summarizes the animal characteristics. The duration of IOP elevation after the first measurement of greater than 25 mmHg varied from 259 to 1599 days. As is typical of this model, the IOPs were variable and moderately high (means from 24.2 to 49.3 mmHg). From 7 to 41 mfERG testing sessions were performed on each animal during the epoch of elevated IOP. At the end of the study, histologic examination of the optic nerves revealed that the axonal loss was moderate to profound. No axons were visible in the optic nerves of the 2 animals that had ONT followed by experimental glaucoma.
Selected individual waveforms for the 2 recovery animals are shown in . These are the unfiltered waveforms from the single stimulus elements at the center of the 103 element field (Ring 1). The waveforms for the right and left eyes as well as the difference (OD-OS) waveform are superimposed. Each right-left pair of traces was obtained on the same test day. Similar, although not identical, traces were found for the right and left eyes at baseline (pre-IOP elevation) for both monkeys for K1 as well as K2.1. During the epoch of high IOP, elevated K1 amplitudes were evident in both Rh3 and Cy3. The K2.1 trace was only modestly affected in Rh3. However K2.1 was clearly reduced in Cy3. After lowering of IOP by trabeculectomy, the K1 and K2.1 waveforms were nearly identical in the 2 eyes for Rh3. Recovery for Cy3 was limited to the earliest portion (N1 and the rising slope of P1) of K1. The later features (N2-P2) showed a phase shift that had a marked effect on the difference waveform. Following trabeculectomy, the K2.1 waveform of the experimental eye of Cy3 did not recover.
Figure 1 Selected individual waveforms for the 2 recovery animals—Rh3 (A) and Cy3 (B). These are the unfiltered waveforms from single stimulus elements at or adjacent to the center (Ring 1) of the 103 element array. The waveforms for the right (solid line, (more ...)
The largest K1 waveforms in both eyes were located in the central (macular) region as expected due to the unstretched stimulus pattern. Compared to the control eye, there was a marked increase in the amplitudes of the early waveform features in the glaucomatous eye that was particularly evident in the response to the central (macular) elements.
Repeated-measures ANOVAs performed separately on each animal’s K1 LFC RMS showed significantly higher amplitudes in the eye with experimentally elevated IOP as evidenced by Eye or Eye × Ring interactions. This pattern was present for all four rhesus and all four cynomolgus monkeys. A similar pattern was also evident in the K2.1 LFC RMS analyses. The K1 HFC RMS ANOVA Eye or Eye × Ring interactions were significant for 3 of 4 rhesus and 3 of 4 cynomolgus monkeys. The K2.1 HFC RMS ANOVA showed only 2 significant Eye × Ring interactions in the rhesus but all 4 cynomolgus monkeys showed significant effects involving eye for this measure. ANOVAs using N1 and P1 measures of mfERG amplitude showed patterns of effects to similar the RMS analyses.
As expected, there was a consistent significant interocular correlation of mfERG measures across testing sessions. To evaluate the consistency of direction of the difference, OD/OS ratios were calculated for each test session for each animal. T-tests (evaluated with Bonferroni correction) comparing the observed ratios versus the null hypothesis of 1.0 showed significant relative elevation of K1 RMS amplitudes for all for rhesus monkeys for each of the 4 ring averages. Only one of the cynomolgus monkeys showed consistent elevation of K1 LFC RMS amplitudes (). The K2.1 LFC RMS amplitudes were elevated in one rhesus, depressed in another and unchanged in the other two. In contrast, all four cynomolgus monkeys’ K2.1 LFC RMS were significantly lower than the expected ratio of 1.0 across all four ring averages ().
Figure 2 A. Comparisons of the K1 low frequency component RMS means and SEMs (over the 9–35 ms interval, Ring 2) of the OD/OS (experimental glaucoma /control eye) ratios for each animal. B. Comparisons of the K2.1 (Ring 2 low frequency RMS values, 0-to-80 (more ...)
Effects of IOP elevation on various specific waveform parameters were confirmed by repeated measures t-tests comparing the experimental and fellow eyes. Two predominant effects of IOP elevation on the mfERG waveforms were evident. There was an increase in amplitude of the early mfERG waveforms (N1 and P1) of K1 in the rhesus monkeys and a decrease in K2.1 in the cynomolgus monkeys.
The difference in K1 OD/OS ratios for the rhesus N1-P1 is much greater that what we typically see for this species. We reviewed the data from 8 other control rhesus animals recorded for a separate study. The mean OD/OS ratio for Ring 2 of the K1 N1-P1 was 0.98 ± 0.127 (s.d.). The average OD/OS ratio for the 4 rhesus in this study was 1.47 during the epoch of elevated IOP. Thus, the experimental glaucoma animals have OD/OS ratios that were 3.7 standard deviations greater that the expected control value of 1.0—well above the 95% confidence interval of 1.96 standard deviations.
