Coupled K+-water flux through the HERG potassium channel measured by an osmotic pulse method
Coupled K+-water flux through the HERG potassium channel measured by an osmotic pulse method
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DOI：
10.1085/jgp.200509377
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发表时间：
2005-11-01
影响因子：
3.8
通讯作者：
Oiki, S
中科院分区：
文献类型：
--
作者：
Ando, H;Kuno, M;Oiki, S
流电位（V流）是离子通道的一个特征，其中渗透离子和水分子以单一的文件运动。V流提供了离子和水通量（水离子耦合比）的定量测量，这是阐明离子渗透机制的先决条件。我们开发了一种用全细胞膜片钳结构测量V流的方法。将稳定表达HERG钾通道的HEK293细胞通过超快溶液切换系统（渗透脉冲[快速跳脱]方法）压紧并暴露于高渗溶液中短时间（< 1 s）。在渗透脉冲之前、期间和之后，通过一系列电压坡道监测反转电位。在渗透跃迁后，反转电位立即发生变化，形成V流。在对称K+溶液（10 mM）中，在不同渗透压下测量的V（流）s呈线性关系，斜率为-0.7 mV/ δ Osm，由此得出水离子耦合比（n，水通量与阳离子通量之比）；Levitt， D. G., s . R. Elias, and J. M. Hautman. 1978。Biochim。Biophys。Acta. 512: 436-451)的计算结果为1.4。在对称的100 mM K+溶液中，耦合比显著降低（n = 0.9），表明离子占据率增加的状态下的渗透过程变得显著。我们提出了一个将水离子耦合比与离子渗透模式联系起来的图表，并提出1的耦合比可能代表了单孔中最少的水合离子通量。
The streaming potential (V stream) is a signature feature of ion channels in which permeating ions and water molecules move in a single file. V stream provides a quantitative measure of the ion and water flux (the water-ion coupling ratio), the knowledge of which is a prerequisite for elucidating the mechanisms of ion permeation. We have developed a method to measure V stream with the whole-cell patch-clamp configuration. A HEK293 cell stably expressing the HERG potassium channel was voltage clamped and exposed to hyperosmotic solutions for short periods of time (< 1 s) by an ultrafast solution switching system (the osmotic pulse [quick jump-and-away] method). The reversal potentials were monitored by a series of voltage ramps before, during, and after the osmotic pulse. The shifts of the reversal potentials immediately after the osmotic jump gave V stream. In symmetrical K+ solutions (10 mM), the V(stream)s measured at different osmolalities showed a linear relationship with a slope of -0.7 mV/Delta Osm, from which the water-ion coupling ratio (n, the ratio of the flux of water to the flux of cations; Levitt, D. G., S. R. Elias, and J. M. Hautman. 1978. Biochim. Biophys. Acta. 512: 436-451) was calculated to be 1.4. In symmetrical 100 mM K+ solutions, the coupling ratio was decreased significantly (n = 0.9), indicating that the permeation process through states with increased ion occupancy became significant. We presented a diagrammatic representation linking the water-ion coupling ratio to the mode of ion permeation and suggested that the coupling ratio of one may represent the least hydrated ion flux in the single-file pore.