Membrane Potential Of Horizontal Cells Research Articles

Horizontal Cell Feedback to Cone Photoreceptors in Mammalian Retina: Novel Insights From the GABA-pH Hybrid Model.

How neurons in the eye feed signals back to photoreceptors to optimize sensitivity to patterns of light appears to be mediated by one or more unconventional mechanisms. Via these mechanisms, horizontal cells control photoreceptor synaptic gain and enhance key aspects of temporal and spatial center-surround receptive field antagonism. After the transduction of light energy into an electrical signal in photoreceptors, the next key task in visual processing is the transmission of an optimized signal to the follower neurons in the retina. For this to happen, the release of the excitatory neurotransmitter glutamate from photoreceptors is carefully regulated via horizontal cell feedback, which acts as a thermostat to keep the synaptic transmission in an optimal range during changes to light patterns and intensities. Novel findings of a recently described model that casts a classical neurotransmitter system together with ion transport mechanisms to adjust the alkaline milieu outside the synapse are reviewed. This novel inter-neuronal messaging system carries feedback signals using two separate, but interwoven regulated systems. The complex interplay between these two signaling modalities, creating synaptic modulation-at-a-distance, has obscured it’s being defined. The foundations of our understanding of the feedback mechanism from horizontal cells to photoreceptors have been long established: Horizontal cells have broad receptive fields, suitable for providing surround inhibition, their membrane potential, a function of stimulus intensity and size, regulates inhibition of photoreceptor voltage-gated Ca2+ channels, and strong artificial pH buffering eliminates this action. This review compares and contrasts models of how these foundations are linked, focusing on a recent report in mammals that shows tonic horizontal cell release of GABA activating Cl− and HCO3− permeable GABA autoreceptors. The membrane potential of horizontal cells provides the driving force for GABAR-mediated HCO3− efflux, alkalinizing the cleft when horizontal cells are hyperpolarized by light or adding to their depolarization in darkness and contributing to cleft acidification via NHE-mediated H+ efflux. This model challenges interpretations of earlier studies that were considered to rule out a role for GABA in feedback to cones.

Calcium-activated BKCa channels govern dynamic membrane depolarizations of horizontal cells in rodent retina.

Large conductance, Ca2+ -activated K+ (BKCa ) channels play important roles in mammalian retinal neurons, including photoreceptors, bipolar cells, amacrine cells and ganglion cells, but they have not been identified in horizontal cells. BKCa channel blockers paxilline and iberiotoxin, as well as Ca2+ free solutions and divalent cation Cav channel blockers, eliminate the outwardly rectifying current, while NS1619 enhances it. In symmetrical 150mm K+ , single channels had a conductance close to 250pS, within the range of all known BKCa channels. In current clamped horizontal cells, BKCa channels subdue depolarizing membrane potential excursions, reduce the average resting potential and decrease oscillations. The results show that BKCa channel activation puts a ceiling on horizontal cell depolarization and regulates the temporal responsivity of the cells. Large conductance, calcium-activated potassium (BKCa ) channels have numerous roles in neurons including the regulation of membrane excitability, intracellular [Ca2+ ] regulation, and neurotransmitter release. In the retina, they have been identified in photoreceptors, bipolar cells, amacrine cells and ganglion cells, but have not been conclusively identified in mammalian horizontal cells. We found that outward current recorded between -30 and +60mV is carried primarily in BKCa channels in isolated horizontal cells of rats and mice. Whole-cell outward currents were maximal at +50mV and declined at membrane potentials positive to this value. This current was eliminated by the selective BKCa channel blocker paxilline (100nm), iberiotoxin (10μm), Ca2+ free solutions and divalent cation Cav channel blockers. It was activated by the BKCa channel activator NS1619 (30μm). Single channel recordings revealed the conductance of the channels to be 244±11pS (n=17; symmetrical 150mm K+ ) with open probability being both voltage- and Ca2+ -dependent. The channels showed fast activation kinetics in response to Ca2+ influx and inactivation gating that could be modified by intracellular protease treatment, which suggests β subunit involvement. Under current clamp, block of BKCa current increased depolarizing membrane potential excursions, raising the average resting potential and producing oscillations. BKCa current activation with NS1619 inhibited oscillations and hyperpolarized the resting potential. These effects underscore the functional role of BKCa current in limiting depolarization of the horizontal cell membrane potential and suggest actions of these channels in regulating the temporal responsivity of the cells.

