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Opioid receptors (ORs) convert extracellular messages to signaling events by coupling to the heterotrimeric G proteins, G α • βγ Classic pharmacological methods, such as [ 35 S]GTP γ S binding and inhibition of cyclic AMP production, allow for general opioid characterization, but they are subject to the varying endogenous G α proteins in a given cell type. Bioluminescence resonance energy transfer (BRET) technology offers new insight by allowing the direct observation of G α subunit-specific effects on opioid pharmacology. Using a Venus-tagged G βγ and nanoluciferase-tagged truncated G protein receptor kinase 3, an increase in BRET signal correlated with OR activation mediated by a specific G α protein. The magnitude of the BRET signal was normalized to the maximum response obtained with 10 µM 2-(3,4-dichlorophenyl)- N -methyl- N -[(1 R ,2 R )-2-pyrrolidin-1-ylcyclohexyl]acetamide (U50,488) for the kappa OR (KOR). Opioids reached equilibrium with the KOR, and concentration-response curves were generated. Although the full agonists U50,488, salvinorin A, nalfurafine, and dynorphin peptides were equally efficacious regardless of the G α subunit present, the concentration-response curves were leftward shifted when the KOR was signaling through G α z compared with other G α i/o subunits. In contrast, the G α subunit distinctly affected both the efficacy and potency of partial kappa agonists, such as the benzomorphans, and the classic mu opioid antagonists, naloxone, naltrexone, and nalmefene. For example, (-)pentazocine had EC 50 values of 7.3 and 110 nM and maximal stimulation values of 79% and 35% when the KOR signaled through G α z and G α i1, respectively. Together, these observations suggest KOR pharmacology varies based on the specific G α subunit coupled to the KOR. SIGNIFICANCE STATEMENT: Opioid receptors couple to various heterotrimeric Gαβγ proteins to convert extracellular cues to precise intracellular events. This paper focuses on how the various inhibitory Gα subunits influence the pharmacology of full and partial agonists at the kappa opioid receptor. Using a bioluminescent assay, the efficacy and potency of kappa opioids was determined. Opioid signaling was more potent through Gαz compared with other Gα proteins. These observations suggest that Gαz may impact opioid pharmacology and cellular physiology more than previously thought., (Copyright © 2020 by The American Society for Pharmacology and Experimental Therapeutics.)

A combination of buprenorphine (BUP) and samidorphan (SAM) at a 1:1 (mg/mg) fixed-ratio dose is being investigated as an adjunctive treatment of major depressive disorder (BUP/SAM, ALKS 5461). Both [ 3 H]BUP and [ 3 H]SAM bound to the μ -, κ -, and δ -opioid receptors (MOR, KOR, and DOR, respectively) with K d values of 3 nM or less. [ 3 H]BUP dissociated from the MOR more slowly than [ 3 H]SAM did. In the [ 35 S]GTP γ S assay, BUP was a partial agonist at the MOR, KOR, and DOR. SAM was an antagonist at the MOR and a partial agonist at the KOR and DOR. The pharmacology of the combination of SAM and BUP was characterized at ratios like the molar ratios of both compounds at steady state in humans. In all assessments, SAM reduced the efficacy of BUP at the MOR without altering its potency. At the KOR, SAM had no significant effect on the activity of BUP. In bioluminescent resonance energy transfer assays, SAM, naltrexone, and naloxone were partial agonists when the MOR was coupled to the G α oB and G α z , and were antagonists when coupled to G α i At the KOR, SAM was a partial agonist activating G α oA and G α oB and a full agonist in stimulating G α z SAM inhibited BUP's recruitment of β -arrestin to the MOR, suggesting an attenuation of BUP's efficacy in activating G proteins correlated with an inhibition of β -arrestin recruitment. The collective data suggest that SAM attenuates the efficacy of BUP under all conditions tested at the MOR and DOR but had little effect on BUP activity at the KOR., (Copyright © 2018 by The American Society for Pharmacology and Experimental Therapeutics.)

A series of levo- and dextromorphinan pairs have been synthesized and evaluated for their affinities to the mu, kappa, and delta opioid receptors, the N-methyl-D-aspartate (NMDA) channel, and sigma 1 and 2 receptors. It was found that levo isomers tended to have higher affinities at the opioid receptors and moderate to high affinities to the NMDA and sigma receptors, while dextro isomers tended to have lower affinities to the opioid receptors but comparatively higher affinities to the NMDA and sigma receptors. This series of compounds have interesting and complex pharmacological profiles, and merit further investigation as potential therapies for drug abuse treatment.

Based on a renewed importance recently attributed to bi- or multifunctional opioids, we report the synthesis and pharmacological evaluation of some analogues derived from our lead μ agonist / δ antagonist, H-Dmt-Tic-Gly-NH-Bzl. Our previous studies focused on the importance of the C-teminal benzyl function in the induction of such bifunctional activity. The introduction of some substituents in the para position of the phenyl ring (-Cl, -CH(3), partially -NO(2), inactive -NH(2)) was found to give a more potent μ agonist / antagonist effect associated with a relatively unmodified δ antagonist activity (pA(2) = 8.28-9.02). Increasing the steric hindrance of the benzyl group (using diphenylmethyl and tetrahydroisoquinoline functionalities) substantially maintained the μ agonist and δ antagonist activities of the lead compound. Finally and quite unexpectedly D-Tic2, considered as a wrong opioid message now; inserted into the reference compound in lieu of L-Tic, provided a μ agonist / δ agonist better than our reference ligand (H-Dmt-Tic-Gly-NH-Ph) and was endowed with the same pharmacological profile.