title: Opioid agonist and antagonist treatment differentially regulates immunoreactive mu-opioid receptors and dynamin-2 in vivo.
abstract: Opioid agonists and antagonists can regulate the density of mu-opioid receptors in whole animal and in cell culture. High intrinsic efficacy agonists (e.g., etorphine), but not lower intrinsic efficacy agonists (e.g., morphine), produce mu-opioid receptor down-regulation and can alter the abundance of mu-opioid receptor mRNA. Conversely, opioid antagonists substantially increase the density of mu-opioid receptors without changing its mRNA. Mu-opioid receptor up-regulation has been associated with decreases in the trafficking protein dynamin-2, whereas mu-opioid receptor down-regulation produces an increase in dynamin-2 abundance. To probe the differences between opioid agonist and antagonist-induced mu-opioid receptor regulation, the current study determined changes in mu-opioid receptor density using a combined radioligand binding ([3H] DAMGO) and quantitative Western blotting approach in mouse spinal cord. Furthermore, the differences between intermittent and continuous dosing protocols were evaluated. Continuous (7-8 days) s.c. infusions of naloxone (5 mg/kg/day) or naltrexone (15 mg s.c. implant pellet) increased mu-opioid receptor density in radioligand binding assays (approximately +80%) in mouse spinal cord and down-regulated dynamin-2 abundance (approximately -30%), but had no effect on the abundance of immunoreactive mu-opioid receptor. Continuous (7 days) s.c. infusion of etorphine (200 microg/kg/day) decreased immunoreactive mu-opioid receptor (approximately -35%) and [3H] DAMGO binding (approximately -30%), and concurrently increased dynamin-2 abundance (approximately +40%). Continuous (7 days) morphine infusion (40 mg/kg/day plus 25 mg s.c. implant pellet) had no effect on any outcome measure. Delivery of the same daily dose of etorphine or naloxone using intermittent (every 24 h for 7 days) s.c. administration had no effect on immunoreactive mu-opioid receptor, [3H] DAMGO binding or dynamin-2 abundance. These data indicate that mu-opioid receptor density, determined in radioligand binding assays, and immunoreactive dynamin-2 abundance are regulated by continuous, but not intermittent, opioid ligand treatment. Furthermore, the differential regulation of mu-opioid receptor abundance by agonists and antagonists in immunoblotting assays contrasts with changes in [3H] DAMGO binding. Taken together, these results suggest that etorphine-induced down-regulation may depend upon mu-opioid receptor degradation and changes in dynamin-2-mediated receptor trafficking. Conversely, antagonist-induced up-regulation does not require an increase in mu-opioid receptor synthesis and may entail conversion of receptors to an appropriate conformation to bind ligand, as well as changes in receptor trafficking.
authors.affiliation: Department of Pharmaceutical Sciences, College of Pharmacy and Allied Health Professions, St. John's University, 8000 Utopia Parkway, Queens, NY 11439, USA. yoburnb@stjohns.edu
authors.forename: Byron C
authors.initials: BC
authors.lastname: Yoburn
authors.forename: Vishal
authors.initials: V
authors.lastname: Purohit
authors.forename: Kaushal
authors.initials: K
authors.lastname: Patel
authors.forename: Qiuyu
authors.initials: Q
authors.lastname: Zhang
mesh_terms: D000701:Analgesics, Opioid / Q000494:pharmacology; D000818:Animals; D001667:Binding, Competitive / Q000187:drug effects; D015153:Blotting, Western; D034285:Dynamin II / Q000378:metabolism*; D020875:Enkephalin, Ala(2)-MePhe(4)-Gly(5)- / Q000378:metabolism; D005048:Etorphine / Q000494:pharmacology; D008297:Male; D051379:Mice; D009020:Morphine / Q000494:pharmacology; D009270:Naloxone / Q000494:pharmacology; D009292:Narcotic Antagonists / Q000494:pharmacology; D011869:Radioligand Assay; D017450:Receptors, Opioid, mu / Q000819:agonists / Q000037:antagonists & inhibitors / Q000378:metabolism*; D013116:Spinal Cord / Q000187:drug effects / Q000378:metabolism
chemical_list: D000701:Analgesics, Opioid; D009292:Narcotic Antagonists; D017450:Receptors, Opioid, mu; D020875:Enkephalin, Ala(2)-MePhe(4)-Gly(5)-; D009270:Naloxone; D005048:Etorphine; D009020:Morphine; D034285:Dynamin II