Demonstration of α-adrenoceptors in the rabbit bladder base and urethra with 3H-dihydroergocryptine ligand binding

The purpose of this work was to demonstrate the presence of α-adrenoceptors in a crude membrane preparation made from rabbit bladder base and urethra. This was achieved by radioligand binding studies, using 3H-dihydro-α-ergocryptine (3H-DHE) as the radioligand. The specific binding, i.e. the binding that could be inhibited by 10-5 M phentolamine, was saturable with 73 fmol 3H-DHE bound per mg membrane protein. Binding was at steady state after 60 min., and reversible. Rate constants for association and dissocation were 3x107 M-1 min.-1, and 2x10-2 min.-1, respectively. A number of compounds were tested for their abilities to complete with 3H-DHE for the binding sites. The relative affinity of some adrenoceptor agonists was (-)-adrenaline>(-)-noradrenaline much larger than (+-)-isoprenaline. Steroselectivity was shown, since (-)-noradrenaline had 42 times higher affinity than (+)-noradrenaline. Adrenoceptor antagonists inhibited 3H-DHE binding in the following order of potency: DHE>phentolamine much larger than (+-)-propranolol. The dissociation constant, Ksub(D), for DHE to the binding sites was estimated in three different ways. The constants were derived from saturation, competition, and kinetic studies, and gave Ksub(D) values of 1.1,1.4 and 0.7 nM, respectively. The results suggest that α-adrenoceptors were labelled by 3H-DHE in the tissue homogenates. (author)