Specific Binding of β-Adrenergic Catecholamines to a Subcellular Fraction from Cardiac Muscle

Abstract

Abstract The 78,000 x g subcellular fraction of canine cardiac tissue has been shown to bind norepinephrine specifically. β-Adrenergic drugs compete with norepinephrine in a potency series parallel to their potency in causing an increase in the force of cardiac muscle contraction. d and l isomers of the catecholamines competed equally well. These subcellular particles also contain adenylate cyclase which may be stimulated by catecholamines with a similar specificity. [3H]Norepinephrine is concentrated 1000-fold in the particles at 10-9m and may be fully dissociated in 1 m HC1 without evidence of structural alteration. Two different binding constants, K = 1.04 x 107 m-1 and 1.33 x 106 m-1, as well as two different dissociation rates for norepinephrine, were observed. The pH optimum for binding was 7.5, with substantial decrease at more acidic pH and little change at more alkaline pH. Divalent cations at 1 to 5 mm diminished binding as did chelating agents, reagents reacting with sulfhydryl groups, urea, temperatures above 50°, and proteolytic enzymes. The monovalent cations, Na+ and K+, and the enzymes ribonuclease, desoxyribonuclease, and phospholipase were without effect. Tris and trimethylamine and increases in temperature up to 50° enhanced binding. Binding of norepinephrine to this subcellular fraction was differentiated from binding to neural vesicles by virtue of a different specificity, a lack of sensitivity to reserpine, the absence of an ATP requirement, and the preservation of norepinephrine binding after chemical sympathectomy.