title: mu-opioid receptor-stimulated synthesis of reactive oxygen species is mediated via phospholipase D2.
abstract: We have recently shown that the activation of the rat mu-opioid receptor (MOPr, also termed MOR1) by the mu-agonist [D-Ala(2), Me Phe(4), Glyol(5)]enkephalin (DAMGO) leads to an increase in phospholipase D2 (PLD2) activity and an induction of receptor endocytosis, whereas the agonist morphine which does not induce opioid receptor endocytosis fails to activate PLD2. We report here that MOPr-mediated activation of PLD2 stimulates production of reactive oxygen molecules via NADH/NADPH oxidase. Oxidative stress was measured with the fluorescent probe dichlorodihydrofluorescein diacetate and the role of PLD2 was assessed by the PLD inhibitor D-erythro-sphingosine (sphinganine) and by PLD2-small interfering RNA transfection. To determine whether NADH/NADPH oxidase contributes to opioid-induced production of reactive oxygen species, mu-agonist-stimulated cells were pre-treated with the flavoprotein inhibitor, diphenylene iodonium, or the specific NADPH oxidase inhibitor, apocynin. Our results demonstrate that receptor-internalizing agonists (like DAMGO, beta-endorphin, methadone, piritramide, fentanyl, sufentanil, and etonitazene) strongly induce NADH/NADPH-mediated ROS synthesis via PLD-dependent signaling pathways, whereas agonists that do not induce MOPr endocytosis and PLD2 activation (like morphine, buprenorphine, hydromorphone, and oxycodone) failed to activate ROS synthesis in transfected human embryonic kidney 293 cells. These findings indicate that the agonist-selective PLD2 activation plays a key role in the regulation of NADH/NADPH-mediated ROS formation by opioids.
authors.affiliation: Department of Pharmacology and Toxicology, Otto-von-Guericke University, Magdeburg, Germany.
authors.forename: Thomas
authors.initials: T
authors.lastname: Koch
authors.forename: Anja
authors.initials: A
authors.lastname: Seifert
authors.forename: Dai-Fei
authors.initials: DF
authors.lastname: Wu
authors.forename: Marija
authors.initials: M
authors.lastname: Rankovic
authors.forename: Jürgen
authors.initials: J
authors.lastname: Kraus
authors.forename: Christine
authors.initials: C
authors.lastname: Börner
authors.forename: Lars-Ove
authors.initials: LO
authors.lastname: Brandenburg
authors.forename: Helmut
authors.initials: H
authors.lastname: Schröder
authors.forename: Volker
authors.initials: V
authors.lastname: Höllt
mesh_terms: D000701:Analgesics, Opioid / Q000494:pharmacology*; D000818:Animals; D002460:Cell Line; D004705:Endocytosis / Q000187:drug effects / Q000502:physiology; D004789:Enzyme Activation / Q000187:drug effects / Q000502:physiology; D004791:Enzyme Inhibitors / Q000494:pharmacology; D006801:Humans; D009243:NAD / Q000187:drug effects / Q000378:metabolism; D009249:NADP / Q000187:drug effects / Q000378:metabolism; D018384:Oxidative Stress / Q000187:drug effects* / Q000502:physiology; D010739:Phospholipase D / Q000187:drug effects* / Q000378:metabolism; D051381:Rats; D017382:Reactive Oxygen Species / Q000819:agonists* / Q000378:metabolism; D017450:Receptors, Opioid, mu / Q000187:drug effects* / Q000378:metabolism; D015398:Signal Transduction / Q000187:drug effects* / Q000502:physiology
chemical_list: D000701:Analgesics, Opioid; D004791:Enzyme Inhibitors; D017382:Reactive Oxygen Species; D017450:Receptors, Opioid, mu; D009243:NAD; D009249:NADP; C105606:phospholipase D2; D010739:Phospholipase D