Gil Z, Magleby K L, Silberberg S D
Department of Life Sciences and The Zlotowski Center for Neuroscience, Ben-Gurion University of the Negev, Beer Sheva, 84105, Israel.
Biophys J. 1999 Jun;76(6):3118-27. doi: 10.1016/S0006-3495(99)77463-1.
To investigate the mechanism for the delayed activation by voltage of the predominant mechanosensitive (MS) channel in Xenopus oocytes, currents were recorded from on-cell and excised patches of membrane with the patch clamp technique and from intact oocytes with the two-electrode voltage clamp technique. MS channels could be activated by stretch in inside-out, on-cell, and outside-out patch configurations, using pipettes formed of either borosilicate or soft glass. In inside-out patches formed with borosilicate glass pipettes, depolarizing voltage steps activated MS channels in a cooperative manner after delays of seconds. This voltage-dependent activation was not observed for outside-out patches. Voltage-dependent activation was also not observed when the borosilicate pipettes were either replaced with soft glass pipettes or coated with soft glass. When depolarizing voltage steps were applied to the whole oocyte with a two-electrode voltage clamp, currents that could be attributed to MS channels were not observed. Yet the same depolarizing steps activated MS channels in on-cell patches formed with borosilicate pipettes on the same oocyte. These observations suggest that the delayed cooperative activation of MS channels by depolarization is not an intrinsic property of the channels, but requires interaction between the membrane and patch pipette.
为了研究非洲爪蟾卵母细胞中主要机械敏感（MS）通道电压延迟激活的机制，我们采用膜片钳技术从细胞膜上的贴壁和切除的膜片记录电流，并采用双电极电压钳技术从完整的卵母细胞记录电流。使用由硼硅酸盐或软玻璃制成的移液管，MS通道可以在膜片外翻、贴壁和膜片外转的配置中通过拉伸激活。在用硼硅酸盐玻璃移液管形成的膜片外翻膜片中，去极化电压阶跃在延迟数秒后以协同方式激活MS通道。对于膜片外转膜片，未观察到这种电压依赖性激活。当硼硅酸盐移液管被软玻璃移液管替换或用软玻璃涂层时，也未观察到电压依赖性激活。当用双电极电压钳向整个卵母细胞施加去极化电压阶跃时，未观察到可归因于MS通道的电流。然而，相同的去极化阶跃激活了在同一卵母细胞上用硼硅酸盐移液管形成的贴壁膜片中的MS通道。这些观察结果表明，去极化对MS通道的延迟协同激活不是通道的固有特性，而是需要膜与膜片移液管之间的相互作用。
