title: Pharmacological characterization of AR-M1000390 at human delta opioid receptors.
abstract: We investigated the pharmacological properties of a newly synthesised delta agonist AR-M1000390, derived from SNC-80 ((+)-4-[(alpha R)-alpha-((2S,5R)-4-allyl-2,5-dimethyl-1-piperazinyl)-3-methoxybenzyl]-N,N-diethyl-benzamide), in the neuroblastoma cell line SK-N-BE expressing only human delta-opioid receptors. Binding and functional experiments showed a weak affinity (K(i) = 106 +/- 34 nM) correlated with a weak potency (EC(50) = 111 +/- 31 nM) to inhibit the forskolin-stimulated cAMP accumulation. Sustained activation of opioid receptors in the presence of the maximal inhibitory concentration of AR-M1000390 produced a rapid and strong desensitization. In order to examine the contribution of internalization and down-regulation in the desensitization processes, binding and functional experiments were conducted in the presence or in the absence of hypertonic sucrose solution to block clathrin-dependent opioid receptor endocytosis. We observed both the inability of AR-M1000390 to down-regulate opioid receptors and the absence of any effect of sucrose on desensitization. The lack of delta-opioid receptor internalization by AR-M1000390 was further corroborated by confocal microscopy using antibodies directed either against the endogenous delta-opioid receptors or the FLAG-tagged delta-opioid receptors stably expressed in the SK-N-BE cells. These data suggest that uncoupling rather than internalization is responsible for delta-opioid receptors desensitization by AR-M1000390.
authors.affiliation: Laboratoire de Pharmacologie moléculaire de la tolérance aux opiacés, Université de Caen, Caen, France.
authors.forename: Nicolas
authors.initials: N
authors.lastname: Marie
authors.forename: Gérard
authors.initials: G
authors.lastname: Landemore
authors.forename: Claire
authors.initials: C
authors.lastname: Debout
authors.forename: Philippe
authors.initials: P
authors.lastname: Jauzac
authors.forename: Stéphane
authors.initials: S
authors.lastname: Allouche
mesh_terms: D000262:Adenylyl Cyclases / Q000378:metabolism; D001549:Benzamides / Q000494:pharmacology*; D015536:Down-Regulation; D004705:Endocytosis / Q000187:drug effects; D006801:Humans; D018613:Microscopy, Confocal; D009292:Narcotic Antagonists / Q000494:pharmacology*; D009447:Neuroblastoma / Q000378:metabolism / Q000473:pathology; D010879:Piperazines; D010880:Piperidines / Q000494:pharmacology*; D017465:Receptors, Opioid, delta / Q000819:agonists* / Q000276:immunology / Q000378:metabolism; D015398:Signal Transduction; D013395:Sucrose / Q000494:pharmacology; D014407:Tumor Cells, Cultured
chemical_list: D001549:Benzamides; C415566:N,N-diethyl-4-(phenylpiperidin-4-ylidenemethyl)benzamide; D009292:Narcotic Antagonists; D010879:Piperazines; D010880:Piperidines; D017465:Receptors, Opioid, delta; C088464:4-(alpha-(4-allyl-2,5-dimethyl-1-piperazinyl)-3-methoxybenzyl)-N,N-diethylbenzamide; D013395:Sucrose; D000262:Adenylyl Cyclases