). The applicant proposes to characterize coronary adenosine receptor(s) and their signal transduction pathways as well as the molecular biology due to the pharmacological difference from the well known adenosine receptors from other tissues and species. There is evidence suggesting the presence of these receptors in coronary smooth muscle and endothelium. These are both the inhibitory (A1) and stimulatory (A2) adenosine receptors. Endothelium may be partly responsible to modulate the action of adenosine through the release of nitric oxide perhaps via stimulation of A2 like receptor. Therefore, the central hypothesis is that the modulation of coronary tone by adenosine is determined by a net effect of these receptors (A1, A2 and/or A2b). Binding of adenosine to these receptors (including endothelium) may involve various effector systems (e.g. adenylate and guanylate cyclases, PKC, PLC) through the activation of different G proteins leading to the changes in Ca++ to modulate the vascular tone. The applicant proposes to study the: (a) further identification and characterization of adenosine receptors (A1,A2) from coronary smooth muscle using organ bath, binding and photoaffinity labeling studies; (b) regulation of vascular smooth muscle adenosine receptor(s) in organ baths by incubating with various relatively selective A1 and A2 agonists and antagonists and measure the activity and levels of G-protein(s), cAMP, binding and Northerns; (c)mechanism(s) of up-regulation of PKC and its isoforms by the activation of A1 vascular smooth muscle adenosine receptor; (d) identification and characterization of coronary endothelial adenosine receptor using binding and photoaffinity labeling studies and measure the release of nitric oxide; (e) possible coupling of coronary endothelial adenosine receptor(s) to G-protein(s) using toxins and to identify the effector systems; and finally (f) clone, sequence, express and conduct functional assays of the pharmacologically different A2 adenosine receptor(s) (possibly A4) from coronary smooth muscle.
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