Reliability and significance of measurements of a-wave latency in rats.

PURPOSE: To determine whether measurements of the a-wave latency of the electroretinogram (ERG) can be made as reliably as that of the implicit time (IT) in rats. In addition, to determine the relationship between the potential level selected for the latency and the baseline potential level. METHODS: ERGs, elicited by different stimulus intensities, were recorded from Long-Evans rats. The a-wave latency was determined by measuring the time between the stimulus onset and the beginning of the negative-going a-wave, and the IT was measured as the time between the stimulus onset and the peak of the a-wave. To test the reliability of the measurements of the latency, the a-wave latency and the IT were measured by three independent observers for the same 15 ERGs. RESULTS: The mean a-wave latency was approximately 14 milliseconds, and the mean a-wave implicit time was approximately 36 milliseconds. The mean of the a-wave latency and the IT, as measured by the three observers, were within 1 millisecond of each other. The coefficient of variation was as good for the latency as for the IT of the a-wave. The potential level selected for the latency was lower than the mean baseline potential level by 1 to 2 standard deviations. CONCLUSIONS: Selection of the a-wave latencies can be made as reliably as that for the IT. Because the a-wave latency is not affected by the activity of the second order neurons, the latency is a better measure than the IT of the time course of the a-wave.

The a-wave latency in control subjects and patients with retinal diseases.

PURPOSE: To determine the a-wave latency of the electroretinograms (ERGs) recorded from control subjects and patients with retinal diseases. METHODS: The a-wave latency and implicit time (IT) were measured retrospectively from the ERGs of 40 control subjects and 99 patients. The patients included 9 with complete congenital stationary night blindness (cCSNB), 13 with achromatopsia or cone dystrophy, 5 with supernormal and delayed rod ERG syndrome, and 72 with retinitis pigmentosa (RP). To assess whether latency measurements can be obtained reliably by different observers from patients with smaller a-wave amplitudes and noisier baselines, the a-wave latency and IT of the ERG of the right eye of 10 control subjects and 10 patients with RP were measured by three observers. RESULTS: The mean a-wave latency measured for the same 10 control ERGs by three observers differed by less than 1 millisecond while the mean IT differed by 1.7 milliseconds. For 10 ERGs from RP patients, the mean for the a-wave latency measured by the three observers differed by less than 2.0 milliseconds and by 1.1 millisecond for the IT. The coefficient of variation varied from 24.8% to 36.7% for the latency and from 11.5% to 16.0% for the IT. The a-wave latencies elicited by the 0-dB stimulus under scotopic and photopic conditions from the 40 control subjects were not statistically different. The a-wave latency in patients with cCSNB did not differ significantly from that in control subjects. The longer a-wave latency in patients with achromatopsia suggested that the rods have a longer latency than cones. The scotopic and photopic a-wave latencies were significantly longer in RP patients. The longer latency in RP patients was not due to smaller a- or b-wave amplitudes. CONCLUSIONS: The a-wave latency can be measured as reliably as the IT in control subjects but the reliability is not as good for the latency as for the IT in RP patients. The larger coefficients of variation in RP patients were most likely due to the measurements being made from RP patients at different stages of their disease. Our results suggest that the a-wave latency in control subjects is determined by cones under both scotopic and photopic conditions. The longer a-wave latency in RP patients suggests that the rods and cones are altered over a significant area of the retina.

Evidence that a-wave latency of the electroretinogram is determined solely by photoreceptors.

PURPOSE: To identify the retinal cells that determine the a-wave latency of rats. METHODS: Electroretinograms (ERGs) were recorded from the rod-dominated (0.85% cones) retinas of Long-Evans rats following an intravitreal injection of 1 microL of 40 mM 2-amino-4-phosphonobutyric acid to block the activity of the ON pathway of the second order retinal neurons. ERGs were also recorded following an intraperitoneal injection of sodium iodate to destroy the retinal pigment epithelial (RPE) cells. Damage to a large area of the retina was produced by constant light exposure, and focal damage to the retina was induced by argon laser photocoagulation. The effects of age and anesthesia level on the a-wave latency were also determined. RESULTS: Blocking the activity of the ON pathway of the second order retinal neurons did not alter the a-wave latency, and destroying the RPE cells also did not alter the a-wave latency. Damage to a large area of the retina resulted in prolonging the latency but focal retinal damage did not alter the a-wave latency. The a-wave latency was longer in young rat pups but was adult-like by 18 days. The level of anesthesia had no effect on the latency except at very deep stages. CONCLUSIONS: The a-wave latency is determined solely by the activity of the photoreceptors. A prolonged latency would indicate that the photoreceptors are damaged over a large area of the retina.

