Morphology of physiologically identified bipolar cells in the retina of the tiger salamander, Ambystoma tigrinum.

Intracellular recordings of the light responses of bipolar cells were made in the isolated perfused retina of the larval tiger salamander by microelectrodes filled with a 1% solution of the enzyme horseradish peroxidase (HRP). Two classes of bipolar cell were identified in terms of their responses to luminous spots and annuli centered upon their receptive fields: on-center cells, which depolarized in response to a centered spot and hyperpolarized in response to a concentric annulus, and off-center cells, which responded with the opposite polarities. Physiologically identified cells were labelled with HRP by iontophoretic injection and prepared for light microscopy. Examples of each of the three classes of bipolar cell described in Cajal's study of the amphibian retina were found amongst the labelled cells. The only morphological feature found to correlate with the physiological classification was the stratum at which the axon of the cell arborized in the inner plexiform layer. The axons of on-center cells arborized in the more proximal sublamina b, while those of off-center cells arborized in the more distal sublamina a. This is consistent with earlier findings in the retinae of the cat and the carp.

Characterization of retinal injury using ERG measures obtained with both conventional and multifocal methods in chronic ocular hypertensive primates.

PURPOSE: To characterize, using both conventional and multifocal electroretinogram (ERG) recordings as well as histologic measures, retinal injury in the chronic ocular hypertensive primate model for experimental glaucoma. METHODS: Ocular hypertension was induced in the right eye of 7 cynomolgous monkeys, Macaca fascicularis, using laser injury to the aqueous outflow tissue at the anterior chamber angle. At 16 months after IOP elevation, ERG recordings were made from both eyes of all animals using both conventional and multifocal methods. After electrophysiological recording, animals were killed and retinal samples were radially sectioned for histologic analysis. RESULTS: Histologic measures showed that ocular hypertensive injury was largely or completely limited to a loss of retinal ganglion cells (RGCs). The degree of RGC loss was similar in central and peripheral retina. Amplitudes of conventional ERG responses were mostly unaffected in eyes having severe loss of RGCs, a finding that is consistent with limited injury to photoreceptors, bipolar cells, and amacrine cells. Peaks in both the first- and second-order multifocal ERG responses were attenuated in ocular hypertensive eyes, and amplitude of these peaks was highly correlated with the density of surviving RGCs. CONCLUSIONS: The results are consistent with a conclusion that both first- and second-order components of the multifocal ERG response from the monkey reflect a significant contribution from activity in RGCs and may provide a useful measure for the clinical diagnosis and management of glaucoma.

Contribution to the kinetics and amplitude of the electroretinogram b-wave by third-order retinal neurons in the rabbit retina.

The ERG b-wave is widely believed to reflect mainly light-induced activity of on-center bipolar cells and Müller cells. Third-order retinal neurons are thought to contribute negligibly to generation of the b-wave. Here we show that pharmacological agents which affect predominantly third-order neurons alter significantly both the kinetics and amplitude of the b-wave. Our results support the notion that changes in the amplitude and kinetics of light-induced membrane depolarization in third-order neurons produce similar changes in the amplitude and kinetics of the b-wave. We conclude that activity in third-order neurons makes a significant contribution to b-wave generation. Our results also provide evidence that spiking activity of third-order neurons truncates the a-wave by accelerating the onset of the b-wave.

Similar effects of carbachol and dopamine on neurons in the distal retina of the tiger salamander.

Though there is considerable evidence that dopamine is an important retinal neuromodulator that mediates many of the changes in the properties of retinal neurons that are normally seen during light adaptation, the mechanism by which dopamine release is controlled remains poorly understood. In this paper, we present evidence which indicates that dopamine release in the retina of the tiger salamander, Ambystoma tigrinum, is driven excitatorily by a cholinergic input. We compared the effects of applying carbachol to those of dopamine application on the responses of rods, horizontal cells, and bipolar cells recorded intracellularly from the isolated, perfused retina of the tiger salamander. Micromolar concentrations of dopamine reduced the amplitudes of rod responses throughout the rods' operating range. The ratio of amplitudes of the cone-driven to rod-driven components of the responses of both horizontal and bipolar cells was increased by activation of both D1 and D2 dopamine receptors. Dopamine acted to uncouple horizontal cells and also off-center bipolar cells, the mechanism in the case of horizontal cells depending only upon activation of D1 receptors. Carbachol, a specific cholinomimetic, applied in five- to ten-fold higher concentrations, produced effects that were essentially identical to those of dopamine. These effects of carbachol were blocked by application of specific dopamine blockers, however, indicating that they are mediated secondarily by dopamine. We propose that the dopamine-releasing amacrine cells in the salamander are under the control of cells, probably amacrine cells, which secrete acetylcholine as their transmitter.

Effects of bicarbonate versus HEPES buffering on measured properties of neurons in the salamander retina.

