title: Antagonism of Dopamine Receptor 2 Long Affects Cannabinoid Receptor 1 Signaling in a Cell Culture Model of Striatal Medium Spiny Projection Neurons.
abstract: Activation of dopamine receptor 2 long (D2L) switches the signaling of type 1 cannabinoid receptor (CB1) from Gαi to Gαs, a process thought to be mediated through CB1-D2L heteromerization. Given the clinical importance of D2 antagonists, the goal of this study was to determine if D2 antagonists could modulate CB1 signaling. Interactions between CB1 and D2L, Gαi, Gαs, and β-arrestin1 were studied using bioluminescence resonance energy transfer 2 (BRET(2)) in STHdh(Q7/Q7) cells. CB1-dependent extracellular regulated kinase (ERK)1/2, CREB phosphorylation, and CB1 internalization following cotreatment of CB1 agonist and D2 antagonist were quantified. Preassembled CB1-Gαi complexes were detected by BRET(2) Arachidonyl-2'-chloroethylamide (ACEA), a selective CB1 agonist, caused a rapid and transient increase in BRET efficiency (BRETEff) between Gαi-Rluc and CB1-green fluorescent protein 2 (GFP(2)), and a Gαi-dependent increase in ERK phosphorylation. Physical interactions between CB1 and D2L were observed using BRET(2) Cotreatment of STHdh(Q7/Q7) cells with ACEA and haloperidol, a D2 antagonist, inhibited BRETEff signals between Gαi-Rluc and CB1-GFP(2) and reduced the EMax and pEC50 of ACEA-mediated Gαi-dependent ERK phosphorylation. ACEA and haloperidol cotreatments produced a delayed and sustained increase in BRETEff between Gαs-Rluc and CB1-GFP(2) and increased the EMax and pEC50 of ACEA-induced Gαs-dependent cAMP response element-binding protein phosphorylation. In cells expressing CB1 and D2L treated with ACEA, binding of haloperidol to D2 receptors switched CB1 coupling from Gαi to Gαs In addition, haloperidol treatment reduced ACEA-induced β-arrestin1 recruitment to CB1 and CB1 internalization. D2 antagonists allosterically modulate cannabinoid-induced CB1 coupling, signaling, and β-arrestin1 recruitment through binding to CB1-D2L heteromers. These findings indicate that D2 antagonism, like D2 agonists, change agonist-mediated CB1 coupling and signaling.
authors.affiliation: Departments of Pharmacology and Ophthalmology and Visual Sciences, Dalhousie University, Halifax, Nova Scotia, Canada.
authors.forename: Amina M
authors.initials: AM
authors.lastname: Bagher
authors.affiliation: Departments of Pharmacology and Ophthalmology and Visual Sciences, Dalhousie University, Halifax, Nova Scotia, Canada.
authors.forename: Robert B
authors.initials: RB
authors.lastname: Laprairie
authors.affiliation: Departments of Pharmacology and Ophthalmology and Visual Sciences, Dalhousie University, Halifax, Nova Scotia, Canada.
authors.forename: Melanie E M
authors.initials: ME
authors.lastname: Kelly
authors.affiliation: Departments of Pharmacology and Ophthalmology and Visual Sciences, Dalhousie University, Halifax, Nova Scotia, Canada emdenova@dal.ca.
authors.forename: Eileen M
authors.initials: EM
authors.lastname: Denovan-Wright
mesh_terms: D000818:Animals; D001095:Arachidonic Acids / Q000494:pharmacology; D002478:Cells, Cultured; D003342:Corpus Striatum / Q000166:cytology*; D017362:Cyclic AMP Response Element-Binding Protein / Q000378:metabolism; D065127:Dopamine D2 Receptor Antagonists / Q000494:pharmacology*; D004705:Endocytosis / Q000187:drug effects; D048049:Extracellular Signal-Regulated MAP Kinases / Q000378:metabolism; D019206:GTP-Binding Protein alpha Subunits, Gi-Go / Q000378:metabolism; D006220:Haloperidol / Q000494:pharmacology; D006801:Humans; D007700:Kinetics; D008297:Male; D008810:Mice, Inbred C57BL; D008954:Models, Biological; D009474:Neurons / Q000187:drug effects / Q000378:metabolism*; D010766:Phosphorylation / Q000187:drug effects; D011485:Protein Binding / Q000187:drug effects; D020033:Protein Isoforms / Q000378:metabolism; D055503:Protein Multimerization / Q000187:drug effects; D043884:Receptor, Cannabinoid, CB1 / Q000378:metabolism*; D017448:Receptors, Dopamine D2 / Q000378:metabolism*; D000071557:beta-Arrestins / Q000378:metabolism
chemical_list: D001095:Arachidonic Acids; D017362:Cyclic AMP Response Element-Binding Protein; D065127:Dopamine D2 Receptor Antagonists; D020033:Protein Isoforms; D043884:Receptor, Cannabinoid, CB1; D017448:Receptors, Dopamine D2; C119325:arachidonyl-2-chloroethylamide; D000071557:beta-Arrestins; D048049:Extracellular Signal-Regulated MAP Kinases; D019206:GTP-Binding Protein alpha Subunits, Gi-Go; D006220:Haloperidol