and development concentratingon neurotrophic factors. We identified 15genes that are altered in the hypothalamus during heat conditioning. Among the induced-genes we decided to further study the induction of M-RAS, 14-3-3, N-CAM and BDNF, which are all related to growth and cellular interaction. The mRNA of M-RAS is maximally induced after 12 hours, and that of 14-3-3, N-CAM and BDNF after 6 hours. We are further analyzing the amount and exact location of the induction of these mRNAs. Keywords: thermotolerance, chicks, differential display Major depression has been shown to involve alterations in noradrenergic transmission. Recent research focuses on intracellular processes leading to synaptic plasticity as involved in the pathophysiology and treatment of depression. We have suggested a role for NE as a possible mediator of these processes. Previously, we reportedNE-indueed differentiation m SH-SY5Y cells accompanied by an increase in the cell adhesion molecule CAM-L1, in laminin and in CREB, all involved in processes of svnaptic plastici.ty and neurite outgrowth. These genes were'also increased in hippocampus and frontal cortices of rats treated with antidepressants, while decreased in a rat model of depression. In the present study we compared the expression of CAM-L1, lamlnin, CREB and p-CREB in post mortem brains of depressed patients (n=15), bipolar patients (n=15), sehizophreme patients (n=14), and normal controls (n=15) obtained from the Stanley Foundation Brain Bank. We found a two-fold increase in mRNA and protein levels of CAM-L in the frontal cortex of the depressed group compared to controls, which was not observed in either bipolar patients or schizophrenic patients. Likewise, CREB expression was increased 2.5 fold in depressed and 1.5 fold in bipolar patients, while pCREB was increased only in depressed patients, by 4 folds. No alterations were observed in laminin. No effect was found for post-mortem interval, age, age of onset, gender or antipsychotic medication. The present results support the hypothesis that noradrenergic dysregulation in depression, lead to abnormal expression of genes volved in synaptie plasticity.
Keywords: depression, CAM-L1, CREB, post-mortem Direction-specific effects of adaptation in motion-selective neurons of cat primary visual cortex ampl 1.1; Priebe N.J. and Ferster D.
Det. of Neurobiology, Weizmann Institute of Science, Rehovot 76100; :Dept. of Neurobiology and Physiology, Northwestern Universitfi, Evanston, IL, USA Adaptation, cause by prolonged visual stimulation, reduces the subsequent visual responses of neurons in primary visual cortex, as measured in both spiking and mean membrane potential. In previous works it was shown that adaptation reduced membrane potential hyperpolarization of V neurons (Carandini et al. 1997). To distinguish between intrinsic mechanisms to network mechanism of adaptation we have studied the specificity of adaptation in visual responses of the neurons. If the mechanisms underlying adaptation are intrinsic to the recorded cell, adaptation should not change the selectivity of the neurons, but scale or reduce its response equally to all stimuli. We examine this hqgothesis on direction selectivity in simple and complex cells. In simple cells we have found that adaptation, induced by drifting gratings, in either the preferred or the non-preferred direction cause a tonic hyperpolarization similar to what was previously described. We did not measure, however, significant effect on the modulation of the response. In Complex cells, on the other hand, we found a direction-specific effect of adaptation: Adaptation in the preferred direction caused a pronounced reduction in the direction selectivity assayed from the mean potential of the response. In contrast, adaptation in the non-preferred direction led to an increase in the direction selectivity of the neuron. The selectivity index of the 96 non-adapted inputs, calculated from the responses to either direction after adaptation with the same direction of motion, was no different than the control index, suggests that cortical adapting inputs do not change the selectivity of the cells. Furthermore, unless a significant fraction of the cortical inputs to direction selective cells are not affected by adaptation, intracortical activity does not make a significant contribution to direction selectivity. The Mismatch negativity (MMN) component and the "F-(fusion) comolex y', theres'lgonse to net-fusion, are brain potentials evoked by a deviant event. In this study we attempted to find out the extent to which these responses reflect the same process, albeit in different contexts.
The stimuli were base (presented to the front), that fused with the formant transitions presented to the front, left or right of the subject (the latter two producing an echo-sensation), resulting in a standard and deviant V-C-V sequences /aga/and/ada/, respectively. Brain potentials were recorded from 10 normal hearing, right handed native Hebrew speakers, whose ages ranged between 19-30 years, that discriminated these V-C-Vs. Low resolution electromagnetic tomography (LORETA) t-test comparison images, that were run on the responses to deviants, showed that in case of front-fusion (no echo-sensation) activity contributing to the MMN was larger between 19 I-218 msec and was localized to the temporal cortices (BA 21,22). In this case, the phonetic identity of the auditory obiect affected the MMN. In contrast, an enhanced "F-complex' was associated with the lateralized fusion conditions that sounded different in spatial attributes (echo). This activity was localized to parieto-temporal (BA 19, 9), occipital (BA 31) and frontal (BA 9, 10) regions in the latency range of 2261296 msec.
These results indicate that the MMN and 'F-complex tap different aspects of deviance: the "F-complex" is affected by properties of the auditory object itself, whereas the MMN is affected by its novelty in relation to other stimuli. Keyw)rds: speech, auditory evoked potentials (AEPs), fusion, MMN. "F (fusion)-complex" antagonist, and has anti-oxidant and anti-inflammatory activities. It was shown to be ne_uroprotective in brain isehemia and traumatic brain injury models. PRS-211,220 is a novel analog of Dexanabinol, with two times higher affinity for the NMDA receptor and 30 times better ability_ to inhibit cyclooxygenase-2 than Dexanabinol. The aim of the present study was to determine whether PRS-211,220 had long-term beneficial effects on functional outcome and on infarct volume following focal brain ischemia. The middle cerebral artery (MCA) of adult Sprague Dawley rats was occluded for 120 minutes by intraluminal suture under halothane anesthesia.
PRS-211 220 (0.1, 0.25, 0.5, 1 2.5, 5 or 10 mg./k IV) and its vehicle were administered by the end of the ishemia. The neuroprotective efficacy of'the compound was evaluated behaviorally by the "staircase test" and morphologically by measuring nfarct volume. Rats were trained for week prior to the insult, twice a day for 15 minutes. Thereafter, rats were tested for 2-3 weeks. By the end of the last test brains were removed, serially sectioned and stained with thionin. Infarct volumes were evaluated using a computerized image analyzer. PRS-211,220 induced a dose related improvement in staircase test performance on the contralateral side. (30-55% compared with vehicle alone, 55% improvement at 0.5 mg/kg p<0.05,). Infarct volume was also reduced with PRS-211,220 (44% at 0.5 mg/kg Recently we have found that human high-order object areas are driven partially by local object fragments, and partially by global completion effects. Here we studied to what extent the local and global effects might emerge at different rates. We studied the temporal dynamics of these effects using brief exposures through a backward masking paradigm. In the first experiment, line drawings of animal shapes _,w,..ere shown that were occluded by parallel, vertical bands ("grid '), or scrambled by randomi,z, ing he relative location of the object stripes ("scrambled), each presented in 2 different exposure durations (60 ms, 250 ms). In the second experiment, the,same objects were presented either without occlusions ("whole ') or with the "grid' condition, again using 60 and 250 ms exposures. The results showed that for both exposure durations, i,n, occipito-temporal (LO and pFs) cortex, the signal in the "grid' condition was higher than the "scrambled" in the first experiment, and the "whole" condition exceeded the "grid" in the second one. However, the ratio between the preferred and non-preferred conditions in each experiment was similar at 60 ms and 250 ms. The activation pattern matched very well the recognition performance of the subjects. In conclusion, our results show that the emergence of global completion effects is as rapid as the emergence of local feature representation. Thus, as early as 60 ms when the visual activation is still low, the relative contribution of completion effects is already similar to that found for longer exposure times. Surprisingly, the upper calcarine sulcus that normally represents the lower wsual field (that the subject cannot see) was activated by the visual stimulus. Thus, the pattern of activation reported here does not conform to the classical patt .ern of neurpnal plasticity found in monk.eys.
The activation in the cortical regions representing the blind field may be interpreted as (1) expectation of the stimulus in the blind field due to its cyclic appearance (2)  Center, Tel Aviv 64239; 5Tel Aviv University, felAviv 69978 The human ventral occipito-temporal cortex exhibits complex activation patterns to different objects. Recent studies revealed some of the principles which may underlie these representations, e.g. resolution needs and expertise level.
However, the representation of objects along these dimensions may be either modular (i.e. each stimulus type activates a distinct region) or continuous (i.e. activations to different stimulus .t .y.s slide smoothly along a certain dimension). The two possibdities lead to different predictions: in the modular case different objects should either activate the same area or different areas, whereas in the continuous case activations to different objects will be partly overlapping and partly offset compared t6 each other. To test these predictions we conducted a functional MRI experiment, in which subjects (9) were presented with head images in front and back views. Images were presented in 9-see single-category epochs, interleaved with 6-sec periods of blank screen. We obtained activations of highly overlapping et slightly offset regions in the fusiform gyrus. Crucially, te direction of this displacement was consistent across subjects and therefore cannot be attributed to random variability; l'n addition, the non-overlapping strips of activation were not the regions of highest actavation to each category, excluding low-resolution, or partial volume effects as the cause of the overlap. We conclude that the activation pattern reflects a continuous topography, in which representations of objects "slide" smoothly along the fusiform uppSrtedbvlSF8009. Results: A significant association of low socio-economic status and IS incidence was found. On univariate analysis a significant positive association was found with spouse low educational level (p=0.05), spouse employment in blue-collar occupation (p=0.047) and with low income (p=0.0017). The IS patients were less property owners (p=0.000) and lived in crowded conditions (p=0.027), without additional medical insurance (p=0.025) and without emotional support (p=0.037) and without friends (p=0.000). On multivariate analysis, systolic hypertension heart disease, diabetes, family history of ertenson and low income were found to be risl factors for Conclusion: Low socio-economic status is associated with an increased risk for IS. People with a low level of education and income is the target population for intervention on known medical risk factors to prevent IS. Introducaon'. Individuals with cognitive impairment as well as the entire population are exposed to painful medical procedures and painful conditions. However, since many of them cannot communicate verbalize with the surrounding, they are frequently undertreate<l for pain. Pain behavior scales that do not necessitate verbal reports have only recently been used for this population. Our aim was to study whether the behavioral responses to pain differ according to the level of cognitive impairment and whether these are in accordance with verbal reethods:rts of pain.
108 cognitively impaired individuals (22 mild; 43 moderate; 23 severe and 24 profound) participated. They were videotaped before and during a flu vaccination. Two examiners, using two behavioral scales, analyzed pain behavior: NCCPC-R (general pain behaviors) and FACS (facial expression of pan). Subjects with mild and moderate cognitive impairment were also asked to rate their perceived pan on a VAS scale.
Results: Pain behavior, measured with NCCPC-R, was significantlv increased in all groups during the injection (p<0.001-0:01). while facial expression of pan (FACS) was only significahtly increased in the mild and moderate retardation groups (p<0.01). 30% of individuals with severe to profound retardation responded with a "frees" during injection compared to only 4.5% of individuals with mild to moderate retardation. There was a good correlation (r=0.76) between the VAS and the behavioral scores only in individuals with mild retardation. In addition, there was a good to high_ correlation between the two methods (r=0.60-0.87) and a high correlation between the two examiners (r=0.91-0.97). Conclusion: Both NCCPC-R and FACS reliably detect changes in pain intensity in individuals with mild to moderate retardation but NCCPC-R is more sensitive to these changes compared to FACS in sever to profound retardation. Both scales have high intra rater reliability. The response to acute in differs according to the level of retardation. eywords: pain, mental retardation, pain behavior Faculty ofLife Sciences, Bar-Ilan UniversiO The term "neurosteroids" refers to steroids that can be synthesized de novo in the nervous system from sterol precursors. The group includes pregn_enolone (PREG), dehydroepiandrosterone (DHEA), their sulfates (PREG-S and DHEA-S) and reduced metabolites such as tetrahydroprogesterone (allopregnanolone). In addition to the well known genomic effect of steroids via nuclear receptors, neuroactive steroids can also act as allosteric modulators of neurotransmitter receptors, such as GABAA, NMDA, and sigma receptors, and thus affect behavior. Indeed, clinical studies in humans have associated some of these hormones with a sensation of "well-being", but also with reward-related processes, mood and motivation. Neurosteroids thus mayplay a role in substance abuse. In the present study, we testedthe hypothesis that the neurosteroid DHEA attenuates cocaine serf-administration. Rats were pretreated with either DHEA (2 mg/kg/day i.p.) or vehicle solution, and were then trained to serf-administer cocaine (1 mg/kg/infusion) on a fixed-ratio schedule of reinforcement, while continuing hormone treatment. DHEA significantly reduced cocaine-seeking behavior. These results suggest that DHEA affects cocaine reward. We further analyzedbrain DHEA levels in non-treated rats after exposure to cocaine. Our data show a 2-3 fold increase in DHEA levels in different mesolimbic brain areas of rats that maintain drug-seeking behavior compared to sham-operated controls.
This suggests that changes in brain concentrations of neurosteroids may play a role in the modulation of psychological states, including cocaine dependence. Although it is known that selective serotonin reuptake inhibitors (SSRIs), as other antidepressants, elevate mood only after 3-4 weeks of treatment, the mechanism responsible for this delay is not understood. SSRIs have been demonstrated to alter the levels of neurosteroids such as allopregnanolone (THP) which possess anxiolytic and mood elevating properties. We compared the effect of 9 and 21 days i.p. admimstration of paroxetine, a potent SSRI, on the synthesis of THP and its precursor, 5c-dihydroprogesterone (DHP) in the mouse cortex, hypothalamus and olfactory bulb. Cortex, olfactory bulb and hypothalamus synthesized levels of DHP were stgnificantly raised after 9 days of paroxetine administration, whereas a significant rise in the THP synthesized level was observed only after 21 days of treatment. Peripheral synthesis of DHP measured by the level in serum, significantly increased after 9 days, but reverted to normal values after 21 days. No increase was detected in serum THP levels either after 9 or 21 days' treatment. Differences in peripheral and brain synthesis indicates independence in brain synthesis. cells against the 13-amyloid (AI3) toxicity. In addition, EGCG significantly increased (-8-fold) the secretion of the non-amyloidogenic soluble form of the amyloid precursor protein (sAPPa) into the conditioned media of SH-SY5Y human neuroblastoma and rat pheochromocytoma PC 12 cells.
The increase was dose-dependent and was blocked by the hydroxamic acid-based metalloprotease inhibitor, Ro31-9790, suggesting that the effect is mediated via ct-secretase activity.
Infubition of protein kinase C (PKC) with the inhibitor GF109203X, or by down-regulation of PKC, blocked the EGCG-induced increase in sAPPtx secretion, indicating the involvement of PKC in the stimulated effect. Indeed, EGCG induced the phosphorylation of PKC, thus identifying a novel mechanism of EGCG action by stimulation of the non-amyloidogenic pathway via PKC activation. In addition, the administration of EGCG (2 mg/kg) to mice, for 7 or 14 days resulted in a significant decrease in membrane-bound holoprotein APP levels, with a concomitant increase in sAPP leve/s in the hippocampus. Consistently, EGCG markedly increased PKCtx and PKCe in the membrane and the cytosolic fractions of mice hippocampus. These f'mdings strongly suggest that EGCG has protective effects against AI3-induced neurotoxicity and regulates secretory processes of  (PD). Several lines of evidence correlate neuroprotection and cell survival processes with stimulation of protein kinase C (PKC) and extracellular signal-regulated kinases (ERK1/2). In this context, we tested the effects of low neuroprotective concentrations of EGCG (0.1-10 IM) on these signaFmg pathways, employing the 6-hydroxydopamine (6-OHDA) model of PD. The present study shows a novel rotective mechanism of EGCG, via restoration of the reduced KC and ERK1/2 activities, caused by 6-OHDA toxicity-, in the human neuroblastoma (NB) SH-SY5Y cell line. Since EGCG was also shown to increase phosphorylated PKC, we suggest that PKC may be necessary for its neuroprotective action against 6-OHDA. This was confLrrned by the selective sensitivity of EGCG action to the inhibitor of PKC, GF109203X (llaM). In addition, gene expression analysis revealed that EGCG prevented the 6-OHDA-induced expression of several pro-apoptotic molecules such as Bax and Bad, as well as the decreased Bcl-2, Bcl-w and Bcl-xL. These results establish a neuroprotective mechanism of EGCG action against oxidative stress-induced cell death, via stimulation of PKC and modulation of cell survival/apoptotic genes.