Electrophysiological studies of the isolated retina involve perfusing the tissue with a physiological Ringer's. Organic pH buffers such as HEPES have become increasingly popular in recent years because for many purposes they offer a convenient and reliable alternative to the more traditional bicarbonate/CO2. In this paper, however, we report that important functional properties of rods, bipolar cells, and horizontal cells in the salamander, Ambystoma tigrinum, are sensitive to the choice of buffer and, in the case of horizontal cells, that sensitivity is acute. In bicarbonate/CO2 Ringer's, the dark potential of the horizontal cell was typically near -50 mV and saturating light caused it to hyperpolarize to about -75 mV. On switching to HEPES-buffered Ringer's at the same pH, horizontal cells depolarized in darkness to about -20 mV, close to the chloride equilibrium potential, and the kinetics of their light responses changed. The cone-driven components of light responses increased in size relative to rod-driven components. Saturating lights still hyperpolarized the cells to -75 mV, however. Horizontal cells, being coupled via gap junctions, form a syncytium and syncytial length constants, measured in bicarbonate/CO2 Ringer's, were generally in the range 150-225 microm. On switching to HEPES-buffered Ringer's, length constants increased substantially to 250-330 microm. All these changes were reversible. We discuss our findings within the context of the cell's ability to regulate its internal pH.

Receptive field of the retinal bipolar cell: a pharmacological study in the tiger salamander.

1. It is widely believed that signals contributing to the receptive field surrounds of retinal bipolar cells pass from horizontal cells to bipolar cells via GABAergic synapses. To test this notion, we applied gamma-aminobutyric acid (GABA) agonists and antagonists to isolated, perfused retinas of the salamander Ambystoma tigrinum while recording intracellularly from bipolar cells, horizontal cells, and photoreceptors. 2. As we previously reported, administration of the GABA analogue D-aminovaleric acid in concert with picrotoxin did not block horizontal cell responses or the center responses of bipolar cells but blocked the surround responses of both on-center and off-center bipolar cells. 3. Surround responses were not blocked by the GABA, antagonists picrotoxin or bicuculline, the GABAB agonist baclofen or the GABAB antagonist phaclofen, and the GABAC antagonists picrotoxin or cis-4-aminocrotonic acid. Combinations of these drugs were similarly ineffective. 4. GABA itself activated a powerful GABA uptake mechanism in horizontal cells for which nipecotic acid is a competitive agonist. It also activated, both in horizontal cells and bipolar cells, large GABAA conductances that shunted light responses but that could be blocked by picrotoxin or bicuculline. 5. GABA, administered together with picrotoxin to block the shunting effect of GABAA activation, did not eliminate bipolar cell surround responses at concentrations sufficient to saturate the known types of GABA receptors. 6. Surround responses were not blocked by glycine or its antagonist strychnine, or by combinations of drugs designed to eliminate GABAergic and glycinergic pathways simultaneously. 7. Although we cannot fully discount the involvement of a novel GABAergic synapse, the simplest explanation of our findings is that the primary pathway mediating the bipolar cell's surround is neither GABAergic nor glycinergic.

Signal transfer from photoreceptors to bipolar cells in the retina of the tiger salamander.

Under conditions of dark-adaptation, in response to weak stimuli, the distal retina behaves as a linear system. During the process of voltage transfer from the rods to the bipolar cells the information encoded in the rod responses is spatially filtered. The spatial filtering is determined by the spatial properties of the receptive field of the bipolar cell. These, in turn, depend upon the spatial properties of three syncytia, those of the receptors, the horizontal cells and the bipolar cells themselves. The response of the bipolar cell to these weak stimuli is a linear difference of two components; a component generated by the receptive field center and a component generated by the receptive field surround. The receptive field surround is misnamed since it extends throughout the receptive field center and contributes to the response of the bipolar cell to stimuli located anywhere within the receptive field. The receptive field surround has the spatial properties that would be expected if it were generated by an input from the horizontal cells to the receptive field center of the bipolar cell. The cellular pathway mediating this input remains unclear though we have evidence that it involves, at least in part, a feedforward pathway from horizontal cells to bipolar cells. If a feedback pathway also exists it is not mediated by the GABAA synapse on the synaptic terminals of the cones.

Spatial organization of the bipolar cell's receptive field in the retina of the tiger salamander.

