Sheets M F, Hanck D A
Department of Medicine, Northwestern University Medical School, Chicago, IL 60611.
J Cardiovasc Electrophysiol. 1993 Dec;4(6):686-94. doi: 10.1111/j.1540-8167.1993.tb01254.x.
Studies of tetrodotoxin-sensitive sodium current (INa) after modification of inactivation by intracellular enzymes in mammalian cells have demonstrated a marked increase in peak INa at test potentials near current threshold causing a large, negative shift of the peak INa conductance-voltage relationship by approximately -20 mV. These findings support a kinetic model in which the unmodified Na channel has rapid and voltage-independent inactivation from the open state. However, the kinetics of cardiac Na channels differ from those of mammalian neuronal Na channels. In particular, inactivation of cardiac Na channels has been proposed to be more voltage dependent than that of tetrodotoxin-sensitive Na channels. To help understand the role of inactivation in cardiac Na channel kinetic behavior, we studied Na currents before and after modification of inactivation by the proteolytic enzyme, alpha-chymotrypsin.
Whole cell INa was measured in single canine cardiac Purkinje cells that were voltage clamped and internally perfused with a large-bore suction pipette. The decay of INa in response to step depolarizations was dramatically slowed after perfusion with intracellular alpha-chymotrypsin consistent with modification of inactivation. In contrast to mammalian tetrodotoxin-sensitive Na current, Boltzmann distribution fits to peak INa conductance-voltage (GNa-V) relationships after alpha-chymotrypsin showed no change in either the potential at half maximum conductance (V 1/2), after correction for the spontaneous background shift of INa kinetics, or in the voltage-dependence of conductance (i.e., slope factor of GNa-V relationships). Maximal peak INa conductance increased by 18%. INa tail-current relaxations at potentials < or = -110 mV, after correction for spontaneous shifts in Na channel kinetics, were also similar before and after modification by alpha-chymotrypsin.
alpha-chymotrypsin modified inactivation of cardiac INa with little or no change in activation, and cardiac Na channel inactivation was slow near threshold and played little role in determining V1/2 for peak INa conductance-voltage relationships.
在哺乳动物细胞中，通过细胞内酶改变失活状态后对河豚毒素敏感的钠电流（INa）的研究表明，在接近电流阈值的测试电位下，峰值INa显著增加，导致峰值INa电导-电压关系出现大幅负向偏移，约为-20 mV。这些发现支持了一种动力学模型，即未修饰的钠通道从开放状态具有快速且电压依赖性的失活。然而，心脏钠通道的动力学与哺乳动物神经元钠通道不同。特别是，有人提出心脏钠通道的失活比河豚毒素敏感的钠通道更依赖电压。为了帮助理解失活在心脏钠通道动力学行为中的作用，我们研究了蛋白水解酶α-糜蛋白酶改变失活状态前后的钠电流。
在单个犬心脏浦肯野细胞中测量全细胞INa，这些细胞通过大口径吸液管进行电压钳制和内部灌注。在用细胞内α-糜蛋白酶灌注后，响应阶跃去极化的INa衰减显著减慢，这与失活的改变一致。与哺乳动物河豚毒素敏感的钠电流不同，α-糜蛋白酶处理后峰值INa电导-电压（GNa-V）关系的玻尔兹曼分布拟合显示，在对INa动力学的自发背景偏移进行校正后，半最大电导电位（V 1/2）或电导的电压依赖性（即GNa-V关系的斜率因子）均无变化。最大峰值INa电导增加了18%。在对钠通道动力学的自发偏移进行校正后，在电位≤ -110 mV时的INa尾电流松弛在α-糜蛋白酶修饰前后也相似。
α-糜蛋白酶改变了心脏INa的失活，而激活几乎没有变化，并且心脏钠通道失活在阈值附近缓慢，在确定峰值INa电导-电压关系的V1/2方面作用不大。
