CARDIAC A1-ADENOSINE RECEPTOR RESERVE

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DESCRIPTION: (Adapted from the Applicant's Abstract) Recent studies in the applicants' laboratory revealed that the potency of adenosine to inhibit isoproterenol-stimulated L-type calcium inward current (beta-ICa,L) of atrial myocytes is 12-fold higher than to activate the inwardly rectifying K+ current (IKAdo). The studies proposed in this application are designed to test the following hypotheses related to the differential potency of adenosine to inhibit beta-ICa,L and to activate IKAdo in atrial myocytes: Aim #1 - distinct A1 receptor subtypes (A1a and A1b) subserve inhibition of beta-ICa,L and activation of IKAdo; Aim #2 - a single receptor is coupled to both responses, but there is a greater receptor reserve for inhibition of beta-ICa,L than for activation of IKAdo; Aim #3 - different A1AdoR agonists will have different magnitudes of receptor reserve. The importance of receptor reserve as a determinant of cardiomyocyte responsiveness to A1AdoR agonists will be resolved. A newly synthesized irreversible A1AdoR antagonist will be used to inactivate A1AdoRs for the analysis of receptor reserve. Pharmacological methods will be used to define A1AdoR subtypes and to determine the receptor occupancy-response relationships for both A1AdoR-mediated responses (activation of IKAdo/inhibition of beta-Ica,L) in atrial myocytes and for the direct shortening of the atrial monophasic action potential (MAP) caused by various A1AdoR agonists. Studies will be carried out with guinea pig freshly isolated atrial myocytes and isolated perfused hearts. Activation of IKAdo and inhibition of beta-ICa,L by adenosine and A1AdoR agonists will be recorded by use of the whole cell patch-clamp technique. Shortening of the atrial MAP caused by adenosine and by A1AdoR agonists will be quantitated by standard electrophysiological methods. Although the main focus of this project is on receptor reserve, they will also investigate (Aim #4) whether the higher potency and potentially greater reserve for the anti beta-adrenergic action may confer tonic inhibition by endogenous adenosine of sympathetic activation of the heart in the anesthetized guinea pig. The hypothesis that endogenous adenosine exerts tonic inhibition of the cardiostimulatory effects of increased sympathetic neural activity will be tested by measuring the magnitude of reflex tachycardia caused by nitroprusside-induced hypotension, in the absence and presence of A1AdoR antagonists and of an allosteric enhancer of agonist binding to the A1AdoR. The studies will have important clinical implications for the design and use of A1AdoR agonists in the treatment of cardiac disease, and for the role for adenosine in regulation of cardiac function. The long-term goal of the studies is to understand mechanisms by which organ and/or response selectivity to A1AdoR agonists is achievable.