1. The spatial properties of rods, horizontal cells and bipolar cells were studied by intracellular recording in the isolated, perfused retina of the tiger salamander, Ambystoma tigrinum. Low stimulus intensities were used in order to keep cell responses close to, or within, their linear intensity/response range. 2. Spatial properties of bipolar cell receptive fields, measured while perfusing with normal Ringer solution, were compared with those measured during exposure to agents that eliminated the bipolar cells' receptive field surround (RFS). In this way, the spatial properties of the receptive field centre (RFC) and those of the RFS could be characterized independently. 3. To a good approximation, the contribution to the horizontal cell's response of unit area of its receptive field declined exponentially with distance from the centre of the receptive field. The (apparent) length constant describing this decay was 200 microns. The one-dimensional length constant of the horizontal cell syncytium was thus 248 microns. The variation of response amplitude with the radius of a centred circular stimulus was consistent with this finding. 4. This was true also of the RFCs of bipolar cells. The one-dimensional length constant of the RFC of off-centre bipolar cells averaged 124 microns. That of the RFC of on-centre cells averaged 62 microns though values were more variable, the RFCs of some on-centre cells being comparable to those of off-centre cells. These values were independent of the class of photoreceptor driving the bipolar cell. 5. The large size of the RFCs of off-centre cells and many on-centre cells cannot by explained by light scatter within the retina or by voltage spread within the rod syncytium. We proposed that off-centre cells are tightly coupled in a syncytium. On-centre cells, on average, are less tightly coupled. 6. The spatial properties of the bipolar cell's RFS were consistent with the notion that the RFS represents a convolution of the horizontal cell's receptive field and the bipolar cell's RFC. 7. The spatial properties of bipolar cell receptive fields were reconstructed from the measured properties of their RFCs and the measured properties of horizontal cell receptive fields. Under the conditions of our experiments, the bipolar cell's response could be described by a linear difference between a component generated by the RFC and a component generated by the RFS. 8. The spatial filtering characteristics of the bipolar cells were calculated from our data.(ABSTRACT TRUNCATED AT 400 WORDS)

Effects of 2-amino-4-phosphonobutyric acid on cells in the distal layers of the tiger salamander's retina.

1. We studied the effects of 2-amino-4-phosphonobutyric acid (APB) on the response properties of rods, horizontal cells and bipolar cells in the isolated, perfused retina of the tiger salamander, Ambystoma tigrinum. A concentration of 100 microM was found to be sufficient to elicit maximal effects. 2. Rods hyperpolarized slightly upon exposure to 100 microM-APB and their response amplitudes were slightly reduced. The amplitude of the cone-generated component of the rod's response to 700 nm light was not significantly affected by APB. 3. Horizontal cells hyperpolarized by 2-5 mV upon exposure to 100 microM-APB. The rod-driven component of the horizontal cell response increased in amplitude while the cone-driven component decreased in amplitude. APB thus causes an increase in voltage gain between rods and horizontal cells and a decrease in cone/horizontal cell gain. These findings can be explained in terms of an APB-induced reduction in transmitter release from the cones. 4. APB at a concentration of 100 microM caused an increase in the length constant of the horizontal cell syncytium. Our analysis shows this to be due primarily to a 50% reduction in the coupling impedance between the cells of the syncytium. 5. The effects of APB on off-centre bipolar cells were qualitatively similar to those on horizontal cells. APB increased the amplitudes of rod-driven responses and reduced those of cone-driven responses. The length constants, both of the receptive field centre and of the surround, were increased and the strength of the surround relative to the centre was reduced by about 20%. 6. APB abolished the depolarizing light responses of the receptive field centres of on-centre bipolar cells. A hyperpolarizing response remained whose spatial properties were similar to those of the receptive field surround. We believe this response to reflect a direct (feedforward) input to on-centre bipolar cells from horizontal cells.

Neuropathology of the hyperkinetic child.

A subgroup of hyperactive children with minimal brain dysfunction may be identified by means of specific physiological testing procedures. These children display a consistent pattern of behavioral symptoms in association with pharmacological, electrophysiological and specific "soft" neurological signs of CNS dysfunction.

Voltage gain of signal transfer from retinal rods to bipolar cells in the tiger salamander.

1. Intracellular recordings of the voltage responses of rods and both functional classes of bipolar cell were made in the isolated, perfused retina of the tiger salamander, Ambystoma tigrinum. 2. Brief, dim flashes of 519 nm light delivered to the receptive-field centres were used to measure the flash sensitivities of twenty-one on-centre bipolar cells and thirty-six off-centre cells. In each experiment the flash sensitivity of a rod was also measured using diffuse illumination of the same duration and wave-length. 3. The mean flash sensitivity of the rods (fifty-nine cells) was 4.47 mV photon-1 micron 2 flash. The mean flash sensitivity of the off-centre bipolar cells was 35.4 mV photon-1 micron 2 flash (thirty-six cells). The mean flash sensitivity of the on-centre bipolar cells was 12.5 mV photon-1 micron 2 flash. 4. The ratio of the flash sensitivity of the bipolar cell to that of a rod recorded in the same retina defined the gain of voltage transfer from rod to bipolar cell. For signal transfer to on-centre bipolar cells the mean value of the voltage gain was 5.05 +/- 1.34 (S.E. of mean). For signal transfer to the off-centre bipolar cells, the mean value of the gain was 10.4 +/- 1.29. 5. The on-centre cell gain in the salamander was smaller by a factor of 27 than that of the on-centre cells in the dogfish retina (Ashmore & Falk, 1980 a), while the off-centre cell gain was comparable in the two species. Possible reasons for the large difference between the voltage gains of on-centre cells in the dogfish and salamander are considered.