Sources of protons and a role for bicarbonate in inhibitory feedback from horizontal cells to cones in Ambystoma tigrinum retina.

In the vertebrate retina, photoreceptors influence the signalling of neighbouring photoreceptors through lateral-inhibitory interactions mediated by horizontal cells (HCs). These interactions create antagonistic centre-surround receptive fields important for detecting edges and generating chromatically opponent responses in colour vision. The mechanisms responsible for inhibitory feedback from HCs involve changes in synaptic cleft pH that modulate photoreceptor calcium currents. However, the sources of synaptic protons involved in feedback and the mechanisms for their removal from the cleft when HCs hyperpolarize to light remain unknown. Our results indicate that Na+ -H+ exchangers are the principal source of synaptic cleft protons involved in HC feedback but that synaptic cleft alkalization during light-evoked hyperpolarization of HCs also involves changes in bicarbonate transport across the HC membrane. In addition to delineating processes that establish lateral inhibition in the retina, these results contribute to other evidence showing the key role for pH in regulating synaptic signalling throughout the nervous system. Lateral-inhibitory feedback from horizontal cells (HCs) to photoreceptors involves changes in synaptic cleft pH accompanying light-evoked changes in HC membrane potential. We analysed HC to cone feedback by studying surround-evoked light responses of cones and by obtaining paired whole cell recordings from cones and HCs in salamander retina. We tested three potential sources for synaptic cleft protons: (1) generation by extracellular carbonic anhydrase (CA), (2) release from acidic synaptic vesicles and (3) Na+ /H+ exchangers (NHEs). Neither antagonizing extracellular CA nor blocking loading of protons into synaptic vesicles eliminated feedback. However, feedback was eliminated when extracellular Na+ was replaced with choline and significantly reduced by an NHE inhibitor, cariporide. Depriving NHEs of intracellular protons by buffering HC cytosol with a pH 9.2 pipette solution eliminated feedback, whereas alkalinizing the cone cytosol did not, suggesting that HCs are a major source for protons in feedback. We also examined mechanisms for changing synaptic cleft pH in response to changes in HC membrane potential. Increasing the trans-membrane proton gradient by lowering the extracellular pH from 7.8 to 7.4 to 7.1 strengthened feedback. While maintaining constant extracellular pH with 1mm HEPES, removal of bicarbonate abolished feedback. Elevating intracellular bicarbonate levels within HCs prevented this loss of feedback. A bicarbonate transport inhibitor, 4,4'-diisothiocyano-2,2'-stilbenedisulfonic acid (DIDS), also blocked feedback. Together, these results suggest that NHEs are the primary source of extracellular protons in HC feedback but that changes in cleft pH accompanying changes in HC membrane voltage also require bicarbonate flux across the HC membrane.

Oral Proton Pump Inhibitors Disrupt Horizontal Cell-Cone Feedback and Enhance Visual Hallucinations in Macular Degeneration Patients

Visual hallucinations (VHs) occur in macular degeneration patients with poor vision but normal cognitive function. The underlying mechanisms are poorly understood. We report the identification of pharmaceutical agents that enhance VH and use these agents to examine the contribution of retinal neurons to this syndrome. We detail clinical observations on VH in five macular degeneration patients treated with proton pump inhibitors having the core structure, 2-pyridyl-methylsulfinyl-benzimidazole. We tested possible retinal mechanisms using paired whole cell recordings to examine effects of these compounds on feedback interactions between horizontal cells and cones in amphibian retina. Five patients with advanced wet macular degeneration described patterned VHs that were induced or enhanced by oral proton pump inhibitors. The abnormal images increased with light, disappeared in the dark, and originated in the retina, based on ophthalmodynamometry. Simultaneous paired whole cell recordings from amphibian cones and horizontal cells showed that 2-pyridyl-methylsulfinyl-benzimidazoles blocked the negative shift in voltage dependence and increase in amplitude of the calcium current (ICa) in cones that is induced by changes in horizontal cell membrane potential. These effects disrupt the negative feedback from horizontal cells to cones that is important for the formation of center-surround receptive fields in bipolar and ganglion cells, and thus for normal spatial and chromatic perception. Our study suggests that changes in the output of retinal neurons caused by disturbances in outer retinal feedback mechanisms can enhance patterned visual hallucinations.

