Specific connectivity between photoreceptors and horizontal cells in the zebrafish retina.

The functional and morphological connectivity between various horizontal cell (HC) types (H1, H2, H3, and H4) and photoreceptors was studied in zebrafish retina. Since HCs are strongly coupled by gap junctions and feedback from HCs to photoreceptors depends strongly on connexin (Cx) hemichannels, we characterized the various HC Cxs (Cx52.6, Cx52.7, Cx52.9, and Cx55.5) in Xenopus oocytes. All Cxs formed hemichannels that were conducting at physiological membrane potentials. The Cx hemichannels differed in kinetic properties and voltage dependence, allowing for specific tuning of the coupling of HCs and the feedback signal from HCs to cones. The morphological connectivity between HC layers and cones was determined next. We used zebrafish expressing green fluorescent protein under the control of Cx promoters. We found that all HCs showed Cx55.5 promoter activity. Cx52.7 promoter activity was exclusively present in H4 cells, while Cx52.9 promoter activity occurred only in H1 cells. Cx52.6 promoter activity was present in H4 cells and in the ventral quadrant of the retina also in H1 cells. Finally, we determined the spectral sensitivities of the HC layers. Three response types were found. Monophasic responses were generated by HCs that contacted all cones (H1 cells), biphasic responses were generated by HCs that contacted M, S, and UV cones (H2 cells), and triphasic responses were generated by HCs that contacted either S and UV cones (H3 cells) or rods and UV cones (H4 cells). Electron microscopy confirms that H4 cells innervate cones. This indicates that rod-driven HCs process spectral information during photopic and luminance information during scotopic conditions.

Physiological and molecular characterization of connexin hemichannels in zebrafish retinal horizontal cells.

Connexin channels mediate electrical synaptic transmission when assembled as cell-to-cell pores at gap junctions and can mediate transmembrane currents when expressed in plasma membranes as hemichannels. They are widely expressed in the vertebrate retina where in electrical synapses they are critical for transmission of visual signals. While the roles of connexins in electrical synapses are well-studied, the function and roles of connexin hemichannels in the nervous system are less well understood. Genetic deletion in zebrafish of connexin (Cx) 55.5 alters horizontal cell feedback to cones, spectral responses, and visual behavior. Here, we have characterized the properties of hemichannel currents in zebrafish retinal horizontal cells and examined the roles of two connexin isoforms, Cx55.5 and Cx52.6, that are coexpressed in these cells. We report that zebrafish horizontal cells express hemichannel currents that conduct inward current at physiological negative potentials and Ca(2+) levels. Manipulation of Cx55.5 and Cx52.6 gene expression in horizontal cells of adult zebrafish revealed that both Cx55.5 and Cx52.6 contribute to hemichannel currents; however, Cx55.5 expression is necessary for high-amplitude currents. Similarly, coexpression of Cx55.5 with Cx52.6 in oocytes increased hemichannel currents in a supra-additive manner. Taken together these results demonstrate that zebrafish horizontal cell hemichannel currents exhibit the functional characteristics necessary to contribute to synaptic feedback at the first visual synapse, that both Cx55.5 and Cx52.6 contribute to hemichannel currents, and that Cx55.5 may have an additional regulatory function enhancing the amplitude of hemichannel currents.