Project Summary The retina is an excellent model for studying the development and function of neural circuitry. The retina contains five major neuronal classes which can be subdivided into well over a hundred neuronal subtypes. Amacrine Cells (ACs), one of the five neuronal classes, comprise only one percent of the retinal cell population, but account for around half of this subtype diversity. Only a handful of AC subtypes have been studied in depth, in part due to a lack of tools to prospectively isolate and manipulate individual populations. Therefore, our understanding of how visual circuits function remains incomplete. For example, recent work shows that the direction selective circuit, one of the most well-studied circuits in the retina, includes glycinergic input from an undefined AC subtype. To fully understand how ACs contribute to visual processing it is imperative to study more AC subtypes and their defining features. In preliminary studies, I identified a Robo3CreER mouse line that genetically labels a population of previously undescribed glycinergic amacrine cells. Based on their morphology and stratification pattern, I hypothesize that these Robo3+ amacrine cells (RACs) may be the cells that supply glycinergic inhibition to ON starburst amacrine cells in the direction selective circuit. I will employ mouse genetics, confocal microscopy, in situ hybridization, electron microscopy, and electrophysiology to test this hypothesis. In Aim 1, I will define the morphological properties and population-level organization of RACs. In Aim 2, I will determine whether RACs form inhibitory synapses onto ON starburst amacrine cells. In Aim 3, I will determine whether Robo3 plays a role in RAC development and/or function. These aims will broaden our knowledge of how individual cellular components function together in retinal circuitry.