Keywords: EGCG, apoptosis, neuroprotection, PKC Exposure to acute stress blocks the induction of long term potentiation at amygdala-prefrontal cortex synapses in vivo Maroun M. and Richter-Levin G. The Brain and Behavior Center, University ofHaifa, Haifa 31905 Most studies of stress-induced synaptic plasticity impairments have focused on the hippocampus. The present study examined whether stress, which impairs hippocampal long-term potentiation (LTP), also affects LTP at amygdala-prefrontal cortex synapses in vivo. After undergoing an inescapable stress experience, rats exhibited markedly impaired LTP without having effects on baseline transmission, in contrast, unstressed control animals showed robust LTP that persisted during the recording period. Similar to stress, LTP at the amygdala-prefrontal cortex synapses was completely blocked when the competitive NMDA receptor antagonist CPP, was injected prior the tetanus. These results demonstrate that similar to the hippocampus, LTP in the amygdala-prefrontal cortex synapses s blocked after exposure to stress and the induction of LTP in this pathway is an NMDA receptor-dependent process. The present data show the amygdala-prefrontal pathway is amenable to long lastingplastic changes, behavioral stress and administration of NMDA-receptor antagonist CPP attenuate LTP in this pathway. Taken together these data suggest that the amygdala-prefrontal cortex pathway is relevant to stress-mediated impairments in LTP, and this pathway may have an important role in emotional memory processes. criteria, yet animal studies include the entire exposed population as the study population. We examined the effect of grouping prestressed rats according to severity of response on the statisttcal analysis of results. The effects of exposure to a cat on behavioral measures in rats were demonstrated. Response severity was assessed and used to divide the animals into "diagnostic" groups. The two extremes were studied, i.e. those clearly "maladapted" and those clearly "well-adapted", using arbitrarily selected "Cut-off Behavioral Criteria" (CBC). The middle group was discarded for reasons of clarity. The hypothalamic-pituitary-adrenal axis and heart-rate variability were subsequently analyzed for the entire expose.d population and then according to "the CBC. A single ten-rmnute exposure to a predator caused anxiety or fear-related behaviors. However, only 25.3% of exposed'rats were affected. Compared to controls and to "well-adapted exposed rats, "maladapted" rats exhibited sign!ficanfl.y htgher plasma corticosterone and ACTH concentrations, ncreased sympathetic activity, diminished vagal tone and increased sympathovagal balance. These differences were significantly more obvious when data were analyzed according to CBC. Animals respond to stress heterogeneously, resembling humans. Overlooking this heterogeneity may significantly affect the results of bit-behavioral data Analvsis, and animals can (and probably should) be divided int6 distinct groups according to their response. Keywords: Post-traumatic Stress Disorder, animal model, anxiety, stress, maladapted, well adapted Imaging dementia using b value q-space analyzed diffusion magnetic resonance imaging Mayzel-Ore 0.1, Assaf.1,2, Gii A.z, Ben-Bashat D. Verchovsky.a, Mordohovitch 3, Graif M.2, Rider-Groswasser If, Hendler T. , Korezyn A.D 3' and Cohen chool of Chemistry Tel-Aviv University Tel-Aviv 69978; 2 Wohl Insatute for "/fdvnced Imagine ant 3Devt. of,Neurology Tel Aviv SouraJkyMedical Center, el Aviv 6239"; 4Sackler Faculty of Medicine, Tel Aviv University, Tel Aviv 69978 High b-value diffusion weighted magnetic resonance imaging is highly sensitive to white matter (WM) changes. Diffusion images, when analyzed using the .q-space approach, show areas of abnormal WM in multiple sclerosis (N-IS) not detected by other MRI methods like T2-, TI-, weighted imaging, FLAIR, and DTI (Assaf et al, Magn. Reson. Med. 47:115-126 [2202]). When applied to vascular dementia(VaD) patients, high b-value q-space analyzed diffusion detected areas of abnormal WM extending further than T2 apparent hyperintensities. In the areas that presented hyperintense signal-in the FLAIR and T2-weighted images the WM density, as detected by q-space images, was reduced. These observations point to extensive nerve fiber loss in the WM in VaD. Analysis of high b-value, q-space analyzed diffusion images of Alzheimer's (AD) patients also contributed information not apparent from other MR] methods. AD patients presented less marked WM density decline in the parietal lobes than VaD patient but similar WM loss in the frontal lobes. Interestingly, diffusion images of AD patients also showed marked effects in gray matter areas, consistent with the pathologic changes in AD that consist of accumulation of neurofibnqlary tangles and senile plaques along with neuronal and synaptic loss leading to atrophy. To summarize, high b-value q-space diffusion MR] shows high sensitivity to WM changes and may be useful to characterize tissue loss in VaD and AD, as well as to the differentiation between those two paolo_gies.
Keywords: Alzheimer s disease, vascular dementia, q-Space, diffusion MR/ Dexanabinoi and its analog PRS-211,092 inhibit LPS-induced brain inflammation Meilin S. Oni E., Lavon I., Sheinin T., Avraham A., Efroni G., Lavie V.,'Greenberg O., Elgamiel R., Bar-Joseph.A. and Fink G. Pharmacology, Pharmos Lid Kiryat Weizmann, Rehovot 76326 Dexanabinol, a nonpsychotropic cannabinoid, and its novel analog, PRS-211,092, are neuroprotective in brain ischemia induced by middle cerebral artery occlusion. Both compounds were previously shown to reduce PGE-2 secretion m LPS activated macrophages in vitro. The aim of the present study was to assess Whether Dexanabinol and PRS-211,092 inhibit central neuroinflammation. C57BL mice were injected intra-cerebro-ventricularly (i.c.v) with either 250 ng LPS (dissolved in 5.! PBS) or 5xl PBS as sham. The mice were immediately m.lected IP with either cremophor-ethanol (vehicle) alone Dexanabinol (20mg/kg) or PRS-211,092 (20mg/kg). The }nice were killed under anesthesia 24 hours and 3 days after LPS injection, and the brains removed. IL-1 beta gene expression was determined in brains extracts from animals killed at 24 hours using RT-PCR. Analysis of activated microglia was performed after 3 days using immunohistochemistry. Treatment with Dexanabinol or PRS-211,092 reduced the number of microglia immunoreactive cells surrounding the hippocampus by 40% and 47% respectively compared with that in animals treated with vehicle alone. IL-113 mRNA level was reduced by 50%  We also describtdthe functl0nal implication of the interaction in Xenopus oocytes. Here we present further investigations that 101 attempt to elucidate the structural basis of such interactions.
We found that nearly complete deletion of the C-terminus of Kv2.1 channel (delC416) abolished Syntaxin 1A induced left shifts of the channel steady state activation and inactivation curves. It also abolished SNAP-25 induced increase of the sustained current and right shift of the steady-state inactivation. The slowing down of the onset of inactivation by SNAP-25 disappeared too. Similarly to delC416, deletion of the last two-thirds of the C-terminus (delC351) abolished the effects of syntaxin on the channel, but in contrast to delC416, only partially affected the effects of SNAP-25, diminishing the mcrease in the sustained current without significant effect on the steady-state inactivation curve. Based on these data we suggest that: i) s.yn,taxin 1A interacts with the proximal half of the C-terminus; li) SNAP-25 has two sites of mteraction, one, high affinity, at the proximal half, deletion of which cancels the right shift ofthe steady-state inactivation, and another site, low affinity, at the distal half of the channel, responsible, mainly, for the sustained current effect. These data concur with in vitro binding assays obtained in our lab (see the abstract of S. Tsuk). Keywords: Kv channels, syntaxin 1A, SNAP-25, C-terminus, SNARE complex Fusion of speech elements: Early auditory cortex involvement Mittelman N.. Bleich N., Laufer I. and Pratt H.

Evoked Potentials Laboratory, Technion, Haifa
Purpose: To define early auditory cortex activity_ to fusion of speech elements, and suggest plausible speech processing stages. Me-thods." Stimuli were binaural formant transition and base, that were presented separately or fused to form the vowel-consonant-vowel sequence/ada/. Eleven right-handed, adult, native Hebrew speakers listened to 2/sec presentations. Brain potentials from Cz during the 250 msec following transition onset in the responses to transition, to the fused word and to base alone were recorded. The net-fusion response was extracted by subtracting the sum of potentials to the base and the formant transition from the potentaals to the fused sound. Results': Auditory middle-latency components (20-45 msec), ing of nine peaks and troughs were recorded in response to the base, to the formant transition and to the fused /adal. In general, the responses to the fused object were significantly smaller in peak amplitude and in total activity (area under the curve) resulting in the difference waveform of the net-fusion response that also included 9 peaks, but with opposite polarities. Conclusion: audito_ry cortex is involved in the defmition and f sounds elements as speech, as early as 30 msec after stihaulus onset. This early processing iiavolves both inhibition and occlusion, and precedes the later stages of discrimination and meaning analysis. Keywords: auditory object, streaming, cortex In vivo two-photon imaging of dendritic stability in identified glomeruli of the mouse olfactory bulb Mizrahi A. and Katz L.C. Howard Hughes Medical Institute and the Dept. o of Neurobiology, Duke University Medical Center, Durham, NC, 27710, USA Ongoing sensory neuron and granule cell neurogenesis resul in turnover of both excitatory and inhibitory inputs to mitral cells, the main output neurons of the mammalian olfactory bulb. To evaluate the long-term stability of mitral cell dendrites in the face of these ongoing changes, we used two-photon microscopy to image mitral cell dendrites over prolonged periods in adult mice in vivo. Using transgenic mice expressing Yellow Fluorescent Protein in roughly I/3 of the m{tral cell population (Feng et al., Neuron 28:41-51 [2000]), we imaged the same dendritic trees in identified glomeruli over intervals of up to two weeks. We first examined the stability of mitral cell dendrites over short and long periods. Randomly chosen areas of the dendritic arbors were reconstructed in three dimensions and compared at different time intervals. The overall arborization of dendritic trees and even the smallest distal dendritic segments remained extremely stable over periods of hours, days and weeks (imaging intervals: hour, n=3; 24 hours, n=6:2 weeks, n=3). We next used intrinsic signal imaging of the dorsal surface of the olfactory bulb to identify individual glomeruli activated by a specific odorant.
The sizes of glomeruli stimulated by 0.1% butanal (n=5) were almost identical to the glomerular size imaged by two-photon microscopy. Given that dendritic stability ofmitral cells serves as a solidbaseline in control mice, we re currently using this combined approach to assess the effects of odor exposure and odor-based learning on morphological stability in identified glomeruli.

102
Keywor.ds: o.lfactory bulb, dendrites, two-photon microscopy, transgenlc mace Pathogenic self-antigens in autoimmune disease are potentially protective: A lesson from the visual system Mizrahi T. and Schwartz M. t. of Neurobiology, Weizmann Institute of Science, Rehovot 0 Axonal injury in the central nervous system (CNS) leads to primary and secondary degeneration of the fibers, with resulting death of the corresponding cell bodies. Our laboratory recently discovered that CNS myefinated axons, after suffering a mechanical insult such as a crush injury, can benefit from the activity of autoreactive T cells directed against myelin antigens (self-antigens associated with an autoimmune disease in the brain andspinal cord). Using a rat model of optic nerve crush or glutamate toxicity, we show here that vaccination with pepfides derived from either interphotoreceptor retinoid-binding protein or s-antigen (a retinal self-antigens that can cause experimental autoimmune uveitinitis) reduces retinal ganglion cells (RGCs) loss resulting from either glutamate toxicity or axonal injury. In the case of glutamate msult, no such protection was obtained by vaccination with myelin antigens. These results suggest that as in the case of myelin-associated antigens, the protective antigen for RGCs is identical to the self-pathogen associated with the common autoimmune disease in this tissue (i.e. uveitis). Based on our results we propose a more general phenomenon: pathogenic-self antigens in autoimmune disease may represent the potential protective autoimmune-evoking antigens. When sounds are presented in pairs with intervals of < 300 ms, the 100-ms cortical response N 100m to the 2nd stimulus is enhanced (Loveless et al., 1989). We studied the build-up of this enhancement effect and its relationship to behavioral performance and task difficulty. Whole-scalp magnetoencephalographic (MEG) signals were recorded from 10 subjects with a 306-channel neuromagnetometer. Sequences of 5 pairs of 50-ms binaural tones were presented with stimulus-onset asynchronies of 150 ms, inter-paar intervals f s and an inter-sequence-interval of 10 s. Subjects were asked to discriminate inter-pair frequencies under 2 conditions: easy (1000 vs. 1100 Hz) and difficult (1040 vs. 1055 Hz). In 50% of the pairs the tones were of the same frequency. All stimuli elicited prominent N100m responses in the auditorv cortex, peaking at 105 + 4 ms to the st tone and at 155 + 15 ins to the 2nd. The response to the 1st tone in a pair decreased by 46 + 3% from the 1st to the following pairs. I-n contrast, the response to the 2nd tone in a pair gradually increased in amplitude from the 2nd to the 5th pair. Consequently, the ratio of the 2nd and 1st responses increased throughout the five pairs from 0.73 + 0.07 to 1.23 + 0.05 (p < 0.02). Task difficulty did not affect the response amplitudes, whereas the response latencies to the 2nd stimulus were 11 + 4 ms longer for the difficult condition across pairs and hemispheres ,p < 0.02); the difference was most prominent (29 ms) in the 5th pair (LH: p < 0.03; RH: p < 0.05). Taken together, our findings suggest that the difficulty of behavioral discrimination is related to the dynamics of response build-up rather than to response strengths as such.
ZDep. Of Neurobiochemistry, Wise Faculty ofLife Sciences. Tel Aviv University, Tel-Aviv 69978; :Molecular Neuroscience, Rudolf Magnus Institute, University of trecht Medical Center, Utrect, Netherlands During repetitive or intense stimulation, neurosecretory cells and neurons undergo use-dependent changes in secretory strength, which are partially mediated bv activation of protein kinase C (PKC). However, the molecular targets of PKC and their mode of action remain elusive. In the present study we investigated the effect of Muncl8 phosphorylation on catecholamine secretion from chromaffm cells by membrane capacitance measurements following flash photolysis of caged calcium. Muncl8 is phosphorylated by PKC in a calcium-dependent manner and this phosphorylation inhibits its interaction with Syntaxin. By mutating the three PKC phosphorylation sites of Muncl 8 we created the Muncl 8-3A mutant, which mimics the non-phosphorylated form of the native protein. Overexpression of Muncl8-3A caused an increase in the number of fusion competent vesicles as well as an increase in vesicle recruitment under high calcium concentration, similar to the effect of wild-type Mtmcl8 overexpression. In contrast, the response to a second stimulation was similar to control cells while Muncl8 wild-type overexpressing cells exhibited a larger response. This suggests that vesicle pool refilling is reduced in Muncl8-3A overexpressing cells. As was shown recently, during the first flash stimulation, high calcium levels activate PKC, which acts on specific targets to enhance vesicle pool refilling. The reduction in vesicle pool refilling observedin the Muncl8-3A cells as compared to Muncl8 wild-type cells suggests that Munc 18 may be one of these targets.-Application of phorbol ester demonstrated that PKC in Muncl8-3A overexpressing cells can still enhance secretion but to a lesser degree than controls. These results indicate that phosphorylation of Muncl8 in chromaffin cells potentiates vesicle recruitment after emptying of the releasable vesicle pools. Keywords: Munc 18, PKC, secretion, membrane capacitance Distinct neural activity following an invalidly cued target in a cued attention task Ofek E. and Pratt H.
