Phosphorylation-Dependent Synchronization of Random Spontaneous Local Diastolic Ca2+ Releases Regulates Action Potential Firing Rhythmicity of Pacemaker Cells

Abstract

Substantial variability in heart beat intervals is detected within EKG time series analyses of numerous species (from mice to humans), and has often been linked to variation in autonomic neural impulses to the heart. Studies in single isolated sinoatrial nodal cells (SANC), however, indicate that mechanisms intrinsic to pacemaker cells not only regulate the average AP beating interval (APBI), but also determine APBI variability (APBIV). Furthermore, in permeablized SANC, which do not generate APs, phosphorylation of SR Ca2+ cycling proteins is one mechanism that regulates the spatiotemporal synchronization of spontaneous local RyR activation resulting in local Ca2+ release (LCRs) of shorter periods and reduced period variability. We tested the idea that the spatiotemporal synchronization of spontaneous local diastolic RyR activation linked to phosphorylation of Ca2+ cycling proteins is a determinant of both the average APBI and APBIV of intact SANC. Reduced SR Ca2+ cycling protein phosphorylation and increased APBI of cultured adult rabbit SANC (c-SANC) were accompanied by reduced kinetics and increased beat-to-beat variation of the SR refilling rate with Ca2+, as reflected in the decay time of AP induced Ca2+-transients; spatiotemporal de-synchronization of spontaneous, local RyR activation, was manifested by increased average LCR period and its variability, and by increased variability of surface membrane AP parameters. Increased protein phosphorylation effected by beta-AR stimulation in both c-SANC and freshly isolated SANC, accelerated the kinetics and reduced beat-to-beat variability of SR Ca2+ refilling, increased the spatiotemporal synchronization of LCR periods, reduced the variability of AP parameters and reduced both APBIV and average APBI. Thus, both the spontaneous AP firing rhythm and average firing rate of isolated SANC are linked to synchronization of random, local spontaneous RyR activation, modulated by SR Ca2+ cycling protein phosphorylation.