Dynamic ion selectivity of K2P channels and paradoxical depolarization

K2P通道的动态离子选择性和矛盾去极化

• 批准号: 8519135
• 负责人: Haijun Chen
• 金额: $ 27.67万
• 依托单位: STATE UNIVERSITY OF NEW YORK AT ALBANY
• 依托单位国家: 美国
• 财政年份: 2012
• 资助国家: 美国
• 起止时间: 2012-08-01 至 2017-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8519135
• 关键词: Accounting; Action Potentials; Alkali Metals; Arrhythmia; Behavior; Blood; Cardiac; Cardiac Myocytes; Cells; Characteristics; Chemicals; Chinese Hamster Ovary Cell; Data; Electrophysiology (science); Equation; Etiology; Exhibits; Family; Genes; Genetic; Goals; Heart; Heart Diseases; Human; Hypokalemia; Ion Channel; Ions; Light; Mediating; Membrane Potentials; Methods; Molecular; Molecular Conformation; Monovalent Cations; Patch-Clamp Techniques; Permeability; Physiological; Potassium; Potassium Channel; Protein Isoforms; Rattus; Research; Role; Stimulus; System; Testing; Tissues; Triplet Multiple Birth; base; extracellular; flexibility; improved; insight; interest; inward rectifier potassium channel; knock-down; member; novel; response

DESCRIPTION (provided by applicant): In hypokalemia or lowered extracellular K+ levels, human cardiac cells can become paradoxically depolarized inconsistent with the Nernst equation for K+. Such paradoxical depolarization has been observed for over thirty years. It is also crucial to the etiology of hypokalemia-induced cardiac arrhythmia. However, its molecular mechanism is not well understood. Ion selectivity of K+ channels is generally considered to be static and not changed in response to physiological stimuli. How K+ channels select K+ over other monovalent cations still remains an unsolved question. Particularly, ion selectivity and the selectivity filter of dimeric two-pore domain K+ channels (K2P) are less understood compared to those in tetrameric K+ channels. Our long-term goal is to understand physiological roles of K+ channels and molecular mechanisms of K+ channel function. The objective of this proposal is to explore functional roles of TWIK-1, the first cloned member of mammalian K2P channels in human cardiomyocytes in hypokalemia, and to characterize dynamic behaviors in ion selectivity and permeability of K2P channels. Based on studies in the past thirty years, the implications that are derived from these studies, and our preliminary studies on cloned TWIK-1 K+ channels, we hypothesize: 1) TWIK-1 K+ channels respond to challenges of lowered extracellular K+ levels and contribute to paradoxical depolarization in human cardiomyocytes in hypokalemia by changing ion selectivity and conducting inward leak Na+ currents; 2) K2P channels can adjust the conformations of the selectivity filter and exhibit dynamic behaviors in ion selectivity and permeability for monovalent cations. By employing standard methods and conventional approaches in the field of ion channels and electrophysiology, we will test these hypotheses in two specific aims: 1) Investigate how TWIK-1 K+ channels regulate the resting potential and action potential of human primary cardiomyocytes in both normal and hypokalemic conditions. 2) Study ion selectivity and permeability of K2P channels for small alkali metal ions and large organic monovalent cations in the absence of intracellular K+. The proposed research will demonstrate physiological roles of TWIK-1 K+ channels in human cardiomyocytes, describe a novel mechanism that regulates cardiac excitability, provide novel insights on the understanding of paradoxical depolarization in the heart in hypokalemia, and shed light on the etiology of hypokalemia-induced cardiac arrhythmias. It will also introduce the concept of dynamic ion selectivity of K+ channels under physiological conditions, provide evidence of a flexible K+ selectivity filter, which supports or supplements well-known hypotheses regarding ion selectivity of K+ channels, and improve the understanding of ion selectivity and the selectivity filter of K2P channels.

描述（由申请人提供）：在低钾血症或细胞外K+水平降低时，人心脏细胞可能出现反常去极化，与K+的能斯特方程不一致。这种反常的去极化现象已经被观察了三十多年。它也是低钾血症引起的心律失常的病因学的关键。然而，其分子机制尚未完全了解。K+通道的离子选择性通常被认为是静态的，并且不会响应于生理刺激而改变。K+通道如何选择K+而不是其他一价阳离子仍然是一个未解决的问题。特别是，离子选择性和二聚体双孔结构域K+通道（K2 P）的选择性过滤器的理解比那些在四聚体K+通道。我们的长期目标是了解K+通道的生理作用和K+通道功能的分子机制。本研究的目的是探讨第一个克隆的哺乳动物K2 P通道成员TWIK-1在低钾血症时人心肌细胞中的功能作用，并表征K2 P通道的离子选择性和通透性的动态行为。基于近30年来的研究成果，结合我们对克隆的TWIK-1 K+通道的初步研究，我们提出以下假设：1）TWIK-1 K+通道响应细胞外低K+水平的挑战，通过改变离子选择性和传导内漏Na+电流，参与低钾血症心肌细胞的反常去极化; 2）K2 P通道可以调节选择性过滤器的构象，对一价阳离子的离子选择性和渗透性表现出动态行为。本研究采用离子通道和电生理学领域的标准方法和常规手段，从两个方面验证这些假设：1）研究TWIK-1 K+通道在正常和低钾条件下如何调节人原代心肌细胞的静息电位和动作电位。2)研究在细胞内K+缺乏的情况下，K2 P通道对小的碱金属离子和大的有机单价阳离子的离子选择性和渗透性。拟议的研究将证明TWIK-1 K+通道在人类心肌细胞中的生理作用，描述一种调节心脏兴奋性的新机制，为理解低钾血症心脏中的反常去极化提供新的见解，并阐明低钾血症诱导的心律失常的病因。它还将介绍生理条件下K+通道的动态离子选择性的概念，提供灵活的K+选择性过滤器的证据，支持或补充有关K+通道离子选择性的已知假设，并提高对离子选择性和K2 P通道选择性过滤器的理解。