Genetic analysis of the Robo3+ glycinergic amacrine cell

项目摘要 视网膜是研究神经回路发育和功能的极好模型。的 视网膜包含五个主要的神经元类别，其可以被细分为远远超过一百个神经元类别。 亚型无长突细胞（AC）是五种神经元之一，仅占视网膜细胞的1%， 但占该亚型多样性的一半左右。只有少数AC亚型 这是因为缺乏前瞻性地分离和操纵个体种群的工具。 因此，我们对视觉回路功能的理解仍然不完整。例如，最近的工作 显示方向选择回路，视网膜中研究最充分的回路之一，包括甘氨酸能神经元。 来自未定义AC子类型的输入。为了充分了解AC如何对视觉处理做出贡献， 研究更多的AC亚型及其定义特征势在必行。 在初步研究中，我确定了一个Robo 3CreER小鼠系，它在遗传上标记了一个种群， 以前未描述的甘氨酸能无长突细胞。根据它们的形态和分层模式， 假设这些Robo 3+无长突细胞（RAC）可能是向ON提供甘氨酸能抑制的细胞 方向选择回路中的星状无长突细胞。我将使用老鼠遗传学，共聚焦显微镜， 原位杂交，电子显微镜和电生理学来验证这一假设。在目标1中，我将定义 RAC的形态学特性和群体水平组织。在目标2中，我将确定RAC是否 在ON星状无长突细胞上形成抑制性突触。在目标3中，我将确定Robo 3是否扮演一个 在RAC开发和/或功能中的作用。这些目标将拓宽我们对个体细胞如何 在视网膜回路中，这些成分一起起作用。

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.