IONIC CHANNELS IN DISSOCIATED SMOOTH MUSCLE CELLS

这项建议的目的是为了研究平滑肌中的 跨膜离子电流的膜通道和 通过膜电位、神经递质、离子、 代谢物和其他制剂。由于这些离子通道控制着 膜电位，低于动作电位，并负责 Ca++进入细胞后，它们在控制细胞的顺畅中起着核心作用 肌肉收缩，这是一个相当重要的治疗主题(例如， 高血压、血管痉挛、子宫和胃肠收缩)。 关于这些离子通道控制的基本问题仍然存在 然而，由于与实验相关的问题，没有回答 将平滑肌细胞偶联为具有功能的电合胞 狭窄曲折的细胞间隙和神经细胞的混合物 组织中的元素。为了避免这些困难，我们将采用 一种新鲜分离的(与培养的相反)平滑肌的制备 我们将应用以下技术的细胞：1.)录音带 “电流钳”条件下的膜电位；电压钳位 整个细胞膜；膜片钳记录电流的方法 通过单一离子通道；分子的平衡和快速动力学结合 放射性标记的神经递质和相关物质。选择性， 整流特性、活化和失活动力学以及 通过膜电位和各种试剂控制这些参数将 对于各种渠道中的每一个都要进行研究。从那里的这些研究 应该会对人类的电活动有一个更基本的理解 以及平滑肌和其他细胞共有的离子通道 单元类型。

The purpose of this proposal is to investigate in smooth muscle the membrane channels responsible for transmembrane ionic currents and the control of these channels by membrane potential, neurotransmitters, ions, metabolites and other agents. Since these ionic channels control the membrane potential, underly the action potential and are responsible for Ca++ entry into the cell, they play a central role in the control of smooth muscle contraction, a topic of considerable therapeutic importance (e.g., in hypertension, vasopasm, uterine and gastrointestinal contractility). Fundamental questions about the control of these ionic channels remain unanswered, however, because of experimental problems associated with the coupling of smooth muscle cells into a functional electrical syncytum with narrow, tortuous intercellular spaces and with the admixture of neural elements in the tissue. To circumvent these difficulties, we shall employ a preparation of freshly dissociated (as opposed to cultured) smooth muscle cells to which we shall apply the following techniques: 1.) recordings of membrane potential under "current clamp" conditions; 2.) voltage-clamp of the entire cell membrane; 3.) patch clamp recording of current flowing through single ionic channels; 4.) equilibrium and rapid kinetic binding of radiolabelled neurotransmitters and related agents. The selectivity, rectifying properties, kinetics of activation and inactivation and the control of these parameters by membrane potential and various agents will be studied for each of a variety of channels. From these studies there should emerge a more basic understanding of the electrical activity of smooth muscle and of those ionic channels common to smooth muscle and other cell types.