Sarcoplasmic reticulum Ca2+ cycling protein phosphorylation in a physiologic Ca2+ milieu unleashes a high-power, rhythmic Ca2+ clock in ventricular myocytes: relevance to arrhythmias and bio-pacemaker design

J Mol Cell Cardiol. 2014 Jan;66:106-15. doi: 10.1016/j.yjmcc.2013.11.011. Epub 2013 Nov 22.


Basal phosphorylation of sarcoplasmic reticulum (SR) Ca(2+) proteins is high in sinoatrial nodal cells (SANC), which generate partially synchronized, spontaneous, rhythmic, diastolic local Ca(2+) releases (LCRs), but low in ventricular myocytes (VM), which exhibit rare diastolic, stochastic SR-generated Ca(2+) sparks. We tested the hypothesis that in a physiologic Ca(2+) milieu, and independent of increased Ca(2+) influx, an increase in basal phosphorylation of SR Ca(2+) cycling proteins will convert stochastic Ca(2+) sparks into periodic, high-power Ca(2+) signals of the type that drives SANC normal automaticity. We measured phosphorylation of SR-associated proteins, phospholamban (PLB) and ryanodine receptors (RyR), and spontaneous local Ca(2+) release characteristics (LCR) in permeabilized single, rabbit VM in physiologic [Ca(2+)], prior to and during inhibition of protein phosphatase (PP) and phosphodiesterase (PDE), or addition of exogenous cAMP, or in the presence of an antibody (2D12), that specifically inhibits binding of the PLB to SERCA-2. In the absence of the aforementioned perturbations, VM could only generate stochastic local Ca(2+) releases of low power and low amplitude, as assessed by confocal Ca(2+) imaging and spectral analysis. When the kinetics of Ca(2+) pumping into the SR were increased by an increase in PLB phosphorylation (via PDE and PP inhibition or addition of cAMP) or by 2D12, self-organized, "clock-like" local Ca(2+) releases, partially synchronized in space and time (Ca(2+) wavelets), emerged, and the ensemble of these rhythmic local Ca(2+) wavelets generated a periodic high-amplitude Ca(2+) signal. Thus, a Ca(2+) clock is not specific to pacemaker cells, but can also be unleashed in VM when SR Ca(2+) cycling increases and spontaneous local Ca(2+) release becomes partially synchronized. This unleashed Ca(2+) clock that emerges in a physiological Ca(2+) milieu in VM has two faces, however: it can provoke ventricular arrhythmias; or if harnessed, can be an important feature of novel bio-pacemaker designs.

Keywords: AP; Ca(2+); Calcium clock; Calcium cycling; Cardiac ventricular myocytes; IBMX; LCR; PDE; PKA; PLB; PP; Protein phosphorylation; RyRs; SANC; SERCA-2; SR; SR Ca(2+) pump; Spontaneous local calcium releases; VM; action potential; calcium; isobutyl-1-methylxanthine; local Ca(2+) releases; phosphodiesterase; phospholamban; protein kinase A; protein phosphatases; ryanodine receptors; sarcoplasmic reticulum; sinoatrial node cells; ventricular myocytes; β-ARs; β-adrenergic receptor stimulation.

Publication types

  • Research Support, N.I.H., Intramural

MeSH terms

  • Animals
  • Antibodies / pharmacology
  • Biological Clocks / genetics*
  • Calcium / metabolism*
  • Calcium-Binding Proteins / genetics
  • Calcium-Binding Proteins / metabolism*
  • Cyclic AMP / metabolism
  • Gene Expression Regulation
  • Heart Ventricles / cytology
  • Heart Ventricles / metabolism
  • Myocytes, Cardiac / cytology
  • Myocytes, Cardiac / metabolism*
  • Pacemaker, Artificial
  • Phosphoprotein Phosphatases / genetics
  • Phosphoprotein Phosphatases / metabolism
  • Phosphoric Diester Hydrolases / genetics
  • Phosphoric Diester Hydrolases / metabolism
  • Phosphorylation
  • Protein Binding
  • Rabbits
  • Ryanodine Receptor Calcium Release Channel / genetics
  • Ryanodine Receptor Calcium Release Channel / metabolism*
  • Sarcoplasmic Reticulum / metabolism*
  • Sarcoplasmic Reticulum Calcium-Transporting ATPases / antagonists & inhibitors
  • Sarcoplasmic Reticulum Calcium-Transporting ATPases / genetics
  • Sarcoplasmic Reticulum Calcium-Transporting ATPases / metabolism
  • Signal Transduction
  • Sinoatrial Node / cytology
  • Sinoatrial Node / metabolism


  • Antibodies
  • Calcium-Binding Proteins
  • Ryanodine Receptor Calcium Release Channel
  • phospholamban
  • Cyclic AMP
  • Phosphoprotein Phosphatases
  • Phosphoric Diester Hydrolases
  • Sarcoplasmic Reticulum Calcium-Transporting ATPases
  • Calcium