Partial characterization of the specific binding of ³H-N-allylnormetazocine (SKF 10047) to rat brain membranes

The specific binding of ³H(±)-N-allylnormetazocine (³H(±)-SKF 10047) to rat brain membranes was defined as that which could be displaced by 100μM (+)SKF 10047. This binding was found to be saturable and Scatchard analysis indicated the presence of three binding components with differing apparent K(D) and B(max) values. These sites varied considerably in their sensitivity to the effects of heat, trypsin, and ionic strength, with the low affinity site (K(D) = 245 nM) being least sensitive to elevated temperature and trypsinization and most sensitive to elevation of the ionic strength of the incubation buffer. A modest degree of selectivity could be attained for each of the three sites by varying the concentration of ³H(±)-SKF 10047, e.g. at 160 nM ³H(±)-SKF 10047 approximately 75% of the total binding was calculated to be due to the low-affinity site. Using this technique it was found that both isomers of SKF 10047 were equi-effective in displacing binding from the low-affinity site, but (-)SKF 10047 was 150 times more potent than (+)SKF 10047 at the high-affinity site. Also, naloxone was very potent in inhibiting binding to the high-affinity site (IC₅₀ = 4.7 nM), but very weak at the low-affinity site (IC₅₀ = 100 μM). Phencyclidine (PCP) and several of its derivatives were found to have potencies similar to those of SKF 10047 in inhibiting binding to the low-affinity site. These data suggest that the low-affinity binding site for this prototypic sigma opiate may be analogous to the putative PCP receptor.