Abstract 13555: Synergism of PDE3 and PDE4 Activity Regulates cAMP-Mediated PKA-Dependent Local Ca2+ Releases to Modulate Basal Spontaneous Firing of Cardiac Pacemaker Cells
The spontaneous beating rate of rabbit sinoatrial node cells (SANC) is controlled by cAMP-mediated, PKA-dependent (cAMP/PKA) local subsarcolemmal Ca2+ releases (LCRs) from ryanodine receptors (RyR). LCRs occur during diastolic depolarization (DD) and activate an inward Na+/Ca2+ exchange current that increases DD rate and regulates spontaneous SANC firing. Since PDE3 and PDE4 represent major PDE activities in rabbit SANC, we tested hypothesis that combined activity of (PDE3+PDE4) impacts on basal spontaneous SANC firing via regulation of cAMP/PKA signaling. Phosphorylation of phospholamban (PLB) at Ser16 site was used as a marker of cAMP/PKA-dependent protein phosphorylation in SANC. Specific PDE3 inhibitor, cilostamide (Cil, 0.3 μmol/L), or PDE4 inhibitor, rolipram (Rol, 2 μmol/L), increased PLB phosphorylation by ∼20%, but combination of Cil+Rol increased PLB phosphorylation by ∼110%, an effect similar to that (∼140%) produced by broad spectrum PDE inhibitor IBMX. L-type Ca2+ current (ICa,L) ensures LCR existence, providing Ca2+ available for pumping in SR. Cil or Rol alone increased the amplitude of ICa,L by ∼60% and ∼4%, respectively, while (Cil+Rol) or IBMX increased ICa,L by ∼100%. Cil increased the spontaneous SANC firing rate (perforated patch-clamp) by ∼20% (from 148±13 to 175±13 beat/min, n=8), while Rol produced no acceleration of spontaneous firing at 2, 20 or 100 μmol/L. Similar to IBMX, (Cil+Rol) increased the spontaneous SANC beating rate by ∼50% (from 134±8 to 197±11 beat/min, n=9), the effect was due to a marked increase in the LCR number, size and decrease in the LCR period that predicted the concomitant decrease in the spontaneous cycle length. When RyR were disabled by ryanodine and LCRs were abolished, both IBMX and (Cil+Rol) failed to accelerate DD rate or increase SANC firing rate indicating key role of Ca2+ cycling for PDE-dependent control of spontaneous beating. We conclude that both PDE3 and PDE4 regulate spontaneous SANC firing, and a crucial role of PDE4 is revealed only when PDE3 and PDE4 are concurrently inhibited. Thus, synergism of PDE3+PDE4 suppresses basal cAMP/PKA-dependent PLB phosphorylation and reduces ICa,L amplitude to decrease RyR Ca2+ release, prolong the LCR period and limit the spontaneous SANC firing rate.
- © 2012 by American Heart Association, Inc.