Selective abolition of OFF responses in kainic acid-lesioned chicken retina

288

Brain Research, 535 (1990) 288-300 Elsevier

BRES 16126

Selective abolition of OFF responses in kainic acid-lesioned chicken retina M.A. Golcich*, I.G. Morgan and D.R. Dvorak** Visual Sciences Group, Research School of Biological Sciences and Centrefor Visual Sciences, Australian National University, Canberra City, A. C. Z (Australia)

(Accepted 26 June 1990) Key words: Retina; Bipolar cell; Amacrine cell; Ganglion cell; Kainic acid; Transience

When ganglion cell responses were recorded from optic axons in the superficial layers of the chicken optic tectum, the responses recorded are predominantly ON-OFF transient, with some ON transient, and rare OFF transient responses. Several weeks after excitotoxic lesion of the retina with 40 nmol of kainic acid injected intravitreally, only ON transient responses could be recorded from the contralateral optic tectum. ON response latency and threshold were not affected. At low light intensities responses in the kainic acid-lesioned retinas showed a sustained component which was not detected in control retinas, but at high light intensities, the sustained component disappeared and the responses were extremely transient. The disappearance of the OFF responses seems to be due to elimination of the OFF component of the responses of cells which are normally ON-OFF transient, rather than the silencing of these cells, leaving only the normally ON transient cells. Morphological evidence suggests that approximately two thirds of the bipolar cells and most amacrine cells are destroyed by the kainic acid lesion (Ingham and Morgan, Neuroscience, 9 (1983) 165-181), and pharmacological logic (Morgan, Prog. Retinal Res., 2 (1983) 247-266) suggests that the missing bipolar cells should be OFF bipolar cells. These results therefore suggest that ON-OFF transient cells receive direct input from bipolar cells, which determines their basic response type. These results also suggest that amacrine cells have little if any role to play in the generation of the basic centre responses of these ON-OFF transient ganglion cells, and that while amacrine cells may have a role in the generation of transient responses in the inner plexiform layer, transient responses can be generated without the intervention of amacrine cells, particularly at high intensities. INTRODUCTION Visual processing within the retina converts the sustained hyperpolarizing ( O F F ) responses of the photoreceptors into a rich variety of responses at the ganglion cell level, including responses which require complex trigger features (for review see ref. 29). Two features of this conversion are the generation of O N responses by sign inversion, and the generation of transient responses. It has long been accepted that in the vertebrate retina, O N and O F F pathways diverge in the outer plexiform layer ( O P L ) , where O F F bipolar cells respond to centre illumination with a sign-conserving hyperpolarization while O N bipolar cells respond to centre illumination with a sign-inverting depolarization 23'64. Since light reduces the output of the p h o t o r e c e p t o r (PR) transmitter, the P R transmitter must depolarize O F F bipolar cells and hyperpolarize O N bipolar cells. Early studies suggested that the divergence of O N and O F F pathways in the O P L was carried through to the ganglion cell C level, by sign-conserving O N and O F F bipolar cell output (for

recent reviews, see refs. 11, 54). Recently, dual push-pull innervation has been p r o p o s e d , where ON and O F F ganglion cells are not only excited by bipolar or amacrine cells of one sign, but are also inhibited by bipolar cells or amacrine cells of the opposite sign 3'4'26'58. In this way the depolarization of a ganglion cell could be g e n e r a t e d by an increase in excitation or a decrease in inhibition or a combination of both, while a h y p e r p o l a r i z a t i o n could result from a decrease in excitation or an increase in inhibition or both. The responses of p h o t o r e c e p t o r s and horizontal and bipolar cells are sustained, but within the inner retina, the responses of amacrine and ganglion cells are often transient 64. T h e r e are several theories as to how transient responses may arise. In general it is agreed that bipolar cell input of the a p p r o p r i a t e sign provides excitatory drive to p r o d u c e the O N , O F F or O N - O F F responses. Given the initial transients often seen in r e c o r d e d bipolar cell responses, T o y o d a et al. 6° suggested that shaping of the bipolar cell responses would be necessary to produce transient responses in postsynaptic cells, while Miller 39

* Present address: National Vision Research Institute, 386 Cardigan Street, Carlton, Vic. 3053, Australia. ** Present address: mmersham, Australia. Correspondence: I.G. Morgan, Visual Sciences Group, Research School of Biological Sciences and Centre for Visual Sciences, Australian National University, GPO Box 475, Canberra City, A.C.T. 2601, Australia. 0006-8993/90/$03.50 © 1990 Elsevier Science Publishers B.V. (Biomedical Division)

289 later suggested that thresholding could also be involved. The necessary shaping could involve inhibitory feed-back from amacrine cells (possibly G A B A e r g i c ) to bipolar cell terminals, thus truncating release of transmitter 6 (see ref. 11 for review). There is both morphological 17'31'32'49'62 and physiological 59 evidence for such G A B A e r g i c feedback synapses. Alternatively, Miller 39 has suggested that inhibitory amacrine cells feed-forward onto the transient cells, shaping the responses at this level. Werblin 63 has invoked the regenerative properties of amacrine cells as a m a j o r determinant of transient responses at the ganglion cell level, providing both transient excitatory and inhibitory inputs to the ganglion cells. There is also some evidence that the properties of the amacrine cell or ganglion cell m e m b r a n e may contribute to the shaping of transient responses 2'3°, or that there may be mechanisms intrinsic to the bipolar cell terminal 25. A recent detailed study of bipolar to transient amacrine cell transmission in the carp retina 27 showed that sustained depolarization of bipolar cells produced by extrinsic current, transiently depolarized O N - O F F amacrine cells, but could not elucidate the mechanism. Irrespective of how sustained responses are converted into transient, a feature common to the models of Dowling 11 and Sakai and Naka 54 is that transience is a property generated in excitatory amacrine cells which is passed on to the ganglion cells. Alternatively MiUer 39 has suggested that transient ganglion cells receive direct input from sustained bipolar cells, and that their responses are generated by thresholding, and by the input from inhibitory amacrine cells. At two key points in the retina, at the photoreceptor terminal and the bipolar cell terminal, glutamic acid is believed to be the transmitter (for review see ref. 41). Both glutamic acid, and the glutamic acid analogue kainic acid depolarize O F F bipolar cells and horizontal cells but hyperpolarize O N bipolar cells in the OPL. They also depolarize amacrine and ganglion cells in the inner plexiform layer (IPL) 5'24'46'55'56'57. Kainic acid is also a potent neurotoxin. It is believed to produce neuronal death by interacting with glutamic acid receptors, causing sustained depolarization and associated ionic and osmotic imbalances 47,53. In the chicken eye, 40 nmol of kainic acid injected intravitreally produces a retinal lesion in which about two-thirds of the bipolar cells and most of the amacrine cells are destroyed 2°. In the chicken retina, most ganglion cells survive exposure to kainic acid 16'2°'61. Since kainic acid depolarizes O F F bipolar cells and hyperpolarizes ON bipolar cells, Ingham and Morgan 2° suggested that it would selectively destroy O F F bipolar cells but spare ON bipolar cells, thus providing an explanation for the partial bipolar cell destruction. D v o r a k and Morgan 14 examined the effects of intravitreal kainic acid on visual-evoked

