Binding of 3H-clonidine to an alpha-adrenoceptor in membranes of guinea-pig ileum.

Abstract

The binding of 3H-clonidine to membrane particles from guinea-pig ileum was investigated. The specific binding, i.e. the binding that could be inhibited by high concentrations of unlabeled clonidine or noradrenaline, was of high affinity, KD3 nM. The number of sites was approximately 25 fmol/mg protein. Rate constants of association and dissociation were 5.3×107 M−1 min−1 and 0.18 min−1, respectively. Affinites of various drugs to the binding site were determined by measuring their effect on the binding of 3H-clonidine. The affinity of adrenergic agonists decreased in the order clonidine = tramazoline > (−)-erythro-α-methylnoradrenaline > (−)-noradrenaline ≫ (−)-phenylephrine. (−)-Noradrenaline had about 20 times more affinity than the (+)-isomer. The affinity of α-adrenoceptor antagonists decreased in the order phentolamine > rauwolscine = yohimbine > WB 4101 > pseudoyohimbine > prazosin = corynanthine. Yohimbine and rauwolscine had about 100 times more affinity than their stereoisomer corynanthine. Serotonin 10 μM and metiamide 10 μM did not affect the binding, and propranolol inhibited it only at high concentrations. — The results indicate that 3H-clonidine labels an β2-adrenoceptor in guinea-pig ileum. The orders of affinity of α-adrenoceptor agonists and antagonists agree well with their orders of potency in functional tests, namely as modulators of cholinergic transmission in the guinea-pig ileum and as modulators of noradrenaline release in the rabbit pulmonary artery. An α-adrenoceptor should be classified as α2 when the affinities of clonidine, tramazoline and α-methylnoradrenaline greatly exceed the affinity of phenylephrine, and when the affinities of rauwolscine and yohimbine exceed those of prazosin and corynanthine.