Evoked Potentials Laboratory, Technion Israel Institute of Technology, Haifa The cued attention task involves presentation of a cue stimulus, which provides information to direct attention to a target stimulus that follows, to which the subiect has to respond. In the minority of cases the cue provides'erroneous information about the target (invalid cues). The cue validity effect on brain response to the tarset was analyzed in the current study, and the brain structures mvolved were estimated.
In previous studies, a P3 component in response to the target was not found. Using a modified cued attention paradigm in the current study, a P3 component was evoked. The distinct neural activity pattern, during P3 period, was analyzed following identical target stimuli, which differed from each other only by the valence of the preceding cue valid or invalid. LORETA t-test results of 10 subjects showed significantly higher brain activity (current density) following invalidly cued targets. This higher activation was especially salient on the background of lack of significant validity effect on P amplitude. Unique brain areas were activated following the invalidly-but not the validly cued, targets. Those areas include attention related areas (BA), as x ell as emotion related areas (BA), and, surprisingly, also visual cortex. In summary,, distinct and significant activation was found, following identical stimuli, depending on whether they were preceded by invalid comparedto valid cues. The additional brain activation in the invalid case may be related to emotional processes, attention shift, and spatial updating. Univergity, Mt. Carmel 31905 Skill is acquired in a gradual manner, i.e., repetitions lead to better performance. However, the effect of time intervals on the course of learning is less clearly understood. Here we show that spacing the same total amount of practice events over days significantly improves performance. Subjects were trained in a mrror-reading paradigm given 12 blocks of practice on a repeated list of 24 words. Practice was given in a single session or in four sessions, separated by an interval of either half an hour or 24 hours. The reaction time data was well fitted by power functions with respect to the number of repetitions. However, while practice gwen in the same day, i.e., in a single session or in sessions spaced with half-hour intervals, resulted in identical slopes, a significantly steeper slope was found for the group in which practice was spaced over 4 days. When three additional groups were given the same amount of training with spacing as desci'ibed above, but with non-repeated words, the groups trained with a short or no time interval displayed a shallower slope compared with the group given spaced training over 4 days. Our results show that the number of repetitions during practice is not the only factor that determines the course of acquiring a .cognitive skill; rather the spacing in time of practice events s mportant. These results are congruent with the notions that: a) that within a session training may be superfluous beyond a certain number of repetitions, b) that time-after-practice contributes to the processes of learning complex tasks. Keywords" procedural learning, distributed practice, mirror reading 103 The metabotropic glutamate G-protein-coupled receptor mGluR3 is voltage sensitive Ohana L., Pamas I. and Pamas H.
Dept. of Neurobiology, Hebrew University, Jerusalem G-protein coupled receptors (GPCRs) comprise the largest superfamily of proteins in mammalians andare involved in most signal transduction processes as well as in re_gulation of many fundamental processes such as release of neurotransmitter from nerve terminals. In spite of GPCRs being transmembrane proteins they are not considered, unlike voltage gated channels, to be voltage sensitive. However, it was shown that muscarinic receptors, GPCRs, bind ACh in a voltage dependent manner (Ilouz et al, J Biol Chem, 274: 29519-29528 [1999]). Here, we examined whether mGluR3, a presynaptic autoreceptor that mediates feedback inhibition of glutamate release in CNS, exhibits voltage sensitivity. Using fresh rat brain synaptosomes we show that pr.esynaptic glutamate receptors at their physiological envtronment, similarly to the muscarinic receptors, brad glutamate in a voltage dependent manner. Deoolarization reduces the maximal binding of [3H]Glu several folds and further analysis revealed that depolarization reduced the fraction of the high affinity glutamate receptors. The experiments with synaptosomes could not discern whether it is the mGluR3 or the NMDA receptors (or both) that is responsible for the voltage dependent binding. This question was resolved using Xenopus oocytes. The mGluR3-mediated G-protein gated potassium channel currents were used to assay flae activity of the mGluR3. We found that the apparent affinity of the mGluR3 toward glutamate was reduced upon depolarization. The results presented here are compatible with the notion that the mGluR3 is by itself voltage sensors. Keywords: G-protein coupled receptors, metabotropic glutamate receptors, voltage sensitivity Preparation and characterization of monoclonal antibodies directed specifically at the apolipoprotein E isoform Ophir G.,_Smordinsky N. andMichaelson D.M. F{culty ofLife Sciences, Tel Aviv University, Tel Aviv 69978 Apolipoprotein E (apoE), the most abundant lipoprotein in the brain occurs as three alleles, termed E2, E3 and E4, which differ from each other by a single amino acid substitution. Genetic and epidemiological studies revealed that the allele E4 (apoE4) is a major risk factor of chronic neurodegenerative diseases such as Alzheimer's disease (AD) and of acute insults such as head injury. The mechanisms underlying the isoform specific pathological effects of apoE4 and the molecular structural features which differentiate between apoE4 and the other apoE alleles are not known. The present study was directed at the development of mAbs which react isoform specifically with apoE4 in its native form and at their use together with mAbs which react equally well with the different apoE isoforms, for structural analysis of the apoE molecule. The first approach used was to immunize apoE knockout mice which express human apoE3 with purified human apoE4. This approach yielded several mAbs Which bound specifically to apoE4 in ELISA experiments but whose binding characteristics varied in different preparations. In order to overcome this problem we undertook an alternative approach in which control mice were injected with synthetic peptides specific to apoE4 followed by screening for mAbs which bind specifically to apoE4 in immunoblot, ELISA and immunohistochemical assays and which specifically immunoprecipitate apoE4. The apoE4 specific and-pan apoE mAbs thus obtained will be described and their application for structural and functional studies of apoE4 will be discussed.
Keywords: apolipoprotein E, monoclonal antibodies, Alzheimer's disease, neurodegeneration Critical calpain-dependent ultrastructural alterations underlie the transformation of an axonal segment into a rowth cone after axotomy of cultured Aplysia neurons ren R., Dormann A., Gitler D. and Spira M.E.
f Neurobiology, Institute ofLife Science, The Hebrew

University of Jerusalem
The transformation of a stable axonal segment into a motile growth cone is a critical step in the regeneration of amputated axons. In earlier studies we found that axotomy of cultured Aplysia neurons leads to a transient and local elevation of the free intracellular Ca2+ concentration, calpain activation, localized proteolysis of the submembrane spectrin and growth cone formation. Inhibition of calpain by calpeptin prior to axotomy inhibits growth cone formation. Here we investigated the mechanisms by which calpain activation participates in the transformation of an axonal segment into a growth cone. To that end we compared the ultrastructural alterations induced by axotomy of cultured Aplysia neurons performed under control conditions and in the presence of calpeptin. We identified critical calpain-dependent cytoarchitectural alterations that underlie the formation of a growth cone after axotomy. Calpain-dependent processes lead to restructuring of the neurofilaments and microtubules to form a specialized cytoskeletal compartment 15-50-pro pro ,xm, al to the tip of the transected axon. This compartment "traps transported vesicles and serves as a locus for microtubule polymerization. As a result, a densepool of vesicles accumulates in close proximity to a segment ofthe plasma membrane along which the spectrin membrane skeleton was proteolyzed by calpain. We propose that this facilitates the fusion of vesicles with the plasma membrane, promoting the extension of the growth cone s lamellipodium. The growth process is further supported by the radial polymerization of microtubules from the growth cone's center.
Keywords: axotomy, growth cone, microtubules calpain, regeneration OlPosing effects apoE3 and apoE4 on APP metabolism owing closed head in,jury Oron L., Moskovitch L., Ezra Y., Shohami E. and lson D.M. 1Faculty of_Life Sciences, Tel Aviv University, Tel Aviv; :School of Pharmacy, The Hebrew University of Jerusalem, Jerusalem 91120 This study examined the possibility that the pathological effects of apoE4 and the beneficial effects of apoE3 following Closed Head Injury (CHI) are mediated by interactions with the amyloid precursor protein APP. Measurements of APP and of its soluble fragment APPs in cortical homogenates revealed that they were the same in sham treated control, apoE-deficient, and apoE3 and apoE4 transgenic mice. CHI induced marked elevation in cortical APP and APPs levels of the apoE3 transgenic mice whereas those of the other mice rueS were not si_gnificantly affected by this treatment. rmore the elevation in APPs in the head injured apoE3 mice was associated with elevation of APPsot .CHI also increased the levels of cortical apoE. There were however no differences in the cortical apoE levels of the apoE3 and apoE4 transgenic mice either prior to or following head injury.
Measurements of the hippocampal APP and APPs levels of sham treated mice revealed that they were the same in the different mice groups. However, CHI induced a significant elevation in the hippocampal APPs and APPsot level-s of the apoE3 transgenic mice and a decrease in the corresponding APPs and APPSot levels of the head injured apoE4 transgenic mice. In contrast the hippocampal APP levels were not affected by CHI. These findings show that APP metabolism is affected by the apoE genotype in a brain area specific manner. These effects may play a role in the pathological effects of apoE4 and the beneficial effects of apoE3 followmgbrain inj ,_ury.
Background: Cannabinoids, acting through specific receptors (CB and CB2) are implicated in the control of neurologicaland endocrine functions. 2-AG, an endogenous cannabinoid was shown to be neuroprotective after closed head inj_ury (CHI) [Panikashvili et al. Nature 413: 527-531, 2001]. This study examines the role of CB1 receptors in the development of secondary brain injury. Methods: CB1 knockout (CB1-/-) mice and their genetic background control C57BL/6J were used. CHI was induced on the exposed skull of mice under ether anesthesia using weight-drop device. Edema was evaluated 24h after CHI using the wet:dry weight ratio. The neurological status was evaluated using a 10-points scoring system (neurological severity score, NSS). ANSS reflects recovery after CHI and is calculated at any time (t) as the difference NSS(lh)-NSS(t). 2-AG, dissolved in ethanol:emulphor:saline (1:1:18) was injected subcutaneously immediately after injury. Results: Interestingly, CBI(-/-) mice suffered some (non-significant) basal clinical impairment, as shown by the pre-inj_ury NSS (1.57+0.3 vs. 0.56+0.3, p=0.08). These mice showed almost no clinical recovery at 24h after CHI, in contrast to the controls (ANSS 0.71+0.29 vs. 1.89+0.26, p=0.016). To further demonstrate the role of CB1 in recovery after CHI, 2-AG was injected to CB (-/-) mice. This treatment, which was effective in the controls, had no effect on the neurological recovery of CB 1(-/-) mice (ANSS 2.86+/-0.55 vs. 0.43+/-0.3, respectively, p=0.0041) and did not reduce edema (78.81+/-0.34 vs. 80.57+/-0.61% water, p=0.0308). These findings strongly indicate that CB1 receptors are mvolved in the attenuation of secondary damage after CHI.

Jerusalem
We previously found hemispheric differences in feature search (Pavlovskaya et al, Spatial Vision 14:151-73, 2001) when search arrays were in one or the other hemifield, rather than when the array was central and target elements lateral central.
In parallel, Ahissar & Hochstein (Nature, 387, 401-6, 1997) foundperceptual learning transfer across position or orientation depends on task condition: Learning effects transfer when the task is easy (large target-distraetor difference; limited target position uncertainty; long test-to-mask delay) and are considerably specific when conditions are harder. These easy vs. hard condition differences were related to cerebral sites of training modification: hard tasks were seen as requiring low-level (specific) representations while easy tasks are performed using high cortical level mechanisms alone. We ask f inter-hemispheric transfer also depends on task difficulty, since high-level receptive fields include eontra-lateral areas. Subjects performed color and orientation feature search, each within one hemifield. Sessions were half easy, half hard. Following training, we switched the sides of color and orientation tasks, and found nearly complete transfer for easy conditions, and considerably less with difficult conditions. Our results support the conclusion that easy search depends on high cortical level mechanisms, while hard search is performed by guided return to low levels when fine discriminations are required. Practice improves performance by modifying high or low levels, accordingly. Thus, easy task perfohnance and learning take place at a cortical area sufficieny high that its receptive fields are not limited to a single hemifield. Psychiatry was the only medical discipline that had no scientific foundation until recent years. Biological psychiatry began .approximately half a century ago when medications were discovered. The so called "bological" effect of medication on mood affect and behavior initialized an enthusiastic sur.ge of biological investigations in psychiatry..
Research was dtrected to bochemistry and thus findings are confined to neurotransmitter and receptor alterations relevant to medications and their clinical effects. This trend of biological psychiatry is at its peak even nowadays. To be a psychiatrist with molecular biology education would advance your career enormously. Disappointing however are the advances for the patients. Even though patients' symptoms have improved, patients are not anywhere near real cure conditions, especially if serious mental disorders such as schizophrenia are targeted. Both the molecular biology as well as the genetic research findings failed to approach the big picture of mental disorders as brain diseases. To put the findings of biological psvchiatrv in any useful context the bra.in as a dynamic orgamztion 6f neural networks has to be addressed. Cutting-edge thinking about mental disorders point to neuroscience as the future science for psychiatry. Mental disorders are thus disturbances in the organization of complex dynamic interacting cell ensembles and ever-changing neuronal networks spread in the brain. The future of psychiatry will require understanding of complex non-linear dynamics. Concepts such as criticality and optimizations, that are currently alien to psychiatrists will be the basis of understtnding mental "conditions. Instead of horrifying nomenclatures such as "schizophrenia" in an ineffective low-reliability diagnostic system, futuristic psychiatry will diagnose "optimization breakdowns" in the brains of patients. Psychiatric treatment will be directed toward re-optimization of neural brain systems for real cure of mental illness.
Keywords: psychiatry, complexity, system theory, functional connectivity Observation of spontaneous cortical layer activity using physiological MRI noise Pelled G. and Goelman G. HBRC, medical biophysics andNuclear Medicine Dept.,

Hadassah Hebrew University Hospital, Jerusalem
Measuring the functional magnetic resonance imaging (fMRI) blood-oxygenation level dependent (BOLD) signal non-invasively from the different layers of the cortex, can expand our Understanding to the current debate in the fMRI community regarding the neuronal source of the fMRI signal, since the cortex is composed of six layers that different in there cell bodies, s!apses and dendrites densi'. Using MRI physiological noise we measure the BOLD fluctuation of the cortical layers during rest. As a model we use the rat's visual cortex, which has been extensively, studied using electrophysiology methods. Two different statistical approaches were used. In the first, for each voxel temporal SD were calculated and averaged together. In the second, the area spatial SD were calculated-and averaged along time. Generally, the first approach measures the level of spontaneous neuronal activity, while the second approach measures the homogeneity behavior of the neuronal compartments. Our results show high temporal fluctuations in the deeper layers, indicating high spontaneous activity in these layers and low spatial fluctuations in these layers, indicating high homogeneity behavior. These results, are in agreement with previous, electrophysiolo,gy studies, and demonstrate that the physiology MRI nose xs sensitive enough to detect localize neuronal activity and neuronal homogeneity behavior.
Keywords: cortical layers, BOLD, fMRI Early coassembly of KCNQ1 with KCNE1 or Yotiao but not with an LQT5 KCNE1 mutant prevents channel inhibition The I:s potassium channel complex consists of the heteromeric assembly of two structurally distinct at and 13 subunits called KCNQ and KCNE1, respectively. Mutations in KCNQ1 and KCNE genes produce the long QT (LQT) ,syndrome, a human genetic cardiovascular disease. We recently identified a small region in the KCNQ1 C-terminus (aa. 589-620) which functions as an assembly domain for KCNQ at subunits. Here we show that co-expression of WT KCNQ1 with. CAD, a KCNQ C-terminus peptide (510-620), suppresses K channel activity. CAD inhibits KCNQ currents probably by inhibiting the tetrameric assembly of WT subunits as it encompasses the channel assembly domain and a at helix structure that is part of a calmodulin-binding domain. Confocal immunocytochemical images confirm the virtual absence of KCNQ1 expression in the plasma membrane. Co-expression of .WT KCNE with WT KCNQ and CAD, fully restores the K* channel activity and the_ plasma membrane labeling of KCNQ1 and KCNE1 proteins. Conversely, a naturally occurring LQT5 mutation of KCNE1 (W87R), located at the C-terminus, is unable to fully restore neither functional K* currents nor the membrane localization of the channel subunits. Yotiao, an adaptor protein that binds to the same KCNQ assembly domain could restore like other I subunits (KCNE2 and KCNE3) functional K* currents when co-expressed with WT KCNQ and CAD. In all, these data suggest that the early co-assembly of KCNQ with KCNE1 or Yotiao, but not with an LQT5 KCNE1 mutant (W87R), prevents channel inhibition by the CAD tetramerizatton peptide.
