title: Effects of the allosteric antagonist 1-(4-chlorophenyl)-3-[3-(6-pyrrolidin-1-ylpyridin-2-yl)phenyl]urea (PSNCBAM-1) on CB1 receptor modulation in the cerebellum.
abstract: 1-(4-Chlorophenyl)-3-[3-(6-pyrrolidin-1-ylpyridin-2-yl)phenyl] urea (PSNCBAM-1) has recently been described as a cannabinoid CB1 receptor allosteric antagonist associated with hypophagic effects in vivo; however, PSNCBAM-1 effects on CB(1) ligand-mediated modulation of neuronal excitability remain unknown. Here, we investigate PSNCBAM-1 actions on CB(1) receptor-stimulated guanosine 5'-O-(3-[(35)S]thio)triphosphate ([(35)S]GTPγS) binding in cerebellar membranes and on CB(1) ligand modulation of presynaptic CB(1) receptors at inhibitory interneuron-Purkinje cell synapses in the cerebellum using whole-cell electrophysiology. PSNCBAM-1 caused noncompetitive antagonism in [(35)S]GTPγS binding studies, with higher potency against the CB receptor agonist (-)-cis-3-[2-hydroxy-4-(1,1-dimethyl heptyl)phenyl]-trans-4-(3-hydroxypropyl)cyclohexanol (CP55940) than for R(+)-[2,3-dihydro-5-methyl-3-[(morpholinyl)methyl]-pyrrolo[1,2,3,-de]-1,4-benzoxazinyl]-(1-naphthalenyl)methanone mesylate] [WIN55,212-2 (WIN55)]. In electrophysiological studies, WIN55 and CP55940 reduced miniature inhibitory postsynaptic currents (mIPSCs) frequency but not amplitude. PSNCBAM-1 application alone had no effect on mIPSCs; however, PSNCBAM-1 pretreatment revealed agonist-dependent functional antagonism, abolishing CP55940-induced reductions in mIPSC frequency but having no clear effect on WIN55 actions. The CB(1) antagonist/inverse agonist N-(piperidin-1-yl)-1-(2,4-dichlorophenyl)-5-(4-iodophenyl)-4-methyl-1H-multipyrazole-3-carboxamide (AM251) increased mIPSC frequency beyond control; this effect was reversed by PSNCBAM-1. PSNCBAM-1 pretreatment also attenuated AM251 effects. Thus, PSNCBAM-1 reduced CB(1) receptor ligand functional efficacy in the cerebellum. The differential effect of PSNCBAM-1 on CP55940 versus WIN55 actions in [(35)S]GTPγS binding and electrophysiological studies and the attenuation of AM251 effects are consistent with the ligand-dependence associated with allosteric modulation. These data provide the first description of functional PSNCBAM-1 allosteric antagonist effects on neuronal excitability in the mammalian central nervous system (CNS). PSNCBAM-1 allosteric antagonism may provide viable therapeutic alternatives to orthosteric CB(1) antagonists/inverse agonists in the treatment of CNS disease.
authors.affiliation: School of Pharmacy, University of Reading, Whiteknights, Reading, United Kingdom.
authors.forename: Xiaowei
authors.initials: X
authors.lastname: Wang
authors.forename: James G
authors.initials: JG
authors.lastname: Horswill
authors.forename: Benjamin J
authors.initials: BJ
authors.lastname: Whalley
authors.forename: Gary J
authors.initials: GJ
authors.lastname: Stephens
mesh_terms: D000494:Allosteric Regulation / Q000187:drug effects / Q000502:physiology; D000818:Animals; D002531:Cerebellum / Q000187:drug effects* / Q000378:metabolism*; D004305:Dose-Response Relationship, Drug; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D010671:Phenylurea Compounds / Q000737:chemistry / Q000378:metabolism* / Q000494:pharmacology*; D011485:Protein Binding / Q000502:physiology; D011725:Pyridines / Q000737:chemistry / Q000378:metabolism* / Q000494:pharmacology*; D043884:Receptor, Cannabinoid, CB1 / Q000037:antagonists & inhibitors* / Q000502:physiology*
chemical_list: C526691:1-(4-chlorophenyl)-3-(3-(6-pyrrolidin-1-ylpyridin-2-yl)phenyl)urea; D010671:Phenylurea Compounds; D011725:Pyridines; D043884:Receptor, Cannabinoid, CB1