[Ca2+]i was measured using the fluorescent indicator indo 1 in voltage-clamped ferret and rat ventricular myocytes. The Ca2+ content of the sarcoplasmic reticulum (SR) was estimated from the integral of the Na(+)-Ca2+ exchange current activated by caffeine. Refilling of the SR after caffeine removal was enhanced by stimulation. As the systolic Ca2+ transient recovered, the integral of the L-type Ca2+ current decreased and that of the Na(+)-Ca2+ exchange tail current increased. For the early pulses, the gain of Ca2+ via the Ca2+ current is greater than the loss via the exchanger, and during steady state stimulation, the fluxes are equal. The difference in the integrals gives a measure of the net gain of cell Ca2+ with each pulse. When these are summed, the calculated gain of cell Ca2+ agrees well with the increase of SR Ca2+ produced by stimulation, as measured from the caffeine-evoked currents. There was a nonlinear relationship between SR Ca2+ content and the magnitude of the systolic Ca2+ transient such that at high SR Ca2+ content a given increase of content had a greater effect on the Ca2+ transient than did an increase at low SR content. In conclusion, the effects of systolic Ca2+ on the Ca2+ current and Na(+)-Ca2+ exchange current provide a means to regulate SR Ca2+ content and thence the systolic Ca2+ transient.