The electroretinogram (ERG) has been commonly used to distinguish various retinal diseases and evaluate their stages in clinical practice [
On the other hand, the determination of the normative values of healthy subjects is of utmost importance because there are no normative values of ERG parameters such as implicit time (msec) and amplitude (
The purposes of the present study were as follows: first, to evaluate the reproducibility of the records with an examiner and the consistency of the records between examiners regarding parameters obtained by the RETeval device; second, to determine the normative values; and, third, to clarify the usefulness of pupil records.
We examined 100 eyes of 50 healthy subjects (12 males and 38 females), ranging in age from 20 to 24 years old (
ERG and pupil recordings were performed during the period from 10 AM to 2 PM, when the condition of the pupil is most stable, from the right to the left eye in a sitting posture. The skin electrode was placed on the orbital rim 2 mm from the margin of the lower eyelid, and the eyes were examined by directing the subject’s gaze at the red fixation spot in the center of the eyecup, with first the left and then the right eye covered with the subject’s hand.
After 20 minutes of light adaptation, the cone-system response was evaluated by the records of response in the order of cone-response to flicker-response. By using the protocol of the DR assessment [
The retinal illuminance energy of cone-response, flicker-response, and maximal-response was 85 Td-s, and the only rod-response was 0.24 Td-s. The background illuminance under the light adaptation was 848 Td-s. The stimulus frequency was 0.5 Hz for 9 times for the rod-response, 2 Hz for 30 times for the cone-response, 28.3 Hz for 141–424 times for the flicker-response, and 0.1 Hz for 5 times for the maximal-response. The oscillatory potential waveform is obtained by applying 85–190 Hz bandpass filter (to the maximal-response). Then, up to 5 cursors are automatically placed on the oscillatory potential peaks and troughs and are indicated on the report as black dots on the waveform by device software. Implicit times (time to peak) and amplitudes (peak to following trough) are reported for each individual cursor. The sums of implicit times and amplitudes for all cursors are also reported.
The signal processing for the flicker tests has previously been described in detail [
These red and blue-light sources used for pupil response measurements, which are feasible for colored-light stimulus under a custom protocol for research purposes, were manufactured by LKC Technologies Inc. and ordered from the MAYO Corporation (Aichi, Japan).
Statistical analysis was performed using commercially available statistical software (SPSS, version 20.0; IBM Corporation, Armonk, NY).
Reproducibility of parameters obtained by the measurement of the right eyes (50 eyes) of all the subjects twice at 1-minute intervals by an examiner (examiner A) was evaluated by the coefficient of variation (CV), calculated by dividing the value of the standard deviation by the mean value.
Regarding the parameters obtained by the measurement of the left eyes (50 eyes) of all the subjects twice at 1-minute intervals, their consistency between 2 examiners (examiners B and C) was evaluated by intraclass correlation coefficients (ICCs). Consistency was expressed as “almost perfect” when ICC was ≥0.81, as “substantial” when it was 0.80–0.61, and as “moderate” when it was 0.60–0.41.
The normative values of ERG parameters were calculated from the mean of two consecutive measurements obtained by the same examiner (examiner A). Then, the mean value and the normal range were analyzed with the upper and lower confidence limits with a 95% confidence interval (CI).
DR assessment to colored-light stimulus was evaluated from the measurement of the right and left eyes of the subjects once at 1-minute intervals by an examiner (white-light stimulus: examiner D; red-light stimulus: examiner E; blue-light stimulus: examiner F), respectively.
Pupil response to white-, red-, and blue-light stimulus for 5 seconds each was automatically analyzed by the area ratio (difference of pupil area under luminance of 4 and 32 Td-s) and constriction ratio (under luminance of 32 Td-s) (%) that were manually calculated by the equation: (initial pupil size before light stimulus − minimal pupil size during light stimulus)/(initial pupil size before light stimulus) × 100
Pearson product-moment correlation coefficient (
Table
Reproducibility of ERG parameters.