field potentials in the optic tectum of the chicken using full field stimuli. Forty nmol of intravitreal kainic acid totally and irreversibly eliminated tectal light-evoked O F F responses, while O N responses recovered substantially within a week. The present study compares the receptive field properties of single ganglion cells in kainic acid-lesioned retinas with those of normal retinas. The use of small spots avoided surround stimulation that can block centre O F F responses, and single unit recordings are superior to tectal evoked field potentials in detecting any weak O F F response that may be present. The main object of this study was therefore to test the following possibilities. 1. That since kainic acid depolarizes O F F bipolar cells and hyperpolarizes ON bipolar cells, O F F bipolar cells should be destroyed and ON bipolar cells spared. Assuming a sign-conserving pathway from bipolar cells to ganglion cells, the kainic acid-lesion should thus result in only ON responses from ganglion cells. 2. That since kainic acid destroys most of the amacrine cells, and most chicken ganglion cells are O N - O F F transient in their responses 8'37, if excitatory transient amacrine cells provide the drive to the O N - O F F transient ganglion cells, they should be silenced in the kainic acid lesioned retina. 3. That since kainic acid destroys most of the amacrine cells, mechanisms of transience involving amacrine cells should be disrupted, and ganglion cell responses should therefore reflect more closely the sustained responses of bipolar cells.

MATERIALS AND METHODS

Animals and lesioning technique Two day-old White Leghorn x Black Australorp male chickens (Gallus gallus) were obtained from Research Poultry Farm, Vic., Australia. The animals were kept on a 12:12 h light-dark cycle and supplied with food and water ad libitum. At 4-7 days post-hatch, chicks were anaesthetized with ether and the right eye was injected intravitreally with 10 ~i of freshly prepared 4 mM kainic acid (brought to pH 7.0 with NaOH) or 10/zl of distilled water as a control, using a 50/d Hamilton syringe fitted with a 2 cm x 26 gauge needle as previously described 44. Surgery Electrophysiological analyses were caried out on fifty 5- to 7-week-old chickens (30 controls/20 kainic acid-lesioned). Deep surgical anaesthesia was induced by intraperitoneal injection of 25% urethane in 0.9% saline. The ambient temperature was maintained between 25 and 30 °C and the animal was wrapped in thermalinsulating material to minimize loss of body heat. The anaesthetized animal was secured in a stereotaxic apparatus and the skin overlying the posterior and left mediolateral cranium was removed. A rigid head clamp was cemented to the skull and a 8 x 5 mm diameter elliptical hole was drilled into the cranium overlying the mediolaterai aspect of the left optic tectum. A small section (1.0 x 2.5 mm) of dura overlying the tectum was removed to expose the tectal surface. The eyelid of the right eye was retracted with suture thread,

290 and both cornea and exposed tectum were irrigated half-hourly with Ringer's solution. Birds were examined for corneal opacity and loss of acuity, but no problems were encountered. Recordings in both control and kainic acid-lesioned birds were always made from similar parts of the tectum, and receptive fields were always found in a roughly corresponding area.

Refraction Before presentation of light stimuli, the refractive state of the eye was determined approximately using the method of reflex reversal. Glass correcting lenses were placed 1 mm from the corneal surface. Once single unit recording had begun, the first suitable unit was used to further adjust the spectacle lens power so as to maximize grating acuity, using a set of hand-held, high contrast, square wave gratings 5°.

i n v o l v i n g p r i m a r i l y the I P L and t h e i n n e r n u c l e a r layer (INL).

in

sections with t h o s e r e p o r t e d by I n g h a m and M o r g a n 2° c o n f i r m e d that 40 n m o l kainic acid p r o d u c e d a lesion similar to that p r o d u c e d by the 20 a n d 60 n m o l doses used by I n g h a m and M o r g a n 2°. T h e m a j o r i t y of c h i c k e n retinal g a n g l i o n cells p r o v e d to h a v e low b a c k - g r o u n d d i s c h a r g e rates, and thus r e q u i r e d visual s t i m u l a t i o n to be d e t e c t e d . ' N o i s e ' b o a r d s w e r e tried b u t did not s t i m u l a t e all units. T h e best stimulus found

Microelectrodes

B o t h the I N L and t h e I P L w e r e r e d u c e d

thickness by a b o u t 4 0 % . C o m p a r i s o n of t h e t r a n s v e r s e

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Single-unit recordings were made from ganglion cell axons and terminal arbours in the superficial layer of the optic tectum using platinum-plated tungsten-in-glass microelectrodes with a tip length of 3-10 pm and a 100 Hz impedance of 0.5-1.0 M ~ 36. The microelectrode was supported in a Huxley micromanipulator with x,y,z control.

that t h e y could not be studied with t h e available stimuli.

Visual stimulation

chicken is s h o w n in Fig. 2 ( u p p e r trace). This r e s p o n s e is

spots flashed in t h e c e n t r e of the r e c e p t i v e field, but a few units d e t e c t e d with the filigree stimulus e i t h e r did not r e s p o n d to spots at all, o r s h o w e d such r a p i d h a b i t u a t i o n T h e r e s p o n s e of a retinal g a n g l i o n cell f r o m a control

Visual stimuli were projected onto a white screen positioned in the right lateral field at a distance of 325 mm from the chicken's right cornea. The screen was curved to minimize distance changes with horizontal displacement of the image in the horizontal axis. At the beginning of the experiments, a hand-held Neitz streak retinoscope was used to produce moving bars of light of variable size, velocity and luminance. This method allowed rapid localization of the receptive field centre and detection of direction selectivity and motion sensitivity. Two optical benches built as double lens Maxwellian view systems 65 allowed spots and annuli of different sizes to be presented independently. White light sources for the two optic benches were converted slide projectors utilizing 150 W quartz-halogen bulbs. Maximum light intensity for the spot was 227 cd/m 2 and for the annulus was 55.4 cd/m 2. Neutral density filters were used to attenuate stimulus intensity. Stimulus duration was controlled with electronic shutters of less than 3 ms response time. The animal was adapted to a background luminance of 0.44 cd/m 2 and receptive fields were mapped using a small spot of white light. (0.5 °, 7.2 cd/m2). Spots were positioned on the screen using a fine vernier X/Y adjustment of the stimulus aperture. At each location, the spot was flashed on and off, and the presence or absence of a response marked directly on the white paper overlying the screen.

