Rate-dependent Ca2+ signalling underlying the force-frequency response in rat ventricular myocytes: A coupled electromechanical modeling study

Rate-dependent effects on the Ca2+ sub-system in a rat ventricular myocyte are investigated. Here, we employ a deterministic mathematical model describing various Ca2+ signalling pathways under voltage clamp (VC) conditions, to better understand the important role of calmodulin (CaM) in modulating the key control variables Ca2+/calmodulin-dependent protein kinase-II (CaMKII), calcineurin (CaN), and cyclic adenosine monophosphate (cAMP) as they affect various intracellular targets. In particular, we study the frequency dependence of the peak force generated by the myofilaments, the force-frequency response (FFR). Our cell model incorporates frequency-dependent CaM-mediated spatially heterogenous interaction of CaMKII and CaN with their principal targets (dihydropyridine (DHPR) and ryanodine (RyR) receptors and the SERCA pump). It also accounts for the rate-dependent effects of phospholamban (PLB) on the SERCA pump; the rate-dependent role of cAMP in up-regulation of the L-type Ca2+ channel (ICa;L); and the enhancement in SERCA pump activity via phosphorylation of PLB.Our model reproduces positive peak FFR observed in rat ventricular myocytes during voltage-clamp studies both in the presence/absence of cAMP mediated -adrenergic stimulation. This study provides quantitative insight into the rate-dependence of Ca2+-induced Ca2+-release (CICR) by investigating the frequency-dependence of the trigger current (ICa;L) and RyR-release. It also highlights the relative role of the sodium-calcium exchanger (NCX) and the SERCA pump at higher frequencies, as well as the rate-dependence of sarcoplasmic reticulum (SR) Ca2+ content. A rigorous Ca2+ balance imposed on our investigation of these Ca2+ signalling pathways clarifies their individual roles. Here, we present a coupled electromechanical study emphasizing the rate-dependence of isometric force developed and also investigate the temperature-dependence of FFR. Our model provides mechanistic biophysically based explanations for the rate-dependence of CICR, generating useful and testable hypotheses. Although rat ventricular myocytes exhibit a positive peak FFR in the presence/absence of beta-adrenergic stimulation, they show a characteristic increase in the positive slope in FFR due to the presence of Norepinephrine or Isoproterenol. Our study identifies cAMP-mediated stimulation, and rate-dependent CaMKII-mediated up-regulation of ICa;L as the key mechanisms underlying the aforementioned positive FFR

The cytosolic phospholipase A2 pathway, a safeguard of beta2-adrenergic cardiac effects in rat.

reserved10We have recently demonstrated that in human heart, β(2)-adrenergic receptors (β(2)-ARs) are biochemically coupled not only to the classical adenylyl cyclase (AC) pathway but also to the cytosolic phospholipase A(2) (cPLA(2)) pathway (Pavoine, C., Behforouz, N., Gauthier, C., Le Gouvello, S., Roudot-Thoraval, F., Martin, C. R., Pawlak, A., Feral, C., Defer, N., Houel, R., Magne, S., Amadou, A., Loisance, D., Duvaldestin, P., and Pecker, F. (2003) Mol. Pharmacol. 64, 1117-1125). In this study, using Fura-2-loaded cardiomyocytes isolated from adult rats, we showed that stimulation of β(2)-ARs triggered an increase in the amplitude of electrically stimulated [Ca2+](i) transients and contractions. This effect was abolished with the PKA inhibitor, H89, but greatly enhanced upon addition of the selective cPLA(2) inhibitor, AACOCF(3). The β(2)-AR/cPLA(2) inhibitory pathway involved G(i) and MSK1. Potentiation of β(2)-AR/AC/PKA-induced Ca2+ responses by AACOCF3 did not rely on the enhancement of AC activity but was associated with eNOS phosphorylation (Ser(1177)) and L-NAME-sensitive NO production. This was correlated with PKA-dependent phosphorylation of PLB (Ser(16)). The constraint exerted by the β(2)-AR/cPLA(2) pathway on the β(2)-AR/AC/PKA-induced Ca2+ responses required integrity of caveolar structures and was impaired by Filipin III treatment. Immunoblot analyses demonstrated zinterol-induced translocation of cPLA(2) and its cosedimentation with MSK1, eNOS, PLB, and sarcoplasmic reticulum Ca2+ pump (SERCA) 2a in a low density caveolin-3-enriched membrane fraction. This inferred the gathering of β(2)-AR signaling effectors around caveolae/sarcoplasmic reticulum (SR) functional platforms. Taken together, these data highlight cPLA(2) as a cardiac β(2)-AR signaling pathway that limits β(2)-AR/AC/PKA-induced Ca2+ responses in adult rat cardiomyocytes through the impairment of eNOS activation and PLB phosphorylation.B. AIT-MAMAR; M. CAILLERET; C. RUCKER-MARTIN; A. BOUABDALLAH; G. CANDIANI; C. ADAMY; P. DUVALDESTIN; F. PECKER; N. DEFER; C. PAVOINEB., AIT MAMAR; M., Cailleret; C., RUCKER MARTIN; A., Bouabdallah; Candiani, Gabriele; C., Adamy; P., Duvaldestin; F., Pecker; N., Defer; C., Pavoin