LOCAL, STOCHASTIC RELEASE OF CA2-CLAMPED RAT-HEART CELLS - VISUALIZATION WITH CONFOCAL MICROSCOPY( IN VOLTAGE)

1. Confocal microscopy and the fluorescent Ca2+ indicator fluo-3 (K+ s alt) were used to measure cytosolic free calcium ion concentration ([C a-i(2+),) during excitation-contraction (E-C) coupling in single, volt age-clamped, rat cardiac ventricular cells. 2. Local [Ca2+](i) transie nts were measured nearly simultaneously in different, separate, subcel lular volumes of similar to 2.0 mu m(3). During depolarization, local [Ca2+](i) transients were distinctly different from each other and fro m whole-cell [Ca2+](i) transients. These differences were particularly apparent during small depolarizations, and were substantially reduced by ryanodine.3. Components of the local [Ca2+](i) transients, particu larly those evoked by small depolarizations, were closely similar, in time course and amplitude, to spontaneous local [Ca2+](i) transients, or 'sparks' (which have been shown previously to be Ca2+ released from sarcoplasmic reticulum). 4. Analysis of local [Ca2+](i) transients in the spatial frequency domain (power spectrum) revealed that high powe r at spatial frequencies of 0.05-0.2 mu m(-1) was always associated wi th spontaneous calcium 'sparks' and with local [Ca2+](i) transients ev oked by small depolarizing pulses (e.g. to -31 mV). Evoked local [Ca2](i) transients in the presence of ryanodine, and those evoked by depo larization to very postive clamp-pulse potentials (+45 mV), were assoc iated with considerably lower power at this frequency. 5. The results suggest that whole-cell [Ca2+](i) transients evoked by voltage-clamp d epolarization, and thus by L-type Ca2+ current, are comprised of local [Ca2+](i) transients that are similar to the spontaneous calcium 'spa rks'. At very positive clamp-pulse potentials, however, the electicall y evoked local [Ca2+](i) transients may be smaller, perhaps as a resul t of smaller unitary L-type Ca2+ current.