Electrophysiological study of the functional structure of the Na channel.

本系列实验采用鱿鱼巨轴突内灌注和电压钳方法，试图利用新发现的生物毒素和化学物质来阐明Na通道的功能结构。实验分为以下几个方面：1）环多胺衍生物从细胞膜内表面阻断Na通道的机制。cyclam及其具有烷基胍侧链的类似物（G-cyclam）已显示仅从内表面阻断Na通道。该块的模式是电压依赖的，也是时间依赖的。在G-cyclam的情况下，阻断的时间过程由单个指数函数控制。Cyclam对Na通道的阻断作用呈双指数函数。当[Na] i从50增加到200 mM时，阻断作用增强，两态三势垒模型很好地解释了钠通道的所有阻断作用。从这些环状多胺ANA的分子考虑， ...更多信息 logues，合理的假设是烷基胍鎓基团在Na通道内自由移动，从而附着带负电荷的位点并实际上阻断Na通道。2)当两种蜘蛛毒素JSTX-3和NSTX-3被施加于细胞内时，也获得了类似的发现。这些蜘蛛毒素有一个共同的2，4-二氢氧-苯乙酰基-麦角酰-cadabelino-ptreamine。分子结构的差异在末端被识别，其中JSTX-3具有gpysine和NSTX-3精氨酸残基。由于NSTX-3比JSTX-3发挥更强的阻断作用，因此再次假定具有永久正电荷的可自由移动的末端基团在阻断Na通道中起重要作用。3)外源性钠通道调节剂灰蝶毒素（GTX）出现在细胞内相，在细胞内相非灌注时引起的去极化明显大于持续灌注时。因此，可以得出结论，作用位点在细胞内相。少

In this series of experiments using the internal perfusion and the voltage clamp method on squid giant axon, an attempt was made to elucidate the functional structure Na channel by using the newly found biological toxins and chemicals. The experiments were categorized as follows; 1) The mechanism of block of the Na channel from the internal surface of cell membrane by cyclic-polyamine derivatives. cyclam and its analogue having the alkylguanidunium side chain (G-cyclam) have shown to block the Na channel only from the internal surface. Mode of the block is voltage- as well as time-dependent. In the case of G-cyclam, time course of the block is governed by a single exponential function. While, cyclam blocks Na channel with a double exponential function. The enhancement of blocking action occurred when [Na]_i increased from 50 to 200 mM. Two state three barrier model well explains all these blocking actions in the Na channel. from the molecular consideration of these cyclic aolyamine ana … More logues, it is reasonable to assume that alkyl guanidinium group freely moves inside the na channel, thereby attaching the negative charged site and in ture blocking the Na channel. 2) Similar finding also obtained, when two spider toxins, JSTX-3 and NSTX-3, were applied intra-cellularly. These spider toxins have a common 2,4 dihydrooxy-phenylacethyl-aspargynil-cadabelino-ptreamine. Difference in molecular structure is recognized in the terminal where JSTX-3 has gpysine and NSTX-3 arginine residues. Since NSTX-3 exerts stronger blocking action than JSTX-3 does, again freely movable terminal group having a permanent positive charge is assumed to play an important role in blocking the Na channel. 3) Externally applied grayanotoxin (GTX), Na channel modifier, appeared in the intracellu lar phase and induced much bigger depolarization in the non-perfusion of intracellular phase than in the continuous perfusion. Thus, it has been concluded that the site of action is in the intracellular phase. Less