Comparison of vascular and respiratory effects of endothelin-1 in the pig

The haemodynamic and respiratory responses caused by i.v. administration of endothelin-1 (ET-1) (20–100 pmol/kg) were studied in anaesthetized spontaneously breathing pigs. Intravenous bolus administration of synthetic ET-1 (40–100 pmol/kg) caused a transient decrease followed by a long-lasting increase in mean pulmonary arterial pressure and dose dependent vasoconstriction both in the systemic and pulmonary circulations. The effect on pulmonary arterial pressure was biphasic, with an initial transient fall followed by a long-lasting dose dependent increase. A biphasic response of the systemic mean arterial pressure was demonstrated only at a high dose of ET-1 (100 pmol/kg). ET-1 administration did not significantly change breathing pattern or phasic vagal input, but caused a significant decrease in passive compliance. Passive resistances or active compliance and resistances of the respiratory system were not modified. These results suggest that in the pig ET-1 is a more potent constrictor of vascular than of bronchial smooth muscle. The vasoconstrictor activity was greater in the pulmonary than the systemic circulations.


Introduction
Endothelin (ET-1), a 21 amino acid peptide recently isolated from the culture medium of endothelial cells, is one of the most potent vasoconstrictors known. In isolated vascular strips of various experimental animals, ET-1 induces a vasoconstriction that is slow to develop and long-lasting. 2 ET-1 administered intravenously to healthy volunteers and pigs, 4 had a plasma half-life of about 1 min. Plasma ET-1 was cleared mainly by the kidney, splanchnic circulation and skeletal muscle. Despite its short plasma half-life, the systemic administration of ET-1 to various animal species induces a long-lasting pressure effect in vivo, an initial pressure response, either contraction or dilation depending on the species, followed by a secondary vasoconstriction of longer duration. -9 The transient hypotensive effect is thought to be mediated by potassium channels 1 and/or by ET-1 evoked release of endothelium derived relaxing factors. 1>5 ET-1 has been shown to contract non-vascular smooth muscle cells also, including guinea-pig tracheal and bronchial strips. 16 '7 In vivo potent bronchoconstrictor activity of ET-1 has been demonstrated in the guinea-pig and in the rat. 18 '19 Although the haemodynamic activity of ET-1 has been extensively studied in various animal species, very few studies have been performed to (C) 1993 Rapid Communications of Oxford ktd evaluate the effects of ET-1 on breathing pattern and compliance and resistances of the respiratory system. Thus, the aim of this investigation was to determine the dose dependence of the haemodynamic and respiratory effects induced by the intravenous administration of ET-1 in the pig. The static and dynamic activities of the tracheobronchial tree were evaluated by studying passive and active pulmonary compliance and resistances.

Materials and Methods
Six Large White pigs of either sex, weighing 20 The right femoral artery was cannulated with a solution and stored at -20C in a stock solution polyethylene catheter to monitor arterial blood of 50/g/ml. Each animal was given  pressure. The right external jugular vein was pmol/kg ET-1 injected as a bolus into the jugular cannulated to administer ET-1. A balloon-tipped vein. The bronchoconstrictor effect of ET-1 was catheter (Swan-Ganz 5F) was introduced into the also evaluated at the concentration of 400 pmol/kg pulmonary artery to monitor pulmonary arterial ET-1. In all pigs, the cardiovascular and respiratory pressure and cardiac output (CO) was evaluated by parameters were monitored before and continthe thermodilution technique (Cardiac Output uously after ET-1 injection for 5 min and again Computer 701 I.L.). Systemic and pulmonary every 5min until 30min. arterial pressure were recorded by connecting the The results are expressed as means q-S.E.M. The catheters to a fluid filled capacitance manometer statistical significance was evaluated by paired (Bell & Howell 4-422). All signals were recorded Student's t-test comparing the value obtained after simultaneously on a multichannel pen recorder (Nec ET-1 administration to the last pre-injection values. San-ei Instruments Polygraph mod. 8K40).
Difference of the values were considered statistically After recording the spontaneous resting breathsignificant at p < 0.05.
ing pattern the passive compliance was evaluated by occluding the tracheal cannula at the end of Results inspiration and at various lung volumes during expiration, in order to plot the passive pressure-As shown in Fig. 1A, the intravenous bolus volume (P-V) relationships of the respiratory administration of ET-1 at 40 and 100 pmol/kg to system. At each volume, changes in pressure were pigs caused long-lasting increases in mean pulmocomputed after 0.5 s, a time that seemed to be nary arterial pressure (MPAP) that were statistically sufficient to accommodate most of the stress significant at 15 min and persisted up to 30 min. In relaxation phenomena. The resistances of the contrast, administration of 20pmol/kg ET-1 did respiratory system (Rrs) were obtained from the not significantly atCfect MPAP. At all doses, the rs/Crs ratio. The passive time constant of the pulmonary vasoconstriction was preceded by a respiratory system was computed from the transient decrease in MPAP that was completely expiratory trace flow obtained after reopening the reversed at 5 min. At 40 pmol/kg, there was a airways closed at the end of inspiration. :rs was the statistically significant increase in total pulmonary time interval from the peak expiratory flow to 64% vascular resistance (TPVR) (Fig. 1B). of its decay. The resistance of the respiratory system As shown in Fig. 1C, there was a statistically was calculated by subtracting the resistance value significant increase in mean systemic arterial of the set-up. Active compliance (C'rs) and active pressure (MAP) only after the administration of resistances (R'rs) of the respiratory system were 100 pmol/kg ET-1. The peptide did not signifievaluated by occluding the airway at the endcantly aect total systemic vascular resistances expiratory lungo volume to obtain tracheal occlusion (TSVR). The administration of ET-1 did not cause pressure. Ptr, / and VT were measured at 0.04 s any significant changes in cardiac output (CO) or intervals after the onset of inspiration, the first stroke volume (SV)( Table 1). At 100 pmol/kg, the during occluded inspiratory effort and the and peptide caused an increase in heart rate that was VT during the immediately preceding spontaneous statistically significant at 100 min. ET-1 did not inspiration. Onset of inspiration was defined as significantly change respiratory frequency, tidal inspiratory flow and/or a negative deflection in Ptr" volume, pulmonary ventilation, inspiratory or Timing of breathing was analysed in terms of expiratory duration of unoccluded breaths (data not duration of inspiration and expiration for spontan-shown) at any of the doses. eous (TI, TE) and occluded breaths obtained by As shown in Fig. 2A, 40 pmol/kg ET-1 caused occlusion of the airway at the end-expiratory level, a significant lengthening of expiratory time of as suggested by Miserocchi et al.2'21 The latter occluded breaths (TEo). In contrast, the lengthening manoeuvre provides evaluation of the timing of of TIO was not statistically significant (Fig. 2B). Both breathing in the absence of lung volume related 20 and 100 pmol/kg of the peptide had no vagal afferents. The vagal inhibitory effect on statistically significant effect on these parameters. In respiratory centres was evaluated as the TIo/TI ratio, control conditions the TIo/T ratio, considered to Heart rate (HR), mean pulmonary arterial pressure be an index of phasic vagal feed-back, was greater (MPAP), mean systemic arterial pressure (MAP), than 1 and it was not significantly changed by ET-1 total pulmonary vascular resistances (TPVR), and administration (Fig. 2C). The arrows indicate bolus ET-1 administration. resistances (Fig. 3B), while 100 pmol/kg of ET-1 did not change the passive compliance or resistances of the respiratory system (Figs 3A and B). The active compliance (C'rs) and resistances were not significantly affected by ET-1 administration (Figs 3C and D).
In order to demonstrate the bronchoconstrictor activity of the peptide, ET-1 was administered to one pig at a higher concentration (400 pmol/kg).
This dose seemed to increase active respiratory resistances without affecting C'rs (data not shown). On the contrary, 400 pmol/kg of ET-1 did not have any greater effects on MPAP and TPVR than those of 40 and 100 pmol/kg (Figs 4A and B).

