TY - JOUR T1 - Agonist-, Antagonist-, and Inverse Agonist-Regulated Trafficking of the δ-Opioid Receptor Correlates with, but Does Not Require, G Protein Activation JF - Journal of Pharmacology and Experimental Therapeutics JO - J Pharmacol Exp Ther SP - 1015 LP - 1020 VL - 298 IS - 3 AU - Paulette A. Zaki AU - Duane E. Keith, Jr. AU - James B. Thomas AU - F. I. Carroll AU - Christopher J. Evans Y1 - 2001/09/01 UR - http://jpet.aspetjournals.org/content/298/3/1015.abstract N2 - In this study, we explored the relationship between ligand-induced regulation of surface δ opioid receptors and G protein activation. G protein activation was assessed with [35S]guanosine-5′-O-(3-thio)triphosphate (GTPγS) binding assays conducted at both 37 and 0°C. Ligand-independent (constitutive) activity of the δ-receptor was readily observed when the [35S]GTPγS binding assay was performed at 37°C. We identified a new class of alkaloid inverse agonists (RTI-5989-1, RTI-5989-23, RTI-5989-25), which are more potent than the previously described peptide inverse agonist ICI-174864 (N,N-diallyl-Tyr-Aib-Aib-Phe-Leu). Treatment with these inverse agonists for 18 h caused up-regulation of surface receptors. Eighteen-hour treatment with etorphine resulted in approximately 90% loss of surface receptor, whereas fentanyl, diprenorphine, and morphine caused between 20 and 50% loss. The abilities of ligands to modulate [35S]GTPγS binding at 37°C showed a strong correlation with their abilities to regulate surface receptor number (r2 = 0.86). Interestingly, the ability of fentanyl to activate G proteins was markedly temperature sensitive. Fentanyl showed no stimulation of [35S]GTPγS binding at 0°C but was as efficacious as etorphine, morphine, and diprenorphine at 37°C. Neither the ligand-induced receptor increases nor decreases were perturbed by pertussis toxin pretreatment, suggesting that functional G proteins are not required for ligand-regulated δ-opioid receptor trafficking. The American Society for Pharmacology and Experimental Therapeutics ER -