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Recording techniques Responses were recorded by means of a purpose-built AC preamplifier with a probe headstage (10 x amplification). The signal from the amplifier was sent to a Tektronix 5113 oscilloscope. Responses were stored in digital form on an Apple lie computer and in analog format using a Vetter Model B FM tape recorder.

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Morphology techniques In 7 cases (2 controls, 5 kainic acid-lesioned), at the end of a recording session the retina from the injected eye was placed in cacodylate-buffered paraformaldehyde: glutaraldehyde fixative. After fixation, a small block of retinal tissue corresponding to the recording site in the tectum was removed and embedded in araldite. Semi-thin sections were cut on an ultramierotome to approximately 1 pm thickness and stained with Toluidine blue, as previously described**. RESULTS H i s t o l o g y of the retina s h o w e d a d r a m a t i c r e d u c t i o n in the retinal thickness in kainic a c i d - t r e a t e d eyes (Fig. 1),

GCL OFL Fig. 1. Semi-thin (1 gm) transverse sections through central areas of control and kainic acid-lesioned chicken retinas. These sections are of retinas from which ganglion cell recordings were made and from the vicinity of the receptive fields of ganglion cells recorded. Note the marked reduction in bipolar and amacrine cell numbers which is reflected in the reduced thickness of the INL and IPL. The outer retina appears relatively unchanged. ONL, outer nuclear layer; OPL, outer plexiform layer; INL, inner nuclear layer; IPL, inner plexiform layer; GCL, ganglion cell layer; OFL, optic fibre layer. Bar = 50 #m.

291 unless stimulated and responded to both the onset and offset of the light stimulus with a short burst of spikes. Control units typically responded with a phasic burst of 1-4 spikes (spike rate of up to 200 Hz) at both stimulus onset and offset. Units like this were classified as transient O N - O F E Fig. 2 (lower trace) shows the response of a unit from a kainic acid-lesioned eye. This unit is typical of units of lesioned retinas showing a robust response at stimulus onset and no response at stimulus offset. Absence of the OFF response was the most striking feature of units from kainic acid-lesioned retinas, and held for all spot intensities and spot sizes used (from threshold to 227 cd/m 2, spot diameters from 0.5 ° to 20°). Stimulus conditions for classification of units as ON, OFF or ON-OFF were standardized to a 0.5 ° spot, in the geometric centre of the receptive field, flashed for 500 ms. The intensity of the stimulus was chosen to be a 1.5 log unit attenuation which corresponds to 7.2 cd/m 2, approximately mid-way in the intensity/response range for dark-adapted chicken retinal ganglion cells. In all, recordings were made from 305 units. Some units were lost before sufficient testing had been done to enable classification, and others were so unusual in their stimulus requirements that characterization was not practical with the stimuli available (total, 151 units), thus leaving 154 well-characterized units (64 control/90 kainic acid-lesioned). 89% of control units responded ON-OFF, with only 9% responding ON only. In contrast, 100% of

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Fig. 2. Response histogams for control and lesioned units. Each histogram shows the cumulative responses to l0 presentations of a 1.0° spot stimulus in the centre of the receptive field. Stimulus duration was 500 ms and stimulus intensity was intermediate between threshold and saturation. The control response was uniformly ON-OFF and the lesioned unit responded only ON. Note also that at this light intensity the lesioned unit shows a less transient response. Maximum light intensity was 227 cd/m2, -2.0 indicates a 2 log unit attenuation from this maximum (background light intensity < 0.00l cd/m2). The smallest vertical bar in each histogram represents 1 spike, bin width = 4 ms.

kainic acid-lesioned units responded ON only. It was found that the ON-OFF response of some control units changed to ON only or OFF only as the stimulus was moved away from the centre of the receptive field to the periphery. Fig. 3 shows how the response of an ON-OFF unit in a control bird changed from ON-OFF in the centre to ON only in the periphery. Note that the latency of the response increased towards the periphery, but that the ON response remained quite robust. The ON only or OFF only periphery was usually, but not always, concentric with the ON-OFF centre. Fig. 4 shows how the response of kainic acid-lesioned units remained ON responsive right across the receptive field. No OFF peripheries were found in kainic acid-treated units. Fig. 4 also shows another feature typical of kainic acid-lesioned units, a sustained component in the ON response (see also Figs. 2 and 6). Control units also changed response as the stimulus brightness was increased from threshold to saturation. Fig. 5 shows the latency of both the ON and OFF responses decreased with increasing stimulus intensity. ON responses of control units generally showed a transient response of 1-4 spikes, becoming more phasic at higher stimulus intensities. A similar pattern was seen with the OFF responses, except that, in some units, the OFF response dropped out altogether at higher intensities. The loss of OFF responses of ON-OFF units at higher stimulus intensities was particularly common with larger spot sizes. The response of kainic acid-lesioned units changed with light intensity in a similar manner to that of control units apart from the total absence of OFF responses at all spot sizes and intensities (Fig. 6). As with control units, when stimulus intensity was increased, not only did the latency decrease to a minimum of about 30 ms but the response also became more phasic. Fig. 6 reveals a sustained component in the ON response. The sustained component dropped out at higher stimulus intensities, but the response remained typically less transient than in controls. For each stimulus intensity, the latencies of the beginning of the ON response to 0.5°centred spots of all units in a treatment were pooled, and the mean latency and standard error calculated (Fig. 7). As the intensity of the flashed spot increased to a maximum, the latency of the ON response decreased to a minimum of around 40 ms for both kainic acid-treated and control groups. The standard error bars overlap at all intensities plotted indicating that there is no significant difference between the groups of units for the latency of the ON response. The ON response threshold to 0.5 ° centred white spots was defined as the lowest stimulus intensity that resulted in a discernible ON response in the peristimulus response

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and, in these, threshold determination was simple. Units with an ongoing discharge required a judgement based on visual inspection of the cumulative response histogram. If judgement was difficult because of the nature of the ongoing discharge (e.g. bursting), the test was repeated. Spike frequency rises steeply from threshold (see Figs. 5 and 6), and thus the error in threshold determination is unlikely to be more than 0.2 log units (corresponding to less than 0.04 cd/m 2 at - 4 . 0 log unit attenuation). The threshold for the ON response of control units was - 2 . 4 + 0.16 log units attenuation compared to - 2 . 3 + 0.15 for the kainic acid-lesioned units. There is no significant difference between thresholds of control and kainic acid-lesioned units (dr = 33, t-statistic = 0.48, P > 0.63; M a n n - W h i t n e y U-statistic = 150.5, P > 0.49). To compare the duration of the ON response in control and kainic acid-lesioned units, the ON response durations were measured from the response histograms. For each stimulus intensity, the O N response durations of all units in a treatment were pooled and the mean duration and standard error calculated. These values were plotted against stimulus intensity to produce Fig. 8. In the control group, as the intensity increased from 'threshold' to maximum, the mean O N response duration increased from 80 ms at 'threshold' to a maximum of about 120 ms and then fell to a minimum of about 55 ms at maximum intensity. The kainic acid-lesioned group showed the

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was measured directly from the individual response histograms and pooled for all units of that treatment. The mean latency at each stimulus intensity is plotted, and the error bars show -+ S.E.M. Stimulus duration was 500 ms, and spot size was 0.5° visual field for all response histograms used to generate this graph. Maximum light intensity was 227 Cd/m2, with attenuation in log units given on the abscissa (background light intensity < 0.001 ed/m2).

