DESCRIPTION (adapted from the applicant's description): This proposal studies calcium regulation in intact ventricular cells with a focus on sarcoplasmic reticulum (SR) calcium release and the sodium/calcium (Na/Ca) exchanger. The proposal has three specific aims. Specific aim 1 determines the amount of calcium released during a calcium spark and the number of SR calcium release channels (ryanodine receptors, RYRs) contributing to a calcium spark (i.e. cluster size). Specific aim 2 measures the flux of SR calcium release during an action potential. Specific aim 3 explores the contribution of Na/Ca exchange current to the action potential and the trigger for SR calcium release. The majority of these studies will be performed on single cardiac ventricular cells isolated from adult rabbits, rats, mice and ferrets. Membrane currents (both whole cell and single channel) will be measured with the patch clamp technique. Whole-cell calcium transients will be measured with indo-1 by standard techniques in epifluorescence microscopy. Local calcium transients (calcium sparks and sarcolemmal calcium transients) will be visualized as fluo-3 fluorescence with laser scanning confocal microscopy.