Spectrum of the mu, delta- and kappa-binding sites in homogenates of rat brain.

1 In homogenates of rat brain, the binding characteristics of tritiated opiates and opioid peptides were examined and the relative capacities of mu-, delta- and kappa-binding sites of the opiate receptor determined by saturation analysis.2 In competition experiments, binding of the selective mu-ligand [(3)H]-[D-Ala(2),MePhe(4),Gly-ol(5)]enkephalin at the mu-site was displaced by [D-Ala(2),D-Leu(5)]enkephalin with rather low affinity (K(I) = 12.6 nM) and more readily by the ketazocine-like compounds (-)-ethylketazocine (K(I) = 3.1 nM) and (-)-bremazocine (K(I) = 0.32 nM), which also displaced the binding of [(3)H]-[D-Ala(2),D-Leu(5)]enkephalin from the delta-site. In contrast, the binding to the kappa-site was easily displaced by ethylketazocine (1.0 nM) and bremazocine (0.37 nM) but not by the mu-ligand [D-Ala(2),MePhe(4),Gly-ol(5)]enkephalin (K(I) = 2000-3000 nM) or the delta-ligand [D-Ala(2),D-Leu(5)]enkephalin (K(I) > 20,000 nM).3 The dissociation equilibrium constant (K(D)) and the binding capacity (pmol/g) of the mu-binding site were determined with the selective mu-ligand [(3)H]-[D-Ala(2),MePhe(4),Gly-ol(5)]enkephalin. For the delta-site, [(3)H]-[D-Ala(2),D-Leu(5)]enkephalin was used in the presence of unlabelled [D-Ala(2),MePhe(4),Gly-ol(5)]enkephalin in order to suppress cross-reactivity to the mu-binding site. For the estimation of kappa-binding, [(3)H]-(+/-)-ethylketazocine or [(3)H]-(-)-bremazocine were used in the presence of unlabelled mu- and delta-ligands for the suppression of cross-reactivities to the mu- and delta-binding sites.4 In rat brain the capacity of the mu-binding site was 7.3 pmol/g brain, that of the delta-binding site 6.7 pmol/g brain and that of the kappa-binding site 2.0 pmol/g brain. Thus, the kappa-binding site had the lowest value whereas in the guinea-pig brain the capacity of the mu-binding site was lower than that of the delta- or kappa-binding site.