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NEUROPHARMACOLOGY

Salvinorin A: Allosteric Interactions at the µ-Opioid Receptor

Clinical Psychopharmacology Section, Intramural Research Program, National Institute on Drug Abuse, National Institutes of Health, Department and Health and Human Services, Baltimore, Maryland (R.B.R., D.L.M., H.X., J.A.G., C.M.D., J.S.P.); and Division of Medicinal & Natural Products Chemistry, College of Pharmacy, The University of Iowa, Iowa City, Iowa (K.T., M.S., T.E.P.)

Salvinorin A [(2S,4aR,6aR,7R,9S,10aS,10bR)-9-(acetyloxy)-2-(3-furanyl)-dodecahydro-6a,10b-dimethyl-4,10-dioxo-2h-naphtho[2,1-c]pyran-7-carboxylic acid methyl ester] is a hallucinogenic -opioid receptor agonist that lacks the usual basic nitrogen atom present in other known opioid ligands. Our first published studies indicated that Salvinorin A weakly inhibited µ-receptor binding, and subsequent experiments revealed that Salvinorin A partially inhibited µ-receptor binding. Therefore, we hypothesized that Salvinorin A allosterically modulates µ-receptor binding. To test this hypothesis, we used Chinese hamster ovary cells expressing the cloned human opioid receptor. Salvinorin A partially inhibited [³H]Tyr-D-Ala-Gly-N-Me-Phe-Gly-ol (DAMGO) (0.5, 2.0, and 8.0 nM) binding with E_MAX values of 78.6, 72.1, and 45.7%, respectively, and EC₅₀ values of 955, 1124, and 4527 nM, respectively. Salvinorin A also partially inhibited [³H]diprenorphine (0.02, 0.1, and 0.5 nM) binding with E_MAX values of 86.2, 64, and 33.6%, respectively, and EC₅₀ values of 1231, 866, and 3078 nM, respectively. Saturation binding studies with [³H]DAMGO showed that Salvinorin A (10 and 30 µM) decreased the µ-receptor B_max and increased the K_d in a dose-dependent nonlinear manner. Saturation binding studies with [³H]diprenorphine showed that Salvinorin A (10 and 40 µM) decreased the µ-receptor B_max and increased the K_d in a dose-dependent nonlinear manner. Similar findings were observed in rat brain with [³H]DAMGO. Kinetic experiments demonstrated that Salvinorin A altered the dissociation kinetics of both [³H]DAMGO and [³H]diprenorphine binding to µ receptors. Furthermore, Salvinorin A acted as an uncompetitive inhibitor of DAMGO-stimulated guanosine 5'-O-(3-[³⁵S]thio)-triphosphate binding. Viewed collectively, these data support the hypothesis that Salvinorin A allosterically modulates the µ-opioid receptor.

Address correspondence to: Dr. Richard B. Rothman, Clinical Psychopharmacology Section, 5500 Nathan Shock Drive, Baltimore, MD 21224. E-mail: rrothman{at}mail.nih.gov