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An Analysis of the Variations in Potency of Grayanotoxin Analogues in Modifying Frog Sodium Channels of Differing Subtype

木杨毒素类似物修饰不同亚型青蛙钠通道效力的差异分析

基本信息

批准号：
09680814
负责人：
SEYAMA Issei
金额：
$ 1.92万
依托单位：
HIROSHIMA UNIVERSITY
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (C)
财政年份：
1997
资助国家：
日本
起止时间：
1997 至 1998
项目状态：
已结题

项目摘要

Responses of TTX-sensitive (TTX-s) and insensitive (TTX-i) Na^+ channels, in frog dorsal root ganglion (DRG) cells and frog heart Na^+ channels, to two grayanotoxin (GTX) analogues, GTX-I and alpha-dihydro-GTX-II, were examined, using the patch clamp method. GTX-evoked modification occurred only when repetitive depolarizing pulses preceded a single test depolarization ; modification, during the test pulse, was manifested by a decrease in peak Na^+ current accompanied by a sustained Na^+ current. GTX-evoked modification of whole-cell Na^+ currents was quantified by normalizing the conductance for sustained currents through GTX-modified Na^+ channels to that for the control peak current through unmodified Na^+ channels. The dose-response relation for GTX-modified Na^+ channels was constructed by plotting the normalized conductance against GTX concentration. With respect to TTX-i Na^+ channels found in DRG cells, the EC_<50> (helf-maximal concentration) and maximal normalized conductance were estimated to be 35 mu M and 0.23 for GTX-I, and 54 mu M and 0.37 for alpha-dihydro-GTX-II.By contrast, TTX-s Na^+ channels in DRG, and Na^+ channels in ventricular cells were found to have a much lower sensitivity to both GTX analogues. Currents through single Na^+ channels were recorded from cell-attached membrane patches to elucidate, the factors responsible for the marked differences in potency of the GTX analogues in modifying DRO TTX-i and ventricular Na^+ channels. In DRG cells and ventricular myocytes, single Na^+ channels modified by the two GTX analogues (both at 100 mu M), had similar relative conductances (range : 0.25-0.42) and open channel probabilities (range : 0.5-0.71). From these observations, we conclude that the differences in responsiveness of DRG TTX-i, and ventricular whole cell Na^+ currents to the GTX analogues studied are related to the number of Na^+ channels modified.

采用膜片钳技术，研究了蛙背根神经节（DRG）细胞和蛙心脏Na^+通道中TTX敏感（TTX-s）和不敏感（TTX-i）Na^+通道对两种灰蝶毒素（GTX）类似物GTX-I和α-dihydro-GTX-II的反应。GTX诱发的修饰仅发生在重复去极化脉冲先于单次测试去极化时;在测试脉冲期间，修饰表现为峰值Na^+电流的降低，并伴有持续的Na^+电流。通过将GTX修饰的Na^+通道的持续电流的电导与未修饰的Na ^+通道的对照峰电流的电导归一化，可以定量GTX对全细胞Na^+电流的修饰。通过绘制归一化电导对GTX浓度的曲线，构建了GTX修饰的Na^+通道的剂量-反应关系。对于在DRG细胞中发现的TTX-i Na^+通道，<50>GTX-Ⅰ的EC_（半最大浓度）和最大归一化电导估计为35 μ M和0.23，α-二氢-GTX-Ⅱ的EC_（半最大浓度）和最大归一化电导估计为54 μ M和0.37。相比之下，发现DRG中的TTX-s Na^+通道和心室细胞中的Na^+通道对这两种GTX类似物的敏感性要低得多。从细胞贴附的膜片上记录通过单个Na^+通道的电流，以阐明GTX类似物在修饰DRO TTX-i和心室Na^+通道方面的效力存在显著差异的原因。在DRG细胞和心室肌细胞中，两种GTX类似物（均为100 μ M）修饰的单个Na^+通道具有相似的相对电导（范围：0.25-0.42）和开放通道概率（范围：0.5-0.71）。从这些观察结果中，我们得出结论，DRG TTX-i和心室全细胞Na^+电流对所研究的GTX类似物的反应性差异与修饰的Na^+通道数量有关。