Demonstration of alpha-adrenoceptors in the rabbit heart by [3H]-dihydroergocryptine binding.

For direct identification of alpha-adrenoceptors in a membrane fraction of the rabbit heart the potent alpha-adrenoceptor antagonist [3H]-dihydroergocryptine ([3H]-DHE) was used. 1. The binding of [3H]-DHE was saturable with 80 fmol of [3H]-DHE bound/mg protein and of high affinity with an equilibrium dissociation constant (KD) of 11.5 nM. Binding of [3H]-DHE (6 nM) was rapid (t 1/2 = 2 min) and readily reversible. From the ratio of the rate constants for forward (K1 = 1.97 X 10(7) M-1 min-1) and reverse (K2 = 0.206 min-1) reactions a KD-value of 10 nM was calculated, which is in good agreement with that obtained by equilibrium studies. 2. Adrenergic agonists compete for [3H]-DHE binding in an order to potency: (-)adrenaline greater than (-)phenyleprine greater than (-)isoprenaline and adrenergic antagonists in the order: phentolamine greater than yohimbine greater than (-)propranolol. Binding is stereospecific as indicated by the greater potency of (-)adrenaline than (+/-)adrenaline in displacing [3H]-DHE from the binding sites. 3. For comparison the binding of the potent beta-adrenoceptor antagonist (-)[3H]-dihydroalprenolol ((-)[3H]-DHA) was measured in the same membrane fraction. The number and affinity of beta-adrenoceptors amounted to 115 fmol of (-)[3H]-DHA bound/mg protein at saturation and KD = 7.9 nM. Adrenergic agonists compete for (-)[3H]-DHA binding in an order of potency: (-)isoprenaline greater than (-)adrenaline greater than (-)phenylephrine; and adrenergic antagonists in the order: (-)prapranolol greater than phentolamine. 4. It is concluded that in a membrane fraction of the rabbit heart there exist binding sites for [3H]-DHE which have characteristics indistinguishable from alpha-adrenoceptors. Thus the present results are in agreement with previously reported data on the existence of cardiac alpha-adrenoceptors in the rabbit heart (Schümann et al., 1974; Endoh et al., 1976b).