The force of contraction of the heart is controlled by neurotransmitters and other chemicals. This control is initiated at the level of specific extracellular receptors that regulate the intracellular levels of cyclic AMP and Ca++. Catecholamines increase contractile force through a mechanism involving increases in intracellular cAMP and CA++, while acetylcholine and adenosine decrease contractile force through mechanisms believed to involve decreases in intracellular cAMP and Ca++. Our overall aim is to obtain a clear and complete biochemical and pharmacological analysis of Beta-adrenergic, muscarinic cholinergic and adenosine receptors throughout cardiac development, from the embryonic to adult stages. For each receptor we will determine receptor number and affinity for agonist and antagonist ligands using radioligand binding studies in preparations of embryonic, newborn, young adult and adult chicken hearts. We will determine the molecular basis of developmentally related changes in receptor number and/or affinity. The mechanisms responsible for """"""""endogenous desensitization"""""""" of receptors that occurs at the onset of cholinergic and adrenergic neurotransmission will be probed. In addition, the role of endogenous adenosine in regulating the activity of the adenosine receptor during cardiac development will be investigated. The number and affinity of Ca++ antagonist binding sites will be determined in immature and mature hearts. Furthermore, the interaction of Ca++ antagonists with muscarinic, adenosine and Beta-adrenergic receptors in the developing heart will be assessed. The efficiency of receptor: Effector coupling for muscarinic, adenosine, and Beta-adrenergic receptors will be determined at all developmental stages. These studies will quantitate the relationship between receptor occupancy and the abilities of the receptors to modulate the activity of adenylate cyclase. The effect of simultaneous occupancy of another receptor on this relationship will also be probed in order to determine the nature of receptor interactions. Similar studies will be performed to relate receptor occupancy to effects on contractile force. Finally, studies will be performed to assess basal and drug-induced efflux of adenosine from preparations of immature and mature hearts in order to gain an understanding of the role of this important modulator during cardiac development. The proposed studies will significantly contribute to our understanding of the molecular basis of action of acetylcholine, adenosine and catecholamines on developing cardiac tissue.
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