CHARACTERIZATION OF [H-3] NALTRINDOLE BINDING TO DELTA-OPIOID RECEPTORS IN MOUSE-BRAIN AND MOUSE VAS-DEFERENS - EVIDENCE FOR DELTA-OPIOID RECEPTOR HETEROGENEITY

Naltrindole (NTI) is a potent and selective nonpeptide delta opioid re ceptor antagonist. This study reports on the binding characteristics o f [H-3]NTI (specific activity = 30.5 Ci/mmole) for mouse brain and vas deferens (MVD) tissues. In brain, [H-3]NTI had unusually high specifi c binding to delta receptors (80% at its Kd concentration) relative to other selective delta receptor radioligands. Saturation Kd values wit h 95% confidence intervals for mouse brain and MVD tissue preparations were 56.2 (41.8-75.7) and 104 (25.8-420) pM, respectively. These Kd v alues were significantly different (P = .028) and [H-3]NTI binding to both tissues was best fit by a one-site model. Receptor densities were 83.9 (66.8-106) fmol/mg of protein for mouse brain and 14.8 (7.03-31. 2) fmol/mg of protein for the MVD. Binding inhibition studies showed t hat NTI and the delta opioid receptor agonists [4'-CI-Phe(4)]DPDPE and [D-Ala(2), Glu(4)]deltorphin had high affinity for the sites labeled by [H-3]NTI in both tissue preparations whereas mu [Tyr-Pro-psi-MePhe- D-Pro-NH2 (PL-17)] and kappa (U-69593) agonists had micromolar affinit y. Both agonists recognized multiple sites in mouse brain under contro l (with 5 mM Mg++) and treatment (with 50 mu M guanylyl-5'-imidodiphos phate and 100 mM NaCl) conditions but only single-site binding was obs erved for MVD (only control condition tested). [D-Ala(2), Glu(4)]delto rphin showed about 6.5-fold selectivity for a portion (approximate to 33%) of mouse brain sites (Ki = 130 pM) compared to sites labeled by [ H-3]NTI in MVD (Ki = 1200 pM) under control conditions. No significant difference was observed for [4'-CI-Phe(4)]DPDPE binding affinity to b oth tissues (Ki = 450-680 pM) under control conditions. The affinity o f opioid agonists, but not antagonists at [H-3]NTI binding sites in mo use brain, was substantially reduced by the presence of guanylyl-5'-im idodiphosphate and sodium ions consistent with guanine nucleotide-bind ing protein regulation of the delta receptors. The portions of high- a nd low-affinity sites recognized by [4'-CI-Phe4]DPDPE and [D-Ala(2), G lu(4)]deltorphin in mouse brain labeled by [H-3]NTI under treatment co nditions were not significantly different (each subtype represented ap proximate to 50% of the total population) suggesting delta receptor he terogeneity in this tissue. It is concluded that [H-3]NTI binds to del ta opioid receptor affinity states and subtypes with equal affinity an d can be used for their characterization in conjunction with different treatment conditions and ligands.