Fohlmeister J F, Adelman W J
Biophys J. 1986 Sep;50(3):489-502. doi: 10.1016/S0006-3495(86)83486-5.
Internally perfused squid giant axons with intact sodium inactivation gating were prepared for gating current experiments. Gating current records were obtained in sinusoidally driven dynamic steady states and as dynamic transients as functions of the mean membrane potential and the frequency of the command sinusoid. Controls were obtained after internal protease treatment of the axons that fully removed inactivation. The nonlinear analysis consisted of determining and interpreting the harmonic content in the current records. The results indicate the presence of three kinetic processes, two of which are associated with activation gating (the so-called primary and secondary processes), and the third with inactivation gating. The dynamic steady state data show that inactivation gating does not contribute a component to the gating current, and has no direct voltage-dependence of its own. Rather, the inactivation kinetics appear to be coupled to the primary activation kinetics, and the coupling mechanism appears to be one of reciprocal steric hindrance between two molecular components. The mechanism allows the channel to become inactivated without first entering the conducting state, and will do so in about 40 percent of depolarizing voltage-clamp steps to 0 mV. The derived model kinetics further indicate that the conducting state may flicker between open and closed with the lifetime of either state being 10 microseconds. Dynamic transients generated by the model kinetics (i.e., the behavior of the harmonic components as a function of time after an instantaneous change in the mean membrane potential from a holding potential of -80 mV) match the experimental dynamic transients in all details. These transients have a duration of 7-10 ms (depending on the level of depolarization), and are the result of the developing inactivation following the discontinuous voltage change. A detailed hypothetical molecular model of the channel and gating machinery is presented.
制备具有完整钠失活门控的内部灌注鱿鱼巨轴突用于门控电流实验。门控电流记录是在正弦驱动的动态稳态下以及作为动态瞬变获得的，作为平均膜电位和指令正弦波频率的函数。在对轴突进行内部蛋白酶处理以完全去除失活后获得对照。非线性分析包括确定和解释电流记录中的谐波含量。结果表明存在三个动力学过程，其中两个与激活门控相关（所谓的初级和次级过程），第三个与失活门控相关。动态稳态数据表明，失活门控对门控电流没有贡献，并且其自身没有直接的电压依赖性。相反，失活动力学似乎与初级激活动力学耦合，并且耦合机制似乎是两个分子成分之间相互空间位阻的一种。该机制允许通道在不首先进入导通状态的情况下失活，并且在约40％的去极化电压钳制步进到0 mV时会这样做。推导的模型动力学进一步表明，导通状态可能在开放和关闭之间闪烁，任何一种状态的寿命为10微秒。由模型动力学产生的动态瞬变（即谐波成分作为平均膜电位从 - 80 mV的保持电位瞬间变化后的时间函数的行为）在所有细节上都与实验动态瞬变匹配。这些瞬变的持续时间为7 - 10毫秒（取决于去极化水平），并且是不连续电压变化后失活发展的结果。提出了通道和门控机制的详细假设分子模型。