294 same general trend as controls, but the duration was nearly twice as long at all stimulus levels, and the between unit variability was higher. It should be noted that ' t h r e s h o l d ' is not the mean threshold, as these graphs are of p o o l e d data, and some units did not respond at all at the lower intensities. Such units did not contribute to the p o o l e d means at intensities below each unit's threshold. The p s e u d o - O F F response (see below) was not included in this measurement. It can be seen from Fig. 10 that the p s e u d o - O F F response did not run into the p r i m a r y O N response, and so, in all cases, there was no difficulty in separating the two m a j o r response components. The i n t e n s i t y - r e s p o n s e spike function was also obtained using the same O N response p e r i o d described in the intensity/duration results, by pooling the n u m b e r of spikes p e r stimulus during the O N response period (see Fig. 9). In the control group, the m e a n n u m b e r of spikes p e r stimulus increased from about 2 at 'threshold' to a m a x i m u m of 5 at m a x i m u m light intensity. The kainic acid-lesioned group also began at about 2 spikes p e r stimulus but rapidly increased to 6 with the first 0.5 log unit increase in intensity, and continued to rise thereafter to a m a x i m u m of about 9 near maximum light intensity. Except at ' t h r e s h o l d ' , the m e a n n u m b e r of spikes per stimulus in the kainic acid-lesioned group was about twice that for the controls, and the between unit

variability was greater. The consistent lack of overlap of the error bars shows that the kainic acid treatment increased the n u m b e r of spikes p e r stimulus relative to the control group. O n e unit recorded from a kainic acid-lesioned animal warrants a separate description of its response nature, because it was difficult to decide if it had an O F F response. This unit had a high m a i n t a i n e d discharge, and r e s p o n d e d to increased or d e c r e a s e d b a c k g r o u n d illumination with a sustained decrease or a sustained increase in discharge rate respectively, but did not respond to small spots or the b a r stimulus. Large spots (10 °) generated a sustained decrease in m a i n t a i n e d discharge during light O N , but offset of spot illumination resulted merely in a return to the pre-stimulus rate. D u e to the unusual nature of the receptive field and unusual responses of this unit, it was not classified. This unit showed the p r o p e r t i e s required of a unit detecting dimming of the general a m b i e n t illumination. U n d e r certain stimulus conditions, some kainic acidlesioned units showed spike activity during the O F F stimulation p e r i o d (see Fig. 10, u p p e r histogram). This spike activity was unlike the characteristic O F F response of O N - O F F units (Fig. 5), in its e x t e n d e d , bursting nature. To test if this O F F activity was a true O F F response or a response to the initial O N stimulus, the period of spot presentation was increased to longer than

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Log stimulus intensity Fig. 9. Pooled intensity-number of spikes per stimulus curves for control and kainic acid-lesioned units, The average number of spikes per stimulus presentation during the ON response was calculated from the individual response histograms and pooled for all units of that treatment. The mean (pooled data) number of spikes per stimulus at each stimulus intensity is plotted, and the error bars show _+ S.E.M. (calculated per histogram). Stimulus duration was 500 ms, and spot size was 0.5* visual field for all response histograms used to generate this graph. Maximum light intensity was 227 cd/mz, with attenuation in log units given on the abscissa (background light intensity < 0.001 cd/m2).

295 the standard 500 ms. Fig. 10 shows that when the ON stimulation period was increased, the bursting response occured before the offset of stimulation. This demonstrates that the response is in fact post-excitatory rebound and is not generated by stimulus offset. We have called this phenomenon the pseudo-OFF response. Although the pseudo-OFF response was a common feature of kainic acid-lesioned units, a few control units also showed this phenomenon. In kainic acid-lesioned units, the pseudo-OFF response tended to be more robust, and occurred over a wider range of stimulus conditions than in control units, but the stimulus conditions producing the pseudo-OFF response varied for different units and were not studied in detail. Each animal used for recordings was refracted by the method of maximization of grating acuity. There was no statistically significant difference in grating acuity between control (2.78 + 0.62, cycles/°, n = 8) and kainic acid-lesioned units (2.7 + 0.54 cycles/°, n = 9) (dr = 13, t-statistic = 0.15, P > 0.88). Nor was there a significant difference in supplementary refraction (refractive error plus screen distance correction) required between control (1.38 + 3.2 dioptres, n = 8) and kainic acid-lesioned eyes (0.44 + 0.48 dioptres, n = 9) (df = 13, t-statistic = 1.58, P > 0.135). The decrease by one dioptre or a factor of 3 for kainic acid-lesioned eyes may seem large, despite the lack of statistical significance. However, considering that the ON

OFF

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Fig. 10. The pseudo-OPF response. This kainic acid-lesioned unit shows spike activity during the OFF stimulation period (upper histogram). Extending the period of stimulus presentation demonstrates that this spike activity is not a true OFF response (lower

histograms). Each histogram shows the cumulative responses to ten presentations of a 1.0" spot, 500 ms duration. (Luminance of stimulus was 3.3 log units attenuated from the maximum intensity of 227 cd/m2, background luminance was 0.027 cd/m2, bin widths = 4 ms, 8 ms, and 16 ms, for the upper, middle, and lowest histograms respectively).

total optical power of the 6-week-old chicken eye is about 120 D, one dioptre difference is small. Even for units with the finest grating acuity (5-6 cycles/°), it was found that supplementary lens power could be varied by about 2 D without impairing acuity. The distance to the screen was approximately one-third of a meter and thus should require 3 D to focus an emmetropic eye. This indicates about 1.5 D myopia (on average) for control eyes, and 2.5 D myopia (on average) for kainic acid-lesioned eyes. DISCUSSION The majority of control units recorded in this study responded to the onset and offset of light with transient bursts of activity. In the absence of stimulation they had very low maintained discharge rates. This picture is in general agreement with that obtained in a number of other studies on both chicken and pigeon retina s'l°' 12,13,19,34,35,37,48