Both the increase in K1 RMS (9–35 ms) OD/OS ratio in the rhesus and the decrease in the K2.1 RMS (0–80 ms) ratio for in the cynomolgus monkeys were stable over the relatively long time period of this study. shows the individual ratios versus days following IOP elevation to greater than 25 mmHg. In each case, the ratios were close to the mean even at the earliest test dates. In other words, the linear regression lines were essentially flat with a non-significant adjusted R2 for the rhesus K1 of 0.005 and an adjusted R2 for the cynomolgus K2.1 of 0.000.
Figure 3 Ratios of OD/OS (experimental glaucoma/control eye) at each test point for all animals plotted against days from the time that the IOP was first elevated above 25 mmHg in the experimental glaucoma eye. Linear regression lines (dashed) and 95% confidence (more ...)
A significant effect of species was evident in the OD/OS ratio analyzed with mixed-model ANOVAs based on the average ratio obtained across all sessions. The K1 LFC RMS ANOVA revealed significant effect of Species, [F(1,6)=65.3, p < .0001] with no effect of Ring or Species × Ring interaction. The same Species effect held for K2.1 LFC RMS [F(1,6)=41.6, p<.001] with no interaction with Ring. Similarly, an effect of Species was present in the K1 HFC RMS [F(1,6)=14.2, p<.01, and K2.1 HFC RMS (F(1,6)=21.9, p<.01] analyses.
The two monkeys that underwent ONT in the experimental eye prior to LTD showed effects on their waveforms similar to those seen in the monkeys with experimental glaucoma alone (). For example, both of these animals had increased early (N1 and P1) K1 components and the K2.1 was markedly reduced in the cynomolgus (CY4) compared to a K2.1 closer to baseline in the rhesus (Rh4).
Despite Eye × Ring interactions in many of the individual ANOVAs, no overall trend in the OD/OS ring ratios. There were no significant effects for Ring or Eye × Ring in the ANOVAs evaluating the effect of Species. Thus, there was no evidence for a consistent effect of retinal eccentricity in these data.
One rhesus (Rh3) and one cynomolgus (Cy3) monkey underwent lowering of the intraocular pressure in the experimental eye by a trabeculectomy procedure. illustrates the IOP recordings of the treated (OD) and control (OS) eyes over the course of the experiment in the rhesus (Rh3). Superimposed on the IOP recordings are OD/OS ratios of the K1 low frequency component RMS for the 9 to 35 ms portion of the waveforms for Ring 2. Even with considerable test/retest variability, the mfERG amplitudes were correlated with experimentally elevated and reduced IOP. As shown in , the baseline (pre-IOP elevation) ratios (filled circles, prior to IOP elevation by LTD) varied around 1.0 (0.93 ± 0.10). Following 2 sessions of LTD (back arrows) the OD/OS ratios increased to 1.43 ± 0.05(open circles). The ratios were relatively constant over this ensuing interval of elevated IOP. After 58 weeks of elevated IOP, a trabeculectomy was performed (gray arrow), after which, the IOPs in the experimental eye were lower than the fellow eye. At the 3 and 5 week test points following trabeculectomy, the OD/OS ratios remained high (1.51 and 1.46 respectively, black diamonds). However, from 7 weeks post trabeculectomy onward, the ratios dropped back to nearly unity (1.06 ± 0.03).For both of the monkeys that underwent trabeculectomy, there was a strong correlation between IOP (IOP difference between OD and OS) vs. K1 N1-P1 amplitude. The correlation coefficient for Rh3 was 0.62 (analysis of variance p <0.001) and for Cy3 it was 0.54 (analysis of variance p <0.002).
Figure 4 IOP recordings (right scale) of the treated (solid line) and control (dotted line) eyes over the course of the experiment in Rh3. Superimposed on the IOP recordings are the OD/OS (experimental glaucoma/control eye) ratios of K1 RMS (Ring 2, 0–80 (more ...)
Means and standard errors of the means (SEMs) of K1 RMS (9–35 ms, low frequency component) and K2.1 RMS (0–80 ms, low frequency component) of Ring 2 for Rh3 and Cy3 for the 3 epochs (baseline, high IOP and post-trabeculectomy) are shown in . The means were fairly constant over time for all parameters of the control eyes (black dots). However, changes were evident in many of the parameters of the experimentally glaucomatous eyes. K1 was markedly increased during the period of high IOP for both Rh3 and Cy3. Following trabeculectomy, the mean K1 returned to levels similar to the control eyes in both monkeys (). The K2.1 response differed markedly between the two animals. Rh3 showed a mild increase in K2.1 during the epoch of high IOP and then a moderate reduction following IOP lowering by trabeculectomy. Cy3, by comparison, showed a marked decrease in K2.1 with high IOP. There was no recovery of the waveform after IOP lowering. The mean K2.1 after IOP lowering was even less than during the time of high IOP ().
Means and SEMs of the K1 RMS (9–35 ms, low frequency component) (A) and K2.1 RMS (0–80 ms, low frequency component) (B) of Ring 2 for Rh3 and Cy3 for the 3 epochs (baseline, high IOP and post-trabeculectomy).