Dept. of Physiology and Pharmacology, Sackler School of Medicine, Tet/lviv University, Tel Aviv 69978 Voltage-dependent K channels play a major role in brain functions. Among them, the M potassium current has profound effects on brain excitability as ts low-threshold gating and its slow activation and deactivation act as a brake for neuronal firing. Recently, the KCNQ2/3 channel complex belonging to the KCNQ family of voltage-dependent K channels was identified as the molecular correlate of the M-current.
Furthermore, the KCNQ2 and KCNQ3 channel at subunits are 105 mutated in families with benign familial neonatal convulsions, a neonatal form of epilepsy. In this study, we characterize two novel openers of KCNQ2/3 channels, meclofenamic acid and 1-EBIO. Using CHO cells transfected with the KCNQ2/KCNQ3 cDNAs and the patch-clamp technique, we found that meclofenamic acid and 1-EBIO activate KCNQ2/3 channels, by causing a marked leftward shift in the voltage dependence of activation (-23 mV and -8 mV, respectively). In addition, both compounds slow down the channel deactivation kinetics. These openers increase the KCNQ2/3 current amplitude at physiologically relevant potentials (-70 mV to 0 mV). Recording of membrane potentml in Xenopus oocytes expressing KCNQ2/3 channels indicates that incubation with 30 laM meclofenamic acid within 1-2 min produces a hyperpolarization from a resting potential of-62 mV to -78 mV. Interestingly, these openers potently and reversibly blocked the evoked and spontaneous electrical activity of rat cortical neurons. In view of the crucial role of KCNQ2 and KCNQ3 channel subunits in epilepsy and neuronal excitability, enhancement of KCNQ2/3 potassium currents by these openers may prove to be an important target for future anti-epileptic and neuroprotective therapy. Keywords: K* channels, epilepsy, KCNQ Visual deficits associated with object images revealed by fMRI in ,human amllyopia Pianka p.l, Lerner Y.", Azmon B.', Leia H. Stolovitch H. , tein A. , Harel M.2, Malach R. and Hendler T. lTel-Aviv Sourasky Medical Center, Iel Aviv; 2tYeizmann Institute of Science, Rehovot 76100; Kaplan Hospital, Rehovot Amblyopia is a visual developmental disorder characterized by abnormal foveal vision due to aberrant early visual experience.
In normal subjects foveal vision is associated with the representation 6f faces (Levy et al. Nature Neurosci 2001). Here we explored possible relationship between the amblyopic deficit and fMRI activation to various object categories. Ntne unilateral amblyopic and 3 healthy subjects with ngrmal vision were studied. Three fMRI experiments were conducted using red-green filters for monocular stimulation: (1) Retinotopic mapping of the sound eye (2) Presentation of small and large pictures correlating to visual acuity of 6/6 and 6/60 respectively (3) Line drawings of faces and houses of equal size presented to each eye. In low order visual areas (e.g. retinotopic) the sound eye's activation was similar to normal subjects. However, in the amblyopic eye, the activation for small pictures was markedly reduced compared to the sound eye, while large pictures' activation was only slightly reduced.
In high order visual areas the sound eye's activation was similar to normal subjects while the amblyopic eye showed marked activation reductions that appeared to be more emphasized in the fusiform gyrus compared to the collateral sulcus. We conclude that abnormal early visual experience resulting in amblyopia affects in a selective manner the central representa.tion of objects both in low and high order visual Practicing certain visual tasks leads to a significant improvement in performance, a process termed "perceptual learning". Learning was shoaa to be specific for basic stimulus features such as local orientation, rettnal location and eye of presentation, suggesting plasticity of the primary visual cortex m adults. However, it has never been showh that such a low-level learning may have impact on higher-level visual tas"ks, like recogmtion. Amblyopia is characterized by several functional abnormalities in spatial vision, including reduction in visual acuity (VA), contrast sensitivity, function (CSF), and vernier acuity, as well as spatial distortion, abnormal spatial interactions and impaired contour detection. The visual deficiencies are thought to be irreparable after the first decade of life once the developmental maturation window has been terminated. The vision loss is thought to result from abnormal operation of the neuronal networks within the primaD.' visual cortex, most notably, orientation selective neurons and their interactions. The perceptual learning procedure described here was designed to train this network by efficiently stimulating these neuronal populations and effectively prtmoting their spatial interactions. Here, using a systematic low-level training of a malfunctioning adult visual szstem, we show that inducing low level changes may yield significant perceptual benefits and transfer to higher visual tasks. The training proced.u[.e produced a two-fold improvement in CSF and letter recogrutton tasks in the treatment group (N=63). No improvement found in the control group (N=14). The results demonstrate that perceptual learning can improve basic representations within the adult visual system that did not develop during the critical period. Keywords: vision, perceptual learning, development,

amblyopia, lateral interactions
The adverse effects of repeated intraperitoneal injections are reversed by chronic treatment with the antidepressant drug imipramne Pollak Y?, Ovadia H. and Yirmiya R.
Dept. of lr'sychology, The IT[ebrew University of Jerusalem, Mount Scoous, Jerusalem; Dept. of Neurology, Hadassah:Hebrew University Hospital, Jerusalem Previous research suggested that repeated handling and intraperitoneal (i.p.) injections constitute a chronic stressor, producing depressive-like physiological consequences. This present study was aimed to characterize further the effects of repeated _saline injections in SJL/J mice, partieularly in the context or experimental autoimmune encephalomyelitis (EAE).
Mice were injected daily with saline (10 mt/kg) for 3 weeks or not handled. Alterations in body weight, spleen weight and blood interleukin-1 (IL-1) protein levels were measured, as well as the susceptibility to the induction of EAE by myelin auto-antigen, Repeated injections were associated wth lower body weight and higher levels of IL-1. No effect on spleen weight was recorded. Following immunization with PLP, a common myelin-associated Protein, chronically injected mice presented with aggravated clinical symptoms of EAE and higher mortality rate. Chronic treatment wth daily injections of th6 tricyclic antidepressant drug imipramine (10 mg/kg/day) imipramine, completely reversed the effects of repeated injections on bod3) weight and EAE-associated mortality, and significantly attenuated the effects on IL-1 production and EAE-assocated symptoms severity. Oral administration of imipramine (10 mg/kg/day dissolved in the drinking water) had no effect on non-injected EAE mice, indicating that the effects of imipramine on EAE were limited to counteracting the consequences of repeated injections. In conclusion, repeated injections in mice are associated with imipramine-suppressible adverse physiological consequences that may be related to higher production of pro-inflammatory cytokines. These results highlight the importance of control non-handled animals in any study design consisting of repeated i.p. injections. Keyword: repeated injections, stress, experimental autoimmune e.ncephalomyelitis, interleukin-Chronic treatment from adolescence with TV-3326, a brain-selective MAO-cholinesterase inhibitor, abolishes hyoeranxiety in adult orenatally-stressed rats tunes in animals in which anxiety was induced by. acute stress and was sensitive to behzodiazepines failed to demonstrate an anxioljctic effect of TADs. Chronic hyperanxiety and depressive-like behaviour can be induced in primates and rodents by prenatal stress. The aim of the present study was to see whether amitrip'line and TV-3326, a novel MAO-eholinesterase inhibitor, had maxiolytic effects in this model. Male and female offspring [PSI. of rats (at. least 9/group) that had been stressed by daffy restraint during gestation and of control rats [C], were given water, amitriptyline (4.5 mg/kg/day) or TV-3326 (17 mg/kg/day) in the drinking water from the age of 6 to 12 wee'ks. TV-3326 inhibited MAO-A and B in the brain by 70% and 60%, respectively. During the 12th week, PS and C rats were tested in the elevated plus-maze, a validated test for anxiety. TV-3326 selectively increased the time spent by PS but not C rats in open arms from 11.9+3.4 to 73.4+15.7 sec (P<0.001), indicating a decrease in anxiety. It was only possible to detect an anxiolytic effect of arnitriptyline in PS rats 2 weeks after cessation of treatment, when its depressant effect on exploration in C rats under continuous treatment was no longer seen.
In conclusion, chronic administration of TV-3326, selectively abolishes hyper-anxiety induced by prenatal stress without suppressing behviour of controls. Keywords: antidepressant, chronic anxiety, prenatal stress, plus maze Reversible internalization of voltage gated channels accompany brefeldin A-induced structural remodeling of cultured Aplysia neurons Praer-Khoutorskv M, and Spira M.E.

Li Sciences Insiitute, The Hebrew Univers#y of Jerusalem
Eukaryotic cells constitutively internalize the plasma membrane. Nevertheless, the morphology and membrane properties of the cells are maintained constant by compensatory constitutive exocytosis. Recently, we reported that the neurites and axons of cultured Aplysia neurons are internalized within 24-48 hr of 51xg/ml brefeldin A application (Prager-KhoutorskT and Spira 2000, Neuroscience Letters supplement 55, $43).
When brefeldin is washed away, the Golgi apparatus reassembles, and the neuron extends new neurites. Here we report that the passive and active membrane properties of the neurons undergo alterations during these restructuring processes. Control experiments revealed that both the cell body and axons are excitable and that the action potentials are not blocked by BFA. We found, that for as long as the axons are not totally internalized (in BFA) depolarization of the neuron evokes action potentials. Nevertheless, when the axons are internalized depolarization of the formally excitable cell body fails to generate action potentials. We thus conclude that the cell bod looses it excitability during the axon's membrane internalization. Cytochalasin B inhibits the effects of BFA.
Under these conditions the cell's input resistance and excitability are not altered. When BFA is washed and the naked cell body begins to extend growth cones, depolarization generates normal action potentials. These observations suggest that the newly formed vesicles carry to the plasma membl:ane the normal repertoire of voltage-gated channels to generate action potentials. We conclude that regulation of the balance between constitutive exocytosis and endocytosis leads to remodeling of the morphology and the biophysical properties of adult neurons. Keywords: neuroplasticity, Golgi, endocytosis exocytosis,

Aplysia
The effect of eternal noise on ord span Putter-Katz H. and Ahissar M. 'z Interdisciplmary Center fo_r Neural Co_mputalion, Hebrew University of Jerusalem; "Dept. of Psychology. Hebrew University of Jerusalem Previous studies documented strong correlations between auditory perception and working memory in the general population, learning impaired su6jects and hearing impaired Individuals. We hypothesized that the items retained in working memory are those perceived with sufficiently high signal to noise ratio, with whole words as default candidates. When signal to noise ratio is low, either due to external or to internal noise, the items retained in memory, will be sub-word elements, and consequently word span will decrease dramatically. We tested this hypothesis in the general population. Subjects were asked to recall sequences of 1-6 two-syllable pseudowords that were presentedin quiet and in noise. For the majority of subjects the addition of noise decreased single word'repetition accuracy by 10-20%. For these subjects, word span in noise was significantly lower than one would expect hadspan (memory) and repetition accuracy been independent. When sequences were longer than two words their repetition accuracy dropped dramatically. These results are consistent with our hypothesis that decreased perception (signal to noise ratio) affects level of representation m working memory and consequently decreases memory span.
Ke.ywords: speech perception, verbal memory span, signal to noise ratio Linking morphology and homeostatic synaptic plasticity using a CA1 ovramidai neuron model University Co'liege, London A recently discovered plasticity effect, termed "homeostatic plasticity', takes place when syhaptic conductance is adjusted to maintain what seems to be a stable level of activity of the cell. This process is much slower than LTP/LTD (on the order of hours to days) and has been found to negatively correlate with the level of depolarization. In this study, a detailed model of a CA neuron was used to explore the mpact of dendritic morphology on synaptic homeostatic plasticity. The model consists ofnumerous excitatory synapses that are evenly spread on the dendrites, together with uniformly, distributed active ion channels, giving rise to back-propagatmg action potentials. Synapses were activated by stationary stochastic inputs. A smple local rule was applied to iteratively scale synaptic conductance so as to keep the average local dendritic membrane potential at a predetermined level. The synapses progressively reached an average steady state conductance with a bell-shaped profile along the apicaltree, peaking at about midway from soma to distal rafts. This is reminiscent of recent fmdings on the distribution of synaptic conductance in CA1 neurons. In the shorter basal tree, only the initial increase in synaptic conductance with distance from soma was apparent.
We demonstrate that there is an interesting interaction between specific dendritic morphology, membrane excitability and synaptic plasticity. Together they shape local voltage perturbati, on as well as spatial interaction (the "region of influence ') of one synapse on others; this in turn affects the global activity of the neuron (e.g., axonal spikes). In consequence, very local plastic rules result in a global cooperative effect. Keywords: synapse, plasticity, homeostasis, morphology Protection against soman-induced brain damage and cognitive deficits by an antidotal treatment with anticholinergic and antillutamatergic agents Raveh L., Brandeis R., Gilat E., Cohen G., Alkalay D., Rabinovitz I. and Weissman B.A.

Dept. of Pharmacology, Israel Institute for Biological
Research, P. O. Box, 19, Ness Ziona The protective effects of post exposure antidotal treatments combined with p)Tidostigrnine pretreatment on the deleterious consequences of soman poisoning were studied. TAB (a mixture of TMB4, atropine and benactyzine) was injected concomitant with the development of toxic signs whereas scopolamine (0.1 mg/kg) or caramiphen (10 mg/kg) were administered five nun after soman (1.2 LDs0). Atropine (4 mg/kg) was given at onset of toxic symptoms to the latter two gro.ups. It was found that caramiphen and TAB completely abohshed electroaphic seizure activity_ while scopolamine treatment exhibited only partial protection. Additionally, no significant alteration in the density of peripheral benzodiazepine receptors were noted ensuing the former therapies whereas scopolamine application resulted in a complex outcome: while part of the animals demonstrated no change in the number, of these sites, the other exhibited markedly higher densities. Cognitive functions (learning and memory processes, evaluated using the Morris Water Maze) were unproved by the three treatments compared to soman-injected animals, with the following rank order: The areas devoted to vision in the primate brain constitute -25% of the cortical sheet. Does loss of vision render these regions useless. Recently using fMRI mapping, we showed that the occipital cortex of the congenitally blind is activated during various high level cognitive tasks, including auditory verb generation and verbal memory. This activation was found in all blind subjects (N=10), in regions corresponding to retinotopic visual areas in sighted subjects. If the occipital cortex of the blind has any functional role, one might expect the level of occipital activation to be correlated with the subject's pe ,rformance in similar tasks. We therefore measured the subjects verbal and memory skills, using the standard Wechsler tests. Interestingly, the blind as a group were far superior in their memory performance compared to the sighted controls (for example digit span-blind." 14.3+1.36 SEM compared with the grand population norm of 10). Second, the extent of fMRI activation and its significance level differed substantially between individual subjects. Crucially, strong positive correlation (r0.8-0.9) was also observed between the mdividual subjects" fMRI signal during verbal memory" recall and their performance in the Wechsler's memory test. These 107 results suggest that the occipital cortex of the congenitally blind may undergo a radical change to be seemingly involved in language and memory functions. The only clinically proven therapeutic approach for the treatment of Alzheimer's disease is inhibition of acetylcholinesterase (ACHE) which includes alkylcarbarnates, such as physostigmine, rivastigmine and heptylphysostigmine. Several researchers have noted the surprisingly low potency of ethyl-substituted carbamates compared with methyl substituents, but there have been very few studies on AChE inhibitory activity of longer chain alkyl substituents. The aim of the present study was to measure the kinetics of AChE inhibition (ki) at 37C in different homologous series of carbamates with an H or methyl group as one substituent (R) and a methyl, hyl, n-propyl, n-butyl, n-hexyl or methoxyphenyl as the other (Re), using human erythrocyte ACHE, according to the scheme: Our results showed that ki was highest when,R1 d R were methyl in all series, (e.g. 4081 [M"min'] in the 6-c,arbaq3oylaminoindan series). This was reduced to 102 M-'min when R was ethyl and gradually rose with increasin chain length to 3055 M-Xmin for n-hexvl and 7111 Mmin 'r for methoxyphenyl. A very similar relationship was found for carbamate derivatives of 3-hydroxyphenethylamines.