The absence of maintained discharge in most avian ganglion cells is quite unlike the situation in cat retina where ganglion cells usually show a high maintained discharge of 10-70 Hz depending on background illumination 1. Urethane anaesthesia was used in this preparation, and it is possible that this reduced the maintained discharge to below normal 12, but this effect is unlikely to be large, as Miles 37 found that retinal ganglion cell (RGC) recordings from decerebrate, unanaesthetized chicks rarely showed maintained discharge in the absence of stimulus presentation. One question which needs to be addressed is how representative the recordings were of ganglion cell responses as a whole. In this study, ganglion cell responses were recorded from axons and terminal arbours in the optic fibre layer of the contralateral optic tectum. The optic fibre layer of the chicken is composed predominantly of myelinated optic tract fibres, more than 95% of which are 0.5-1.0/~m in diameter, and does not contain nerve cell bodies 28. Thus, with recording sites superficial in the tectum, there should be no contaminating recordings from visual neurons in the tectum. Morphological studies indicate that the optic tectum is the principal site of termination of retinal axons in chickens, and that this projection is entirely crossed 9'5t. Duff and Cohen 12'13 compared ganglion cell responses recorded from the optic chiasm and optic tectum of the pigeon, finding no significant differences. The encounter rates for responses recorded in the tectum in this study fit well with those obtained by Miles 37 for responses recorded from the chicken retina using intraocular techniques. Thus its seems likely that the tectal recording give a reliable sample of the majority of ganglion cell responses, although the encounter rate of units of a given

296 type may not represent the anatomical proportion of all types within the retina (see ref. 7). The tectal recordings may however miss minor populations of ganglion cells which do not project to the tectum, but which project to minor visual areas 15. For example, a small group of ganglion cells - - the displaced, probably directionally selective ganglion cells - - project not to the tectum, but to the nucleus of the basal optic root 52, and units which project to the anterior dorsolateral nucleus of the thalamus are predominantly sustained 2~. Another possibility which has to be considered is that there were optical differences between control and kainic acid-lesioned eyes. Wildsoet and Pettigrew 65 have shown that there is slight myopia in kainic acid-lesioned eyes. After correction for the artefact of retinoscopy ~8 our results are in close agreement with those of Wildsoet and Pettigrew 66. However this difference should not affect the interpretation of our results, since each eye was refracted appropriately, and after refraction, grating acuities were not significantly different between control and kainic acid-lesioned units. One caveat which needs to be applied to the interpretation of our results concerns the efficacy and selectivity of the lesions induced by kainic acid. Since we have no independent marker for ON and OFF bipolar cells in the absence of intracellular recordings, the evidence that OFF bipolar cells are destroyed is the morphological observation of partial bipolar cell destruction 2°, the known effects of kainic acid on hyperpolarizing ON bipolar cells and depolarizing OFF bipolar cells (see ref. 41), and the pharmacological logic of excitotoxicity which implicates depolarization as an obligatory step 47'53. The results presented in this paper and in a previous paper on the responses of kainic acid-lesioned retinas TM are striking in that OFF responses were never recorded, suggestive of complete abolition of the OFF bipolar cells. Whether some ON responses were impaired or eliminated is difficult to ensure, but the results obtained in this paper show that encounter rates were not lower in kainic acid-lesioned retinas, and that the ON responses observed were close to normal. This is in some ways a surprising result given the degree of disruption of the IPL observed acutely after the injection of kainic acid 43 (unpublished results). Some intact bipolar cell terminals are observed, and these are presumably those of the ON bipolar cell terminals. Other synaptic ribbon-containing terminals in the IPL are clearly swollen and disrupted, presumably the terminals of the OFF bipolar cells. Both amacrine and ganglion cell processes are disrupted, but the present results imply that appropriate synaptic relationships between the surviving (presumably ON) bipolar cell terminals and ganglion cell dendrites seem to be re-established.

Amacrine cell destruction is also quite high, although not complete. Ingham and Morgan 2° found that 85% of amacrine cells were destroyed. So far the only type of amacrine cell which appears to survive the lesion at a rate above 15% survival are the dopaminergic amacrine cells 2°. Destruction of cholinergic and GABAergic amacrine cells, measured both biochemically'~ and morphologically 3s is greater than 90% and enkephalin-, somatostatin-, vasoactive intestinal polypeptide- and substance P-immunoreactive amacrine cells are also almost completely eliminated. The depletion of glycine levels suggests that the glycinergic amacrine cells are also almost completely eliminated (unpublished results). Nevertheless, it is possible that even a few surviving amacrine cells from a given population, or an as yet untested surviving population, could ensure maintenance of transience, or pass on ON-OFF transient responses to ganglion cells. This is a possibility which needs to be remembered in interpreting our results, even though disruption of the amacrine cell network is extensive. It should be noted that these lesioning techniques need to be extensively validated before they are applied to other species. In particular, loss of ganglion cells needs to be carefully monitored, since misinterpretation of results due to selective loss or retention of ganglion cells of specific response types is possible (for discussion, see ref. 42). Comparison of control and kainic acid-lesioned responses - - ON-OFF dichotomy No ON-OFF units were found in kainic acid-lesioned retinas, despite a high encounter rate of 89% in control retinas. Not all stimuli generated an OFF response in control ON-OFF units, so an effort was made with kainic acid-lesioned units to explore all stimulus conditions found to bring out OFF responses in control units. Variation of stimulus intensity, size and position was carried out with all well-characterized kainic acid-lesioned units, but no stimulus used elicited OFF responses. The change in response properties encountered in kainic acid-lesioned retinas relative to those in control retinas could in principle be due to destruction or silencing of the ON-OFF ganglion cells encountered ~n control retinas, leaving only the ON units active. This is unlikely to be the case. Loss of ganglion cells, apart from the displaced ganglion cells which do not project to the tectum 16'52 is limited to less than 20% z°'61, whereas the encounter rate for ON-OFF ganglion cells is 89% in control retinas. The validity of this argument is limited by the possible lack of correspondence between encounter rate and anatomical proportion (see ref. 7), and the possibility that the ON-OFF units are silenced, but not