Discussion
This study examined the haemodynamic and respiratory effects of ET-1 in anaesthetized and spontaneously breathing pigs. The results show that the peptide is a potent vasoconstrictor mainly for the pulmonary circulation, and a mild bronchoconstrictor. ET-1 bolus administration increased pulmonary arterial pressure and vascular resistances. A maximum effect was obtained at 40pmol/kg of ET-1. The failure of a higher dose to induce more marked effects is probably due to saturation of lung ET-1 receptors by repeated administration of the peptide to the same animals. After all doses there was an initial transient vasodilation followed by a long-lasting vasoconstriction. In the systemic vascular bed, ET-1 caused a similar biphasic response only at 100 pmol/kg. The biphasic vascular response has been previously demonstrated in various animal species and in the pig by Pernow et al. 4 The long-lasting vasoconstrictive activity may be due to irreversible interaction of ET-1 with its specific receptors. The greater vascularization and high density of ET-1 receptors in the lung might be responsible, at least in part, for the greater response of the pulmonary than of the systemic circulation.
At 100 pmol/kg significant vasoconstriction was found in the systemic circulation. Saturation of ET-1 binding sites in the lung by previous administration of the peptide might reduce its pulmonary clearance thus favouring the systemic effect. 4 The transient hypotensive effect in both the pulmonary and systemic circulations may reflect ET-1 induced release of such vasodilators as NO and PGI2 . [11][12][13][14][15] At variance with other animal species, ET-1 did not alter cardiac activity in the pig to any statistically significant extent. The late increase in heart rate, not correlated with the pressure effect, is probably due to the release of ET-1 dependent vasoactive mediators. [11][12][13][14][15] The results show that ET-1 does not change the breathing pattern or the Tr/T ratio, in the pig, but causes a significant lengthening of TE Because the TIo/TI ratio is an index of vagal inhibitory activity on respiratory centres, the results show that ET-1 does not alter the vagal input, but causes a change in the bulbo-pontine rhythm probably by reduction of cerebral vascular flow. 22'23 The decrease in Crs was not correlated with a change in passive resistances observed in pigs, which suggests that ET-1 (20-100 pmol/kg) does not alter bronchomotor tone but causes only a decrease in lung distensibility, probably through increases in MPAP, or in lung capillary permeability. The results show that ET-1 does not statistically significantly change C'rs and ,R'rs probably because of induction of various compensatory mechanisms of both nervous and vascular origin. 24 Studies have shown that ET-1 acts as a potent bronchoconstrictor when studied in vitro 16'7 or in vivo in guinea-pigs, a8 In contrast, our results demonstrate that in spontaneously breathing pigs ET-1 is a mild bronchoconstrictor. In fact, constrictive activity on bronchial smooth muscle was caused only by a very high dose of ET-1 (400 pmol/kg). The reason for these discrepancies is not apparent. However, differences in ET-1 catabolism by the lungs of different animal species might be involved.
In summary, the results suggest that the effects on the vascular and respiratory system are different after ET-1 is administered i.v. to spontaneously breathing pigs. ET-1 is a more potent constrictor of vascular than of bronchial smooth muscle. Its vasoconstrictive effect is more marked in the pulmonary than in the systemic vascular bed. Pharmacological studies with selective inhibitors of ET-1 biosynthesis or action should help to clarify the role of ET-1 in the development of pulmonary disease.