Voltage-dependent gating of the cystic fibrosis transmembrane conductance regulator Cl- channel.
Voltage-dependent gating of the cystic fibrosis transmembrane conductance regulator Cl- channel.
复制标题
囊性纤维化跨膜电导调节器CL-通道的电压依赖性门控。
DOI：
10.1085/jgp.200308921
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发表时间：
2003-11
影响因子：
3.8
通讯作者：
Sheppard, David N
中科院分区：
文献类型：
--
作者：
Cai, Zhiwei;Scott-Ward, Toby S;Sheppard, David N
当切除的由内而外的膜斑浸泡在对称的富含Cl−的溶液中时，宏观囊性纤维化跨膜电导调节因子（CFTR）Cl−电流的电流-电压（I-V）关系在大的正电压下向内整流。为了研究内向整流的机制，我们研究了CFTR Cl−通道，使用电压斜坡和阶梯方案，从表达野生型人类和小鼠CFTR的细胞中切除的由内而外的膜补丁。使用电压斜坡方案，+100 mV时人体CFTR Cl−电流的幅度为− 100 mV时的74 ± 2%（n = 10）。宏观CFTR Cl−电流的这种整流完全通过对由相同的电压斜坡协议引起的单通道电流进行平均而产生的系综电流来再现。然而，使用电压阶跃方案，在+100 mV时，人CFTR的单通道电流幅度（i）是−100 mV时的88 ± 2%（n = 10）。基于这些数据，我们假设电压可能会改变人类CFTR的门控行为。利用线性三态动力学模型，我们证明了电压对通道门控有显著的影响。膜去极化缩短了爆发的持续时间和爆发间隔，但增加了爆发内的间隙的持续时间。然而，由于不同速率常数的电压依赖性在相反的方向上，电压对人CFTR的开放概率（Po）没有大的影响。相反，在正电压下，鼠CFTR的Po显著降低，表明鼠CFTR的整流比人CFTR的强。我们的结论是向内整流CFTR是由减少i和门控动力学的变化。我们认为，内向整流是CFTR Cl−通道的内在特性，而不是孔隙阻塞的结果。
When excised inside-out membrane patches are bathed in symmetrical Cl−-rich solutions, the current-voltage (I-V) relationship of macroscopic cystic fibrosis transmembrane conductance regulator (CFTR) Cl− currents inwardly rectifies at large positive voltages. To investigate the mechanism of inward rectification, we studied CFTR Cl− channels in excised inside-out membrane patches from cells expressing wild-type human and murine CFTR using voltage-ramp and -step protocols. Using a voltage-ramp protocol, the magnitude of human CFTR Cl− current at +100 mV was 74 ± 2% (n = 10) of that at −100 mV. This rectification of macroscopic CFTR Cl− current was reproduced in full by ensemble currents generated by averaging single-channel currents elicited by an identical voltage-ramp protocol. However, using a voltage-step protocol the single-channel current amplitude (i) of human CFTR at +100 mV was 88 ± 2% (n = 10) of that at −100 mV. Based on these data, we hypothesized that voltage might alter the gating behavior of human CFTR. Using linear three-state kinetic schemes, we demonstrated that voltage has marked effects on channel gating. Membrane depolarization decreased both the duration of bursts and the interburst interval, but increased the duration of gaps within bursts. However, because the voltage dependencies of the different rate constants were in opposite directions, voltage was without large effect on the open probability (Po) of human CFTR. In contrast, the Po of murine CFTR was decreased markedly at positive voltages, suggesting that the rectification of murine CFTR is stronger than that of human CFTR. We conclude that inward rectification of CFTR is caused by a reduction in i and changes in gating kinetics. We suggest that inward rectification is an intrinsic property of the CFTR Cl− channel and not the result of pore block.