297 destroyed. But, if the ON responses in kainic acidlesioned retinas were being recorded from the ganglion cells responsible for the i0% encounter rate for ON units in control retinas, it should have been harder to find units in kainic acid-lesioned retinas, multiple units should have been more rarely encountered, and .single units should have been easier to isolate. This was not the case. Thus it seems most likely that the ON-OFF units recorded in control retinas have been converted to ON units by elimination of the OFF input. Only 1 out of 64 control units responded OFF only. This very low encounter rate for OFF only units means that the result for the kainic acid-lesioned group of 0 out of 90 is not very significant taken on its own. Several multiple unit recordings were made in kainic acidlesioned preparations in order to reveal OFF units that might have been present but rarely isolated, but none were found. This is consistent with the results of Dvorak and Morgan 14 on total field potentials. The unit detected in a kainic acid-lesioned preparation, which responded to dimming of the general ambient illumination, was continuously active, even under bright light, and did not respond to small spots and thin bar stimuli. Its response features were thus quite different to those of the OFF units, or the OFF component of ON-OFF units encountered in control retinas. The absence of OFF responses is consistent with the hypothesis that 40 nmol kainic acid destroys OFF bipolar cells and spares ON bipolar cells, leaving a retina which generates only ON responses, and is consistent with the hypothesis that bipolar cell input to cells in the inner retina is sign-conserving. It could be argued that the amacrine cells are not essential for the generation of ON responses, but are vital for OFF responses. However studies with N-methyl-D-aspartic acid, which eliminates amacrine cells, but spares bipolar cells show that the characteristic ON-OFF transient responses remain 22. The results are not consistent with the push-pull model of Sterling 5s, since all ON bipolar cells would be expected, on pharmacological grounds, to survive the kainic acid lesion, and, in this model, approximately half of them would be inhibitory, and capable of generating OFF responses. Push-pull models involving internuncial amacrine cells 3'4 are not inconsistent with these results, since such amacrine cells may have been destroyed by kainic acid. However, while they are not inconsistent with amacrine cell-mediated push-pull models, our results do not give any support to them. If there are tonic inhibitory amacrine cell inputs to the ganglion cells in the dark, the decrease in which helps to generate ON responses under normal conditions, then their elimination by the kainic acid lesion does not seem to lead to increased background firing rates, i.e. disinhibition does not seem to be

able alone to bring the cell above threshold. However the removal of inhibitory inputs may contribute to the increase in duration and spike number in the kainic acid-lesioned ON responses. Experiments were carried out under dark-adapted conditions, with stimuli ranging from threshold to well into the photopic range. Miiller et al. 45 have presented strong evidence that, in the cat, under dark-adapted conditions, depolarizing rod bipolar cells drive both the ON and OFF ganglion cells, by driving AII amacrine cells, which in turn drive ON ganglion cells via electrical synapses and OFF ganglion cells via glycinergic synapses by disinhibition. If we apply this model to the chicken retina under dark adapted conditions, and if the hypothetical AII-type amacrine in chicken retina were destroyed by the kainic acid lesion, then both ON and OFF scotopic responses should have been lost. This was not the case. Indeed, thresholds for the ON component were not changed, which suggests that the AII-type amacrines might have been spared by the lesion. But if AII-type amacrine cells had been spared, then according to the model of Mfiller et ai. 45, with ON bipolar cells and AII-type amacrine cells, OFF ganglion cell responses should have been generated under scotopic conditions. This was not the case either. We therefore conclude that the model of Mfiller et al. 45 for the mammalian retina does not apply to the chicken retina, and that there are probably separate ON and OFF bipolar cells involved in generating ON and OFF responses under scotopic conditions, as well as photopic conditions. Given the disruption of the amacrine cell system, our results also suggest that bipolar cell input provides the major drive to ON/OFF ganglion cells and is thus not consistent with the models of Dowling 11 and Sakai and Naka 54, although it is possible that the surviving 10% of amacrine cells includes excitatory internuncial amacrine cells. Our results are not however in conflict with the experimental results of Naka and colleagues (for review see Sakai and NakaS4), since a ganglion cell receiving input from an ON-OFF amacrine cell should be affected by that cell, whether the interaction is sign-conserving or sign-inverting. The question we are addressing is somewhat different, namely what is the major determinant of the response of the ganglion cell under normal conditions. Our results suggest that the major determinant of the responses of ON-OFF transient ganglion cells is direct bipolar cell input, a view which is more consistent with the model of Miller 39. The latter model was largely based on the lack of evidence for excitatory amacrine cells, but it should be noted that cholinergic amacrine cells appear to provide excitatory drive to directionally selective ganglion cells 33. Our results thus support the idea that ON bipolar cell input alone defines the ON

298 responses, and OFF bipolar cell input the OFF responses of inner retinal neurons. This idea is also supported by the selective abolition of ON responses from retinal neurons in chloride-free media 4°, and the selective pharmacological inhibition of ON responses by 2-amino4-phosphonobutyric acid 56. Comparison o f control and kainic acid-lesioned responses - - response transience

The great majority of avian ganglion cell recordings show phasic responses of 2-5 action potentials within 20-100 ms of stimulus presentation. Phasic (transient) ganglion cell responses could be generated during continuing stimulus presentation if the excitatory drive to the ganglion cell was rapidly turned off or was rapidly overcome by a more powerful inhibitory drive that is slightly delayed relative to the excitation. The decrease in latency of ON responses with increasing stimulus strength shows that the ON drive increases with increasing stimulus strength. Not only does the transience mechanism(s) match the increasing strength of excitation, but the rapidity of onset of the transience mechanism increases faster than that of excitation, producing an increasingly transient response with increasing stimulus strength. The differential effects of the kainic acid-lesion at different stimulus intensities may suggest that there are two distinct mechanisms operating. One of these may operate at low to intermediate light intensities, and saturate at higher light intensities. Its removal may lead to the sustained component of the responses of kainic acid-lesioned units at these intensities. This mechanism may be associated with the surround inhibitory mechanism, which in the difference-of-Gaussian model reaches a maximum in the centre, since surround inhibition is severely disrupted in the kainic add lesion (Golcich et al., in preparation), and since it has a narrow operating range (Yang et al., unpublished results). In other words, the sustained component of the responses of kainic acid-lesioned retinas could be explained by the removal of centre-driven surround pathway inhibition mediated by amacrine cells. Its sensitivity to kainic acid suggests that it is mediated by amacrine rather than horizontal cells (Golcich et ai., in preparation). However this disruption is also consistent with feed-back or feedforward models of transience generation. There also appears to be another mechanism which truncates the REFERENCES 1 Barlow, H.B. and Levick, W.R., Changes in the maintained discharge with adaptation level in the cat retins, J. Physiol., 202 (1969) 699-718. 2 Barnes, S. and Werblin, F., Gated currents generate single spike

