Regulation of retinal gap junctions

这项研究的长期目标是确定调节大脑中电耦合的机制。 视网膜，并确定哪些调节模式对观察到的适应过程最重要 N个不同的电耦合神经电路。由缝隙连接介导的电耦合有助于 大多数类型的视网膜神经元的信号处理功能。视觉过程中缝隙连接的调制 适应对许多神经元的敏感度和感受野特性有深远的影响和影响 哺乳动物视杆细胞回路中的信号流路径。在视网膜内的不同回路中，非常不同 对由相同的连接蛋白组成的缝隙连接提出了要求，并且调节反应 观察到的结果是完全不同的。先前的结果表明，有几种不同的机制可以 调节偶联，我们假设这些作用将根据细胞类型的不同而不同。 这个项目的重点是描绘调节Cx35/36缝隙连接的主要机制， 在视网膜神经元和整个中枢神经系统中分布最广泛的连接蛋白。具体的 这个项目要检验的假设是：1)特定残基的磷酸化状态 描述了Cx35缝隙连接的调节状态；2)钙调素参与钙依赖 缝隙连接偶联的调节；以及3)偶联的调节受密切相关蛋白质的影响 Cx35在一个综合体中。各种体外生物物理和生化技术将被用于 研究Cx35的翻译后修饰和蛋白质结合作用。扰乱世界的突变 这些过程将在细胞培养表达系统中进行，并研究它们对调控的影响。这个 视网膜神经元中调控通路的组成部分与Cx35的关联将通过以下方式进行检查 免疫染色。这些研究将确定可能受到不适当监管的主要地点 影响缝隙连接偶联的视网膜疾病。

The long-term objectives of this study are to identify the mechanisms that regulate electrical coupling in the retina, and to determine which modes of regulation are most important for the adaptive processes observed n different electrically coupled neural circuits. Electrical coupling, mediated by gap junctions, contributes to the signal processing functions of most types of retinal neurons. Modulation of gap junctions during visual adaptation has profound effects on sensitivity and receptive field properties of many neurons and influences the path of signal flow in the mammalian rod circuit. In different circuits within the retina, very different demands are placed on gap junctions made of the same connexin proteins, and the regulatory responses observed are quite disparate. Previous results suggest that there are several different mechanisms that regulate coupling, and we hypothesize that the contributions of these will differ depending on the cell type. This project focuses on delineating the major mechanisms that regulate gap junctions made of Cx35/36, the most widespread connexin in retinal neurons and throughout the central nervous system. The specific hypotheses to be examined in this project are that: 1) the phosphorylation state of specific residues characterizes the regulatory state of Cx35 gap junctions; 2) calmodulin is involved in calcium-dependent regulation of gap junction coupling; and 3) regulation of coupling is influenced by proteins closely associated with Cx35 in a complex. A variety of in vitro biophysical and biochemical techniques will be used to characterize posttranslational modifications and protein binding interactions of Cx35. Mutations that disrupt these processes will be made and their effects on regulation studied in cell culture expression systems. The association of components of the regulatory pathways with Cx35 in retinal neurons will be examined by immunostaining. These studies will identify the major sites that could be subject to improper regulation in retinal disorders that affect gap junction coupling.