Low-frequency damped electroretinographic wavelets in young asymptomatic patients with dominant retinitis pigmentosa: a new electroretinographic finding.

OBJECTIVE: To describe a previously unreported electroretinographic (ERG) pattern in early retinitis pigmentosa (RP). DESIGN: Two case reports. PARTICIPANTS: Two unrelated young asymptomatic patients with autosomal-dominant retinitis pigmentosa were studied. MAIN OUTCOME MEASURES: Clinical findings and ERG responses were assessed. RESULTS: No ERG responses were detected scotopically with low-luminance stimuli. With increasingly brighter stimuli, a series of three to five low-frequency damped wavelets developed under both scotopic and photopic conditions. The period of the wavelets was 25 to 37 msec. CONCLUSIONS: Low-frequency damped ERG wavelets occur in some young asymptomatic patients with autosomal-dominant RP. The ERG pattern suggests that these wavelets are predominantly cone-generated.

The c-wave of the electroretinogram recorded under clinical conditions from rabbits.

To determine whether large and repeatable c-waves can be recorded from rabbits with equipment already in use in clinical electroretinographic laboratories, the Burian-Allen electrode, connected bipolarly or monopolarly, was used to record electroretinograms from pigmented rabbits. The Jet electrode was also used. The c-waves elicited by long-duration (4-second) stimuli were compared to those elicited by stroboscopic stimuli. In addition, the c-waves recorded with direct-coupled amplification were compared to those recorded with condenser-coupled amplification (one-half-amplitude bandpass=0.1 Hz). The b-wave amplitude was not altered by the amplifier coupling or by the two stimulus durations. The largest c-waves were elicited by 4-second-duration stimuli and recorded with direct-coupled amplification. Although the c-wave amplitude was reduced by stroboscopic stimuli and by condenser coupling, large and repeatable c-waves were elicited by stroboscopic stimuli and recorded with condenser-coupled amplification. A comparison of stimulus duration and amplifier coupling showed that the stimulus duration was more important in recording large-amplitude c-waves. Similar results were obtained with the Jet electrode. We conclude that repeatable and large c-waves can be elicited by a stroboscopic stimuli and can be recorded with condenser-coupled amplification with good low-frequency response from rabbits.

Effects of basic fibroblast growth factor on the retinal degeneration of the mi(vit)/mi(vit) (vitiligo) mouse: a morphologic and electrophysiologic study.

Basic fibroblast growth factor (bFGF) has been shown to rescue dying photoreceptor cells in the RCS rat, a model with a genetic defect of the RPE that impairs outer segment phagocytosis. The purpose of the present study was to determine whether intravitreal injection of bFGF would have a similar effect on photoreceptor cell death in the vitiligo (C57BL/6-mi(vit)/mi(vit)) mouse. This mutant mouse loses photoreceptor cells slowly over many months. Experimental evidence suggests that the primary cellular site of the defect is the RPE. In the present study, bFGF was prepared with and without heparin in PBS and injected intravitreally into vitiligo mice at ages 2, 4, 6, 8 and 13 weeks, surrounding the onset of photoreceptor cell death. Non-injected, PBS-injected and heparin/PBS injected mice served as controls. Scotopic ERG's were performed on one group of mice prior to killing. Mice were killed 4, 6 or 10 weeks following the injection and the eyes were processed for histology and analysed. The amplitude of the b-wave was significantly smaller in mice injected with bFGF/PBS than in PBS-injected and non-injected eyes regardless of the time of injection or duration following injection. Histological examination revealed that the number of rows of photoreceptor cells did not differ significantly between bFGF-injected, vehicle- or non-injected mice. Although slight improvement in the attachment of outer segments to RPE was observed in 4 week mutants injected with bFGF/heparin/PBS, a similar result was obtained in heparin/PBS injected mutants. In mice injected with bFGF without heparin, detachment was severe and gross disruption of neural retina was observed. There were significantly more macrophages and photoreceptor cells in the subretinal space in bFGF injected mice. It appears that at the dosages and times administered, bFGF does not improve the electrophysiological functioning of the retina nor retard the degeneration of the retina in the vitiligo mouse as it does in the RCS rat.

