Seminar Abstract

“Intracellular [Na+] modulates synergism between Na+/Ca2+ exchanger and Ca2+ current in cardiac excitation-contraction coupling during an action potential”

Contraction of cardiac myocytes is initiated by Ca2+ influx through L-type Ca channels, which induces Ca2+ release from the sarcoplasmic reticulum (SR). The contribution of Ca2+ entry via the Na+/Ca2+ exchanger (NCX) to triggering Ca2+ release during excitation-contraction coupling (ECC) during an action potential (AP) remains uncertain. In order to isolate the contribution of NCX to SR Ca2+ release, independent of its effects on SR Ca2+ load, Ca2+ release was determined by recording Ca2+ spikes using confocal microscopy of patch-clamped rat ventricular myocytes with [Ca2+]i fixed at 150 nmol/L. In response to AP clamps, normalized Ca2+ spike amplitudes (ΔF/F0) increased (P<0.005) sigmoidally from 0.40±0.08 to 0.80±0.06 as [Na+]i was elevated from 0 to 20 mmol/L with an EC50 of 9.08±0.97 mmol/L. The [Na+]i dependent enhancement of SR Ca2+ release was independent of INa or SR Ca2+ load. However, NCX inhibition using either 5 μmol/L KB-R7943 or 30 μmol/L XIP reduced (P<0.05) ΔF/F0 amplitudes in myocytes with 20 mmol/L [Na+]i but not with 5 mmol/L [Na+]i. Since ICa,L inhibition with 50 mmol/L Cd2+ totally abolished Ca2+ spikes, our results demonstrate that, during a cardiac AP at elevated [Na+]i, NCX enhances SR Ca2+ release, by synergistically increasing the efficiency of ICa,L-mediated ECC. Additionally, the slope of the initial repolarization phase of the cardiac AP is a key regulator of ECC. We hypothesize that this synergy between NCX and ICa,L is responsible for AP dependent modulation of cardiac ECC. In voltage-clamped rat ventricular myocytes, we measured Ca2+ spikes triggered by a family of APs with varying slopes of initial repolarization. In myocytes containing 20 mmol/L [Na+]i, ΔF/F0 displayed a biphasic relationship with AP duration at 50% repolarization (APD50), rising initially then decreasing as APD50 of the triggering AP increased from 4 to 52 ms. Maximal Ca2+ release flux was achieved when APD50 was 16 ms (ΔF/F0=0.77±0.06, n=17). In the presence of XIP the APD-dependence of Ca2+ spikes was blunted, ΔF/F0 decreased (P<0.02) to 0.60±0.06 at 16 ms APD50. In myocytes containing 5 mmol/L [Na+]i, the APD-dependence was also blunted (ΔF/F0=0.53±0.06, at 9 ms APD50, n=16) and was unaffected by XIP. We conclude that the biphasic enhancement of SR Ca2+ release with prolongation of APD is achieved through functional synergy between NCX and L-type Ca2+ channel trigger sources of CICR.