Coronary artery vasospasm is a key factor in sudden cardiac death, myocardial infarction and certain other manifestations of ischemic heart disease. The major premise of the proposed project is that alpha and beta adrenergic and serotonergic receptors play a pathogenetic role in coronary vasospasm. In order to test this, in vitro pharmacological methods will be used to systematically analyze the properties of these receptors along the entire lengths of all major epicardial coronary arteries from dog, pig and cow. Human coronary arteries with and without atherosclerotic lesion, obtained at autopsy, will also be analyzed. Vessels will be cleaned, cut into 3 mm rings and mounted in tissue baths for the measurement of isometric tension development. Dose-contractile response curves to alpha adrenergic and serotonergic agonists will be obtained with and without pretreatment with phenoxybenzamine to cause partial receptor inactivation. Dose-relaxation response curves to beta adrenergic agonists will be obtained in carbachol- or histamine-contracted coronary arteries with and without pretreatment with FM24 to cause partial beta receptor inactivation. Dose-response curves will also be obtained in the presence and absence of specific reversible antagonists to alpha and beta adrenergic and serotonergic receptors. The following parameters will be calculated: potency (ED50), agonist (KA) and antagonist (pA2 or KB) dissociation constants and the relationship between fraction of receptors occupied by agonist (RA/Rt) and magnitude of contractile or relaxation response. The agonists and antagonists used will allow differentiation between postsynaptic alpha 1 and alpha 2 and beta 1 and beta 2 adrenoceptors.
The specific aims are to: 1. Determine orders of agonist potency and affinity (1/KA) and antagonist pA2 values at adrenergic and serotonergic receptors. 2. Determine relative receptor density along the length of coronary arteries, based on the relationship between contractile response and fractional receptor occupancy. 3. Determine vasoconstrictor (alpha) and vasodilator (beta) responses to endogenous norepinephrine released by tyramine. 4. Assess the ability of 5-hydroxytryptamine and other vasoconstrictor agents to sensitize coronary arteries to alpha receptor stimulation. 5. Assess adrenergic nerve density and localization by measuring the neuronal uptake of 3H-norepinephrine and by glycoxylic acid histochemical fluorescence of adrenergic nerves. 6. Identify whether activator calcium for contraction comes from extracellular fluid or intracellular stores. 7. Determine if the vascular geometry in the region of an atherosclerotic lesion amplifies the contractile response and leads to a focal spasm. Fulfilling these aims will provide the knowledge necessary to establish the rational basis for drug therapy in the prevention and treatment of coronary vasospastic disorders.
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