ALPHA(2)-ADRENOCEPTOR AND CATECHOLAMINE-INSENSITIVE BINDING-SITES FOR [H-3] RILMENIDINE IN MEMBRANES FROM RAT CEREBRAL-CORTEX

The kinetic and pharmacological characteristics of the binding of the oxazoline antihypertensive drug, [H-3]rilmenidine, to membranes of rat cerebral cortex have been determined. Computerised resolution of curvi-linear, equilibrium binding isotherms was consistent with the existence of two distinct binding sites for [H-3]rilmenidine: K(d) 17.3 +/- 7.41 nM, B(max) 0.197 +/- 0.06 pmol/mg protein and K(d) 254 +/- 48 nM. B(max) 1.59 +/- 0.08 pmol/mg protein. Moreover, the resolution of two association and dissociation rates also suggested the existence of two binding site populations. Drug inhibition studies revealed that specific binding of [H-3]rilmenidine (2 nM) was only inhibited by a maximum of 50% by the catecholamines, adrenaline and noradrenaline, but was completely inhibited by some oxazolines, by guanabenz (a guanidino drug) and by several imidazoline compounds including naphazoline, oxymetazoline and clonidine. Binding isotherms for these drugs were also best fit by a two-site model. The relative K(i) values at the high affinity site for [H-3]rilmenidine and the number of these high affinity sites are consistent with this site being an alpha-2-adrenoceptor. The high affinity of oxymetazoline and low affinity of prazosin for high affinity [H-3]rilmenidine binding sites together with the rank order of potency of oxymetazoline > phentolamine > SKF 104078 > ARC-239 > prazosin suggest that [H-3]rilmenidine binds to the alpha-2A sub-type of adrenoceptor. Computer-resolved K(i) values for drugs at the larger number of lower affinity binding sites were very similar to K(i) values determined in the presence of 10-mu-M adrenaline (used to block alpha-2-adrenoceptor binding). The catecholamine-insensitive binding site did not share thc pharmacological characteristics of previously described, high affinity imidazoline-guanidinium receptive sites or high affinity imidazole sites, but more closely resembles the so-called 'idazoxan receptor'.