The high poten.cy of the aromatic carbamates may be explained by :x-r nteractlons with phenylalanine residues near the active ste of ACHE, however, the tmusual SAR of the alkvl substituents is more difficult to justify and may involve interactions at the peripheral anionic site (Lin el. al, Bioorganic & Medicinal Chem. 7:2683-26891999).

Dept. of Molecular Genetics, Weizmann Institute of Science
Formation of the brain structure in human is a complex process that occurs during several months ofprenatal development.
One of the most striking features of the human brain is its characteristic convolutions. These convolutions are lacking in a severe human brain malformation known as lissencephaly (lissos smooth, encephaly brain). About one in 30,000 life births are affected with this disease. Lissencephalv patients have a reduced life expectancy and a severe mental rrtardation. So far, two genes have been found to be mutated in lissencephaly; LIS1 located on chromosome 17 and Doublecortm an X-linked gene. LIS is a WD repeat protein and is known to be involved in several protein complexes and microtubule regulation by direct interactions with tubulin or via interactions with the dynein/dynactin motor complex. We describe the phenotype observed in Lisl mutant mce in the context of LIS proten interactions. In addition, we discuss an interaction with the cytoplasmic linker protein CLIP-170. CLIP-170 is required for in vitro binding of endocytic transport vesicles to microtubules. We suggest that CLIP-170 and LIS are both required for cargo transport via the dynein motor complex. Using L-alpha-amino-3-hydroxy-5-methylisoxazole-4-propionate (AMPA) receptor activity_ is required for long-term potentiation (LTP), a postulated model of cellular mechanisms underlying learning and memory.. GluR1 (GluR-A), one of four AMPA receptor subtypes, as thought to mediate a postsynaptic mechanism of LTP expression. Previous studies have demonstrated that GluRl-deficient mice lack LTP in the afferent pathways to the CA1 area of the hippoeampus, yet their spatial reference memory is intact (Zamanillo, D. et al. 1999. Science 284:1805-1811. One way of explaining the intact learning ability in the GIuR1-/mice is that they employ extra-hippoeampal structures to solve the spatial learning tasks. We tested ths hypothesis by cytotoxie lesion of the hippocampus using an injection of NMDA. The mice were then tested in the standard version of the Morris water maze task and on a reference memory task on the elevated Y-maze. Performance in the water maze and on the Y-maze was impaired by lesion but not by GluR1 deletion. This suggests that extra-hippocampal compensation cannot account f6r the spared ability of the GIuR1-/mice in spatial learning tasks such as the Morris water maze and the Y-maze. Keywords: GluRl, hippocampus, LTP, working memory Utilization of genetic tools for studying neural development in avian embryos Rendelman Z., Hazanov S., Guy U., Marom K., Feinstein Y. and Klar A.
Dept. of Anatomy and Cell Biology, Hebrew University-Hadassah Medical School, Jerusalem 91120 The employment of genetic tools for studying biology in general, anddevelopmental biology in particular, has advanced our knowledge tremendously over the last decades. The ability to inactivate a specific gene, or to nis-express it, thereby generating a loss or gain-of-function mutant, is the ultimate way to vei-ify its biological role. So far these t6ols were applied successfully in invertebrates and mice.
The avian embo is the preferred organism for studving embryonic development of vertebrate. The embryo is "flat, develops in ovo, and accessible for surgical manipulations.
Hence, the inability to introduce genes in vivo has hampered the progress of research with this organism. In the last few years, a new method, electroporation, was introduced to deliver genes in vivo, and been used effectively in avian embryos.
However, the site of expression is dictated by the topography of the embryo and the electrode. In order to achfeve site-specific expression we have used electroporation of tissue specific enhancers. The activity of the specific enhancers was further amplified by the use of DNA site-specific recombinase. In this way, we are able to achieve high level eetopie expression of an active form or a dominant negative form of a protein, at a specific site. For generating a null phenotype, we are currently applying the RNAi method in chick embryos in vivo.
The genetic tools will be presented, and their potential use for studying axon guidance and cell migration during early development will be demonstrated. Sackler Medical School, Tel-Aviv Umversily Post-traumatic stress disorder affects only 20-30% of the exposed population. Clinical studies of acute and chronic responses to stress generally employ stringent criteria for inclusion in the study population, and yet in animal studies the data are usually .expressed as a function of the entire exposed versus the entire unexposed populations, regardless of individual variation in response. Prior data support an approach to animal models analogous to inclusion criteria in clinical studies.
This series of studies sought to assess prevalence rates of maladaptive versus adaptive responses determined according to two successive behavioral tests (elevated plus maze/open field and acoustic startle response tests), amongst rats exposed to a variety stress paradigms, in the acute and chronic phases, for single and repeated exposures, early in life and/or in adulthood. The results shed light on the pattern of prevalence rates of maladaptive responses to stress over time, with rates dropping from 90% acute responders to a plateau at about 25% enduring/chronic response. This plateau is attained at seven days and remains unchanged at 30 days in two distinct study models. Different types oftrauma are associated with different degrees of response, possibly reflecting a "dose-response" relationship to severity of trauma. Re-traumatization is associated with increasing prevalence of maladaptive responses, all the more so for early-life trauma repeated in adulthood. Setting the affected individuals apart from the unaffected and focusin.g on them clarifies the overall picture and more closely roxmaates clinical studies. The parasitoid wasp Ampulex compressa hunts cockroaches to serve as a living food supply for its larvae. The wasp injects its venom directly into the bran to induce a lethargic state lasting about five weeks. While in this state, the coc'kroach does not escape. Our hypothesis is that the injected venom affects neurons located in the brain, which send descending tonic input to bioaminergic neurons; these, in turn, control the thoracic premotor circuitry. Octopamine is a bioamine, which is secreted by DUM (dorsal unpaired median) neurons in the prey's thoracic portion of the nervous system. It is known to regulate the expression of specific motor patterns by modulating the excitability of specific neurons.
In this work, we show that the activity of DUM neurons is altered in stung animals. Identified DUM neurons in the metathoracic ganglia of stung animals have a lower spontaneous firing rate and their responsiveness to specific sensory stimuli decreases. For example, DUM234 neuron does not produce a burst of action potentials in response to a wind stimulus as it does in control animals.
Given these results, wepropose that the venom injected into the brain affects descending tonic input to the thoracic DUM neurons. The decrease in activity of the DUM neurons could be responsible for the decrease in the responsiveness of thoracic motor circuits and consequently the expression of specific motor patterns such as the escape behavior. Keywords: ampulex compressa, cockroach, octopamine, dorsal unpaired median neurons We performed association studies in 109 sporadic AD cases and 111 aged healthy controls. We investigated natural genetic variations (polymorphisms) in 5 genes selected by pathobiological criteria: p53 and TNFRSF6 (Fas), both apoptosis-related genes which are upregulated in AD brains; complement component (Clr) and interleukin-l-beta (ILl-B), both inflammation-associated genes upregulated in AD brains; and heparane sulfate proteoglycan (HSPG2), which is associated with amyloid plaques and neurofibrillary tangles. We tested whether these genes act as susceptibility genes affecting the risk for AD or as modifier genes influencing its clinical features. While our study population demonstrated a strong association between AD and the APOE-e4 risk factor (p=0.000002), no such association was found with the  19=0.486]. No association was detected also for age at disease onset and disease progression, and no interactive effect was found with APOE-e4. This is the first report of association studies of p53, C lr and HSPG2 with AD or any neurodegenerative disorder at all. Our results of Fas and IL1-B are in agreement with initial reports, published during our work. Further intensive association studies are needed for identifying genetic risk factors that will allow constructing a profile of individuals prone to AD, and may assist in developing novel therapies.
Microglia and macrophage activation in the injured nervous system Rotshenker S. Reichert F., Slobodov U. and Makranz C.  Fc-receptor (FcR). We presently document that CR3/MAC-1 is expected to play the major role in mediating mye.lin phagoc.ytosis after trauma (70% to 80%). However, its ln-vlvo expression does not indicate necessarily the occurrence of myelin phagocytosis. CR3/MAC-1 expressing microglia are fully activatedand phagocytose myelin efficiently in EAE but not after injury to CNS axons. These observations led us to suggest thatCR3/MAC-1 mediated myelin phagocytosis is su_bjected to modulation such that it may range between efficient and inefficient states.
We tested the functional plasticity of CR3/MAC-1 mediated myelin phagocytosis in macrophages and microglia by anti-CR31MAC-1 mAbs, documenting inhibition and augmentation of myelin phagocytosis by anti-CR3/MAC-1 mAbs. We suggest that mAbs binding to distinct extracellular epitopes in t/i3 subunits of CR3/MAC-1 can modulate (inhibit or augment) CR3/MAC-1 mediated myelin phagocytosis by inducing and/or stabi!izin8 distinct conformational changes that correspond with inactive or active functional states CR3/MAC-I. We further suggest that anti-CR3/MAC-1 mAbs may reveal a mechanism by which native extracellular molecules bind to and modulate CR3/MAC-I mediated myelin agocytosis in microglia and macrophages. eywords: trauma, regeneration, microglia, macrophages, myelin 109 Brain reward system and depression: A tool for testing hedonia in an animal model Roth Deri I.. Gispan-Herman I. and Yadid G.

F-7l--ff f i fe Sciences, Bar-Ilan University
Anhedonia is one of the main parameters that distinguish a depressed patient from a normal one. However, the brah mechanisms that underlie anhedonia remain elusive. Data suggest that there are overlaps between depression/anhedonia and neural reward systems. Therefore, the brain reward system may be a potentially important therapeutic target to relieve anhedonic symptoms.
The Flinders sensitive line of rats (FSL) is a valid mad reliable model of depression with strong face-, construct-and predictive-validity. Abnormalities in the reward system of FSL rats were tested behaviorally and neurochemically using the cocaine self.administration paradigm in parallel with microdialysis measurement of doparnine levels in the nucleus accumbens. Our results show that cocaine consumption was significantly higher in control than in FSL rats. Treatment with the antidepressant desipramine increased the cocaine self-administration in FSL rats. Basal dopamine secretion ratio (using GBR12909) and extracellular dopamine in response to cocaine were also abnormal in the nucleus accumbens of FSL rats. Both of these measures were normalized following treatment with desipramine. Taken together, we sutgest that the brain reward system may be a tool for stulying the anhedonia expressed in depressive behavior. Keywords: brain reward system, cocaine, hedonia, animal model A one-way ANCOVA with age and sex as covariants was performed on plasma total homocysteine levels of 190 schizophrenic patients vs 762 controls (evaluated in a screening program for employee health). The effect of schizophrenia was marked (F=130.5, df 1; 902, p<0.0001) and mean homocysteine level was 16.7 + 11.8 (SD) in schizophrenic patients vs 10.6 + 3.6 (SD) in healthy controls (covariance adjusted means 16.5 vs 10.7 in controls). The increase was almost entirely in young male schizophrenic patients. Homocysteine has been shown to be neurotoxic and it has been shown that stress can open the blood brain barrier to some neurotoxic substances. It is possible that the stress of acute psychosis allows high homocysteine levels to enter the brain and cause neurodegeneration, clinical deterioration and chronicity. This hypothesis does not make any assumption as to the origin of high homocysteine in young male schizophrenics. It could be caused by smoking, lack of exercise or by poor diet. Yet via a biochemical mechanism it could negatively affect the course of illness. Keywords: homocysteine, neurodegeneration, schizophrenia Nature 387:598-603 [1997]). This duality, which has been attributed to changes in intracellular C1-concentration, is due to changes in chloride regulation (Wagner et al. J. Physiol. 537: 853-869 [2001]). We investigated the role of the GABA transporter, GAT1, in regulating chloride concentration in SCN neurons. This transporter, which in the SCN is located on axon terminals and glia, as well as on some dendrites, translocates GABA along with chloride and sodium. We studied the effect of the GAT1 inhibitor SKF-100303A on the decay time of GABA currents, induced by either inhibitory synaptlc activity or by local application of GABA. The blocker caused prolongation of tile decay time constant by an average of35%, and had a larger effect on inward (39%) than on outward (31%) currents. We next investigated the effect of the blocker on the recovery of intracellular CI concentration. Loading or depletion of intracellular CI" was induced by a prolonged GABA application, which generated either influx or efflux of chloride, depending on the membrane holding potential.
Recovery was deduced from the change in chloride reversal potential calculated from the response to a test GABA pulse presented at 30sec delay after the first GABA pulse. Wh!le recovery from chloride loading was nearly unaffected by the blocker, recovery from chloride depletion was reduced, particularly during subjective day. Understanding the functions of a single neuron, such as, propagation and generation of synaptic or action potentials, a detailed description of the kinetics and distribution of the underlying iomc conductances is essential. In voltage-clamp experiments, incomplete space c!a.mp distorts the recorded currents, rendering accurate analyss mpossible. We developed a simple numerical algorithm that corrected such distortions. The method performs a stepwise approximation of the conductance density at the site of a local voltage clamp. This is achieved by estimating membrane conductances in a simulation that yields simulated clamp currents, which are then fitted to the distorted recordings from the non-space-clamped structure, relying on accurately reconstructed cell morphology" and experimentally determined passive properties. The algorithm was tested using channels with simplified kinetics and on published realistic potassium-channel models. For all tested conditions, the methrd enabled accurate retrieval of the local densities and kinetics of somatic and dendritic channels at the site of the voltage clamp, rather than averaging these parameters across the structure. Furthermore, correct calculation of conductance gradients required only a few recordings. Neither the addition of noise nor variation of assive parameters significantly reduced the performance of e correction algorithm. The correction method was applied to two-electrode voltage-clamp recordings of K currents fom the apical dendrite of layer 5 neocorticalpyramidal neurons. The generality and robustness of the algorithm make it a valuable tool for voltage-clamp analysis of voltage-gated currents in structures of any morphology that is amenable to the voltage-clamp technique. Key"words: voltage-Clamp, potassium channels, cable-theory The representation of objects in the human lateral occipital complex Schiff M. and Zohary E..
of Neurobiology, Hebrew University, derusalem 91904 Regions in the lateral and ventral occipital cortex, known as LOC, play an important role in human object recognition. LOC is characterized by a stronger response to objects compared to scrambled objects or textures, regardless of the visual cues used to define object shape (luminance, texture or motion differences). Can this cue invariance be observed at the level of single neurons, or are different neuronal populations within LOC sensitive to different object cues. To test this we utilized the fact that LOC shows fMRI-adaptation, a decreased activation in response to repeated stimulus presentations. Our subjects watched two types of three-dimensional objects: (1) stattonary, in which object sha_pe was defmed byits contours, shading, etc, and (2) structure from motion (SFM) stimuli, in which the perception of an object is induced solely from motion cues. Every SFM object was made of 18 different frames, depicting the rotating object as it is seen from sequential viewpoints. The object was unidentifiable from any single frame. Both .types of sttmuli activated LOC, and in both cases fMRI-adaptaton was seen following stimulus repetition. Since there is little physical similarity between successive SFM images, adaptation during the SFM repetition indicates that LOC is involved with holistic object identity. However, when stationary pictures and SFMs of the same object were alternated within a block, no adaptation was apparent. This may imply that while on the macroscopic level LOC is indeed processing object information in a cue invariant manner, separate neuronal groups within LOC process different object cues.
