The Inotropic Peptide βARKct Improves βAR Responsiveness in Normal and Failing Cardiomyocytes Through Gβγ-Mediated L-Type Calcium Current Disinhibition

Rationale: The G(beta gamma)-sequestering peptide beta-adrenergic receptor kinase(beta ARK)ct derived from the G-protein-coupled receptor kinase (GRK)2 carboxyl terminus has emerged as a promising target for gene-based heart failure therapy. Enhanced downstream cAMP signaling has been proposed as the underlying mechanism for increased beta-adrenergic receptor (beta AR) responsiveness. However, molecular targets mediating improved cardiac contractile performance by beta ARKct and its impact on G(beta gamma)-mediated signaling have yet to be fully elucidated. Objective: We sought to identify G(beta gamma)-regulated targets and signaling mechanisms conveying beta ARKct-mediated enhanced beta AR responsiveness in normal (NC) and failing (FC) adult rat ventricular cardiomyocytes. Methods and Results: Assessing viral-based beta ARKct gene delivery with electrophysiological techniques, analysis of contractile performance, subcellular Ca2+ handling, and site-specific protein phosphorylation, we demonstrate that beta ARKct enhances the cardiac L-type Ca2+ channel (LCC) current (I-Ca) both in NCs and FCs on beta AR stimulation. Mechanistically, beta ARKct augments I-Ca by preventing enhanced inhibitory interaction between the alpha 1-LCC subunit (Cav1.2 alpha) and liberated G(beta gamma) subunits downstream of activated beta ARs. Despite improved beta AR contractile responsiveness, beta ARKct neither increased nor restored cAMP-dependent protein kinase (PKA) and calmodulin-dependent kinase II signaling including unchanged protein kinase (PK)C epsilon, extracellular signal-regulated kinase (ERK)1/2, Akt, ERK5, and p38 activation both in NCs and FCs. Accordingly, although beta ARKct significantly increases I-Ca and Ca2+ transients, being susceptible to suppression by recombinant G(beta)gamma protein and use-dependent LCC blocker, beta ARKct-expressing cardiomyocytes exhibit equal basal and beta AR-stimulated sarcoplasmic reticulum Ca2+ load, spontaneous diastolic Ca2+ leakage, and survival rates and were less susceptible to field-stimulated Ca2+ waves compared with controls. Conclusion: Our study identifies a G(beta gamma)-dependent signaling pathway attenuating cardiomyocyte I-Ca on beta AR as molecular target for the G(beta gamma)-sequestering peptide beta ARKct. Targeted interruption of this inhibitory signaling pathway by beta ARKct confers improved beta AR contractile responsiveness through increased I-Ca without enhancing regular or restoring abnormal cAMP-signaling. beta ARKct-mediated improvement of I-Ca rendered cardiomyocytes neither susceptible to beta AR-induced damage nor arrhythmogenic sarcoplasmic reticulum Ca2+ leakage. (Circ Res. 2011;108:27-39.)