The retinal dopamine network alters the adaptational properties of retinal ganglion cells in the cat.

1. Single-unit extracellular recordings of optic tract fibers were used to study ganglion cell (GC) response properties of the intact cat eye before and after the intravitreal injection of haloperidol or SCH23390, dopamine-specific antagonists. Nearly all of the dopaminergic cells in the cat retina are amacrine cells (ACs); thus the dopamine antagonists are thought to primarily block the postsynaptic effects of these dopaminergic amacrine cells. All GCs encountered were subjected to a battery of receptive-field (RF) tests, including classification as X or Y, and as ON or OFF. 2. The effects of haloperidol were greatest in the light-adapted OFF-center pathways and especially in the OFF-center Y-cell. Within 30 min of haloperidol injection, both the spontaneous and light-evoked activity of the OFF-center Y-cell fell to zero, but when the same cell was exposed to lower levels of steady-state background illumination (scotopic levels), the response of the cell once again became robust. 3. OFF-Center Y-cells that had partially recovered from the drug effects and OFF-center X-cells recorded when the drug effect was maximal both possessed intensity-response curves that were shifted to the right of normal. 4. Recovery from the drug effects reflect supranormal responses after the initial response reductions and may be due to haloperidol's action on the dopamine autoreceptor. 5. Of the ON-center cells, only the Y-cells showed response alterations; possessing higher spontaneous activities and slightly reduced amplitudes to RF center (RFC) illumination. 6. The effects of SCH23390 paralleled those of haloperidol except that the onset was faster and the duration of the action of SCH23390 was much shorter, and no supranormal responses followed the initial effects. 7. Dark-adaptation functions of OFF-center GCs revealed a normal rod-cone shift; however, SCH23390 eliminated the rod-cone break, and threshold quickly fell to that of the rod mechanism. 8. The dopaminergic neurons of the cat retina appear to play an important role in regulating the activity of retinal OFF-center pathways in the photopically adapted eye, and one of its functions may be to control the relative contributions of the rod and cone systems to the response properties of light-adapted OFF-center GCs. 9. It is argued that dopamine is released in the light and enhances cone pathway activity, perhaps in the outer retina at bipolar and horizontal cells, and suppresses rod pathway activity, perhaps in the inner retina at amacrine cells.

Electroretinographic study of the C57BL/6-mivit/mivit mouse model of retinal degeneration.

PURPOSE: The C57BL/6-mivit/mivit mouse model of retinal degeneration is characterized by slow progressive loss of photoreceptor cells, concomitant loss of rhodopsin, and uneven pigmentation of the retinal pigment epithelium. The purpose of this study was to determine how these alterations affected the electroretinogram (ERG). METHODS: Scotopic ERGs were measured in two litters of mivit/mivit mice beginning at 4 weeks and continued in the same animals at 2-week intervals through 18 weeks. RESULTS: The mean of the maximum b-wave amplitude (Vmax) at 4 weeks was 234 +/- 14 microV in mivit/mivit mice, which did not differ significantly from controls (266 +/- 26 microV). With increasing age, all components of the ERG decreased and by 12 weeks, the mean of the Vmax had decreased to 170 microV. At 18 weeks, the mean Vmax was 75 microV, and the b- to a-wave ratio was still > 1.0. Comparison of these physiologic data to previously reported morphologic and biochemical data showed a high correlation between the b-wave amplitude and the number of photoreceptor cell nuclei (r = 0.9772) as well as the b-wave amplitude and rhodopsin levels (r = 0.9019). CONCLUSIONS: The loss of all components of the ERG and the lack of a negative-type ERG suggested that the primary cells altered in the mivit/mivit mouse were the photoreceptors. The high correlations between the ERG amplitude and the number of photoreceptor nuclei indicate that the Vmax of the ERG is a good measure of the degree of photoreceptor loss.

Correlation between the physiologic and morphologic changes in experimental autoimmune uveitis induced by peptide G of S-antigen.