Keywords: LOC, structure from motion, adaptation In the present study, we tested whether excitotoxic lesion of BLA and orbitofrontal cortex (OFC), which are reciprocally connected, will induce LI perseveration, which would become evident under conditions that do not allow for LI in controls. LI was measured in a thirst motivated conditioned emotional response procedure by comparing suppression of drinking in response to a tone in rats which received 0 (nonpre-exposed) or 40 tone presentations (pre-exposed) followed by either 2 or 5 tone-sh6ck pairings.
Control rats showed LI with 40 pre-exposures and 2 conditioning trials, but raising the number of conditioning trials to five disrupted LI. In contrast, both OFCand BLA-lesioned rats persisted in exhibiting LI under the latter conditions. PFC and amygdalar pathology has been extensively reported to be an essentml feature of sthizophrenia, thus, our results that OFC and BLA damage lead to persistent rather then disrupted LI, indicate that persistent LI provides an animal model of such Alterations in the opioid system in the hippocampus as well as somepossible functional consequences were ixivestigated by several, methods in adult male rats who were prenatally exposed to either saline-or morphine (10 mg/kg twice daily on gestational days 11-18). In situ hybridization and Northern blots were used to measure proenlephalin and prodynorphin mRNA, and radioimmunoassays were used to quantify proenkephalin-andprodynorphin-derived peptide levels in the dentate gyrus (DG), CA3, and CA1 subfields of the hippocampus. Prenatal morphine exposure in male rats decreases proenkephalin and increases prodynorphin mRNA selectively in the granule cell layer of the DG. Similarly, met-enkephalin peptide levels are decreased and dynorphin B peptide levels are Increased in the DG but not CA3 or CA of prenatally morphine-exposed males. In addition, there are decreases in some dynorphin-derived peptides in the CA3 subfield. Receptor autoradiographv revealed increases in the density of Is but not 5 receptor ldbeling in discrete strata of specific hippocampal subfields in prenatally morphine-exposed males. Prenatal morphine exposure, however, does not alter the latency to onset or number of episodes of wet dog shakes r clonic ,seizures duced by infusion of 10 nmol [D-Ala, MePhe , Gly-oF] enkephalin (DAMGO) into the ventral hippocampus. Interestingly, a naloxone (5 mg/kg) injection 30 min before bicuculline reverses the increased latency to onset of clonic and tonic-clonic seizures in prenatally morphine-exposed males. Thus, the present study suggests that prenatal morphine exposure alters the hippocampal opioid system, suggesting possible alterations in the excitability of the hippocampus. experience-induced changes of synaptic efficacy may be alterations of synaptic shape. In the present study we searched for differences in the fine morphological parameters of dendritic spines on the basal dendrites of Lucifer Yellow-injected layer III pyramidal neurons in the ACd of parentally deprived (3xl hour per day between postnatal days and 21), 45 days old degus in comparison with age-matched socially undisturbed control pups. The 3D-measurements of spines were performed in image stacks produced by a confocal laser-scanning microscope applying a newly developed image analysis software (Herzog et al., SPIE 2984:146-158, 1997. Although the spine volumes remained unchanged, a signiOcant reduction of spine length, accompanied by a significant increase of mean spine diameter was observed in the parentally deprived animals. These overall effects were due to a reduction of spine length on dendritic branch 2 and 4, and a significant increase of spine diameter on branch 3. This is the first evidence that the disturbance of the parent-child contact during an early postnatal phase results in lasting changes of distinct shape parameters of presumed excitato spine synapses, which most likely reflect a experience-related fnie-tunmg of synaptic strength within limbic cortical circuits. Recent studies in our laboratory showed that glutamate evokes T cell-mediated protective mechanisms,. The aim of the present study was to examine the nature of the glutamate receptors and signaling pathways that participate in immune protection against glutamate toxicity. We show, using the mouse visual system, that glutamate-induced toxicity is strain dependent, not only with respect to the amount of neuronal loss t causes, but also in the pathways it activates. In strains that are genetically endowed with the ability to manifest a T cell-dependent neuroprotective response to glutamate insult, neuronal losses due to glutamate toxicity were relatively small, and treatment with NMDA-receptor antagonist worsened the outcome of exposure to glutamate. In contrast, in mice devoid of T cell-dependent endogenous protection, NMDA receptor antagonist reduced the glutamate-induced neuronal loss. In all strains, blockage of the AMPA/KA receptor was beneficial. The results suggest that glutamate-induced toxicity involves multiple glutamate receptors and that the types and relat!ve contributions of receptors vary among strains. We suggest that a multi-factorial protection, based on a mechanism independent of the specific pathway through which glutamate exerts its toxicitv, is likely to be a safer, more comprehensive, and hence more dffective strategy for neuroprotection. Carolina, USA Changes in the cholinergic system in the brain were one of the first neurochemical alterations discovered in depressive disorders. Since then, alterations in many more neurotransmitters were recognized. Using the Flinders sensitive Line (FSL) rats as a model of depressipn, we have found major changes in the neuronal monoalmnerglc systems. Recently, we discovered that specific and local molecular changes relate to the dynamics of depression onset in these rats. It was surprising to find that molecular changes in the limbic system were at terminal sites of the monoaminergic system and not in the cell body regions. This finding suggested that interneurons are involved in the pathology of depression and may serve as the common denominator in neurotransmitter alterations that are involved in the disease onset. The nucleus accumbens (Nacc) is likely to have a role in depression and contains cholinergic interneurons, which may mediate lasting behavioral changesby regulating monoamine release. To date, there has been no Stemattc investigation of the role of the cholinergic system in Nacc in animal models of depression. Acute (45 min) infusion of 5 lam pirenzepine, a muscarinic-1 receptor anta$onist, into the Nacc by a microdialysis probe returned the FSL s increased immobility in the swim test to the level of controls. Pirenze.pine in FSL rats also normalized the basal levels of dopamme and serotonin in the extracellular space in the Nacc. These results may suggest that the cholinergic system is not just another neurotransmltter altered in depression, but is critical for controlling the interaction between several neurotransmitter systems. The convergence of these systems trigger the behavioral alterations foundin depression.
Keywords: cholinergic svstemo nucleus accumbens, muscarinic receptor, dep:ession The same reduction has been found in yeast where it was also observed that unlike lithium, that causes a parallel increase in intracellular inositol-l-phosphate, valproate causes a decrease in this metabolite. This suggested to us that myo-inositol-l-P (MIP) synthase is the site of valproate's action in inositol metabolism. MIP synthase is the rate-limiting step in inositol biosynthesis and in yeast is highly regulated in response to inost"ol. We therefore hypothesized that human brain M1P synthase is a factor in the psychopharmacology of mood stabilizers. It has been previously shown in psychophysical studies that detection threshold ofa Gabor patch, presented at the fixation point is reduced when flanked by co-aligned high contrast alike'stimuli. In the periphery, increased nhibition is found.
However physiological studies in V of cats and monkeys do show increased firing rates for co-aligned stimuli at 4+.
Here we attempt to resolve this apparent conflict. We show that the absence ofperipheral facilitation is not a result of improper scaling. We observed that collinear facilitation was reduced or gone as soon as targets were placed outside fixation (1+), still within the fovea. This suggested the involvement of attentional mechanisms in peripheral facilitation. It has been previously shown that lateral interactions are modulated bv attention in the fovea. We assessed the role of attention in pel:ipheral surround modulation by directing attention to the flankers in a double task paradigm. Subjects performed a Vernier acuity task on the flankers and a detection task on a Gabor target located in between. Collinear thresholds were lower than orthogonal ones in the double task. When performing only the detection task, collinear and orthogonal thresholds were" similar. This result shows that under conditions that force attention to critical parts of the stimulus, facilitation is present at 4+ and further suggests that differences in allocation of attention between fovea and eriphery may underlie the lack of peripheral collinear cilitation. The orientation selectivity of this attention-induced facilitation may imply that it is mediated by lateral interactions, as suggested for foveal facilitation. Keywords: attention, lateral interactions, periphery We are interested in the molecular mechanisms of olfactory rule learning in the piriform cortex. We have shown previously that post-burst atter-hyperpolarization (AHP) and paired-pulse facihtation (PPF) is reduced in pamidal neurons, days after olfactory rule learning. Assuming that these long-term cellular modifications subserve odor rule learning, we are looking into their possible molecular mechanisms. (See also Cohen et al). PKC has been implicated in the process of learning and memory formation. Here we study the involvement of PKC in long-tei'm AHP modulation following odor rule learning. The specific PKC inhibitor GF109203x (10 M) caused an increase in AHP amplitude in neurons from trained rats. Consequently, the difference in AHP amplitude between trained and control rats was diminished. Moreover, activation of PKC by OAG (10 ,tM) significantly reduced the AHP in neurons from naive and pseudotrained rats, but not in neurons from trained rats, abolishing the difference between groups. Using sub-cellular fractionaton and quantitative immunoblotting analysis, we are examining the role of the different PKC isoenzymes in rule learning in the piriform cortex. Phosphorylation of PKC/[3II on Thr638/641 was decreased by 15% n the trained group compared with pseudotrained and naive groups (n=7). In the cytosolic fraction, phosphorylation of PKCct/13II on Thr638/641 was reduced in the trained group by 23 % compared to controls (n=7). We conclude that PKCt/15II It is well known that subjects can detect rapidly an element that differs greatly from surrounding elements in a single dimension such as color or orientation. With attention spread across the entire array, subiects detect presence or absence of such a target with a respbnse time that does not depend on the number of distractor elements. We now asked what will be the speed and accuracy of detecting 2 such elements simultaneously. Subjects viewed a briefly presented 8x8 array of pink lines oriented at 55 (or 60) followed by a masking stimulus after a variable Stimulus-to-mask Onset Asynchrony. On some trials, or 2 of the elements were replaced by a pale green line of the 3 r same orientation, a pink lne of orientation 35-40 (or 0 '), o a line with both these changes. Subjects reported the number of odd lines, and their nature. Surprisingly, we found that subjects were more accurate at detecting targets in arrays with 2 odd elements than arrays with one despite the requirement of accurately reporting the nature of the change. This result was true for all types ofodd elements. In addition, we found that it was easier to report presence of two odd elements one with an odd color and one with an odd orientation than to report the presence of one odd element which differed from the distractor elements both in color and orientation. Analysis of these results as a function of distance between the odd elements suggests that oddity is detected as a Gestalt and that arrays with a paxr of targets are perceived as distinct structures. However, SK3 has not been mapped in mouse brain. The present study documents SK3 distribution in brain of BALB/c mice. At the forebrain level, SK3 staining was documented in thalamic ventromedial complex and laterodorsal lamina, in the medial habenula, and in the central and corticomedial mNYCgdala.
In the mesencephalon, there was staining in the and VTA and in superficial lamina of the superior colliculus. At the pontine level there was staining in the locus coeruleus and parabrachial nucleus. All staining patterns were blocked by pre-incubation of the antibo.dy with the antigen. The SK3 disixibution in mice documented m thepresent study parallels the distribution reported in rats, with the additional presence of SK3 in the amygdala and ventral thalamus. Thus, the present study supports the possibility that SK3 in ventral mesence.phalic dopaminergic neurons is ubiquitous in mammahan species and that this localization could be preserved in humans. SK3 distribution in mice suggests possible involvement of SK3 in pathways that modulate sensory information processing. The presence of SK3 in the centralamygdala nucleus may suggest that in mice, SK3 also participates in neural pathways for integration of sensory stimuliin visceral-emotional reactions.
Keywords: SK3, calcium, potassium, channels A system for rapid uncaging in neural tissue using spatially defined patterns Shoham S. and Wang S. S.-H. oolecular Biology, Princeton University, USA Two-photon laser scanning microscopy has enabled fast high-resolution imaging deep into brain tissue, which is strongly light scattenng. The ongoing development of functanal fluorescent probes and high-throughput techniques for dye loading promise an increasing range of applications of multphoton .optical methods in neuroscience.
The use of hf, ht-sensitive "caged" compounds complements two-photon rmcroscopy by providing a means of optically mampulating biochermcal signals. We are developing a flexible new system that allows controlled photo-release of caged neurotransmitters in defined spatial and temporal patterns With submicron and submillisecondresolution. Our system relies on the steering of an ultraviolet laser beam in two dimensions using acousto-optical devices made from TeO.. The ultraviolet light pattern is projected into the same plane scanned by a custom-made two-photon microscope. The beam can be steered to up to 100,000 locations per second. We are currently beginning to apply the system to neural systems.
1Dept. of Pharracology, School of tharmacy and "Pediatric Intensive Care Unit, Hqdassah Medical Center, The Hebrew University, Jerusalem, Neurobiochemistry, Tel Aviv University, Neurosurgery, Sheba Hospital, and Lawerence Berkeley National Laboratory, Berkeley CA, USA Ras proteins are involved in receptor-mediated signaling pathways, including those regulating cell death and survival. Ras antagonists may therefore be protective when these pathways are activated. S-trans, transfamesylthiosalicylic acid (FTS), a Ras inhibitor, acts primarily on the active, GTP-bound, form of Ras. This study investigates whether: 1) closed head injury, (CHI) activates Ras; 2) FTS interferes with Ras activation; 3) FTS affects functional outcome, NMDA receptor function and lesion volume after ,CHI. Mice were subjected to CHI or sham operation, treated with vehicle or FTS (5mg(kg), and sacrificed2h, 24h or 7d after CHI. Total Ras, Ras-GTP, ERK and Ras-dependent active phospho-ERK 115 were determined using specific Ab. Neurological deficits were assessed by a Neurological Severity Score (NSS) at h-7 d post CHI. Recovery was defined by ANSS(t)=NSS(lh)-NSS(t).
NMDA receptor binding and lesion volume were measured by autoradiography and morphom_etry of brain sections, respectively. Ras-GTP was significantly increased in the contused hemisphere at 2h post-CHI and this increase was inhibited by FTS and by the NMDA antagonist MK-801. Total Ras was not affected and Ras-GTP levels returned to baseline by 24h post CHI. Both drugs reduced phospho-ERK levels after CHI. ANSS and neurological deficits were significantly improved (p<0.0001, 60%) by FTS, which also prevented the CHI-induced reduction in NMDA receptor binding in cortical, striatal and hippocampal regions and produced a trend towards smaller lesions. We propose that Ras activation after CHI contributes to neuronal cell death and its suppression by FTS has a long-lasting beneficial effect.
Keywords: traumatic brain injury, Ras-GTP, S-trans transfamesyl.thiosalicylic acid (FTS), NMDA receptors, neuroprotectaon Activation of Matrix metalloproteinases by TNF-cx in astrocytes in, vitro: a model for brain injury Shoshany Y.', Shohami E. , Brenner T. and Reich R.' Dept. ofPharmacoloev, Hebrew University of Jerusalem: :Dept. of Neurology, trladassah University Hospital, Jerusalem Following head trauma several processes take place including inflammation, neurotransmitter release and cell death. Cytokine action followed by specific gene's activation play a key role in the pathophysiology of brain injury. Among the earliest events (hours) is the disruption of the blood-brain barrier (BBB). Matrix metalloproteinases (MMP) are involved in the disruption of the blood-brain barrier as well in the repair process, taking place in the injured brain. TNFx, .a cytokine upregulated wtln h after trauma was shown to play a major role in the processes leading to BBB disruption. In experimental model of closed head injury, administration of dexanabinol prevented TNFt mediated BBB breaching. In the present study, we undertook to investigate the role of TNFc in reduction of MMP secretion and dexanabinol effect in astroc)es in culture, as a model of brain damage.
Our results indicate that TNF, dose de.pendently, induces the secretion of MMP-2 and MMP-9 in astrocytes culture. Furthermore, dexanabinol prevented the elevation of both MMPs. RT-PCR experiments indicate that the site of action of dexanabinol is at the transcriptional level. Stimulation of astrocytes with LPS led to elevation of TNFx activity that was inhibited by dexanabinol. The inhibitory effect of the drug was on the release of TNFx from mem'branal site by TACE.
