Synaptic transmission in the rod pathway of the mammalian retina

 描述（由申请人提供）：我们研究计划的主要目标是了解神经回路功能如何依赖于组成细胞和突触的内在特性。这个建议的具体目标是确定视杆细胞介导的视觉处理在视网膜上的视杆双极（RB）细胞突触与突触后AII无长突细胞的信号的时间依赖。RB→AII突触处的传递由内在的（例如，囊泡释放和再循环的突触特异性动力学）和外在的（例如，对突触的抑制）形成。 RBs由interneuron电路）因素。该应用程序包括两个具体的目标，整合RB→AII传输的突触和电路级分析。具体目标1测试的新假设，钙离子流入RB终端调节强度和时间的传输在突触的钙调蛋白（CaM）依赖性机制：一个调节膜脂质成分通过调节钙离子非依赖性磷脂酶A2（iPLA 2）;第二个影响自发释放的组成性主动制动。调制的传输进行了研究，通过电生理实验，和时间编码检查视网膜中的RB缺乏一个快速的胞吐模式：复合蛋白（clpx）3敲除。具体目标2决定了聚集在RB上的平行抑制通路的组织和生理功能。扫描块面电子显微镜（SBEM）重建识别抑制性无长突细胞（AC）突触前RB。AC→RB突触的功能研究使用各种电生理技术：配对AC-RB记录，AC的光遗传学刺激，和双极细胞的光遗传学刺激-BCs-突触前AC。与公共卫生的相关性：了解时间编码是如何实现视网膜突触通知视网膜假体的设计和人类视网膜疾病的动物模型的研究。视觉研究的一个目标是开发用于治疗由光感受器变性引起的失明的基于基因的疗法，并且一种有前景的疗法是通过病毒介导的通道视紫红质-2（ChR 2）（一种光门控阳离子通道）的表达在视网膜中间神经元中产生光敏性。我们将在中间神经元中表达ChR 2，以研究视网膜回路中的突触相互作用，从而生成有关视网膜工作范围的关键信息，其中ChR 2是唯一的光传感器。我们提出了国家眼科和视力研究计划中视网膜疾病项目的三个目标：1）确定治疗视网膜色素变性的潜在治疗策略，2）增加对视网膜色素变性的了解， 感光后适应（即兴奋性和抑制性突触之间的相互作用），以及3）增加对神经网络中细胞间相互作用如何产生可解释为视觉图像的信号的理解。

 DESCRIPTION (provided by applicant): The broad goal of our research program is to understand how neural circuit function depends on the intrinsic properties of component cells and synapses. The specific goal of this proposal is to determine how rod-mediated visual processing in the retina depends on the timing of signaling at rod bipolar (RB) cell synapses with postsynaptic AII amacrine cells. Transmission at the RB→AII synapse is shaped by intrinsic (e.g. synapse-specific dynamics of vesicle release and recycling) and extrinsic (e.g. inhibition of RBs by interneuron circuits) factors. This application comprises two specific aims that integrate synapse- and circuit-level analyses of RB→AII transmission. Specific Aim 1 tests the novel hypothesis that Ca2+ influx into the RB terminal regulates the strength and timing of transmission at the synapse by two calmodulin (CaM)-dependent mechanisms: one modulates membrane lipid composition by regulating a Ca2+-independent phospholipase A2 (iPLA2); the second affects a constitutively active brake on spontaneous release. Modulation of transmission is studied by electrophysiological experimentation, and temporal coding is examined in a retina in which RBs lack a fast mode of exocytosis: the complexin (clpx) 3 knockout. Specific Aim 2 determines the organization and physiological function of parallel inhibitory pathways converging on the RB. Scanning block face electron micrographic (SBEM) reconstruction identifies inhibitory amacrine cells (ACs) presynaptic to RBs. AC→RB synapses are studied functionally using various electrophysiological techniques: paired AC-RB recording, optogenetic stimulation of ACs, and optogenetic stimulation of bipolar cells-BCs-presynaptic to the ACs. Relevance to Public Health: Understanding how temporal coding is implemented by retinal synapses informs the design of retinal prosthetics and the study of animal models of human retinal diseases. A goal of vision research is the development of gene-based therapies for treating blindness caused by photoreceptor degeneration, and a promising therapy is the generation of light sensitivity in retinal interneurons by virally-mediated expression of channelrhodopsin-2 (ChR2), a light-gated cation channel. We will express ChR2 in interneurons to study synaptic interactions in retinal circuits and thereby generate critical information about the operating range of a retina in which ChR2 is the only light sensor. We address three goals of the Retinal Diseases Program in the National Plan for Eye and Vision Research: 1) determining potential therapeutic strategies for treatment of retinitis pigmentosa, 2) increasing understanding of post-photoreceptor adaptation (i.e. interactions between excitatory and inhibitory synapses), and 3) increasing understanding of how inter-cellular interactions in neural networks generate signals that are interpretable as visual images.