α2-adrenoreceptors on human platelets: Selective labelling by [3H]clonidine and [3H]yohimbine and competetive inhibition by antidepressant drugs

Abstract

[3H]Clonidine and [3H]yohimbine were used to characterize .alpha.2-adrenoreceptors on human platelets. At 25.degree. C binding was rapid (t1/2 [half-life] of association, 1.8 and 2.7 min) and reversible (t1/2 of dissociation, 0.5 and 8.2 min). The binding sites for [3H]clonidine and [3H]yohimbine showed the specificity required for an .alpha.2-adrenoceptor. The rank order of potency of inhibitors of [3H]clonidine binding was clonidine > yohimbine .mchgt. phenylephrine > prazosin and of [3H]yohimbine binding was yohimbine > clonidine .mchgt. phenylephrine > prazosin Scatchard analysis of [3H]yohimbine binding indicated the existence of a single population of noninteracting sites (Kd = 3.0 nM; Bmax [maximum binding] = 188 fmol[femtomole]/mg protein). The high-affinity binding of [3H]clonidine had a lower affinity and a lower number of sites (Kd = 5.0 nM; Bmax = 35 fmol/mg protein). [3H]Clonidine binding also showed evidence of a 2nd site of much lower affinity and greater number (Kd = 18.6 nM; Bmax = 77 fmol/mg protein) in 40% of the normal population. In vitro, antidepressant drugs competed with [3H]clonidine and [3H]yohimbine for the platelet .alpha.2-adrenoreceptor. The rank order of potency of inhibitors of [3H]clonidine binding was mianserin > amitriptyline .mchgt. iprindole > desipramine and of [3H]yohimbine binding was mianserin > amitriptyline .mchgt. desipramine > iprindole. The inhibition constants (Ki) of adrenergic drugs and of various antidepressant drugs in competing with [3H]clonidine were correlated with the inhibition constants of these drugs in competing with [3H]yohimbine (r = 0.970; P < 0.001) which suggests that both radioligands labeled the same .alpha.2-adrenoreceptor on the human platelet. The inhibition of binding induced by all antidepressant drugs was competitive. In contrast, long-term administration of tricyclic antidepressant drugs to patients is associated with a decrease in the number of binding sites for [3H]clonidine on platelet membranes. Both [3H]clonidine and [3H]yohimbine are useful tools for the quantification of .alpha.2-adrenoreceptors on blood platelets and suggests that the specific binding of radiolabeled .alpha.2-adrenoreceptor ligands to human platelet membranes might be used to monitor changes in .alpha.2-adrenoreceptors during tricyclic antidepressant drug treatment.