title: CB1 cannabinoid receptor-mediated increases in cyclic AMP accumulation are correlated with reduced Gi/o function.
abstract: BACKGROUND
  CB1 cannabinoid receptors (CB1Rs) stimulate Gi/o-dependent signaling pathways. CB1R-mediated cAMP increases were proposed to result from Gs activation, but CB1R-stimulated GTPγS binding to Gs has not heretofore been investigated.
  
  
  METHODS
  Three models of CB1R-stimulated cAMP production were tested: pertussis toxin disruption of Gi/o in N18TG2 cells; L341A/A342L-CB1R expressed in Chinese hamster ovary (CHO) cells; and CB1 and D2 dopamine receptors endogenously co-expressed in MN9D cells. cAMP was assayed by [3H]cAMP binding competition. G protein activation was assayed by the antibody-targeted scintillation proximity assay.
  
  
  RESULTS
  In L341A/A342L-CB1-CHO cells, cannabinoid agonists significantly stimulated cAMP accumulation over vehicle; (-)-3-[2-hydroxyl-4-(1,1-dimethylheptyl)phenyl]-4-[3-hydroxyl propyl] cyclohexan-1-ol (CP55940)-stimulated [35S]GTPγS binding to Gi1/2/3 was reversed, whereas binding to Gs was not different from CB1R. In MN9D cells, CB1 agonist HU210 or D2 agonist quinpirole alone inhibited forskolin-activated cAMP accumulation, whereas HU210 plus quinpirole increased cAMP accumulation above basal. HU210 alone stimulated [35S]GTPγS binding to Gi1/2/3, whereas co-stimulation with quinpirole reversed HU210-stimulated [35S]GTPγS binding to Gi1/2/3.
  
  
  CONCLUSIONS
  CB1R couples to Gs but with low efficacy compared to Gi/o. The L341A/A342L mutation in CB1R reversed CP55940 activation of Gi to an inhibition, but had no effect on Gs. Combined CB1 plus D2 agonists in MN9D cells converted the CB1 agonist-mediated activation of Gi to inhibition of Gi. In these models, the CB1 agonist response was converted to an inverse agonist response at Gi activation. Cannabinoid agonist-stimulated cAMP accumulation can be best explained as reduced activation of Gi, thereby attenuating the tonic inhibitory influence of Gi on the major isoforms of adenylyl cyclase.
authors.forename: Khalil
authors.initials: K
authors.lastname: Eldeeb
authors.forename: Sandra
authors.initials: S
authors.lastname: Leone-Kabler
authors.forename: Allyn C
authors.initials: AC
authors.lastname: Howlett
mesh_terms: D000818:Animals; D016466:CHO Cells; D002186:Cannabinoids / Q000494:pharmacology; D002460:Cell Line; D045744:Cell Line, Tumor; D006224:Cricetinae; D003412:Cricetulus; D000242:Cyclic AMP / Q000378:metabolism*; D003511:Cyclohexanols / Q000494:pharmacology; D013759:Dronabinol / Q000031:analogs & derivatives / Q000494:pharmacology; D019204:GTP-Binding Proteins / Q000378:metabolism; D006801:Humans; D043884:Receptor, Cannabinoid, CB1 / Q000378:metabolism*; D017448:Receptors, Dopamine D2 / Q000378:metabolism; D015398:Signal Transduction / Q000187:drug effects
chemical_list: D002186:Cannabinoids; D003511:Cyclohexanols; D043884:Receptor, Cannabinoid, CB1; D017448:Receptors, Dopamine D2; D013759:Dronabinol; C054649:3-(2-hydroxy-4-(1,1-dimethylheptyl)phenyl)-4-(3-hydroxypropyl)cyclohexanol; D000242:Cyclic AMP; D019204:GTP-Binding Proteins; C062018:HU 211