RT-PCR analysis showed no significant effect on TNF mRNA whereas TACE mRNA levels were reduced significantly. To our knowledge, this is the first demonstration of a direct'effect of dexanabinol on TNFt production/release via inhibition of TACE production in astrocytes. Keywords: Tumor necrosis factor (TNFx), matrix metalloproteinases (MMP), dexanabinol, cultured astroc.vtes, blood-brain barrier Synaptic dynamics determine the functional correlations between neocorticai neprons in excited slices lberber G. , Wu C.Z." and Markram H. of Neurobiology, Weizmann Insttmte of Science, Rehovot Stereotypical microcircuits have long been recognized as computational modules of neocortical function, but the principles that determine the emergence of activit3, patterns within these microcircuits are tmknoaa. We obtained multiple simultaneous whole-cell voltage recordings from neurons in neocortical slices. A cross-correlation analysis of subthreshold membrane voltage between different types "of neurons revealed the emergence of characteristic 'nctional relationships between neurons when the slice was activated bv an excitant extracellular solution. The functional relations'hip between neuronal .pairs was correlated to the nature of the excitatory, synaptic nput to these neurons, specifically to the governing synaptic dynamics. In simulations, differences in cross-correlation delays emerged when the excitatory, discharge was patterned and deviated from purely random spike activi;.
In this study we show that synaptic dynamics determine the relative timing of activit3' of different neurons and thereby impose stereotypical response characteristics in the neocorticl microcircuit as"a function of the excitatoD" activit3. pattern.
Keyw.ords: neocortical microcircuits, svnaptic dynamics, multineuron recordings, subthreshold crrelation's Perception of happy and sad facial expressions in chronic schizophrenia: Evidence for two evaluative svstems Silver H. 'z, Shlomo N.s, Travis T. and Gur R. 4" tSha 'ar Menashe MHC and Technion, 3Mazra MHC; University of Pennsylvania Background." Schizophrenia patients have impaired perception of emotional expressions, but it is not clear whether this is part of a generalized deficit in cognitive function. Aim." To test for existence of emotion-specific deficits by studying the effects of valence on reeogniuon of facial emotional expressions. Method: 24 male subjects suffering from chronic schizophrenia were examined with two tests of perception of emotion: the Penn Emotion Acuity Test (PEAT40) and the Emotion Differentiation Task (EMODIFF). Clinical state was assessed with the SANS and SAPS scales, visual with motor the Benton Visual Retention Test (BVRT) and memtction with the finger tapping test. Results: Identification of happy facial expressions showed significant negative correlation with age, cumulated time in hospital and length of current hospitalization; positive correlations were found with visual retention and finger tapping scores. Identification of sad facial expressions showed significant correlation only with cumulated time in hospital while identification of neutral facial expressions showed no significant correlations. Discrimination between degrees of happy but not sad facial expression, showed a positive correlation with negative symptoms.
Conclusion" Perception of happy and sad emotion relates differently to significant illness parameters. This differentiability supports the existence of an emotion-specific deficit in perception of emotions in schizophrenia and of separate channels for processing positive and negative emotions.
Keywords: emotions, schizophrenia, valence, cognitive Non-auditory brain structures involved in processing auditory linguistic material Sinai A. and Pratt H.
Evoked Potentials Laboratory, Technion-Israel Institute of Technology, Haifa 32000 The purpose of the study was to localize non-auditory cortical areas activated during first and second language processing.
Brain potentials were recorded from 17 normaIhearing, right handed, native Hebrew (first language) speakers that also speak English (second language). Subjects performed a lexical decision task with speech stimuli from both languages. Brain sources of activity were estimated by calculating current densities from th scalp distribution "of the ERPs, using low-resolution electromagnetic tomography analysis (LORETA). Current densities in brain voxels (cubicles f 7 mm spatial resolution) were compared with pre stimulus baseline by non-parametric t-tests adjusted for multiple comparisons. The time course of activity in each Brodman area (BA) was plotted separately for the right and left hemisphere.
Brain reions outside the traditional auditory and language-related regions demonstrated different temporal patterns of activity that varied with the type of stimulus. These regions included, among others, frontal lobe motor regions pre-frontal associative region cingulate gyrus and pre-cuneus. The structure-specific spatio-temporal pattern of activity is compatible with the idea that language processing occurs in spread out neural networks involved with processing different aspects of language. Keywords: hearing language functional imaing Event-Related Potentials The neural representation of odor pleasantness and intensity in the human brain Sobel N. and Anderson K.N.
Helen Wills Neuroscience Institute, l]niversity ofCalifornia at Berkeley, Berkeley, CA, 94720 USA Imaging studies have shown increased amygdaloid complex (AC) activation in response to aversive odors. Odor quality is dependent on odor concentration (intensity) and odorhedonics (pleasantness/unpleasantness). Here we dissociate these dimensions, and ask whether AC activity reflects odor hedonics intensity, or both. An olfactometer generated 5 stimuli; !) PH pleasant high intensity, 2) PL pleasant low intensity, 3) UH unpleasant high intensity, 4) UL unpleasant low intensity, 5) clean air. 16 young healthy subjects participated in an event-related fMRI study at 3T (T2* spiral, TE 30, TR ls, 64x64 FOV, 17 slices, slice thickness 4mm, stimulus IS 20s, stimulus repetition 3(/). A first-pass ana.lysis using SPMgP to compare for regions of increased acuvity related to odor intensity regardless of pleasantness revealed a pronounced locus of activity in the AC (P<0.001). In contrast, an analysis comparing for increased activity related to the unpleasantness of an odorant regardless of intensiW, revealed a locus of activity in the orbitofrontal gyri (P<0.001). In-depth ROI analysis corroborated these findings, demonstrating an increase in signal change in the AC in response to ncreases in intensity, not unpleasantness, of an odor. Activity in the AC was significantly correlated with individual intensity estimates (r=0.35, p=0.001) but not with pleasantness estimates (r=0.04, p=0.7). These findings suggest that the AC may be encoding more of the immediate physical dimensions of a stimulus (intensity) rather than the later SyChological dimensions" ofthe stimulus (pleasantness). ywords: olfaction; fMRI; amygdala The molecular neurobiology basis of physiological stress responses Soreq H.
Institute ofLife Sciences, The Hebrew University of Jerusalem Physiological symptoms reminiscent of post-traumatic stress disorder (PTSD) occur following exposure to anticholinesterases, however, the origin ofthis phenomenon is unknown. Recently, we found a dominant activating polymorphism in the ACHE promoter, carried by over 5.0% of the Israeli and compared with 0.5% o.f the North American population. This polymorphism s associated with acetylcholinesterase (ACHE) overproduction and hypersensitivity to cholinergic agonists and antagonists (Shapira et al., Hum. Mol. Genetics, 9:1273-1281[2000). Its consequences could be attributed to AChE's hydrolytic activity, the complexity of ACHE gene regulation and/or the "non-classical" AChE activities (Soreq & Seidman, Nature Neurosci. Rev. 2:294-302. [2001]). There are three 3' splicing options for AChE mRNA, and 3 corresponding AChE vai,ants with different C-terminal peptides, multimenc assembl) and non-classical function(s). Of these, the normally rare "readthrough" variant, AChE-R, accumulates following stressful stamuli and under acetylcholinesterases (Kaufer et al., Nature 393:373-377 [1998]) or under neuromuscular diseases (e.g. myasthenia gravis) (Brenner et al., FASEB J., in press [2002]). The AChE-R mRNA, usually located in neuronal cell bodies, translocates to dendritic processes following stress.
This rapid yet long-lasting change may suppress the initial insult, but leads to adverse consequences under long-term conditions (Meshorer et al., Science 295:508-512 [2002]).
AChE-R involvement in these adverse symptoms was proven by using partially 2'-oxymethylated oligonucleotides inducing AChE-R mRNA destruction (Shoham et al., J. Mol. Med. 78:228-236 [2000]). The variant selectivity of these antisense agents, currently tested in clinical trials, prevents the stress-induced physiological impairments while protecting cholinergic neurotransmssion by maintaining the synaptic ACHE. Keywords: stress, pharmacogenomics, mRNA translocation, acetylcholinesterase On line confocai imaging of processes underlying the dedifferentiation of an axonai segment into a motile growth cone after axotomy Spira M.E., Erez H., Khoutorsky A. and Prager-Khoutorsky M.
Dept. of Neurobiology, Life Sciences Institute, The Hebrew University of Jerusalem, Jerusalem Regeneration of neuron after mechanical injury requires structural and functional dedifferentiation of the cut axonal end into a motile growth cone (GC). Here we analyzed the events leading to the formation of a growth cone (GC) and examined the mechanisms that set the dedifferentiation process into motion.
To that end we fluorescentlv labeled actin by chimeric EGFP-actin, microtubules (lVfTs) by tetrameth371rhodamine conjugated tubulin, and vesicles by the styril dye RH-237, in primary cultures of identified Aplysia neurons. Confocal imaging revealed that: (a) axotomy triggers a wave of MTs depolymenzation. The depolymerizing wave stops at a point 100-150 m from the cut axonal end, forming a transition zone (TZ) between the depolymerized segment and a proximal segment in which the MTs maintain normal structure. (b) In parallel, actin reach adhesion plaques disappear only to reform minutes later. (c) Anterogradlly transported vesicles are trapped within the TZ. Thereafter, (d) actin bundles assemble along the axons perimeters surrounding the vesicles trap. Once the above-described structure is formed an actin reach GC lamellipodium, supported y radial polymerization of MTs, begins to extend. Axotomy in the presence of nacodasol prevents the restructurin,g of the depol me zed,...MTs and the. tra pin of vesicles, but does not a/fYect e ,n,t,al extens,on oPa C's lamellipodium. In jusplakinolid the MTs based vesicles trap is formed, but the extension of a GC's lamellipodium is blocked. Calpeptin totally inhibits the restructuring oi"the axon (Oren et al., ths meeting). We conclude that a common calpeptin sensitive event triggers the dedifferentiation process. Keywords: axotomy, growth cone, regeneration, calpain, mierotubules Color adaptation models and their predictions to color induction effects, color constancy and color contrast Spitzer H. and Barkan Y.
Dept. of Biomedical Engineering, Faculty of Engineering, Tel Aviv University, Tel Aviv A new biological model and algorithm for color adaptations, which includes the color constancy (first order) and color contrast modulation (second order), is presented. (Color constancy (CC) is a psyeh0physieal phenomenon in which a system has a partial capability to discount the chromaticity of the illumination). The goal of the model was to achieve computational psychophysical predictions to different color appearance effects such as color constancy, color contrast and different induction effects. The model predicts the above human visual performance, including the modulation of perceived color due to surrounding color. It predicts also the dual effects of enhancement and the suppression of the central perceived contrats due to the relation between the central and surround contrasts. The model is based on the properties of retinal ganglion cells (opponent cells) and cortical eells (double opponent cells), as well as on chromatic adaptation mechanisms in these double opponent color-coded cells: remote chromatic adaptation. The suggested color adaptation mechanisms are mo,deled as gain control mechanisms based on the "curve-shifting' effect. This effect is the transition from one response curve to another, due to a change in the light intensity (or color, or contrast) of the local receptive field and its remote area, in order to obtain a higher gain in the new color or contrast. The simulations calculated also on real images. The results indicate that the contribution of adaptation meclaanisms to color constancy and color contrast are significant, robust, and enables color enhancement, color contrast enhancement and color constancy of still and video images. Keywords: visual system, color vision, adaptation, remote effects Omega-3 fatty acid treatment of depressive breakthrough during unipolar maintenance Stahl Z., Nemets B. and Belmaker RH.
Ministry of Health Mental Health Center, Faculty of Health ,Sciences, Ben Gurion University of the Negev, Bee-Sheva Objective: Studies have reported that countries with high intke of fish oil have low rates of depression. We studieda specific omega-3 fatty acid, the ethyl ester of eicosapentaenoic acid (E-EPA) as an adjunct to antidepressant treatment for breakthrough depression in recurrent unipolar patients on maintenance therapy. Methods: Design was four-week parallel group double-blind add-on to ongoing antidepressant therapy. Twenty patients participated, seventeen females and three males, all with diagnosis ot current major depression. Results: Highly significant benefits were found by week 3 of t for eicosapentaenoic acid compared to placebo.

Conclusions:
It is not possible to distinguish whether eicosapentaenoic acid augments antidepressant action in the manner of lithium or has independent antidepressant properties of its own. Keywords: omega-3, depression, antidepressant Gustatory thalamus: Where does it connect to? Stehbere,_J. and Dudai Y.
Dept. of Neurobiology, Weizmann Institute of Science, Rehovot 76100 The role of the gustatory thalamus in gustatory function has been controversial, due mainly to the fact that unlike other senses, gustatorv projections from the parabrachial nucleus (PBN) go eithefdirectly to the cortical taste area or travel via the thalamus. A combination of retrograde (fluorogold) and anterograde (BDA and FDA) neuronal tracers was used to map the afferents and efferents of the gustatory thalamus Ventroposteromedial parvicellular nucleus. Anterograde and retrograde tracers were injected into the insular cortex and PBN. We found that the PBN afferents to the thalamus do not colocalize with CGRP and that the CGRP axons have a non-overlapping more ventral localization from the PBN axons. Thalamlc neurons projecting to the cortex localize 117 indistinctively along the nucleus, covering areas that receive "_input from the PBN, CGRP and areas that do not receive input from neither of the above areas. Corticothalamic projections also do not show a clear mapping, terminals being found all over the nucleus. We were also able to determine the cellular location of the PBN synaptic terminals in the thalamic neurons. In search for a functional topographic map of the thalamus, we combine thalamic retrograde labeling from the cortex and pCREB immunohistochemistry as an activity marker. We test whether there is a distinctive activation of the thalamocortical Ojections when the rat drinks novel taste vs. a familiar one. orted by the Minerva Foundation and the Human Frontiers Science Program Organization. Keywords: thalamus, taste, cortex, pCREB, tracers.
Role of GIy in the regulation of adenylyl cyclase activity following acute and chronic receptor activation Steiner D., Schallmach E., Butovsky E., Nevo I., Saya D. and Vogel Z.
Dept. of Neurobiology, Weizmann Institute of Science, 76100 Rehovot Acute activation of opiate (and other Gi/o) receptors inhibits adenylyl cyclase (AC) activity, while prolonged activation leads to an increase in AC activity, known as AC sensitization or superactivation. This phenomenon has been proposed to play a role in opiate addiction and has been shown for AC eS I, V, VI and VIII, but not for AC-II, IV or VII, onstrating that superactivation is isozyme-specific. To investigate the roles of G and Gi in AC regulation, we produced several point mutations in AC-V and AC-I. We found that a mutation in the Cx region of AC-V (F481 Y) and of AC-I (F314Y) led to reduced inhibition by acute agonist treatment and to the loss of chronic agonist induced superactivation, and in parallel to loss of AC sensitivity to G dimers and to constitutively active G, (demonstrating that this amino acid in C plays an important role in the regulation of AC activity and its interaction with G protein subumts. To distinguish between the roles of Gi and G, three other mutations in C region (shown previously to reduce affinity of Gi to AC) were prepared. Indeed the activity of these mutants was not inhibited ty constitutively active Gai (compared with 70% inhibition of AC-V wild-type). On the other hand, these mutants behaved identically to wild-type AC-V with respect to G stimulation and 13y inhibition and showed normal opiate acute inhibition and chronic induced superactivation. These results suggest that the inhib.itio.n of AC by GIv, and not by Gi, is essential for the superactivaton process. Supported by NIDA and the US-Israel Binational Science foundation.
Central pathways between sacrocaudal afferents and locomotor circmts in the spinal cord of the newborn rat Strauss I. and Lev-Tov A.
