The role of adrenergic receptor mechanisms in understanding the effects of catecholamines on the mammalian myocardium is critical to predicting the physiologic response to drugs in the management of myocardial and other disease. Specifically, the role of alpha-adrenergic receptors in inotropic mechanisms in mammalian heart is poorly understood. I propose to study the pharmacology of alpha-adrenergic receptor activation using a highly purified preparation of intact, hormonally responsive ventricular cardiomyocytes. I will employ radioligand binding techniques to identify and characterize the alpha receptor on the intact cardiomyocytes and to detail its possible interactions with regulatory proteins involved in regualtion of adenylate cyclase. The presence of alpha- and beta-adrenergic receptors on the same cell will allow direct physiologic assessment of the interactions of these receptors in the modulation of adenylate cyclase activity and its consequences in these purified cells. By employing alpha and beta subtype selective agonists and antagonists, I propose to show that the effects of guanine nucleotides on alpha-adrenergic receptor affinity for agonists reflect the capacity of alpha receptors to directly modulate the synthesis of cyclic AMP in respoonse to beta-adrenergic receptor stimulation. This investigation will profit from my previous work on the hormonally specific compartmentation of cyclic AMP action in the cardiomyocyte which provides a specific framework in which to study alpha receptor effects, both at the level of intermediate signal transduction following receptor-agonist interactions and with respect to functional substrates (enzyme activity, phosphoprotein formulation) of the consequence of cyclic AMP elevation within the particulate fraction of the myocyte. The role of receptor coupling proteins and other receptor mechanisms (e.g., cholinergic muscarinic) working in concert to modulate the activity of adenylate cyclase in heart can be directly assessed in the purified myocyte without interference from different cell types present in other models of heart. The myocyte preparation technique also lends itself to studies of myocardial disease where receptors and their effects may be altered. Ischemia can be modeled easily in these myocytes, and cells prepared from diabetic animals and animals with cardiac failure will permit a direct assessment of the effects of disease on alpha-adrenergic mechanisms in the heart.
