Calcium sparks in muscle cells: interactive procedures for automatic detection and measurements on line-scan confocal images series.

In muscle cells, force development is controlled by Ca2+ ions, which are rapidly released from the sarcoplasmic reticulum (SR) during sarcolemmal depolarization. In addition to this synchronized spatially homogeneous calcium signal, localized quantal Ca2+ release events (sparks) have been recorded using laser scanning confocal fluorescence microscopy. Previously, algorithms without user intervention have been developed to automatically detect and analyse sparks on confocal line-scan (space-time: 512 x 512 pixels) single images. Here we present a computer program that we called "HARVest of Elementary Events" (HARVELE) developed to both analyse events on series of confocal images and follow sparks morphology from one site during several seconds. HARVELE, coded in the image-processing language IDL 5.3., can be applied on series of n images (512 x 512 x n) obtained from the same scanning line. Computing simultaneously entire series of images allows to measure, for each release site, the frequency and the morphology of release with the conventional amplitude, size, time and duration parameters defined for sparks analysis. The use of these procedures rapidly provides much information about the properties of calcium release sites in muscle cells population and can be applied on any elementary calcium events whatever the type of cell.