Dept. of Anatomy & Cell Biology, The Hebrew University School of Medicine, Jerusalem The central pathways mediating sacrocaudal afferent (SCA) activation oflocomotor circuits in the lumbar spinal cord were studied in isolated spinal cords of neonatal rats. The lumbar rhythm induced by SCA stimulation was abolished after blocking synaptic transmission in the sacrococcygeal (SC) cord and restored when the synaptic block of specific SC segments was temporarily alleviated. Thus, synaptic activation of SC relay neurons is required for generation of locomotor activity by SCA stimulation. The transmission across these relays required activation of non-NMDA excitatory amino acid receptors, and it was not impaired in the presence of specific antagonists of NMDA O-methyltransferase (COMT) gene that encodes a low enzyme activity variant. While this finding has been replicated by one group, others have shown discrepant findings which may be due to different diagnostic criteria used to assess violent behavior, or, considering the complexity of asgressive behavior, either type and/or type II statistical errors. Consequently, additional studies are needed. In this study, we assess 122 patients with schizophrenia for violent behavior using the Lfetime History of Aggression (LHA) scale, an 11-item questionnaire that is subdivided into Aggression, Self Directed Aggression, and Consequences/Antisocial Behavior subscales. DNA was genotyped for the COMT 158 polymorphism, as well as a functional polymorphism in the monoamine oxidase A gene (MAOA) promoter. Similar to previous findings reported bv our group, a statistically significant association was foutd between violent behavior in schizophrenia and the COMT 158 polymorphism. Mean LHA scores were higher in subjects homozygous for the low enzyme activity COMT variant, 158Met (p-0.005). Analysis ofthe major LHA subscales revealed that the association with 158Met was due to high scores on the Aggression, and Self Directed Aggression subscales, but not the Consequences/Antisocial Behavior subscale. No significant association was detected for the MAOA gene alone. These observations further suggest that COMT is a modifying gene that plays a role in determining interindividual variabditv in the proclivity for outward and self directed violent behavior found in a subgroup of schizophrenic patients.
Keywords: schizophrenia, aggression, COMT, polymorphism In addition, studies have suggested many neurotransmitters and neuromodulators as candidate systems involved in depression.
Neurotransmission changes in the brain should be monitored dynamically and the cross-talk between these neuronal systems should be at multiparametrical levels: neurophysiofogical.
neurochemical and neurogenetic. The present review will discusses recent findings concerning alterations in the neurobiology of the brain that accompany depressive behavior and effective antidepressant treatment in h unique animal model of depression (Flinders Sensitive Line of rats We studied how monkeys hold several familiar stimuli in working memorv. Two Macaque monkeys were trained on a modified delayet-match-to-sample task with multiple samples.
Monkeys viewed a sequence of stimuli on a computer monitor and released a lever when any one of the sample stimuli in the sequence was presented for a" second time within the trial i.e. a "'match" stimulus. Trials had 2-8 stimuli with 1,2 or 3s inter-stimulus intervals. Two .type of errors occurred: 1. "Misses" when the monke;, failed to respond to a "'match" presentation. Distribution ot" these errors depended on the position of the "'cue'" (first presentation of the stimulus which was later matched). The greater the cue-match separation, the poorer the performance. This finding indicates that working .memory.of a given item erodes with time and/gr the number of mtervemng stimuli. However, for a given cue-match separation, performance improved with sequence length, i.e. with number of irrelevant stimuli preceding the cue. This may reflect the rising expectation of reward. 2. "False positives'-when the monkey erronouslv responded to an image which was not a repetition of a 0revic;uslv presented stimulus within the trial (a "'non-match'" sthnulus). Distribution of these errors depended on the time and number of stimuli since the beginning of the trial. Taken together, these tvo findings suggest that noise, associated with the neural dynamics, spontaneously erases items from working memory And induces ghost memories of familiar, unseen stimuli. A dtailed model has corroborated this eywords: working memory Pro-inflammatory brain cytokines, released hours after closed head inj.ury (CI). have deleterious effect. Anti-inflammatory and antl-apoptotlc modalities ameliorate CHI outcome. Erythropoietin (EPO), a kidney-derived cytokine regulates hematopoeisis, acts as a growth factor and apoptotic inhibitor. Cultured brain cells produce .EPO in response to oxidative stress, and EPO receptor is present on neuronal and brain capillary endothelial cells. This study examines the protective effects of EPO in rats and mice undergoing controlled CHI. Methods: CHI was induced using a weight-drop device. Clinical status was evaluated by Neurological Severity Score (NSS), which tests 10 or 17 tasks for mice and rats respectivelv. Animals were treated with 2 doses of i.p. 5,000 umts/kg/dose (1 ml) of HuEPO, or vehicle lh and 24h after CHI. NSS was evaluated at 1, 3 and 7d post CHI, and compared using a two-tailed student t-test. Brains were analyzed for apoptosis at 7d. CHI was of similar severity in both rat groups. Recover), was facilitated in the treated mice starting at 24h and reachmg statistical significance at 7d post CHI (NSS 5.0 vs 6.625 in treated and control mice, respectively, p=0.037). Similar recovery pattern was found in CHI mice. While a considerable neuronal apoptosis and minimal apoptosis of the leukocvtes of the mflammatory infiltrate cells was found at the injui3,' site in control animals, a markedly reduced neuronal apoptosis with increased inflammatory cell apoptosis were found after EPO treatment. We propose that EPO-treated animals recovered faster than controls with less neuronal and more inflammatory cell apoptosis compared to controls, probably through a cell-specific anti-apoptotic mechanism.
Keywords: traumatic brain injury, e,rythropoietin, apoptosis, neuroprotecton Tomosyn is involved in the last stages of the synaptic vesicle cycle izhar 0.1, ra/galili y.l Melamed R. Matti U.2, Rettig 3. a-fi-Xfi-e-r, Dep. Ofeurobiochemistry Wise Faculty ofLiJb Sciences Tel Avtv Untverstty, Tel-Aviv 69978: "Phystologisches Institut, Universitat des S'aarlandes, Homburg/Saar, Germany The process of neurotransmitter release is coordinated by a large number of svnaptic proteins and depends on proper protein-protein intei'actions, m-Tomosyn is a brain-specific, 130kD protein that was identified as a novel binding partner for Syntaxinl. Tomosyn possesses a Synaptobrevin-homology domain and is capable of dissociating Muncl8 from Syntaxinl.
To investigate the role of Tomosvn in catecholamine secretion, we overexpressed Tomosyn in aenal chromaffin cells. Using photolysis of caged-calcium, we studied the effect of Tomosyn on the different kinetic components of exocytosis with membrane capacitance measurements. While overall secretion under overexpression of Tomosyn was not different than control, we observed a pronounced difference in the kinetics of secretion. The exocytotic burst was significantly reduced in Tomosvn-overexpressing cells, indicating a decrease in the numbe? of release-competent vesicles. Furthermore, a detailed examination of secretion rates revealed that under Tomosyn overexpression, higher Ca2]i is required to achieve normal secretion rates. To elucidate the function of the Svnaptobrevin-homology domain, we overexpressed a mutated f6rm of Yomosyn, which lacks this domain (TomosynACC). Under steady-state conditions, deletion of the coiled coil domain slitlatlv reduced the inhibitory effect observed with wild-tvpe "lom'osvn. However, following elevation of [Ca2]i, the in}iibito' effect was abolished altogether. These results suggest that Tomosvn acts at the priming step of the synaptic vesicle cycle. Tom6syn activity might reduce the number of fusion-competent vesicles through its interaction with Syntaxin, bv interfering with the formation of fusion-competent SNARE complexes.
Keywords: tomosyn, syntaxin, chromaffin cell, exocytosis iications of APP processing, PKC and CKS-phosp,horation by rasagiline Yozev-Falach M. , Ami, T. , Bar We have recently reported that the anti-Parkinson drugs rasagiline [N-propargyl-l(R)-aminoindan] regulates the secretion of the neuroprotective-neurotrophic non-amyloidogenic soluble form of the amyloid precursor protein (sAPPer) and initiates cell proliferation in PC-12 and neuroblastoma SH-SY5Y cells. In the present study we provide evidence that protein kinase C (PKC) is critically involved in its APP processing activity, both in vitro and in vivo. Inhibition of PKC by GF109203X (2.5M) attenuated rasagiline induced sAPPer release in PC 12 and SH-SY5Y cells. Indeed, rasagiline (1-10M) promoted PKC posphorylation, as well PKC translocation in PC 12 cells at lh incubation. Administration of rasagiline (0.1 mg/kg) to mice for 14 days significantly decreased (-65%) membrane bound holoprotein APP level in the hippocampus. Concomitantly rasagiline markedly increased PKCx and e in the membrane (55%) and the cytosolic (60%) fractions of mice hippocampus. Additionally, rasagiline treatment significantly elevated the levels of myrstoylated alanine-rich C kinase substrate (MARCKS), a major substrate for PKC, as well as the levels of receptors for actavated C kinase (RACK1). Similar effects on the holo APP and PKC levels were also demonstrated for the two-cholinesterase inhibitor derivatives of rasagiline, TV3326 and TV3279. These results indicate that rasagiline; TV3326 and TV3279 regulate APP processing by a PKC dependent mechanism. The activation and induction of PKC and MARCKS by these drugs may have a crucial role not only in their neuroprotective activity, but also a crucial role in their ability to affect neuronal plasticity and spatial learning processes. Keywords: rasagiline, amyloid precursor protein, protein kinase C, MARCKS, learning process Regulation of sAPP release via MAPK hosphorylation-dependent mechanism byrasagiline ogev-Falach M., Amit T., Bar-Am O.  Rasagiline (N-propargyll(R)-aminoindan), a neuroprotective monoamine oxdase B inhibitor in now under phase III clinical trials for Parkinson's disease (PD), and it rescues dopamine neurons from cell death in animal and cellular models of PD. We now show that rasagiline (1 and 10tM) significantly protected rat PC 12 cells against the 13-amyloid (A1.42) toxicity. In addition, rasagiline sgnificantly increased (--3 fold) the secretion of the non-amyloidogenic soluble form of the amyloid precursor protein (sAPPc) from PC 12 and SH-SY5Y neuroblastoma cells in a dose-dependent fashion. This was blocked by the hydroxamic acid-based metalloprotease inhibitor, Ro31-9790, suggesting mediation via t-secretase activity. Specific inhibitor of the mitogen-activated protein kinase (MAPK) pathway, PD98059,.prevented rasagiline induced sAPPc release, indicating the involvement of MAPK-dependent pathway. Furthermore rasagiline dose dependently (0.1-10tM) increased the phosphorylation of p44 and p42 MAPK, which was abolished by PD98059 and by the protein kinase C inhibitor, GF109203X. Structure activity studies with rasagiline derivatives have indicated that the propargylamine moiety, present in rasagiline initiates sAPPer release, as well as MAPK phosphorylation, indicating that the propargylamine moiety is crucial for these effects. These data suggest another novel pharmacological activity for rasagiline.
Rasagiline might prove useful in processing APP in favor of non-amyloidogenic pathway, thus reducing the risk of amyloidogenic derivatives formation and therefore it may have potential value in Alzheimer's disease. Keywords: rasagiline, neuroprotection, amyloid precursor protein, -secretase, MAPK Neuroprotective action of rasagiline is dependent on activation of Bcl-2, PKC and proteasome complex and inhibition of miochondrial lerim_eability transition (PT) Youdim M.B.H., MaruyamaW. and Naoi M.

Eve Topfand NPF Centers ofExcellence for
Neurode'generative Diseases Jesearch, Technion, Haifa," International Institute of Biotechnology, Gifu, Japan Our cell culture (PC-12, SH-SY5Y) studies have established that the antiParkinson drug, rasagiline and its optical isomer TVP1022 have relatively potent neuroprotective-antiapoptotic activities in response to several dopaminergic neurotoxins (N-methyl-R-salsolinol and SIN-I). These actions are not dependent on monoamine oxidase (MAO) inhibition, since TVP1022 and other derivatives of rasagiline, which do not inhibit MAO, have similar actions. Some of the events in their neuroprotective action include suppression of the decline in mitochondrial membrane potential, activation of caspase 3 and nuclear translocation of GAPDH. We have now confirmed their ability to stabilize mitochondrial membrane potential in isolated mitochondrial and prevent mitochondrial swelling. This would suggest that rasagiline and its derivatives directly. affect the mitochondria, since they inhibit mitochondrial permeability transition (PT), a phenomenon similarly observed m SH-SY5Y cells with over expressed Bcl-2. Rasagiline increases antiapoptotic Bcl-2 gene expression in -SY5Y cells. The rnRNA and protein levels ofBcl-2 and Bcl-xL are also increased significantly, with out affecting those of Bax and Bad. Furthermore rasagiline, similar to Bcl-2 over expression, activates PKC<x and and prevents neurotoxin-induced inhibition of proteasome complex, which in turn results in inhibition of cytochrome c release. In addition, rasagiline increases the gene expression and protein level of glia cell derived neurotrophic factor (GDNF). Our studies have shown that the mechanism of neuroprotective-antipoptotic action of rasagiline depends on inhibition of mitochondrial PT, increased expression of antiapopt.otic-cyto-protective (Bcl-2, Bcl-xL and PKC) and neurotroplaic genes. Keywords; rasagiline, neuroprotection, mitochondrial permeability transition, Bcl-2and PKC gene expression Intrinsic bursting i hippocampal pyramidal neurons lacking apical dendrites Yue C., and Yaari Y.
Dept. of Physiology, Institute of Medical Sciences, Hebrew Unive:Mty-Hadassah School of Medicine, Jerusalem 91120 The intrinsic discharge modes of hippoca_mpal CA1 py(_amidal cells vary along a gradient of 'burstmess' from regular firing to spontaneous bursttng. The propensity to generate a somatic burst is positively related to the size of the spike afterdepolarization (,ADP), generated by a slow inward current that activates during the spike and deactivates slowly thereafter. However, it is not known whether the ADP is locally generated by the somatic spike, is generated remotely at the apical dendrite by the baekqaropagating spike, or both. To differentiate between these alternatives, we have severed the apical dendrites of CA1 pyramidal cells near the soma and examined the firing modes of the amputated neurons in different experimental conditions. Their spectrum of firing modes was similar to that of intact neurons (mostly regular firing cells with a small proportion of bursters). Blocking the muscarinic-sensitive K+ current (IM) with linopidine markedly enhanced the propensity of amputated neurons to generate bursts, whereas blocking the Ca2+-activated K+ currents with intracellular (BAPTA) did not. The propensity to generate bursts was also augmented by raising extracellular K+]. or by removing, extracellular Ca2+. In all conditions, urstmg was readily suppressed by phenytoln or by PKC activation, but was not sensitive to Ni2+ (1 mM), implicating the persistent Na+ current (INaP) in its generation. Thus, somatic spike backpropagation into apical dendrite is not required f6r somatic bursting in hippocampal pyramidal cells. Rather, large spike ADPs that trigger somatic bursts are generated at or near the soma once INaP is sufficiently larger than IM. upported  We recently discovered a small area within the rat mesopontine tegmentum (MPTA) at which bilaterally symmetrical mlcroinjection of minute quantities of pentobarbital induces a transient, reversible anesthesia with non-responsiveness to noxious stimuli, flaccid atonia, and absence of the righting reflex. The behavioral suppression is accompanied by slow-wave EEG and, presumably, loss of consciousness (Devor and Zalkind, Pain 94:101-12 [2001] ). Here we ask whether microinjection into the MPTA on one side is capable of inducing behavioral signs of general anesthesia on both sides of the body. Using 22 rats that showed anesthesia on bilateral microinjection of pentobarbital, we microinjected this drug, in doses of 100 g in 0.5 tl or 200 xg in 1 into MPTA on one side (46 and 44 experiments, respectively). Behavioral results were quantified on motor (posture and righting reflex), and sensory scales (responses to pinching the left and right foot, and the tail). Microinjection sites were located histologically. There were two principal observations. First, anesthesia was induced by unilateral microinjection (44 trials).
The behavioral suppression was bilaterally symmetrical, and indistinguishable from anesthesia induced by bilateral MPTA or by systemic pentobarbital administration. Second, anesthesia duration, recovery time and mean score following bilateral 100 xg pentobarbital were statistically indistinguishable from those resulting from microinjection of 200 tg unilaterally, butgreater than 100 rtg unilaterally Conclusion: MPTA on either side of the brainstem controls both ascending and descending pathways, and both motor functions and pain sensation, bilaterally. Keywords: intracerebral, general anesthesia, unconsciousness, barbiturates, atonia, analgesia, unilateral microinjections