TRANSGENIC MANIPULATION OF BETA-ARK1 MODIFIES BETA1-AR RESPONSIVENESS

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Agonist-induced phosphorylation of beta-adrenergic receptors (beta-AR's) by the beta-adrenergic receptor kinase 1 (betaARK1) in cell suspensions can result in a rapid disruption of the normal coupling between receptors and the stimulatory guanine nucleotide-binding protein Gs. To determine the direct functional significance of betaARK1 on myocardial performance, we measured the contractile responses to acute beta1-AR stimulation in left ventricular myocytes isolated from control and transgenic mice overexpressing either betaARK1 (TGbetaK12) or a betaARK1 inhibitor (TGMini27). Contractile response to five concentrations (.01 to .10 _M) of the beta1-AR agonist norepinephrine (NE) plus prazosin (1 _M) was measured after a 60 second rest, i.e. the rested state contraction (RSC), and during steady state (SSC) stimulation at 0.5 Hz (23_). At baseline, there were no significant differences in RSC or SSC between control, TGbK12 or TGMini27 mice. On the other hand, both the dose-response and kinetics for the NE-induced SSC/RSC response were significantly different among experimental groups (p lesser than 0.001). Specifically, the maximal contractile response induced by NE in myocytes isolated from the TGbetaK12 group was only 70% of the response observed in control cells, and 50% of the response measured in TGMini27 cells. The evolution of the NE contractile response in TGMini27 cells was 37% and 67% faster vs control and TGbK12 cells, respectively (p lesser than 0.03). Thus, substantial functional betaARK1 modulation of beta1-AR signalling occurs in cardiac myocytes, even during short term 1-AR stimulation. These results are consistent with a role for agonist-induced phosphorylation and desensitization of cardiac beta1-AR's by betaARK1 in single myocytes, and highlight the potential role of betaARK1 as an important determinant of the cardiac beta1-AR contractile response. The findings of this study have particular clinical relevance as they provide further support for the use of betaARK1 inhibition as a potential therapeutic approach to improve cardiac function in those humans with chronic heart failure, where betaARK1 levels are known to be elevated.