There is increasing evidence that elevated levels of the beta-adrenergic receptor kinase1 (betaARK1) contribute to impaired catecholamine responsiveness observed in heart failure. Using a gene targeted mouse model of heart failure (MLP-/-), we have recently shown that reversal of betaAR desensitization achieved through betaARK1 inhibition, could lead to a marked improvement in cardiac function. The overall hypothesis of this proposal is that increased beta1AR phosphorylation, as a consequence of elevated betaARK1 levels, not only contributes to, but is a primary cause of heart failure. The following specific aims are proposed 1) To determine the role beta1AR phosphorylation and betaARK1 play in the development of the failing heart and whether high levels of circulating catecholamines are required for the pathogenesis of heart failure; MLP(-/-) mice will be crossed with betaARK1 heterozygous knockout mice and dopamine-beta-hydroxylase knockout mice. 2) To determine whether the salutary effects of betaARK1 inhibition represent a ubiquitous pathologic mechanism; another model of severe cardiac dysfunction and premature death achieved through cardiac targeted overexpression of calsequestrin will be crossed into transgenic mice overexpressing the betaARK inhibitor 3) To determine whether betaARK1 phosphorylation of the beta1AR is a mechanism for the pathogenesis of the failing heart, the carboxyl terminal phosphorylation sites for betaARK1 will be mutated by site directed mutagenesis and tested both in vitro and in vivo for the capacity to prevent phosphorylation and desensitization of beta1AR's 4) To determine whether beta1AR desensitization by betaARK1 is a primary pathologic mechanism for the development of heart failure; beta1AR phosphorylation mutant mice will be crossed into the MLP (-/-) heart failure mouse. Using an integrative approach with novel gene targeting strategies and in-depth physiological analysis of gene targeted mice, we will directly test the hypothesis that desensitization of beta1AR's, mediated through heightened betaARK1 levels, play a causative role in the pathogenesis of the failing heart. The collaborations with the other R01's should provide a unique opportunity to understand the relationships between betaAR signaling, E-C coupling and myocyte cell survival.
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