ERG parameters | Implicit time (ms) | Amplitude ( | |||
---|---|---|---|---|---|
CV | CV | ||||
Test | Retest | Test | Retest | ||
Cone-response | a-wave | 9.5 | 9.7 | 37.0 | 34.9 |
b-wave | 3.8 | 4.0 | 33.3 | 30.2 | |
Flicker-response 28.3 Hz | 2.6 | 2.5 | 30.1 | 30.3 | |
Rod-response | Scotopic b-wave | 14.6 | 12.8 | 31.9 | 29.8 |
Maximal-response | a-wave | 7.5 | 7.1 | 34.0 | 33.9 |
b-wave | 11.8 | 9.3 | 30.9 | 32.3 | |
Oscillatory potentials | 8.5 | 3.3 | 40.8 | 32.2 |
CV: coefficient of variation.
A summary of the interexaminer consistency for the two examiners that performed the ERG in 50 eyes (left eye) is shown in Table
Consistency of ERG parameters.
ERG parameters | Implicit time (ms) | Amplitude ( | |
---|---|---|---|
ICC | ICC | ||
Cone-response | a-wave | 0.48 | 0.71 |
b-wave | 0.76 | 0.87 | |
Flicker-response 28.3 Hz | 0.91 | 0.92 | |
Rod-response | Scotopic b-wave | 0.77 | 0.83 |
Maximal-response | a-wave | 0.51 | 0.72 |
b-wave | 0.63 | 0.92 | |
Oscillatory potentials | 0.85 | 0.87 |
ICC: intraclass correlation coefficients.
Figure
Normative values of ERG parameters.
ERG parameters | Implicit time (ms) | Amplitude ( | |||
---|---|---|---|---|---|
Mean | Range (95% CI) | Mean | Range (95% CI) | ||
Cone-response | a-wave | 12.1 | 12.0–12.2 | 5.8 | 5.6–6.0 |
b-wave | 28.2 | 28.1–28.3 | 21.1 | 20.4–21.7 | |
Flicker-response 28.3 Hz | 24.6 | 24.6–24.7 | 21.6 | 21.0–22.2 | |
Rod-response | Scotopic b-wave | 96.5 | 95.4–97.7 | 44.4 | 43.2–45.6 |
Maximal-response | a-wave | 14.8 | 14.7–14.9 | 40.5 | 39.3–41.8 |
b-wave | 46.7 | 46.3–47.2 | 68.7 | 66.7–70.6 | |
Oscillatory potentials | 152.5 | 151.8–153.2 | 49.0 | 47.3–50.6 |
CI: confidence interval.
All waveforms of the 50 eyes (red line shows mean waveform).
Cone-response
Flicker-response
Rod-response
Maximal-response
A summary of the mean values and the 95% CI of the pupil parameters are shown in Table
Pupil response of DR assessment to colored-light stimulus.
DR assessment | Pupil response | ||||||
---|---|---|---|---|---|---|---|
White | Red | Blue | |||||
Mean | Range (95% CI) | Mean | Range (95% CI) | Mean | Range (95% CI) | ||
AR | RE | 2.4 | 2.2–2.6 | 2.3 | 2.1–2.5 | 1.9 | 1.8–2.1 |
LE | 2.2 | 2.1–2.3 | 2.1 | 1.9–2.2 | 1.8 | 1.7–1.9 | |
CR | RE | 37.4 | 35.9–38.8 | 34.3 | 32.3–36.3 | 39.2 | 37.9–40.6 |
LE | 38.8 | 37.6–40.0 | 36.1 | 34.4–37.7 | 38.6 | 37.2–40.0 | |
DR | 21.2 | 20.7– 21.8 | 23.0 | 22.3–23.7 | 21.9 | 21.2–22.6 |
CI: confidence interval, AR: area ratio, CR: constriction ratio, and DR: diabetic retinopathy scale.