PURPOSE: The authors followed and correlated the physiologic and morphologic changes occurring in experimental autoimmune uveitis (EAU) induced by the peptide G of S-antigen. METHODS: EAU was induced in Lewis rats by footpad inoculation of a 13-amino acid synthetic peptide (peptide G) in complete Freund's adjuvant. Electroretinography (ERG) was used to follow the physiologic changes, and light and electron microscopy were used to examine the morphologic changes. RESULTS: Serial ERG recordings showed a progressive decrease in the b-wave amplitude and a depression of retinal sensitivity beginning on day 18-21 postinoculation (PI). By day 35 PI, the b-wave was decreased by 91%, and the sensitivity was depressed by 4.68 log units. Negative ERG were recorded during the intermediate and late stage. Light and electron microscopy of the retina showed better correlation of the pathologic changes with b-wave depression than with PI day. CONCLUSIONS: ERG recordings were a good method to detect, follow, and quantify the severity of EAU. Their good correlation with the morphologic changes showed that this method can be used to assess the condition of the retina noninvasively.

Bilateral electroretinographic changes induced by unilateral intra-visual cortex inoculation of herpes simplex virus type 1 in BALB/c mice.

Intra-visual cortex inoculation of 10(2) plaque-forming units of herpes simplex virus type 1 (KOS-63) induced physiologic and morphologic retinal changes in 62.3% (33/53) of infected animals; of these, 91% were bilateral. In contrast, inoculation of the same viral titers into the frontal lobe induced retinal alterations in only 13.3% (2/15). Initially, there was a decrease of the b-wave amplitude and retinal sensitivity and necrotic changes of the ganglion cells and nuclei in the inner nuclear layer. Immunoperoxidase staining for virus-specific antigens showed positive staining of the same cell type. Over time, there was a progressive decrease in the electroretinogram until it was extinguished and the retina was replaced by gliotic tissue. Parallel viral recovery studies demonstrated detectable infectious virus in one of eight eyes on day 2 after inoculation and in three of eight eyes on day 4. Thereafter, there was an increase in the percentage of eyes with infectious virus and a concomitant increase in viral titers. Immunoperoxidase staining of brain sections obtained on days 6 through 8 demonstrated virus-specific antigens on cells in the lateral geniculate nuclei and the suprachiasmatic nuclei bilaterally.

Modulation of the receptive field center response of cat retinal ganglion cells by the shift response signal through amacrine cells.

The effect of a stationary and an oscillating grating situated greater than 30 degrees from the RFC on the receptive field center response was examined. The transient firing rate of the center response was suppressed by the oscillating grating for all types of cells. The stationary grating also suppressed the transient firing but the degree of suppression was significantly less than that with the oscillating grating. There was also a significant elevation of the sustained firing rate in Y-cells. The on-going discharges were elevated in all types of units except on-center X-cells when the grating was oscillated. An increase in the area of a grating annulus did not increase the degree of suppression of the center response. This lack of spatial summation of the shift response was related to the properties of the on-off transient amacrine cells.

Physiologic and morphologic retinal changes induced by murine cytomegalovirus in BALB/c and severe combined immune deficient mice.

Anterior chamber inoculation of murine cytomegalovirus (MCMV, 10(4) and 10(5) plaque-forming units) induced both physiologic and morphologic changes in the retinas of immunocompetent BALB/c and B- and T-cell-deficient severe combined immune deficient (SCID) mice. In BALB/c mice, the depression of the b-wave began on days 3-4 postinoculation (PI) and a further depression was recorded on day 7 PI. The electroretinograms (ERGs) remained depressed 1-2 weeks PI after which there was a recovery of the amplitude of the ERG 2-6 weeks later. The recovery was not complete; the maximum amplitude at 6 weeks was significantly lower than the preinoculation value. There was a greater loss in the amplitude than in the sensitivity of the ERG. Histologic examination of retinas with depressed ERGs showed swelling of the retinal pigment epithelium and distortion and shortening of the outer segment of the photoreceptors. With recovery of the ERG, there was normalization of the retinal histology. In SCID mice, the ERGs were extinguished, and there was no recovery. Histologically, there was a complete loss of the photoreceptors in the SCIDs, and electron microscopic examination showed viral particles in the retinal pigment epithelium and inner nuclear cells. These results demonstrate that MCMV can induce retinal pathology as reported in patients and show the importance of B- and T-lymphocytes in controlling the progression of this disease process.

