REGULATION OF SODIUM CHANNEL PHOSPHORYLATION IN BRAIN

离子通道介导细胞膜上的离子流动， 电激励 许多神经递质和激素调节离子 通道活性通过第二信使激活磷酸化。 尽管已知几种第二信使依赖性蛋白激酶 直接磷酸化离子通道，对这种蛋白质知之甚少， 终止磷酸化作用的磷酸酶。 总 该提案的目的是确定磷酸酶在控制 离子通道磷酸化的状态，并确定是否 磷酸酶对离子通道的作用也受到神经递质的调节 或荷尔蒙。 这项建议的重点是确定磷酸酶和调节 控制电压敏感钠通道磷酸化的途径 在老鼠的大脑中。 这些通道被cAMP依赖性蛋白磷酸化 原位激酶。 环腺苷酸依赖性磷酸化可能会改变 神经元钠通道的电生理特性。 因为它的 结构表征良好且与生理学相关 磷酸化位点已被确定，电压敏感的钠 通道是研究去磷酸化的一个很好的候选者。 将进行生化研究，以确定脑磷酸酶 去磷酸化依赖cAMP的磷酸化位点， 频道 成年大鼠脑的离散区域的切片将用于 研究这种磷酸酶在完整的神经元中是如何调节的， 大脑中的特定神经递质通路控制cAMP依赖性 钠通道的磷酸化或去磷酸化。 最后 钠通道磷酸酶的区域和亚细胞定位 将使用生物化学和免疫细胞化学方法进行检查， 酶的分布与钠通道的分布相比较。 通过使用这一策略，将开发一个详细的图片如何钠 通道磷酸化在脑中被控制。 这可能会产生新的见解 和生物化学工具来检查功能性后果， 钠通道磷酸化。 此外，本研究将探讨 磷酸酶是否受神经递质调节的一般问题 以及磷酸酶如何参与控制 离子通道和其他神经元磷蛋白的磷酸化。 因为可逆的磷酸化是调节 离子通道的功能和因此细胞的兴奋，这是重要的 了解这个复杂的监管系统是如何运作的， 了解神经元兴奋性在健康和疾病中的作用。

Ion channels mediate the flow of ions across cellular membranes during electrical excitation. Many neurotransmitters and hormones regulate ion channel activity through second messenger-activated phosphorylation. Although several second messenger-dependent protein kinases are known to directly phosphorylate ion channels, little is known about the protein phosphatases that terminate the influence of phosphorylation. The general goal of this proposal is to define the role of phosphatases in controlling the state of ion channel phosphorylation, and to determine whether phosphatase action on ion channels is also regulated by neurotransmitters or hormones. This proposal focuses on identifying the phosphatases and regulatory pathways controlling phosphorylation of voltage-sensitive sodium channels in rat brain. These channels are phosphorylated by cAMP-dependent protein kinase in situ. Cyclic AMP-dependent phosphorylation may modify the electrophysiological properties of neuronal sodium channels. Because its structure is well characterized and physiologically relevant phosphorylation sites have been identified, the voltage-sensitive sodium channel is an excellent candidate for studying dephosphorylation. Biochemical studies will be performed to identify the brain phosphatase(s) that dephosphorylates cAMP-dependent phosphorylation sites in the sodium channel. Slices from discrete regions of adult rat brain will be used to examine how this phosphatase is regulated in intact neurons and whether specific neurotransmitter pathways in the brain control cAMP-dependent phosphorylation or dephosphorylation of sodium channels. Finally, the regional and subcellular localization of the sodium channel phosphatase(s) will be examined using both biochemical and immunocytochemical methods, and the distribution of the enzyme(s) compared with that of sodium channels. By using this strategy, a detailed picture will be developed of how sodium channel phosphorylation is controlled in brain. This may yield new insight and biochemical tools with which to examine the functional consequences of sodium channel phosphorylation. In addition, this study will address the general question of whether phosphatases are regulated by neurotransmitters or hormones and how phosphatases participate in controlling the state of phosphorylation of ion channels and other neuronal phosphoproteins. Because reversible phosphorylation is such an important means of modulating ion channel function and consequently cellular excitation, it is important to learn how this complex regulatory system operates if we are to understand the role of neuronal excitability in health and disease.