For the pupil parameters of the area ratio and the constriction ratio, moderate correlations were found for the white-light stimulus (right eye
Implicit time of DR assessment to colored-light stimulus.
DR assessment | Implicit time (ms) | ||||||
---|---|---|---|---|---|---|---|
White | Red | Blue | |||||
Mean | Range (95% CI) | Mean | Range (95% CI) | Mean | Range (95% CI) | ||
16 Td-s | RE | 26.8 | 26.5–27.2 | 29.1 | 28.7–29.6 | 26.1 | 25.7–26.5 |
LE | 26.7 | 26.4–27.1 | 28.5 | 28.1–29.0 | 26.2 | 25.8–26.6 | |
32 Td-s | RE | 26.1 | 25.8–26.3 | 27.5 | 27.1–27.9 | 25.7 | 25.3–26.1 |
LE | 26.0 | 25.8–26.2 | 27.1 | 26.8–27.4 | 26.1 | 25.8–26.4 |
CI: confidence interval.
Amplitude of DR assessment to colored-light stimulus.
DR assessment | Amplitude ( | ||||||
---|---|---|---|---|---|---|---|
White | Red | Blue | |||||
Mean | Range (95% CI) | Mean | Range (95% CI) | Mean | Range (95% CI) | ||
16 Td-s | RE | 16.8 | 15.3–18.3 | 16.5 | 15.0–18.0 | 20.2 | 18.7–21.7 |
LE | 14.7 | 13.2–16.2 | 14.1 | 12.5–15.8 | 18.1 | 16.5–19.8 | |
32 Td-s | RE | 20.9 | 19.0–22.7 | 21.0 | 19.1–22.9 | 22.0 | 20.1–23.9 |
LE | 18.1 | 16.5–19.7 | 17.8 | 15.9–19.6 | 19.6 | 17.7–21.4 |
CI: confidence interval.
RETeval parameters had generally good interexaminer consistency. However, the amplitude showed a generally lower intraexaminer reproducibility compared to the implicit time. The possible hypotheses include the pupil size recovery and adaptation of photoreceptor cells after a flash stimulus [
For the DR assessment protocol, although it is well known that DR scales are associated with the severity of diabetic retinopathy [
Retinal photoreceptor cells were regarded as rods and cones for many years; however, Provencio et al. [
One difficulty encountered during the study was the lack of availability of a custom protocol, which resulted in the curtailment of the acquisition of a lot of pupil data. We could not evaluate the intraexaminer reproducibility and interexaminer consistency for pupil parameters. However, the device is capable of simultaneous recording of flicker-responses and pupil responses using the protocol of the DR assessment. Therefore, we considered that the reproducibility and consistency for pupil parameters are consistent with the flicker-response parameters.
If the diseases of the optic nerve or retina are discriminatively detected by ERG recording with skin electrodes in combination with the records of pupil response to colored-light stimulus, not only would the burden of unnecessary ophthalmologic tests for patients be reduced but also this most noninvasive simple estimation method would contribute to the early detection of eye diseases.
Although the present study needs a larger range of subjects of different ages, the normative data could be evaluated in combination with those in the pupil response to colored-light stimulus. Consequently, the RETeval device was suggested as a possible screening device to detect signs and symptoms in visual afferent systems.
All procedures performed in studies involving human participants were in accordance with the ethical standards of the institutional research committee and with the 1964 Helsinki Declaration and its later amendments or comparable ethical standards.
Informed consent was obtained from all individual participants included in the study.
All authors certify that they have no affiliations with or involvement in any organization or entity with any financial or nonfinancial interest in the subject matter or materials discussed in this manuscript.
The authors thank Hidetaka Kudo of MAYO Corporation, for many helpful suggestions with this study, and Robert E. Brandt of MedEd Japan, for editing and formatting the manuscript. This study was supported by a grant from Kitasato University School of Allied Health Sciences (Grant-in-Aid for Research Project, no. 2017-1039). This work was supported by JSPS KAKENHI Grant no. JP16K21346.