A Positive Feedback Synapse from Retinal Horizontal Cells to Cone Photoreceptors

Cone photoreceptors and horizontal cells (HCs) have a reciprocal synapse that underlies lateral inhibition and establishes the antagonistic center-surround organization of the visual system. Cones transmit to HCs through an excitatory synapse and HCs feed back to cones through an inhibitory synapse. Here we report that HCs also transmit to cone terminals a positive feedback signal that elevates intracellular Ca2+ and accelerates neurotransmitter release. Positive and negative feedback are both initiated by AMPA receptors on HCs, but positive feedback appears to be mediated by a change in HC Ca2+, whereas negative feedback is mediated by a change in HC membrane potential. Local uncaging of AMPA receptor agonists suggests that positive feedback is spatially constrained to active HC-cone synapses, whereas the negative feedback signal spreads through HCs to affect release from surrounding cones. By locally offsetting the effects of negative feedback, positive feedback may amplify photoreceptor synaptic release without sacrificing HC-mediated contrast enhancement.

Horizontal cell feedback regulates calcium currents and intracellular calcium levels in rod photoreceptors of salamander and mouse retina.

We tested whether horizontal cells (HCs) provide feedback that regulates the Ca(2+) current (I(Ca)) of rods in salamander and mouse retinas. In both species, hyperpolarizing HCs by puffing a glutamate antagonist, 6,7-dinitro-quinoxaline-2,3-dione (DNQX), onto HC processes caused a negative shift in the voltage dependence of rod I(Ca) and increased its peak amplitude. Conversely, depolarizing HCs by puffing kainic acid (KA) into the outer plexiform layer (OPL) caused a positive voltage shift and decreased rod I(Ca.) Experiments on salamander retina showed that these effects were blocked by addition of the pH buffer, Hepes. Intracellular calcium concentration ([Ca(2+)](i)) was examined in rods by confocal microscopy after loading salamander and mouse retinal slices with Fluo-4. Rods were depolarized to near the dark resting potential by bath application of high K(+) solutions. Hyperpolarizing HCs with 2,3-dihydroxy-6-nitro-7-sulphamoylbenzo[f]quinoxaline (NBQX) enhanced high K(+)-evoked Ca(2+) increases whereas depolarizing HCs with KA inhibited Ca(2+) increases. In both species these effects of NBQX and KA were blocked by addition of Hepes. Thus, like HC feedback in cones, changes in HC membrane potential modulate rod I(Ca) thereby regulating rod [Ca(2+)](i) at physiological voltages, in both mouse and salamander retinas. By countering the reduced synaptic output that accompanies hyperpolarization of rods to light, HC feedback will subtract spatially averaged luminance levels from the responses of individual rods to local changes. The finding that HC to rod feedback is present in both amphibian and mammalian species shows that this mechanism is highly conserved across vertebrate retinas.

Feedback Effects of Horizontal Cell Membrane Potential on Cone Calcium Currents Studied With Simultaneous Recordings

Horizontal cell (HC) to cone feedback helps establish the center-surround arrangement of visual receptive fields. It has been shown that HC activity influences cone synaptic output by altering the amplitude and voltage dependence of the calcium current (ICa) in cones. In this study, we obtained voltage-clamp recordings simultaneously from cones and HCs to directly control the membrane potential of HCs and thereby measure the influence of HC membrane potential changes on ICa in adjacent cones. Directly hyperpolarizing voltage clamped HCs produced a negative activation shift and increased the amplitude of ICa in cones. Both of these effects were abolished by enhancing extracellular pH buffering capacity with HEPES. In contrast, addition of the gap junction blocker, carbenoxolone, did not significantly alter the shifts or amplitude changes in cone ICa produced by changes in HC membrane potential. These results support the hypothesis that changes in the HC membrane potential alter the voltage dependence and amplitude of cone ICa by altering extracellular pH levels at the synapse.

Modulation of horizontal cell function by GABA(A) and GABA(C) receptors in dark- and light-adapted tiger salamander retina.

