Zong X G, Dugas M, Honerjäger P
Institut für Pharmakologie und Toxikologie Technischen Universität München, München, Germany.
J Gen Physiol. 1992 May;99(5):683-97. doi: 10.1085/jgp.99.5.683.
Veratridine modification of Na current was examined in single dissociated ventricular myocytes from late-fetal rats. Extracellularly applied veratridine reduced peak Na current and induced a noninactivating current during the depolarizing pulse and an inward tail current that decayed exponentially (tau = 226 ms) after repolarization. The effect was quantitated as tail current amplitude, Itail (measured 10 ms after repolarization), relative to the maximum amplitude induced by a combination of 100 microM veratridine and 1 microM BDF 9145 (which removes inactivation) in the same cell. Saturation curves for Itail were predicted on the assumption of reversible veratridine binding to open Na channels during the pulse with reaction rate constants determined previously in the same type of cell at single Na channels comodified with BDF 9145. Experimental relationships between veratridine concentration and Itail confirmed those predicted by showing (a) half-maximum effect near 60 microM veratridine and no saturation up to 300 microM in cells with normally inactivating Na channels, and (b) half-maximum effect near 3.5 microM and saturation at 30 microM in cells treated with BDF 9145. Due to its known suppressive effect on single channel conductance, veratridine induced a progressive, but partial reduction of noninactivating Na current during the 50-ms depolarizations in the presence of BDF 9145, the kinetics of which were consistent with veratridine association kinetics in showing a decrease in time constant from 57 to 22 and 11 ms, when veratridine concentration was raised from 3 to 10 and 30 microM, respectively. As predicted for a dissociation process, the tail current time constant was insensitive to veratridine concentration in the range from 1 to 300 microM. In conclusion, we have shown that macroscopic Na current of a veratridine-treated cardiomyocyte can be quantitatively predicted on the assumption of a direct relationship between veratridine binding dynamics and Na current and as such can be successfully used to analyze molecular properties of the veratridine receptor site at the cardiac Na channel.
在来自晚期胎儿大鼠的单个离体心室肌细胞中研究了藜芦定对钠电流的修饰作用。细胞外施加藜芦定可降低钠电流峰值，并在去极化脉冲期间诱导出一种非失活电流，以及复极化后呈指数衰减（时间常数τ = 226毫秒）的内向尾电流。该效应通过尾电流幅度Itail（复极化后10毫秒测量）相对于同一细胞中100微摩尔藜芦定和1微摩尔BDF 9145（消除失活）组合诱导的最大幅度进行定量。基于藜芦定在脉冲期间与开放钠通道可逆结合的假设，预测了Itail的饱和曲线，反应速率常数先前在相同类型细胞的单个与BDF 9145共修饰的钠通道中确定。藜芦定浓度与Itail之间的实验关系证实了预测结果，表现为：（a）在具有正常失活钠通道的细胞中，藜芦定浓度接近60微摩尔时达到最大效应的一半，在300微摩尔时未达到饱和；（b）在用BDF 9145处理的细胞中，藜芦定浓度接近3.5微摩尔时达到最大效应的一半，在30微摩尔时达到饱和。由于其对单通道电导的已知抑制作用，藜芦定在存在BDF 9145的情况下，在50毫秒去极化期间诱导非失活钠电流逐渐但部分降低，其动力学与藜芦定结合动力学一致，当藜芦定浓度分别从3微摩尔提高到10微摩尔和30微摩尔时，时间常数从57毫秒降至22毫秒和11毫秒。正如解离过程所预测的，尾电流时间常数在1至300微摩尔范围内对藜芦定浓度不敏感。总之，我们已经表明，基于藜芦定结合动力学与钠电流之间的直接关系的假设，可以定量预测经藜芦定处理的心肌细胞的宏观钠电流，因此可成功用于分析心脏钠通道上藜芦定受体位点的分子特性。
