Partial Inhibition of Ca 2+ Current by Methoxyverapamil (D600) Reveals Spatial Nonuniformities in Ca 2+ i During Excitation-Contraction Coupling in Cardiac Myocytes

The laser scanning confocal microscope was used in conjunction with the Ca2+ indicator fluo 3 to examine the spatiotemporal properties of free Ca2+ (Ca2+i) transients in isolated rat cardiac myocytes. We show that localized increases in Ca2+i (Ca2+ sparks) can be triggered by membrane depolarization in cardiac myocytes when the sarcolemmal Ca2+ current amplitude is reduced by methoxyverapamil (D600). These depolarization-evoked Ca2+ sparks are similar in amplitude and spatiotemporal properties to spontaneous Ca2+ sparks previously observed at rest. These observations support the idea that Ca2+ sparks are the result of the activation of functional elementary units of sarcoplasmic reticulum (SR) Ca2+ release. The synchronous activation of a large number of Ca2+ sparks can explain the increased amplitude and slower time course of the electrically evoked Ca2+i transient as well as the presence of spatial nonuniformities in Ca2+i during its rise. The data shown here suggest a model for excitation-contraction coupling in which the amplitude of the Ca2+i transient is regulated by variations in the probability of recruitment of elementary SR Ca2+ release units as well as the amount of Ca2+ released by each unit. Since the activation of each release unit will depend on the local amplitude of the Ca2+ current, this model can explain the regulation of the amplitude of the Ca2+i transient by the Ca2+ current. In addition, these data indicate that caution should be applied to the interpretation of signals obtained with nonlinear Ca2+ indicators during the rising phase of the Ca2+i transient, when the nonuniformities in Ca2+i are largest.