Connexins and electrical synapses in the retina

视网膜中的连接蛋白和电突触

• 批准号: 8038927
• 负责人: DAVID L PAUL
• 金额: $ 42.25万
• 依托单位: HARVARD MEDICAL SCHOOL
• 依托单位国家: 美国
• 财政年份: 2004
• 资助国家: 美国
• 起止时间: 2004-08-01 至 2013-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8038927
• 关键词: Address; Amacrine Cells; Antibodies; Blindness; Cells; Connexins; Coupling; Darkness; Data; Detection; Disease; Dyes; Electrical Synapse; Gap Junctions; Genetic; Glycine; Human; Injection of therapeutic agent; Knock-out; Life; Light; Measurement; Methods; Microelectrodes; Modeling; Monkeys; Movement; Mus; Neural Retina; Oryctolagus cuniculus; Pathway interactions; Physiological; Proteins; Recombinant adeno-associated virus (rAAV); Retina; Retinal; Retinal Cone; Signal Pathway; Signal Transduction; Synapses; Testing; Therapeutic; Tissues; Tracer; Transgenic Mice; Transgenic Organisms; Vertebrate Photoreceptors; base; ganglion cell; glycine transporter; neural circuit; neurobiotin; novel; prevent; research study; response; retinal rods; segregation; subretinal injection

DESCRIPTION (provided by applicant): A compelling body of evidence indicates that signals from rod photoreceptors use multiple pathways to reach ganglion cells. In the canonical primary pathway, rod signals gain access to downstream cone circuitry through the AII amacrines which form electrical synapses (gap junctions) with ON cone bipolar and glycinergic synapses with OFF cone bipolar. In the secondary pathway, electrical synapses between rods and cones provide a direct entry for rod signals into cone circuits. However, a credible direct demonstration of rod-cone coupling has been made only in monkey. Because technical issues prevent a conventional approach to measurement of rod-cone coupling in the mouse (i.e. injections of junction-permeant tracers using microelectrodes), we developed a novel method to evaluated coupling. The method is based on transgenic, cell-specific expression of a transporter, the movement of the transported molecule through gap junctions to neighboring cells and the detection of that molecule with specific antibodies. This method eliminates the need for any physical manipulation of the live cells. Despite the widely held assumption that rod-cone coupling underlies the secondary pathway, we found no evidence of rod-cone coupling in the mouse. Thus, we propose a set of experiments to determine the generality of the canonical secondary pathway model by testing rod-cone coupling in the rabbit and to evaluate cone-cone and possible rod-rod coupling in mouse and rabbit retinas. In contrast, our data strongly support the basic tenets of the rod primary pathway. However, while the prevailing model postulates that AII amacrines express Cx36 and cone ON bipolar express Cx45, forming a 'heterotypic' electrical synapse, our data indicate more complexity. We propose there are two types of glycinergic amacrine cells, those expressing Cx45 and those expressing Cx36 and that each forms homotypic junctions with a subset of cone bipolar cells expressing the same connexin. We hypothesize that the expression of incompatible connexins is mechanism to allow segregation of amacrine- cone bipolar interactions according to cell subtype. We will determine the types of cone bipolar involved in rod primary pathway signaling and which connexins they employ. In addition, we will determine if different retinal connexins can functionally interact. PUBLIC HEALTH RELEVANCE: Our studies address fundamental questions about the neural circuitry employed by rod photoreceptors, which contribute to retinal responses over a range of light inputs from near total darkness to bright moonlight. Disorders of the neural retina are a primary cause of human blindness and a rational pursuit of therapeutic strategies requires a full understand of mammalian retinal circuitry.

描述（由申请人提供）：大量令人信服的证据表明，杆状光感受器发出的信号通过多种途径到达神经节细胞。在典型的初级通路中，杆信号通过AII神经元进入下游锥体回路，AII神经元与ON锥双极形成电突触（间隙连接），与OFF锥双极形成甘氨酸能突触。在次级通路中，杆状细胞和锥状细胞之间的电突触为杆状细胞的信号提供了进入锥状细胞回路的直接通道。然而，仅在猴子身上对杆锥耦合进行了可靠的直接演示。由于技术问题阻碍了测量小鼠棒锥耦合的传统方法（即使用微电极注射连接渗透示踪剂），我们开发了一种评估耦合的新方法。该方法基于转运体的转基因细胞特异性表达，转运分子通过间隙连接移动到邻近细胞，并用特异性抗体检测该分子。这种方法消除了对活细胞进行任何物理操作的需要。尽管人们普遍认为杆-锥体耦合是次要通路的基础，但我们在小鼠中没有发现杆-锥体耦合的证据。因此，我们提出了一组实验，通过测试家兔视网膜的杆-锥耦合来确定典型次级通路模型的普遍性，并评估小鼠和家兔视网膜的锥-锥和可能的杆-杆耦合。相反，我们的数据强烈支持杆主通路的基本原理。然而，虽然主流模型假设AII amacrines表达Cx36，而cone ON双极性表达Cx45，形成“异型”电突触，但我们的数据表明更复杂。我们提出有两种类型的甘氨酸能无分泌细胞，表达Cx45和表达Cx36的细胞，每一种细胞都与表达相同连接蛋白的锥体双极细胞亚群形成同型连接。我们假设，不相容连接蛋白的表达是允许根据细胞亚型分离腺-锥体双极性相互作用的机制。我们将确定参与杆初级通路信号的锥双极类型以及它们使用的连接蛋白。此外，我们将确定不同的视网膜连接蛋白是否可以在功能上相互作用。