SPARE ALPHA1-ADRENERGIC RECEPTORS AND THE POTENCY OF AGONISTS IN RAT VAS-DEFERENS

Abstract

The existence of spare .alpha.-adrenergic receptors in rat vas deferens was examined directly using radioligand binding assays and contractility measurements. .alpha.-Adrenergic receptors in homogenates of rat vas deferens were labeled with [125I]BE 2254 [2-[.beta.(4-hydroxyphenyl)ethylaminomethyl]tetralone] (125IBE). Norepinephrine and other full .alpha.-adrenergic receptor agonists [epinephrine, phenylephrine, (t)-.alpha.-methylnorephinephrine and methoxamine] were much less potent in inhibiting 125IBE binding than in contracting the vas deferens in vitro. Treatment with 300 nM phenoxybenzamine for 10 min to irreversibly inactivate .alpha.-adrenergic receptors caused a large decrease in the potency of full agonists in causing contraction of this tissue and a 23-48% decrease in the maximal contraction observed. Using those data, equilibrium constants for activation (Kact values) of the receptors by agonists were calculated. These Kact values agreed well with the equilibrium binding constants (KD values) determined from displacement of 125IBE binding. The reduction in .alpha.-adrenergic receptor density following phenoxybenzamine treatment was determined by Scatchard analysis of specific 125IBE binding sites and compared with the expected reduction (q values) calculated from the agonist dose-response curves before and after phenoxybenzamine treatment. Exposure to 300 nM phenoxybenzamine for 10 min resulted in a 39% decrease in specific 125IBE binding sites, which did not agree with the 93% decrease expected from the calculated q values. Treatment of vas deferens with a dose of phenoxybenzamine (10 .mu.M for 15 min) that completely abolished the contractile response to .alpha.-adrenergic agonists caused an 82% decrease in the density of 125IBE binding sites. Tissues exposed to 300 nM phenoxybenzamine in the presence of 100 .mu.M phentolamine or 3 .mu.M prazosin showed no change in the dose-response curves for agonist-induced contraction or in the density of 125IBE binding sites when compared with controls. Phenoxybenzamine apparently functionally inactivates .alpha.-adrenergic receptors at or near the receptor binding site and the potencies of agonists in activating .alpha.-adrenergic receptors in rat vas deferens agree well with their potencies in binding to the receptors. The greater potency of agonists in causing contraction may be due to spare receptors in this tissue. Phenoxybenzamine evidently irreversibly inactivates .alpha.-adrenergic receptors in rat vas deferens, but the decrease in receptor density is much smaller than that predicted from receptor theory.