Distinguishing surface effects of calcium ion from pore-occupancy effects in Na+ channels
Distinguishing surface effects of calcium ion from pore-occupancy effects in Na+ channels
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DOI：
10.1073/pnas.96.7.4158
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发表时间：
1999-03-30
影响因子：
11.1
通讯作者：
Armstrong, CM
中科院分区：
文献类型：
--
作者：
Armstrong, CM
本文研究了钙离子对鱿鱼巨轴突Na+激活门的影响，用石房蛤毒素（STX）阻断离子进入Na+通道而不阻碍离子进入膜表面，从而区分钙离子的表面效应和孔占据效应。当STX存在时，外部钙浓度变化的动力学效应很小。在不存在STX的情况下，降低钙浓度（从100 mM至10 mM）使Na+通道的关闭（从I-Na尾测量）减慢超过2倍。令人惊讶的是，关闭动力学的电压敏感性随钙浓度而变化，并且它被STX修饰。电压敏感性显然部分取决于当电压被驱动为负时钙进入并阻断通道的能力。在没有添加钙的外部介质中，I-Na尾电流最初随着钙阻滞的解除而振幅增加数倍，然后逐渐减慢并变得更小，因为钙从轴突的层中扩散出来，在电流消失之前看到的I-Na尾表明在没有通道阻滞的情况下关闭非常缓慢或不发生。当钙返回时，I-Na幅度和动力学完全恢复。结果强烈表明，钙占用是通道关闭的要求，未占用的通道折叠成非功能性构象可逆。
The effects of calcium ion on the Na+ activation gate were studied in squid giant axons, Saxitoxin (STX) was used to block ion entry into Na+ channels without hindering access to the membrane surface, making it possible to distinguish surface effects of calcium from pore-occupancy effects, In the presence of STX, gating kinetics were measured from gating curl ent (I-g). The kinetic effects of external calcium concentration changes were small when STX was present. In the absence of STX, lowering the calcium concentration (from 100 to 10 mM) slowed the closing of Na+ channels (measured from I-Na tails) by more than a factor of 2. Surprisingly, the voltage sensitivity of closing kinetics changed with calcium concentration, and it was modified by STX. Voltage sensitivity apparently depends in part on the ability of calcium to enter and block the channels as voltage is driven negative. In external medium with no added calcium, I-Na tail current initially increases in amplitude severalfold with the relief of calcium block, then progressively slows and gets smaller, as calcium diffuses out of the layers investing the axon, I-Na tails seen just before the current disappears suggest that closing in the absence of channel block is very slow or does not occur. I-Na amplitude and kinetics are completely restored when calcium is returned. The results strongly suggest that calcium occupancy is a requirement for channel closing and that nonoccupied channels fold reversibly into a nonfunctional conformation.