Gonoi T, Hille B
J Gen Physiol. 1987 Feb;89(2):253-74. doi: 10.1085/jgp.89.2.253.
Macroscopic Na currents were recorded from N18 neuroblastoma cells by the whole-cell voltage-clamp technique. Inactivation of the Na currents was removed by intracellular application of proteolytic enzymes, trypsin, alpha-chymotrypsin, papain, or ficin, or bath application of N-bromoacetamide. Unlike what has been reported in squid giant axons and frog skeletal muscle fibers, these treatments often increased Na currents at all test pulse potentials. In addition, removal of inactivation gating shifted the midpoint of the peak Na conductance-voltage curve in the negative direction by 26 mV on average and greatly prolonged the rising phase of Na currents for small depolarizations. Polypeptide toxins from Leiurus quinquestriatus scorpion and Goniopora coral, which slow inactivation in adult nerve and muscle cells, also increase the peak Na conductance and shift the peak conductance curve in the negative direction by 7-10 mV in neuroblastoma cells. Control experiments argue against ascribing the shifts to series resistance artifacts or to spontaneous changes of the voltage dependence of Na channel kinetics. The negative shift of the peak conductance curve, the increase of peak Na currents, and the prolongation of the rise at small depolarization after removal of inactivation are consistent with gating kinetic models for neuroblastoma cell Na channels, where inactivation follows nearly irreversible activation with a relatively high, voltage-independent rate constant and Na channels open only once in a depolarization. As the same kind of experiment does not give apparent shifting of activation and prolongation of the rising phase of Na currents in adult axon and muscle membranes, the Na channels of these other membranes probably open more than once in a depolarization.
采用全细胞电压钳技术记录N18神经母细胞瘤细胞的宏观钠电流。通过细胞内应用蛋白水解酶（胰蛋白酶、α-糜蛋白酶、木瓜蛋白酶或无花果蛋白酶）或浴槽应用N-溴乙酰胺来消除钠电流的失活。与在乌贼巨轴突和青蛙骨骼肌纤维中所报道的情况不同，这些处理通常会在所有测试脉冲电位下增加钠电流。此外，去除失活门控平均使峰值钠电导-电压曲线的中点向负方向移动26 mV，并极大地延长了小去极化时钠电流的上升相。来自金蝎和角孔珊瑚的多肽毒素，它们在成体神经和肌肉细胞中减缓失活，在神经母细胞瘤细胞中也会增加峰值钠电导，并使峰值电导曲线向负方向移动7 - 10 mV。对照实验表明，不能将这些移动归因于串联电阻伪迹或钠通道动力学电压依赖性的自发变化。去除失活后，峰值电导曲线的负向移动、峰值钠电流的增加以及小去极化时上升相的延长，与神经母细胞瘤细胞钠通道的门控动力学模型一致，在该模型中，失活跟随几乎不可逆的激活，具有相对较高的、电压不依赖的速率常数，并且钠通道在一次去极化中仅开放一次。由于同样的实验在成体轴突和肌肉膜中未使钠电流的激活出现明显移动和上升相延长，这些其他膜的钠通道在一次去极化中可能开放不止一次。
