Imaging single-channel calcium microdomains.
The Ca(2+) microdomains generated around the mouth of open ion channels represent the basic building blocks from which cytosolic Ca(2+) signals are constructed. Recent improvements in optical imaging techniques now allow these microdomains to be visualized as single channel calcium fluorescence transients (SCCaFTs), providing information about channel properties that was previously accessible only by electrophysiological patch-clamp recordings. We review recent advances in single channel Ca(2+) imaging methodologies, with emphasis on total internal reflection fluorescence microscopy (TIRFM) as the technique of choice for recording SCCaFTs from voltage- and ligand-gated plasmalemmal ion channels. This technique of 'optical patch-clamp recording' is massively parallel, permitting simultaneous imaging of hundreds of channels; provides millisecond resolution of gating kinetics together with sub-micron spatial resolution of channel locations; and is applicable to diverse families of membrane channels that display partial permeability to Ca(2+) ions.
Demuro
Angelo
A
Department of Neurobiology and Behavior, University of California, Irvine, CA 92697-4550, USA.
Parker
Ian
I
D000818:Animals; D015220:Calcium Channels / Q000648:ultrastructure*; D020864:Calcium Channels, N-Type / Q000502:physiology; D020013:Calcium Signaling / Q000502:physiology*; D002462:Cell Membrane / Q000648:ultrastructure; D005456:Fluorescent Dyes; D015640:Ion Channel Gating; D021962:Membrane Microdomains / Q000648:ultrastructure*; D008856:Microscopy, Fluorescence / Q000379:methods*; D009865:Oocytes / Q000378:metabolism; D055095:Optics and Photonics; D018408:Patch-Clamp Techniques / Q000379:methods; D014982:Xenopus laevis
D015220:Calcium Channels; D020864:Calcium Channels, N-Type; D005456:Fluorescent Dyes