Ca2+ sparks evoked by depolarization of rat ventricular myocytes involve multiple release sites

AIM: To investigate the fundamental nature of calcium release events (Ca2+ 'sparks') evoked in rat ventricular myocytes during excitation-contraction (E-C) coupling. METHODS: High-resolution line-scan confocal imaging with the fluorescent calcium indicator and patch-clamp techniques were used to study the spontaneous Ca2+ sparks and sparks evoked by depolarization. RESULTS: 1) Line scans oriented along the length of the cell showed that both spontaneous sparks and sparks evoked by depolarization to -35 mV appeared to arise at single sites spacing about 1.8 mum apart (ie, the sarcomere. length), and measurements of their longitudinal spread (full-width at half-maximal amplitude: FWHM) followed single Gaussian distributions with means of 2.6 mum. 2) Different to this, transverse line scans often revealed spontaneous and evoked sparks that appeared to arise near-synchronously from paired sites. Measurements of transverse FWHM of both spontaneous and evoked sparks showed bimodal distributions, which were fit well by the sums of two Gaussian curves with means of 1.8 and 2.9 mum for spontaneous sparks and with means of 1.9 and 3.1 mum for evoked sparks. Relative areas under the two Gaussian curves were 1.73:1 and 1.85:1, respectively, for spontaneous and evoked sparks. CONCLUSIONS: Ca2+ sparks evoked by depolarization are not 'unitary' events, but often involve multiple sites of origin along Z-lines, as previously shown for spontaneous sparks. Thus, Ca2+ released during sparks directly triggered by influx through L-type Ca2+ channels may, in turn, trigger neighboring sites. The restricted involvement of only a few transverse release sites preserves the essential feature of the 'local control' theory of E-C coupling.