Horizontal cells from the white perch were isolated by enzymatic treatment and trituration of the retina and were maintained in culture for 1-5 days.

The action of many neuromodulators is mediated by intracellular second messengers such as cyclic AMP. In the retina, exogenously applied dopamine alters the conductance of gap junctions between cultured horizontal cells and this effect is mediated by cyclic AMP. However, it is not known how cyclic AMP modulates horizontal cell gap junction function. Here I report that cyclic AMP works by way of a cyclic AMP-dependent protein kinase. Cyclic AMP-dependent protein kinase injected into coupled horizontal cells from white bass (Roccus chrysops) rapidly and reversibly uncoupled the cells, mimicking the actions of dopamine. The threshold for the effect was between 0.06 and 0.03 FM. Injection of Walsh inhibitor of protein kinase

Horizontal cells were enzymatically isolated from the carp retina and maintained in culture for 2-7 days. Cultured horizontal cells typically had resting membrane potentials of -50 to -70 mV and input resistances of 100-150mt. The cells were treated with a number of neurotransmitter agents and their analogues. Significant responses were evoked only by 3,4-dihydroxyphenylalanine (dopamine), L-glutamate, and certain glutamate analogues. The responses to dopamine were inconsistent; most often, the membrane hyperpolarized and input resistances increased. However, highly characteristic responses to L-glutamate and its analogues, quisqualate and kainate, were observed in virtually all of the cells tested. The responses consisted of an initial graded depolarization accompanied by a resistance increase, followedin most cases by a prolonged (1-to 2-min) regener, ative depolarization. The regenerative component ofthe response appears to be Ca2+ dependent, while the underlying graded potential may be due to a decrease in K+ conductance of the membrane.Horizontal cells are second-order neurons believed to mediate lateral inhibitory effects in the outer plexiform layer ofthe retina (1-3). They respond to light with sustained graded potentials (4); in situ, these neurons have never been observed to generate action potentials. In the dark-adapted retina, horizontal cells are maintained in a partially depolarized state by neurotransmitter continuously released by the photoreceptors (5,-6). During illumination, horizontal cells hyperpolarize because transmitter flow from the receptors decreases.' Some horizontal cells respond differentially to color, however; certain wavelengths hyperpolarize the cells whereas other wavelengths depolarize them (4, 7, 8).The identity of the photoreceptor transmitters has not been established. It has been found that the acidic amino acids, Lglutamate and L-aspartate, depolarize horizontal cells in several species and block light-evoked responses (9-11). A recent study in the carp found L-aspartate to be more potent than L-glutamate in this regard and showed that an aspartate antagonist, aaminoadipic acid, blocked both the effects of the natural photoreceptor transmitter and exogenously applied aspartate on many horizontal cells (12). However, in most studies, high concentrations (5-50 mM) of either aspartate or glutamate were used to elicit responses in horizontal cells, and this has led several authors to question whether either of the substances could be the natural photoreceptor transmitter (9)(10)(11)13).The horizontal cells of fish are especially large, which has made them. particularly useful for physiological and pharmacological studies. Fish .horizontal cells can also be readily isolated by enzymatic dissociation ofthe retina, and this technique has been used for biochemical studies of these cells (14,15). As yet, few physiological studies of isolated horizontal cells have been reported (16)(17)(18), and no information concerning the pharmacological properties of isolated horizont...

Horizontal cells from the retinas of white perch were isolated and maintained in cell culture for 3 days to 3 wk. Four morphologically distinct types of horizontal cells could be identified in culture and were labeled types H1, H2, H3, and H4. Whole-cell patch-clamp techniques were used to study the ionic currents present in the four cell types. In all cells, depolarizing commands above threshold elicited a fast-inward current followed by an outward current. The fast-inward current was abolished by tetrodotoxin (TTX) or 0 Na+ Ringer's, indicating the current was carried by Na+. In H1, H2, and H3 cells, the outward current, carried by K+, consisted of two components: a transient current (IA), blockable with 4-aminopyridine (4-AP), tetraethylammonium (TEA), or intracellular cesium and a sustained current that could be blocked with TEA. The H4 cell had only the sustained current. An inward rectifying K+ current (anomalous rectifier) was observed in the four cell types. The current was sensitive to the extracellular K+ concentration. Its activation showed two components: an instantaneous component and a slower component. The slow component becomes faster with greater hyperpolarizations. The four cell types possessed a small, sustained Ca2+ current that, under normal conditions, was masked by the inward Na+ current and outward K+ currents.

1. Retinal bipolar cells were isolated from white bass retinas and maintained in a cell culture preparation. Two morphological types of bipolar cells were observed in cell culture. These were labeled large- and small-bipolar cells based mainly on the size of their somata and primary dendrites. Two types of small-bipolar cells were observed. Isolated bass bipolar cells are very similar to those described in the intact retina. 2. Under current clamp, to depolarizing current injection, small-bipolar cells produced a spike followed by a plateau. Large-bipolar cells showed a slow depolarization to a plateau level. 3. Voltage-gated membrane currents were studied using whole-cell patch-clamp techniques. Channel blocking agents were used to define the ion channels found in the membranes of these cells. 4. The large-bipolar cells were found to possess an A-current, a calcium current, and a calcium-dependent potassium current. 5. Large bipolar cells also possessed an inward rectifier that did not correspond to any previously described. 6. The two types of small-bipolar cells were found to have very similar membrane properties to one another. They lacked a large A-current but possessed a slowly activating, outward rectifying potassium current. Similar to the large-bipolar cells, they showed a calcium current and a calcium-activated potassium current. 7. The inward rectifier of small-bipolar cells was characterized as an H-current. 8. The results suggest that the membrane currents of bipolar cells set a narrow operating range about which the cells function in the intact retina. In addition these currents help shape the responses of bipolar cells to light stimuli but do not confer ON or OFF properties.