Functional Circuitry of Long-Range Connections in the Retina

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Our broad goal is to understand how neural circuits in the retina perform the visual computations that generate healthy vision. Our specific goal in this proposal is to reveal the role of wide-field amacrine cells (WACs) in retinal processing. WACs, a class of GABAergic interneuron, are the largest cells in the retina, and in the mouse they can span nearly half of the retinal area. A conventional model proposes that WACs encode contrast locally over their central dendrites and then spread inhibition globally by action potential (spike)- dependent signaling down millimeter-long axons. This global GABAergic inhibition would tune excitatory (bipolar cell → ganglion cell) retinal circuits and thereby shape ganglion cell responses to local features based on the global statistics of the visual scene. It has been difficult to evaluate WAC function systematically, however, because the field lacks tools for targeting specific WAC types for functional and structural studies. Here, we solve this problem by developing intersectional genetic strategies to label a diverse family of WACs that express the transcription factor Bhlhb5 in the mouse retina. Using a combination of Flp and Cre recombinase expression, in conjunction with dual-reporter alleles, we developed methods to identify Bhlhb5- expressing WAC (B5 WAC) subtypes, which together comprise ~20% of all amacrine cells. Our preliminary studies challenge the conventional model of WAC function. Specifically, we identified a subset of B5 WACs that are non-spiking cells. Ca2+ indicator imaging demonstrates that non-spiking B5 WACs exhibit localized changes in [Ca2+] in dendritic compartments that can be tuned to stimulus properties (e.g., orientation tuning), leading to the hypothesis that despite their large size, these cells are involved primarily in local processing of visual features at multiple points within their wide arbors. Proposed experiments will identify spiking and non-spiking B5 WAC types based on combined genetic, structural and functional criteria and reveal novel roles for these interneurons in retinal function. We will test specific hypotheses about B5 WAC synaptic organization using intersectional optogenetic experiments combined with high-resolution Scanning Block-face Electron Microscopy (SBEM) and STochastic Optical Reconstruction Microscopy (STORM). Our multidisciplinary approach, with unique contributions from four laboratories, will yield new insights into the circuit functions of the retina's largest neurons.