Optical single-channel recording:: imaging Ca2+ flux through individual N-type voltage-gated channels expressed in Xenopus oocytes
Optical single-channel recording:: imaging Ca2+ flux through individual N-type voltage-gated channels expressed in Xenopus oocytes
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DOI：
10.1016/s0143-4160(03)00154-4
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发表时间：
2003-12-01
期刊：
影响因子：
4
通讯作者：
Parker, I
中科院分区：
文献类型：
--
作者：
Demuro, A;Parker, I
膜片钳技术的引入使得单膜离子通道的功能研究成为可能，该技术允许以前所未有的分辨率测量单通道电流。然而，膜片钳也有一些局限性：包括需要物理接触膜片移液管、可能破坏局部细胞结构、无法监测多个通道以及缺乏空间信息。在这里，我们演示了使用共焦荧光显微镜作为一种非侵入性技术来光学监测单个 Ca2+ 通道的门控。近膜荧光信号以约 10 ms 的动力学分辨率跟踪 N 型 Ca2+ 通道的门控，提供来自多个通道的同时且独立的读数，并允许以亚微米空间分辨率绘制它们的位置。光学单通道记录应适用于不同的电压门控和配体门控 Ca2+ 通透通道，并具有单通道高通量功能分析的潜力。 (C) 2003 Elsevier Ltd. 保留所有权利。
Functional studies of single membrane ion channels were made possible by the introduction of the patch-clamp technique, which allows single-channel currents to be measured with unprecedented resolution. Nevertheless, patch clamping has some limitations: including the need for physical access of the patch pipette, possible disruption of local cellular architecture, inability to monitor multiple channels, and lack of spatial information. Here, we demonstrate the use of confocal fluorescence microscopy as a non-invasive technique to optically monitor the gating of individual Ca2+ channels. Near-membrane fluorescence signals track the gating of N-type Ca2+ channels with a kinetic resolution of about 10 ms, provide a simultaneous and independent readout from several channels, and allow their locations to be mapped with sub-micrometer spatial resolution. Optical single-channel recording should be applicable to diverse voltage- and ligand-gated Ca2+-permeable channels, and has the potential for high-throughput functional analysis of single channels. (C) 2003 Elsevier Ltd. All rights reserved.