Essential role of the RIα subunit of cAMP-dependent protein kinase in regulating cardiac contractility and heart failure development
Résumé
Background: The heart expresses two main subtypes of cAMP-dependent protein kinase (PKA type I and II) differing in their regulatory subunits, RIα and RIIα. Embryonic lethality of RIα knockout mice limits the current understanding of type I PKA function in the myocardium. The objective of this study was to test the role of RIα in adult heart contractility and pathological remodelling.
Methods: We measured PKA subunit expression in human heart and developed a conditional mouse model with cardiomyocyte-specific knockout of RIα (RIα-icKO). Myocardial structure and function were evaluated by echocardiography, histology, ECG and in Langendorff-perfused hearts. PKA activity and cAMP levels were determined by immunoassay and phosphorylation of PKA targets was assessed by western blot. L-type Ca2+ current (ICa,L ), sarcomere shortening, Ca2+ transients, Ca2+ sparks and waves and subcellular cAMP were recorded in isolated ventricular myocytes (VMs).
Results: RIα protein was decreased by 50% in failing human heart with ischemic cardiomyopathy and by 75% in the ventricles and in VMs from RIα-icKO mice but not in atria or sino-atrial node. Basal PKA activity was increased ~3-fold in RIα-icKO VMs. In young RIα-icKO mice, left ventricular ejection fraction was increased and the negative inotropic effect of propranolol was prevented, whereas heart rate (HR) and the negative chronotropic effect of propranolol were not modified. Phosphorylation of phospholamban, ryanodine receptor, troponin I and cardiac myosin-binding protein C at PKA sites were increased in propranolol-treated RIα-icKO mice. Hearts from RIα-icKO mice were hypercontractile, associated with increased ICa,L, and [Ca2+]i transients and sarcomere shortening in VMs. These latter effects were suppressed by the PKA inhibitor, H89. Global cAMP content was decreased in RIα-icKO hearts whereas local cAMP at the phospholamban/SERCA2 complex was unchanged in RIα-icKO VMs. RIα-icKO VMs had an increased frequency of Ca2+ sparks and pro-arrhythmic Ca2+ waves and RIα-icKO mice had an increased susceptibility to ventricular tachycardia. Upon aging, RIα-icKO mice showed progressive contractile dysfunction, cardiac hypertrophy and fibrosis, culminating in congestive heart failure (HF) with reduced ejection fraction that caused 50% mortality at 1 year.
Conclusions: These results identify RIα as a key negative regulator of cardiac contractile function, arrhythmia and pathological remodelling.