Functional coupling between sarcoplasmic reticulum and Na/Ca exchange in single myocytes of guinea-pig and rat heart

It has been proposed that the cardiac Na/Ca exchanger is primed by high Ca2+ concentrations derived from the sarcoplasmic reticulum (SR) in a recently identified subsarcolemmal, "Na/Ca exchange-dependent Ca2+ compartment". We tested this hypothesis by investigating the effect on Na/Ca exchange of interventions affecting Ca2+ flux through SR. Experiments were performed in single, isolated myocytes of guinea-pig and rat hearts loaded with Indo 1-AM and free Ca2+ concentration was assessed by measuring the ratio of fluorescence at 405 and 495 nm wavelength. In guinea-pig 1.0 microM ryanodine (Ry), expected to increase Ca2+ flux through the SR, decreased the amplitude of electrically stimulated Ca2+ transients by 35% and inhibited their initial, rapid phase. Responses of these cells to brief superfusions with 15 mM caffeine were inhibited which suggests that 1.0 microM Ry depleted the SR Ca2+ stores. Diastolic Ca2+ concentration was slightly increased, but it dropped below control during prolonged rest. Decrease of the amplitude of the transients in these ryanodine-treated cells were reversed by 2 x 10(-7) M thapsigargin (TG), an inhibitor of the SR Ca(2+)-ATPase, by 1.0 mM Ry, a blocker of the SR Ca2+ release channels and by low Na+ (50.0 mM) superfusion. This suggests that the decreased transients in 1.0 microM Ry result from uptake of Ca2+ by the SR and its rapid release to the subsarcolemmal (cleft) space where a fraction is diverted out of the cell via Na/Ca exchange before it can diffuse to the myofilaments. Removal of the SR from the pathway (addition of TG on 1.0 mM Ry) or reversal of Na/Ca exchange diverts more transsarcolemmal Ca2+ influx to the myofilaments and increases the Ca2+ transient. Decrease of the resting Ca2+ concentration was blocked by 2 x 10(-7) M TG, 1.0 mM Ry, and by 5.0 mM Ni2+, a blocker of Na/Ca exchange. This result suggests that the effect of 1.0 microM Ry on resting Ca2+ concentration also resulted from increase of flux of Ca2+ through the SR and out of the cell by Na/Ca exchange. In rat 1.0 microM Ry decreased amplitude of the transients by approximately 75% but did not affect their kinetics. TG and 1.0 mM Ry decreased the rate of rise of the transients and greatly delayed their decay. This result suggests that normal kinetics of the transients in the cells treated with 1.0 microM Ry depended on the preserved Ca2+ flux through the SR. As SR was not able to retain Ca2+ in these cells, decay of the transients must have depended on stimulated Na/Ca exchange. The results in guinea-pig and rat taken together are compatible with the proposal that Ca2+ released from the SR interacts with the cell's Na/Ca exchanger most probably within the newly defined subsarcolemmal "Na/Ca exchange-dependent Ca2+ compartment".