ION CHANNELS IN THE NERVOUS SYSTEM

电压门控钠通道（NAChs）在神经系统的电信号和传导中起着至关重要的作用。啮齿类动物中至少有十个NaCh基因，每个基因在人类中都有一个已知的同源基因。每种NaCh亚型的具体作用尚不清楚。一种假设是，亚型是有差异分布和调节的。这一建议集中在脑NaCh亚型的调节和定位的分子基础上。Aim 1的目标是鉴定与脑NaChs（特别是Nav1.6）细胞质结构域结合并负责(a)通道调节和(b)靶向不同亚细胞位点的蛋白质。目前有两种互补的方法用于发现和分离与NaChs相关的蛋白质：(1)酵母双杂交测定法和(2)蛋白质纯化/质谱法。目的2集中于NaChs与钙调蛋白的相互作用，该蛋白是用酵母双杂交法分离出来的。这种与脑NaChs的相互作用将被生物化学和电生理学表征。Aim 3利用成像技术研究脑钠通道（特别是Nav 1.6）及其结合蛋白的亚细胞分布。在描述了这种分布之后，将研究NaChs或Aim I中鉴定的结合蛋白突变的影响。这项研究具有基础科学和临床意义。离子通道存在于与其他膜、细胞外和细胞内蛋白质的复合物中。要了解这些通道的行为，重要的是要知道这些复合物中存在哪些蛋白质以及这些蛋白质如何与通道相互作用。肌肉NaChs的突变可导致某些骨骼肌疾病和心肌长QT综合征。预计脑钠通道和与NaChs结合的蛋白质的突变将导致人类中枢神经系统疾病，而拟议的研究将有助于我们对这些疾病的理解。

Voltage-gated sodium channels (NAChs) are vital for electrical signaling and conduction in the nervous system. There are at least ten NaCh genes in rodents, and each gene has a known ortholog in humans. The specific roles of each NaCh subtype are unknown. One hypothesis is that subtypes are differentially distributed and modulated. This proposal is focused upon the molecular basis for modulation and localization of brain NaCh subtypes. The goal of Aim 1 is to identify proteins that bind to the cytoplasmic domains of brain NaChs (esp. Nav1.6) and are responsible for (a) modulation of channel fiinction and (b) targeting to different subcellular sites. Two complementary methods are being used to find and isolate proteins associated with NaChs: (1) the yeast two-hybrid assay and (2) protein purification/mass spectrometry. Aim 2 is focused upon the interactions of NaChs with calmodulin, which was isolated with the yeast two-hybrid assay. This interaction with brain NaChs will be characterized biochemically and electrophysiologically. Aim 3 utilizes imaging techniques to study the subcellular distribution of brain sodium channels (esp., Nav 1.6) and their binding proteins. After characterizing this distribution, the effects of mutations in NaChs or in the binding proteins identified in Aim I will be studied. This research has both basic science and clinical relevance. Ion channels exist in complexes with other membrane, extracellular, and intracellular proteins. To understand the behavior of these channels, it is important to know which proteins are present in these complexes and hew the proteins interact with the channel. Mutations in muscle NaChs are responsible for some disorders of skeletal muscle and for long QT syndrome in cardiac muscle. It is expected that mutations in brain sodium channels and in the proteins that bind to NaChs will produce CNS disorders in humans, and the proposed studies will contribute to our understanding of these disorders.