Stochasticity intrinsic to coupled-clock mechanisms underlies beat-to-beat variability of spontaneous action potential firing in sinoatrial node pacemaker cells

Recent evidence indicates that the spontaneous action potential (AP) of isolated sinoatrial node cells (SANCs) is regulated by a system of stochastic mechanisms embodied within two clocks: ryanodine receptors of the "Ca²⁺ clock" within the sarcoplasmic reticulum, spontaneously activate during diastole and discharge local Ca²⁺ releases (LCRs) beneath the cell surface membrane; clock crosstalk occurs as LCRs activate an inward Na⁺/Ca²⁺ exchanger current (I_NCX), which together with I_f and decay of K⁺ channels prompts the "M clock," the ensemble of sarcolemmal-electrogenic molecules, to generate APs. Prolongation of the average LCR period accompanies prolongation of the average AP beating interval (BI). Moreover, the prolongation of the average AP BI accompanies increased AP BI variability. We hypothesized that both the average AP BI and AP BI variability are dependent upon stochasticity of clock mechanisms reported by the variability of LCR period.We perturbed the coupled-clock system by directly inhibiting the M clock by ivabradine (IVA) or the Ca²⁺ clock by cyclopiazonic acid (CPA). When either clock is perturbed by IVA (3, 10 and 30μM), which has no direct effect on Ca²⁺ cycling, or CPA (0.5 and 5μM), which has no direct effect on the M clock ion channels, the clock system failed to achieve the basal AP BI and both AP BI and AP BI variability increased. The changes in average LCR period and its variability in response to perturbations of the coupled-clock system were correlated with changes in AP beating interval and AP beating interval variability. We conclude that the stochasticity within the coupled-clock system affects and is affected by the AP BI firing rate and rhythm via modulation of the effectiveness of clock coupling.