The physiological function of GABA transporters and GABA receptors in retinal horizontal cells (HCs) under dark-and light-adapted conditions were studied by whole-cell voltage clamp and intracellular recording techniques in retinal slices and whole-mounted isolated retinas of the larval tiger salamander. Puff application of GABA in picrotoxin elicited a NO-711 (a potent GABA transporter blocker)-sensitive inward current that did not exhibit a reversal potential in the physiological range, consistent with the idea that these HCs contain electrogenic GABA transporters. Application of GABA in NO-711 elicited a chloride current in HCs; about half of the current was suppressed by bicuculline or I4AA (a GABA(C) receptor antagonist), and the remaining half was suppressed by bicuculline + I4AA or picrotoxin. In whole-mount retinas, NO-711, bicuculline, I4AA, or picrotoxin hyperpolarized the HCs and enhanced the light responses under dark-adapted conditions, and blocked the time-dependent recovery of HC membrane potential and light responses during background illumination. Based on the parallel conductance model, GABA released in darkness mediates a chloride conductance about three times greater than the leak conductance or the glutamate-gated cation conductance. About half of this chloride conductance is mediated by GABA(A) receptors, and the other half is mediated by GABA(C) receptors. These results suggest that GABA released from HCs through the NO-711-sensitive GABA transporters activates GABA(A) and GABA(C) receptors, resulting in chloride conductance increase which leads to a HC depolarization and reduction of the light response. Additionally, GABA transporters also mediate GABA release in background light that is responsible for the recovery of HC membrane potential and light responses.

Effects of glutamic acid and related agents on horizontal cells in a marine teleost retina.

Excitatory amino acids (EAAs) such as glutamic and aspartic acids, considered as the most likely neurotransmitters at the photoreceptor-horizontal cell synapse of teleost retinas, as well as agonists such as kainic acid and several of their antagonists, were applied to isolated and superfused retinas of the teleost Eugerres plumieri. Intracellular recordings from horizontal cells reveal that EAA receptors are of the kainate-quisqualate type. There is competitive inhibition between the agonist and antagonist agents used, and under their combined effect, the synapse under study remains operational, in a functional state, able to modulate the horizontal cell membrane potential upon retinal illumination.

A method for estimating gap junctional conductance between the retinal horizontal cells

AbstractGap junction between retinal horizontal cells plays an important role in generating the center‐surround antagonistic receptive field. However, there are technical difficulties involved in measurement of the gap junctional conductance from membrane potential of horizontal cells using the ionic current model. To evaluate the proposed method, the gap junctional conductance was estimated from test data generated by the horizontal cell layer model. Conductance can be estimated with high accuracy. This method is also applied to experimental data recorded from the L‐type cell of carp retina.

Voltage- and time-dependent potassium conductances enhance the frequency response of horizontal cells in the turtle retina

The contribution of voltage- and time-dependent potassium conductances to visual information processing in the distal turtle retina was studied in the isolated retina preparation. The effects of specific potassium channel blockers; tetraethylammonium (TEA) and 4-aminopyridine (4-AP) on the membrane potential and photoresponses of L-cones and L-type horizontal cells were monitored with intracellular microelectrodes. Both drugs produced a large depolarization of the L-type horizontal cells though the effect of 4-AP was more transient than that of TEA. While TEA produced response augmentation associated with negligible changes in the kinetics of the photoresponses, 4-AP induced profound changes in response kinetics which were seen as an overshoot of the resting potential at stimulus offset and a pronounced slowing down in the return of the membrane potential toward the prestimulus level. The effects of TEA on horizontal cells could be accounted for by the action of the drug on cone photoreceptors. The effects of 4-AP on the horizontal cells could not be attributed to an indirect action mediated by either the cone photoreceptors or by GABAergic and/or glycinergic neurons in the inner retina. These results suggest that voltage- and time-dependent potassium conductances act to speed up the recovery of the turtle horizontal cell membrane potential from the effects of bright light stimuli. Such a role was supported by the effects of potassium channel blockers on the frequency response curves of horizontal cells; the corner frequency was reduced on the average by 25%.