HSV-2 alters retinal physiology and morphology bilaterally in mice.

Anterior chamber inoculation of 10(4) PFU of the MS strain of HSV-2 resulted in physiologic and morphologic changes in the retina of the inoculated and the uninoculated eyes. In the inoculated eyes, electroretinogram (ERG) depression was first detected on day 3 and abolished ERGs on day 8 postinoculation (PI). The decrease in the ERGs was rapid and the time course was similar for all of the eyes. In spite of a 90% decrease in the amplitude of the b-wave, the retinal sensitivity did not change. Of 23 eyes tested on or after day 10 PI, none had normal, 4.3% had reduced, and 95.6% had abolished ERGs. In the uninoculated eyes, ERG depression was first detected on day 8 and abolished ERGs on day 12 PI. The course of the ERG depression was more variable, and some of the eyes showed a decrease in retinal sensitivity. Of the 22 eyes tested on or after day 17 PI, 18% had normal, 32% had reduced, and 50% had abolished ERGs. The majority (17/33) of the retinas of the inoculated eyes showed panretinal necrosis, although 7 of 33 retinas had pathology confined to the outer layers of the retina. In the uninoculated eyes, only 5 of 30 retinas were necrotic and 14 of 30 retinas had pathology limited to the outer layers of the retina. These observations suggested that the physiologic and morphologic changes progress through two stages: an early stage with reduced ERGs and pathology limited to the outer retinal layers, and a second stage in which the ERG is abolished and the pathologic changes extend into the inner retina. Not all retinas progress to the second stage.

Alterations of the cat's electroretinogram induced by the lesioning of the indoleamine-accumulating amacrine cells.

The neurotoxin 5,7-dihydroxytryptamine (5,7-DHT) was used to destroy the indoleamine-accumulating amacrine cells located in the cat's retina. With 100 micrograms of 5,7-DHT, the alterations in the electroretinogram (ERG) were present in all of the treated eyes and the morphological changes were confined to some of the processes in the inner plexiform layer. The ERGs recorded from treated eyes consisted of negative waves at low intensities and depressed b wave amplitude at higher intensities. The duration of the b wave was not altered but the oscillatory potentials were strongly depressed. The changes were probably permanent. The differences in the ERG changes in cats and rabbits were suggested to arise from the differences in signal processing in the inner retina of rabbits and cats.

Bilateral alterations of the ERG and retinal histology following unilateral HSV-1 inoculation.

The physiological condition of the retinas of BALB/c mice inoculated unilaterally in the anterior chamber with the KOS strain of herpes simplex virus type 1 (HSV-1) was monitored by ERG recordings. After the ERG recordings, the retinas were examined for histopathological changes. In the inoculated eye, depressed ERGs were recorded on day 2 PI and abolished ERGs on day 4 PI. The changes in the ERGs were complete by day 5-6 PI. Of the 53 inoculated eyes followed for longer than day 6 PI, four (7.5%) remained normal, 30 (56.6%) had reduced ERGs and 19 (35.8%) had abolished ERGs. In the contralateral eyes, the first changes were noted on day 8 PI, and abolished ERGs were recorded on day 9 PI. Of the 55 contralateral eyes followed for longer than 10 days, 15 (27.3%) remained normal, four (7.2%) had reduced ERGs and 36 (65.4%) had abolished ERGs. The percentage of eyes with depressed ERGs was significantly higher in the inoculated than in the uninoculated eyes, and the percentage of eyes with abolished ERGs was significantly higher in the uninoculated eyes than in the inoculated eyes. The histopathological alterations were different for the two eyes. In the inoculated eyes, the changes were mainly in the outer retina, with characteristic folds in the photoreceptor and outer nuclear layer interspersed with normal appearing retina. The pigment epithelium was also abnormal. In the uninoculated eyes, the changes began in the inner retina but rapidly spread to all layers of the retina. This panretinal necrosis accounted for the higher percentage of abolished ERGs in the uninoculated eyes. The differences in the alterations of the ERG and the histopathological changes may be related to the underlying mechanism of action of the HSV-1 during the evolution of the experimental retinopathy.

Light ON depresses and light OFF enhances the release of dopamine from the cat's retina.

