PROBING THE OPIOID RECEPTOR COMPLEX WITH (+)-TRANSSUPERFIT .2. EVIDENCE THAT MU-LIGANDS ARE NONCOMPETITIVE INHIBITORS OF THE DELTA-CX OPIOID PEPTIDE BINDING-SITE

Previous studies delineated two classes of delta binding sites; a delta binding site not associated with the opioid receptor complex, termed the delta(ncx) site, and a delta site associated with the opioid receptor complex, termed the delta(cx) site. The delta(ncx), site has high affinity for [D-Pen2,D-Pen5]enkephalin, and is synonymous with what is now identified as the delta1 binding site. Pretreatment of membranes with the delta-selective acylating agents FIT. or (+)-trans-SUPERFIT, deplete membranes of the delta(ncx) binding site, which permits the selective labeling of the delta(cx) binding site with [H-3][D-Ala2,Leu5]lenkephalin. The present study compared the properties of the delta(cx) binding site present in brain membranes pretreated with (+)-trans-SUPERFIT with the properties of the delta(cx) site present in untreated membranes. The major findings are: 1) pretreatment of membranes with (+)-trans-SUPERFIT decreased the IC50 values of delta-preferring drugs, and increased the lC50 values of mu-preferring drugs, for the delta(cx) binding site; 2) the degree of delta selectivity was highly correlated with the magnitude of the (+)-trans-SUPERFIT-induced shift in the IC50 values; 3) the ligand-selectivity patterns of the mu and delta(cx) sites present in (+)-trans-SUPERFIT-pretreated membranes were poorly correlated; 4) whereas mu-preferring drugs were noncompetitive inhibitors of [H-3][D-Ala2,Leu5]enkephalin binding to the delta(cx) site, delta-preferring drugs were competitive inhibitors. Viewed collectively, these data support the hypothesis that the mu and delta(cx) binding sites are distinct, provide additional evidence for delta receptor heterogeneity, and suggest that (+)-trans-SUPERFIT-pretreated membranes will provide a useful preparation for studying the delta(cx) binding site.