Effects of GABA on horizontal cells in the tiger salamander retina

Application of gamma-aminobutyric acid (GABA) slows down the horizontal cell response time course (HCRRT) and induces membrane depolarization in horizontal cells (HCs) after synaptic inputs are blocked by Co2+. We present evidence that suggests both effects are probably mediated by GABAA receptors which open chloride channels in the HC membrane. In any given concentration of GABA, ranged from 0 to 100 microM, the HC membrane potential (VHC) in saturating light and in the presence of 100 microM Co2+ are identical. This result suggests that GABA in both light and 100 microM Co2+ opens the same amount of chloride channels (same gCl) so that VHC determined by chloride and leak conductances has the same value. Higher concentrations of Co2+ (> 300 microM) not only blocks synaptic transmission from photoreceptors to HCs, but also acts as an antagonist that suppresses the GABA-mediated depolarization in HCs.

Modulation of cone to horizontal cell transmission by calcium and pH in the fish retina.

The effects of small changes in the calcium and sodium concentrations and in the pH of superfusion medium on the membrane potential and light-evoked responses of cone horizontal cells in the goldfish retina were examined. Conventional intracellular recording, a bicarbonate-based superfusion medium, and a specially designed superfusion apparatus that reduced pressure wave disturbances were used. An increase in the extracellular calcium concentration, [Ca2+]o, from control levels (0.1 mM) to 1.0 mM hyperpolarized cone horizontal cells and reduced the magnitude of their light responses at all stimulus intensities. Addition of 20 mM NaCl to the 1.0 mM Ca2+ Ringer's solution reversed the hyperpolarizing effect of the 1.0 mM Ca2+ but addition of 20 mM choline, a monovalent cation that does not pass through cyclic GMP-activated channels, did not. Reduction of the superfusate pH from 7.6 to 7.2 by switching from a Ringer's solution gassed with 3% CO2 to one gassed with 10% CO2 hyperpolarized horizontal cells and reduced the magnitude of their light responses at all stimulus intensities for both 0.1 and 1.0 mM Ca2+ Ringer's solutions. An increase in pH to 8.2 by gassing the superfusate with 1% CO2 slightly depolarized the cells in 0.1 mM Ca2+ Ringer's solution but slightly hyperpolarized the cells in the 1.0 mM Ca2+ Ringer's solution. Following pharmacological isolation of the horizontal cells from synaptic input with high doses of glutamate (4-5 mM) and/or Co2+ (4 mM) treatment, no effect on horizontal cell membrane potential due to changes in pHo or [Ca2+]o was observed. These findings are discussed with respect to the cellular mechanisms and sites of action in the outer retina that are affected by changes in pHo and [Ca2+]o.

GABA-mediated positive autofeedback loop controls horizontal cell kinetics in tiger salamander retina

Horizontal cells (HCs) appear to release, and also to be sensitive to, GABA. The external GABA concentration is increased with depolarization of the HC membrane via an electrogenic GABA transporter. This extracellular GABA opens a GABAA-gated Cl- channel in the HC membrane. Since the equilibrium potential for Cl- (ECl) is near -20 mV, GABA released by the HC further depolarizes the HC. The GABA transporter and the GABAA receptor thus constitute a positive feedback loop in the HC membrane. This loop can slow down the kinetics of the light responses in HCs. GABA released via the GABA transporter can affect the GABAA receptor, probably because diffusion from the extracellular space is normally restricted by the intact retinal structure. We therefore used retinal slices rather than isolated HCs to maintain that structure. To measure single-cell currents in the slice, HCs were electrically uncoupled by including cAMP in the patch pipette. Under these conditions, bath application of GABA elicited two currents: (1) a picrotoxin-blocked current reversing near ECl, probably mediated by GABAA receptors, and (2) a picrotoxin-insensitive current similar to that elicited by cis-4-hydroxynipecotic acid (NIP) shown in other preparations to act at the GABA transporter. Under physiological conditions, the HC membrane potential is controlled by two major conductances, the GABAA-gated Cl- conductance described above, and the glutamate-gated conductance modulated by photoreceptor input. A bright light flash eliminates the glutamate-gated conductance, leaving only the GABA-gated Cl- conductance to control the membrane. With the Cl- conductance a significant fraction of the overall membrane conductance the GABAergic positive feedback loop can decrease the response kinetics. We increased the ambient extracellular GABA concentration by adding 50 microM GABA to the extracellular medium. This increased the ambient Cl- conductance, but the transporter still modulated Cl- conductance because responses to light stimuli were significantly slowed. The slowdown of the HC response could be reversed by interrupting the loop in two ways: (1) picrotoxin opened the loop and speeded the responses by uncoupling the GABA concentration from control of the membrane conductance, and (2) NIP opened the loop by uncoupling the extracellular GABA concentration from the Cl- conductance and therefore the membrane potential.(ABSTRACT TRUNCATED AT 400 WORDS)