responses at all light intensities, and which becomes more powerful at high light intensities. The correlation of strength plus latency of the excitation and efficacy plus latency of transience mechanisms suggests that this part of the transience mechanism is inherently linked to, and dependent upon, excitation for its activation. We therefore call it centre-pathway inhibition. It is not disrupted by the kainic acid lesion and may be associated with intrinsic properties of the bipolar cell to amacrine or ganglion cell interaction. While it is not powerful enough to suppress the sustained component at low to intermediate light intensities, at high light intensities it could suppress it, or else the centre-driven surround-pathway inhibitory component, while disrupted, may still be able to abolish the sustained component when sufficiently stimulated. This evidence for two components which contribute to the generation of transient ON-OFF responses is consistent with the results of Werblin 63 that inherently transient ON-OFF excitation and slightly delayed transient ONOFF inhibition combine to generate transient ON-OFF responses at the ganglion cell level. Our results however suggest that the transient ON-OFF excitation is likely to be associated with direct bipolar cell input, while the transient ON-OFF inhibition may be associated with amacrine cells, whereas Werblin 63 attributed both to the regenerative properties of amacrine cells. In summary, in terms of the specific hypotheses of this study, the results obtained are quite consistent with simple passage of ON and OFF response properties from ON and OFF bipolar cells respectively to transient ganglion cells, and give no support to push-pull models for the generation of ON and OFF components. Our results give no support to the idea that transient amacrine cells are vital for the generation of transience at the ganglion cell level, but favour models involving direct bipolar cell input to transient ganglion cells. Transience was not as severely disrupted as might have been expected, since at all light intensities responses had a transient component. This might be generated by properties intrinsic to the bipolar to ganglion cell interaction. However, at low to intermediate light intensities, there was a sustained component to the responses of kainic acid-lesioned units which was not detected in controls. It may normally be suppressed by the operation of amacrine cells.

activity in amacrine cells of the tiger salamander retina, Proc. Natl. Acad. Sci. U.S.A., 83 (1986) 1509-1512. 3 Belgum, J.H., Dvorak, D.R. and McReynolds, J.S., Sustained synaptic input to ganglion cells of mudpuppy retina, J. Physiol., 326 (1982) 91-108. 4 Belgum, J.H., Dvorak, D.R. and McReynolds, J.S., Sustained

299 and transient synaptic inputs to ON-OFF ganglion cells in the mudpuppy retina, J. Physiol., 340 (1983) 599-610. 5 Bloomfield, S.A. and Dowling, J.E., Roles of aspartate and glutamate in synaptic transmission in rabbit retina. II. Inner plexiform layer, J. Neurophysiol., 53 (1985) 714-725. 6 Burkhardt, D.A., Effects of picrotoxin and strychnine upon electrical activity in the proximal retina, Brain Research, 43 (1972) 246-249. 7 Cleland, B.G., Levick, W.R. and Wassle, H., Physiological identification of a morphological class of cat retinal ganglion cells, J. Physiol., 284 (1975) 151-171. 8 Cotter, J.R., Visual and nonvisual units recorded from the optic tectum of Gallus domesticus, Brain Behav. Evol., 13 (1976) 1-21. 9 Crossland, W.J., Cowan, W.M., Rogers, L.A. and Kelly, J.P. The specification of the retino-tectai projection in the chick, J. Comp. Neurol., 155 (1974) 127-164. 10 Donner, K.O., The spectral sensitivity of the pigeon's retinal elements, J. Physiol., 122 (1953) 524-537. 11 Dowling, J.E., The Retina: An Approachable Part of the Brain, Harvard University Press, Cambridge, MA. 12 Duff, T.A. and Cohen, D.H., Retinal afferents to the pigeon optic tectum: discharge characteristics in response to whole field illumination, Brain Research, 92 (1975) 1-19. 13 Duff, T.A. and Cohen, D.H., Optic chiasm fibers of the pigeon: discharge in response to whole field illumination, Brain Research, 92 (1975b) 145-148. 14 Dvorak, D.R. and Morgan, I.G., Intravitreal kainic acid permanently eliminates OFF-pathways from chicken retina, Neurosci. Lett., 36 (1983) 249-253. 15 Ehrlich, D. and Mark, R., At atlas of the primary visual projections in the brain of the chick Gallus gallus, J. Comp. Neurol., 223 (1984) 592-610. 16 Ehrlich, D., Teuchert, G. and Morgan, I.G., Specific ganglion cell death induced by intravitreal kainic acid in the chicken retina, Brain Research, 415 (1987) 342-436. 17 Freed, M.A., Smith, G. and Sterling, E, Rod bipolar array in the cat retina: pattern of input from rods and GABA-accumuluting amacrine cells, J. Comp. Neurol., 266 (1987) 445-455. 18 Glickstein, M. and Millodot, M., Retinoscopy and eye size, Science, 168 (1970) 605-606. 19 Holden, A.L., Receptive properties of retinal cells and tectal cells in the pigeon, J. Physiol., 201 (1969) 56-7. 20 Ingham, C.A. and Morgan, I.G., Dose-dependent effects of intravitreal kainic acid on specific cell types in chicken retina, Neuroscience, 9 (1983) 165-181. 21 Jassik-Gerschenfeld, D., Teulon, J. and Ropert, N., Visual receptive field types in the nucleus dorsolateralis anterior of the pigeon's thalamus, Brain Research, 108 (1976) 295-306. 22 Jones, M., Yang, G. and Dvorak, D., Effects of NMDA-induced amacrine cell loss on the receptive field properties of ganglion cells in chicken retina, Neurosci. Lett., Suppl. 27 (1987) $90. 23 Kaneko, A., Physiological and morphological identification of horizontal, bipolar and amacrine cells in goldfish retina, J. Physiol., 207 (1970) 623-633. 24 Kaneko, A. and Shimazaki, H., Synaptic transmission from photoreceptors to bipolar and horizontal cells in the carp retina, Cold Spring Harbor Syrup. Quant. Biol., 40 (1975) 537-546. 25 Kaneko, A., Pinto, L.H. and Tachibana, M., Transient calcium current of retinal bipolar cells of the mouse, J. Physiol., 410 (1989) 613-629. 26 Kolb, H. and Nelson, R., Neural architecture of the cat retina. In Osborne, N. and Chader, G. (Eds.), Progress in Retinal Research, Vol. 3, Pergamon, Sydney. 1984, pp. 21-60. 27 Kujiraoka, T., Saito, T. and Toyoda, J.I., Analysis of synaptic inputs to ON-OFF amacrine cells of the carp retina, J. Gen. Physiol., 92 (1988) 475-487. 28 LaVail, J.H. and Cowan, W.M., The development of the chick optic rectum. II. Autoradiographic studies, Brain Research, 28 (1971) 421-441.

