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Journal of Biomedical Optics

Optical single-channel recording: imaging Ca2+ flux through individual ion channels with high temporal and spatial resolution
Author(s): Angelo Demuro; Ian Parker
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Paper Abstract

Developments in imaging technology now enable visualization of the functioning of individual ion channels in living cells: something previously possible only by the electrophysiological patch-clamp technique. We review techniques that track channel gating via changes in intracellular [Ca2+] resulting from openings of Ca2+-permeable channels. Spatial and temporal resolution are optimized by monitoring Ca2+ close to the channel mouth, and we describe the use of two imaging modalities: confocal laser scan microscopy (linescan CLSM) and total internal reflection fluorescence microscopy (TIRFM). Both currently achieve a kinetic resolution of <10 ms, provide a simultaneous and independent readout from many channels, and enable their locations to be mapped with submicrometer resolution. TIRFM provides 2-D images from a very thin (~100 nm) optical section, but it is restricted to channels in the plasma membrane of cells adhering close to a cover glass. In contrast, CLSM can image channels in intracellular membranes but, to achieve good temporal resolution, has been utilized only in a linescan mode with limited spatial information. We anticipate that imaging techniques will develop as a useful adjunct to patch-clamping for single-channel studies, with capabilities including simultaneous readout from multiple channels, high-resolution mapping of channel location, and mobility that is inaccessible by electrophysiological means. Optical single-channel recording is applicable to diverse voltage- and ligand-gated Ca2+-permeable channels and has potential for high-throughput functional analysis.

Paper Details

Date Published: 1 January 2005
PDF: 8 pages
J. Biomed. Opt. 10(1) 011002 doi: 10.1117/1.1846074
Published in: Journal of Biomedical Optics Volume 10, Issue 1
Show Author Affiliations
Angelo Demuro, Univ. of California/Irvine (United States)
Ian Parker, Univ. of California/Irvine (United States)

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