Molecular Dissection of Neuronal K+ Channel Function

描述（由申请人提供）：在哺乳动物神经系统中， 电压门控钾（K+）通道具有不同的时间和 电压依赖性和药理学敏感性已经被 鉴定这种异质性具有生理学意义， 各种K+通道的功能是控制静息膜电位、动作电位和细胞膜电位。 电位波形，重复放电模式和对突触的反应 输入。克隆了电压门控性K+通道（Ky）孔形成（cc）和 辅助（B，KChAP，KChIP）亚基显示出更大的潜力， 产生的K+通道多样性比预期的要多， 在哺乳动物神经元中，这些亚基和K+通道之间的联系很差， 明白在这里，我们建议利用分子遗传策略， 确定电压门控K+电流的分子相关性，“Al”IA， 大鼠上级颈交感神经元的K和ISS 神经节（SCG），并定义IAF、“As”、“K”和ISS的功能作用。 最初的实验将检验有两个分子 不同类型的IAf通道（由Kv1和Kv4 a亚基编码）， 确定删除所有IAf通道的功能后果 SCG细胞的点火特性。目标2和目标3中的实验将测试 假设Kv2和Kv3亚家族的Ky α亚基是IK的基础 在SCG神经元中，K和IA分别定义了K和IA在SCG神经元中的作用。 形成动作电位波形并在这些细胞中重复放电。的 最后的目的将探讨辅助KChIP蛋白在 在SCG神经元中产生功能性电压门控K+通道。 我们预计，本提案中概述的研究将提供 从根本上重要的新见解的分子基础的功能 哺乳动物交感神经元的电压门控性K+通道多样性 这些钾通道在调节神经元兴奋性中的作用。 重要的是，似乎多方面的实验方法 也可以应用于确定分子组成， 电压门控（和其他）K+通道在其他细胞中的功能作用。

DESCRIPTION (provided by applicant): In the mammalian nervous system, a variety of voltage-gated potassium (K+) channels with distinct time- and voltage-dependent properties and pharmacological sensitivities have been identified. This heterogeneity has a physiological significance in that the various K+ channels function to control resting membrane potentials, action potential waveforms, repetitive firing patterns and the responses to synaptic inputs. The cloning of voltage-gated K+ channel (Ky) pore-forming (cc) and accessory (B, KChAPs, KChIPs) subunits has revealed even greater potential for generating K+ channel diversity than was expected, and the relationships between these subunits and the K+ channels in mammalian neurons is poorly understood. Here, we propose to exploit molecular genetic strategies to identify the molecular correlates of the voltage-gated K+ currents, 'Al' IAs, 'K' and ISS, in sympathetic neurons isolated from the (rat) superior cervical ganglion (SCG), and to define the functional roles of IAF, 'As' 'K' and ISS. Initial experiments will test the hypothesis that there are two molecularly distinct types of IAf channels (encoded by Kv1 and Kv4 a subunits) and determine the functional consequences of removing all IAf channels on the firing properties of SCG cells. Experiments in aims 2 and 3 will test the hypotheses that Ky alpha subunits of the Kv2 and Kv3 subfamilies underlie 'IK and IAs' respectively, in SCG neurons and define the roles 'K and IAs in shaping action potential waveforms and repetitive firing in these cells. The final aim will explore the role(s) of the accessory KChIP proteins in the generation of functional voltage-gated K+ channels in SCG neurons. We anticipate that the studies outlined in this proposal will provide fundamentally important new insights into the molecular basis of functional voltage-gated K+ channel diversity in mammalian sympathetic neurons and into the roles of these K+ channels in the regulation neuronal excitability. Importantly, it seems likely that the multifaceted experimental approach developed here can also be applied to determine the molecular compositions and functional roles of voltage-gated (and other) K+ channels in other cells.