29 Levick, W.R. and Thibos, L.N., Receptive fields of cat ganglion cells: classification and construction. In N. Osbone and G. Chader (Eds.), Progress in Retinal Research, Vol. 2, Pergamon, Oxford, 1983, pp. 267-319. 30 Lukasiewics, P. and Werblin, E , A slowly inactivating potassium current truncates spike activity in ganglion cells of the tiger salamander retina, J. Neurosci., 8 (1988) 4470--4481. 31 Marc, R.E., Stell, W.K., Bok, D. and Lam, D.M.K., GABAergic pathways in the goldfish retina, J. Comp. Neurol., 182 (1978) 221-246. 32 Mariani, A.E and Caserta, M.T., Electron microscopy of glutamate decarboxylase (GAD) immunoreactivity in the inner plexiform layer of the rhesus monkey retina, J. Neurocytol., 15 (1986) 645-655. 33 Masland, R.H. and Ames, A., Responses to acetyicholine of ganglion cells in an isolated mammalian retina, J. Neurophysiol., 39 (1976) 1220-1235. 34 Maturana, H.R., Functional organisation of the pigeon retina. In R.W. Gerard and J.W. Duyff (Eds.), Information Processing in the Nervous System, Vol. III, Proceedings of the International Union of Physiological Sciences, Excepta Medica, Amsterdam, 1962, pp. 170-178. 35 Maturana, H.R. and Frenk, S., Directional movement and horizontal edge detectors in the pigeon retina, Science, 142 (1963) 977-979. 36 Merrill, E.G. and Ainsworth, A., Glass-coated platinum-plated tungsten microelectrodes, Med. Biol. Eng., 10 (1972) 662-672. 37 Miles, E A . , Centrifugal control of the avian retina. I. Receptive field properties of retinal ganglion cells, Brain Research, 48 (1972) 65-92. 38 Millar, T.J., Boelen, M. and Mogan, I.G., Effects of excitotoxins on amacrine cell types in the chicken retina, Proc. Aust. Biochem. Soc., 18 (1986) $73. 39 Miller, R.E, The neuronal basis of ganglion cell receptive field organizaton and the physiology of amacrine cells. In EO. Schmitt and EG. Worden (Eds.), The Neuroscience, Fourth Study Program, MIT Press, Cambridge. 1979, pp. 227-245. 40 Miller, R. and Dacheux, R., Synaptic organization and ionic basis of ON and OFF channels in mudpuppy retina. III. A model of ganglion cell receptive field organization based on chloridefree experiments, J. Gen. Physiol., 67 (1976) 679-690. 41 Miller, R. and Slaughter, M., Excitatory amino acid receptors of the retina: diversity of subtypes and conductance mechanisms, Trends Neurosci., 9 (1986) 211-218. 42 Morgan, I.G., Kainic acid as a tool in retinal research. In N. Osborne and G. Chader (Eds.), Progress in Retinal Research, Vol. 2, Pergamon, Oxford. 1983, pp. 249-266. 43 Morgan, I.G. and Dvorak, D.R., A physiologically active kainic acid-preferring receptor in chicken retina, Neurosci. Lett., 44 (1984) 299-304. 44 Morgan, I.G. and Ingham, C.A., Kainic acid affects both plexiform layers of chicken retina, Neurosci. Lett., 32 (1981) 275-280. 45 Miiller, E, Wassle, H. and Voigt, T., Pharmacological modulation of the rod pathway in cat retina, J. Neurophysiol., 59 (1988) 1657-1672. 46 Murakami, M., Ohtsuka, T. and Shimazaki, H., Effects of aspartate and glutamate on bipolar cells in the carp retina, Vision Res., 15 (1975) 456--458. 47 Olney, J.W., Neurotoxicity of excitatory amino acids. In E. McGeer, J. Olney and P. McGeer (Eds.), Kainic Acid as a Tool in Neurobiology, Raven, New York. 1978, pp. 95-122. 48 Pearlman, A.L. and Hughes, C.P., Functional role of efferents to the avian retina. I. Analysis of retinal ganglion cell receptive fields, J. Comp. Neurol., 166 (1976) 111-122. 49 Pourcho, R.G. and Goebel, D.J., Neuronal subpopulatons in cat retina which accumulate the GABA agonist, 3H-muscimol: a combined Golgi and autoradiographic study, J. Comp. Neurol., (1983) 25-35. 50 Powers, M.K. and Green, D.G., Single retinal ganglion cell

300

51 52

53

54

55

56

57

58 59

responses in the dark-reared rat: grating acuity, contrast sensitivity and defocussing, Vision Res., 18 (1978) 1533-1539. Ramon y Cajal, S., La retine des vertebres, Cellule, 9 (1893) 17-257. Reiner, A., Brecha, N. and Karten, H.J., A specific projection of retinal displaced ganglion cells to the nucleus of the basal optic root in chicken, Neuroscience, 4 (1979) 1679-1688. Rothman, S.M., The neurotoxicity of excitatory amino acids is produced by passive chloride influx, J. Neurosci., 5 (1985) 1483-1489. Sakai, H.M. and Nake, K.-I., Neuron network in catfish retina. In N.N. Osborne and G. Chader (Eds.), Progress in Retinal Research, Vol. 7, Pergamon, Oxford 1988, pp. 149-208. Shiells, R.A., Falka, G. and Naghshineh S., Action of glutamate and aspartate analogues on rod horizontal and bipolar cells. Nature, 294 (1981) 592-594. Slaughter, M.M. and Miller, R.F., 2-Amino-4-phosphonobutyric acid: a new pharmacological tool for retina research, Science, 211 (1981) 182-185. Slaughter, M.M. and Miller, R.F., The role of excitatory amino acid transmitters in the mudpuppy retina: an analysis with kainic acid and N-methyl aspartate, J. Neurosci., 3 (1983) 1701-1711. Sterling, P., Microcircuitry of the cat retina, Annu. Rev. Neurosci., 6 (1983) 149-185. Tachibana, M. and Kaneko, A., Gamma-aminobutyric acid

60 61

62

63

64

65 66

exerts a local inhibitory action on the axon terminal of bipolar cells: evidence for negative feedback from amacrine cells, Proc. Natl. Acad. Sci. U.S.A., 84 (1987) 3501-3505. Toyoda, J., Hashimoto, H. and Ohtsu, K., Bipolar-amacrine transmission in the carp retina, Vision Res., 13 (1973) 295-307. Tung, N.N., Morgan, I.G. and Ehrlich, D., A quantitative analysis of the effects of excitatory neurotoxins on retinal ganglion cells in the chick, Vis. Neurosci., in press. Vaughn, J.E., Famiglietti, E.V., Barber, R.P., Saito, K., Roberts, E. and Ribak, C.E., GABAergic amacrine ceils in rat retina: immunocytochemical identification and synaptic connectivity, J. Comp. Neurol., 197 (1981) 113-127. Werblin, ES., Regenerative amacrine cell depolarization and formation of ON-OFF ganglion cell response, J. Physiol., 264 (1977) 767-785. Werblin, F.S. and Dowling, J.E., Organisation of the retina of the mudpuppy, Necturus maculosus. II. Intracellular recording, J. Neurophysiol., 32 (1969) 339-355. Westheimer, G., The Maxwellian view, Vision Res., 6 (1966) 669-682. Wildsoet, C.F. and Pettigrew, J.D., Kainic acid-induced eye enlargement in chickens: differential effects on anterior and posterior segments, Invest. Ophthal. Vis. Sci., 29 (1988) 311319.