Terbium simulates the action of calcium on electrophysiological activity in the isolated fish retina

It has been suggested that terbium (Tb) could be substituted for calcium (Ca) in biological tissue and, because of its useful spectroscopic properties, provide an assessment of calcium distribution in the tissue. In order to assess their physiological effects, we have examined the action of free terbium ions on the membrane potential and light-evoked responses of horizontal cells, and on the massed light evoked response (ERG) of the neural retina. Measurements were performed by intracellular and extracellular recording from the isolated fish retina, superfused with Ringers of various ionic compositions, and certain technical difficulties which occurred in the use of terbium are discussed. We show that Tb ions mimic Ca ions in suppression of the ERG and in hyperpolarization of the horizontal cell membrane potential. The effects of Tb persist after its removal from the superfusate for a longer period than do those of calcium, which indicates that terbium binds more firmly at its site(s) of activity. Tb and Ca both suppress the light-evoked S-potentials from the horizontal cells, although some differences were observed in the waveform of the S-potentials during the onset of suppression. We conclude that the physiological effects of Tb on the retina are essentially similar to those of Ca, and that Tb could, therefore, prove a useful marker of Ca distribution in neural tissue.

Modulation of cone-to-horizontal-cell signal transmission in the turtle retina by magnesium ions

The modulation by magnesium ions of cone-to-horizontal-cell signal transmission was studied in the turtle superfused everted eye-cup preparation using solutions which contained various concentrations of divalent cations and/or chelating agents (EDTA and EGTA). Removal of magnesium ions from ‘normal’ solutions had no apparent effect on the horizontal cells provided that ‘normal’ levels of calcium ions were maintained. Solutions which actively removed calcium but contained ‘normal’ levels of magnesium hyperpolarized horizontal cells, reduced the amplitude of their photoresponses but also changed the character of these photoresponses: dim stimuli evoked purely depolarizing responses while bright flashes elicited triphasic responses. Active removal of both calcium and magnesium from the superfusate and from retinal stores with chelators depolarized the horizontal cell and eliminated light-evoked responses. These solutions also depolarized the cones but augmented their light-evoked responses. We conclude that magnesium ions can modulate the membrane potential of horizontal cells, but this role can be revealed only in the absence of calcium ions. Further, the depolarizing responses seen in low calcium, ‘normal’ magnesium superfusion indicate that the ionic mechanisms underlying the horizontal cell photoresponse are more complex than has been previously described.

GABA release from Xenopus retina does not correlate with horizontal cell membrane potential

The relationship between horizontal cell membrane potential and the release of GABA was explored in the retina of Xenopus laevis. The intracellularly recorded membrane potential of horizontal cells was monitored while the retina was exposed to different concentrations of depolarizing agents. The dose-response curves obtained revealed a rise from 5 to 95% maximum depolarization in 0.5–1.5 log unit concentration change. The molar concentrations that elicited a 20 mV depolarization were 40 mM (potassium), 0.8 mM (glutamate), 0.8 mM (glycine), 5 μM (kainate) and 1.3 μM (quisqualate). Autoradiography revealed that radiolabel was accumulated almost exclusively by horizontal cells when isolated retinas were incubated in medium containing 1 μM [ 3H]GABA. Thus, retinal release of radioactivity was used as a measure of [ 3H]GABA release from horizontal cells. Endogenous GABA released from retinas was measured using high performance liquid chromatography and was taken to reflect both amacrine and horizontal cell GABA pools. The release of both [ 3H]GABA and endogenous GABA was stimulated by glutamate, kainate and potassium, but not by glycine or quisqualate. Similar dose-response curves for GABA release and for depolarization were obtained in the case of potassium and kainate but not for glutamate. Potassium-evoked release either of endogenous GABA or [ 3H]GABA was both calcium- and sodium-dependent, whereas kainate- or glutamate-evoked GABA release was sodium-dependent but calcium-independent. The results indicate that depolarization per se is not necesarily associated with transmitter release in Xenopus retinal horizontal cells. It is suggested that the action of a given neurotransmitter upon the efflux of GABA from horizontal cells may depend on the degree to which it modifies the sodium conductance of the horizontal cell.

