Contraction of the mammalian cardiac myocyte is activated by the release of Ca2+ from sarcoplasmic reticulum (SR), triggered by a rapid influx of a small amount of Ca2+ through voltage-gated Ca2+ channels of the surface membrane. The SR Ca2+ release is determined both, by characteristics of the trigger, ie. the transmembrane Ca2+ current (ICa) and by SR Ca2+ load. This project focuses on the effects of SR Ca2+ load on amplification of Ca2+ influx via ICa by SR Ca2+ release, ie. the gain of release. We also ask the question whether control of SR Ca2+ release by voltage-dependent Ca2+ influ is modulated by changes in the SR Ca2+ load. The relation between the exten of SR Ca2+ loading and release selectively by ICa is examined under experimental conditions in which both factors can be simultaneously measure in intact cardiac cells. Isolated rat ventricular myocytes are voltage clamped and loaded with Ca2+ indicator, indo-1/salt. Ca2+ influx via the sarcolemmal Na+-Ca2+ exchanger is inhibited by 0 Na+ in the patch pipette. Potassium currents and the fast Na+ current, which interfere with measurements of ICa, are suppressed by the composition of the patch pipette filling solution and of the extracellular medium. Under these conditions, Ca2+ current is measured directly and provides an exclusive trigger for SR Ca2+ release, and a sole source of changes in the SR Ca2+ load. The SR Ca2 load is graded by the duration of conditioning voltage clamp steps and verified by the caffeine-dependent Ca2+i transients. The SR Ca2+ release i assessed from amplitudes and rates of rise of the ICa-dependent Ca2+i transients. The gain of SR Ca2+ release is indexed by the ratio between changes in the intracellular Ca2+ activity and the magnitude of the integrated Ca2+ current. The results obtained so far show that within the extent of changes in the SR Ca2+ loading achieved under the present experimental conditions, changes of the gain index linearly correlate with changes in the SR Ca2+ releasable with caffeine. We also find that the SR Ca2+ load modulates the effect of the duration of Ca2+ influx via ICa on th Ca2+i transient and contraction. Thus, the SR Ca2+ release by a given Ca2+ current to activate cardiac contraction is highly dependent on the SR Ca2+ load. The linear dependence of gain on SR Ca2+ loading provides further evidence against the existence of strong positive feedback of released Ca2+ on the Ca2+ release triggering mechanism and has theoretical implications for force-interval and staircase phenomena in the heart.
