Abstract
Background— G-protein–coupled receptor kinase 2 (GRK2) is a primary regulator of β-adrenergic signaling in the heart. G-protein–coupled receptor kinase 2 ablation impedes heart failure development, but elucidation of the cellular mechanisms has not been achieved, and such elucidation is the aim of this study. Methods and Results— Myocyte contractility, Ca 2+ handling and excitation-contraction coupling were studied in isolated cardiomyocytes from wild-type and GRK2 knockout (GRK2KO) mice without (sham) or with myocardial infarction (MI). In cardiac myocytes isolated from unstressed wild-type and GRK2KO hearts, myocyte contractions and Ca 2+ transients were similar, but GRK2KO myocytes had lower sarcoplasmic reticulum (SR) Ca 2+ content because of increased sodium-Ca 2+ exchanger activity and inhibited SR Ca 2+ ATPase by local protein kinase A–mediated activation of phosphodiesterase 4 resulting in hypophosphorylated phospholamban. This Ca 2+ handling phenotype is explained by a higher fractional SR Ca 2+ release induced by increased L-type Ca 2+ channel currents. After β-adrenergic stimulation, GRK2KO myocytes revealed significant increases in contractility and Ca 2+ transients, which were not mediated through cardiac L-type Ca 2+ channels but through an increased SR Ca 2+ . Interestingly, post-MI GRK2KO mice showed better cardiac function than post-MI control mice, which is explained by an improved Ca 2+ handling phenotype. The SR Ca 2+ content was better maintained in post-MI GRK2KO myocytes than in post-MI control myocytes because of better-maintained L-type Ca 2+ channel current density and no increase in sodium-Ca 2+ exchanger in GRK2KO myocytes. An L-type Ca 2+ channel blocker, verapamil, reversed some beneficial effects of GRK2KO. Conclusions— These data argue for novel differential regulation of L-type Ca 2+ channel currents and SR load by GRK2. G-protein–coupled receptor kinase 2 ablation represents a novel beneficial Ca 2+ handling phenotype resisting adverse remodeling after MI.
Highlights
G-protein– coupled receptor kinase 2 (GRK2) is a primary regulator of -adrenergic signaling in the heart
Our current study has revealed that GRK2 can influence myocyte Ca2ϩ homeostasis and that its absence in cardiac myocytes[10,13] causes a novel Ca2ϩ handling phenotype that is resistant to cardiac function deterioration after MI
Loss of GRK2 in cardiac myocytes enhances excitation-contraction coupling (ECC) efficiency in the presence of a lower than normal sarcoplasmic reticulum (SR) Ca2ϩ loading condition and better -adrenergic responsiveness in unstressed hearts. This enhancement of ECC occurs through differential regulation of sarcolemmal versus SR Ca2ϩ handling, with the net result being improved Ca2ϩ transients leading to the amelioration of the heart failure (HF) phenotype
Summary
Conditional mice bearing floxed GRK2 alleles were described previously.[10,13] G-protein– coupled receptor kinase 2 KO (␣ myosin heavy chain Ϫ Cre-recombinase ϫ GRK2flox/flox) and wild-type (WT) (GRK2flox/flox) mice were maintained on a C57BL6 genetic background. G-protein– coupled receptor kinase 2 KO and WT mice were 8 to 10 weeks of age when entering the study. Unstressed normal mice and mice with coronary artery ligation (myocardial infarction [MI]) or sham operation were studied. Myocardial infarction was induced by ligating the left anterior descending coronary artery at 2 to 3 mm below its origin as described previously,[10,14] and animals were studied 28 days post-MI or sham operation. Verapamiltreated mice received oral supplementation of verapamil (Sigma-Aldrich, St. Louis, MO) as described previously[15] and were evaluated by echocardiography. Detailed description of experimental procedures is available in the online-only Data Supplement
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