Opposite effects of ammonia and carbon dioxide on dye coupling between horizontal cells in the carp retina

Effects of ammonia (NH 3) and carbon dioxide (CO 2) on the membrane potential of horizontal cells and on dye coupling between the cells in isolated retinas of the carp ( Cyprinus carpio) were investigated. Ammonia (<300 ppm NH 3 in air) initially depolarized and subsequently hyperpolarized, while CO 2 (10% in air) hyperpolarized the membrane potential of horizontal cells, accompanied by a diminution of both center and surround responses to spot and annular light stimuli. During the course of amplitude diminution, the center response consistently became smaller with NH 3 and larger with CO 2 than the surround response. In the presence of intravitreally applied DA (50 μM) or amphetamine (100 μM), a fluorescent dye Lucifer Yellow CH (LY) was found to be restricted to single injected horizontal cells. The presence of intravitreal haloperidol (100 μM) for 20–25 min or an exposure of the retina to NH 3 for 5–10 min diffused the restricted LY from single injected cells to numerous neighboring cells. On the other hand, CO 2 was found to restrict the injected dye to single cells, an effect similar to that of DA and opposite to that of NH 3 and haloperidol. The results suggest that NH 3 appears to act as a coupler while CO 2 acts as an uncoupler on gap junctions between horizontal cells in the carp retina, presumably by changing the intracellular pH. In addition, a brief exposure of cells, marked with LY in the presence of DA, to the exciting light 426 nm was found to prevent the NH 3-induced dye diffusion from single cells to their neighbors; the reason is unknown.

Depolarization of retinal horizontal cells by excitatory amino acid neurotransmitter agonists

We have recorded the intracellular membrane potential of horizontal cells, second-order interneurones of the vertebrate retina, from fish retinae perfused with Ringer solution containing agonist drugs of the excitatory neurotransmitters, l-glutamate and l-aspartate. We show that at concentrations greater than about 10 μM, kainate and quisqualate have a potent depolarizing effect on horizontal cells and suppress their light evoked electrical responses (S-potentials). In contrast, a third agonist, N-methyl- d-aspartate, hyperpolarizes horizontal cells. The depolarizing action of kainate and quisqualate persists in the presence of the synaptic blocker, cobalt chloride, which implies that they bind directly on the horizontal cell membranes. Two kainate-related drugs, which are active on invertebrate neurones, were also examined, one, α-ketokainate, simulates the action of kainate, but the other, dihydrokainate, is ineffective on horizontal cells. l-Glutamate binds with high affinity at quisqualate sites, whereas l-aspartate binds with high affinity at NMDA sites, thus we conclude that l-glutamate is the likely neurotransmitter at the photoreceptor-horizontal cell synapse.

Histochemical studies on catecholaminergic cells in the carp retina.

Histochemical studies on catecholaminergic cells were conducted with the carp (Cyprinus carpio) retina. Catecholamine (CA)-containing cell bodies appear sparsely distributed among amacrine cells in the innermost cellular row of the inner nuclear layer (INL) and occasionally in the outer half part of the inner plexiform layer (IPL); only exceptionally are they found among ganglion cells. The fluorescent cells interspersed with the amacrine cells and in the IPL send their fiber processes toward both the outer plexiform layer (OPL) and the IPL; the fine fibers form dense networks in the INL and IPL. Pretreatment of the fish with intramuscular injection of reserpine (20 hr prior to enucleation) completely depleted CA from the retina. The fluorescence of catecholaminergic cells was enhanced, and the number of fluorescent cells visible was increased, by intravitreous injection of L-DOPA, DA, and NA (3 prior to enucleation). A combination of pretreatment with intramuscular reserpine and intravitreous NA was particularly effective. These results indicate that catecholamines may play an important role in the modulation of the membrane potential of horizontal cells.