The release of [3H]dopamine from the cat's retina was determined by measuring the level of radioactivity in a series of vials in which the retina was incubated. At light ON; there was a sustained decrease and at light OFF there was an enhanced release of dopamine. The results also demonstrate that dopamine is released continuously in the dark.

Intranuclear rodlets in the rabbit retina following treatment with MPTP.

Retinas from pigmented rabbits treated with N-methyl-4-phenyl-1,2,3,6- tetrahydropyridine (MPTP, a chemical inducer of Parkinsonism in man and monkeys) were studied using light- and electron microscopy. The nuclei of many cells in the inner nuclear layer and the ganglion cell layer of the treated retinas contained crystalloids (intranuclear rodlets) of varying length (0.5-8 microns) which were composed of bundles of 12 nm filaments and which were seen only rarely in untreated retinas. The induced rodlets are oval to round cylinders, 15-20 filaments across (although they are smaller in untreated retinas). Similar intranuclear inclusions have been described under varying conditions in neurons and glia in the central nervous system of several animal species. In rabbits injected acutely with MPTP, most of the affected cells are in the inner nuclear layer in the position of bipolar cells, while in the chronically injected animals, clearly identifiable amacrine cells, and the nuclei of some cells in the ganglion cell layer also contained the inclusions. Evidence is presented that the rodlet-containing cells in the ganglion cell layer include both ganglion cells and displaced amacrine cells. These anatomic findings are relevant to understanding the physiological and biochemical effects of the drug on the retina which we reported previously (Wong, Ishibashi, Tucker and Hamasaki, 1985).

Destruction of the indoleamine-accumulating amacrine cells alters the ERG of rabbits.

The indoleamine-accumulating amacrine cells in the rabbit's retina were destroyed by intravitreal injections of 5,7-DHT according to the technique of Ehinger and Florén. One week after the injections, histofluoresence microscopy failed to show the IA-cells in the retina. At this time, the b-wave of the treated eyes was significantly smaller, the OPs were more prominent, and the duration of the b-wave was longer. The a- and c-waves and the off-response were not affected. The loss of the IA-cells resulted in a reduction in the range of dark-adaptation but did not affect the rate of recovery of the threshold. The loss of the IA-cells also had no effect on the response of the retina to flickering light. In a double-flash experiment, the suppression of the response to the second flash was significantly weaker in the treated eyes. These results can be explained by the loss of a negative feedback circuit that has been proposed for the IA-cells from morphologic studies.

Abnormal Y/X ratio in the area centralis of the Siamese cat.

The responses of X- and Y-type retinal ganglion cells were recorded in the optic tract of Siamese cats in order to investigate the encounter rate of Y-cells with respect to retinal eccentricity. The percentage of Y-cells in Siamese cats was highest in or near the area centralis and it decreased with eccentricity. This is in contrast to the proportion of Y-cells in normally pigmented cats, which was lowest in the area centralis and increased with eccentricity. Thus, the Y/X ratio in Siamese cats is higher in the area centralis, but significantly lower in the peripheral retina compared to those in normal controls. The lower percentage of Y-cells in the periphery parallels an additional finding that Y-cells exhibiting very high conduction velocities were missing from the Siamese cat optic tract. Finally, the receptive-field center (RFC) size of X-cells in the area centralis was larger in Siamese cats, and the correlation between the RFC size and conduction velocity was weak in these animals. The results are discussed in terms of behavioral deficits in Siamese cat vision.

A neural pathway for the shift response in the cat.

The shift response (McIlwain effect) was elicited by moving a large grating situated greater than 15 degrees from the conventional receptive field center (RFC). We examined the change in the amplitude of the shift response induced by placing a steady target on the RFC or the RF surround. We found that appropriate stimulation of the RFC or the RF surround will increase and inappropriate stimulation will decrease the amplitude of the shift response in a graded manner. The amplitude of the shift response was not correlated with the maintained activity but was correlated with the transient peak firing rate which is evoked by flashing the enhancing stimulus. A shift stimulus which elicits a strong shift response can be blocked by inappropriate stimulation. The results suggest that the shift signal is modulating a tonic signal which is present in the RF. Because the shift response is a transient excitation, we suggest that the shift response results from a disinhibitory process. A possible neural pathway for the shift response is presented.