title: Evidence for cannabinoid receptor-dependent and -independent mechanisms of action in leukocytes.
abstract: Cannabinoids exhibit immunosuppressive actions that include inhibition of interleukin-2 production in response to a variety of T cell activation stimuli. Traditionally, the effects of these compounds have been attributed to cannabinoid receptors CB1 and CB2, both of which are expressed in mouse splenocytes. Therefore, N-(piperidin-1-yl)-5-(4-chlorophenyl)-1-(2,4-dichlorphenyl)-4-methyl-H-pyrazole-3 carboxyamidehydrochloride (SR141716A), a CB1 antagonist, and N-[(1S)-endo-1,3,3,-trimethyl-bicyclo[2,2,1]heptan-2-yl]-5-(4-chloro-3-methylphenyl)-1-(4-methylbenzyl)-pyrazole-3-carboxamide (SR144528), a CB2 antagonist, were used to investigate the role of cannabinoid receptors in the cannabinoid-induced inhibition of phorbol ester plus calcium ionophore (PMA/Io)-stimulated interleukin-2 production by mouse splenocytes. PMA/Io-stimulated interleukin-2 production was inhibited by cannabinol, cannabidiol, and both WIN 55212-2 stereoisomers with a rank order potency of R-(+)-[2,3-dihydro-5-methyl-3-[(morpholinyl)methyl]pyrrolo[1,2,3-de]-1,4-benzoxazinyl]-(1-napthanlenyl) methanone mesylate (WIN 55212-2) approximately cannabidiol > S-(-)-[2,3-dihydro-5-methyl-3-[(morpholinyl)methyl]pyrrolo[1,2,3-de]-1,4-benzoxazinyl]-(1-napthanlenyl) methanone mesylate (WIN 55212-3) approximately cannabinol. Cannabinoid-induced inhibition of PMA/Io-stimulated interleukin-2 was not attenuated by the presence of both SR144528 and SR141716A. Using pertussis toxin to address the role of G protein-coupled receptors in this response, it was determined that pertussis toxin treatment did not attenuate cannabinol-induced inhibition of PMA/Io-stimulated interleukin-2. With the demonstration that cannabinoid-induced inhibition of PMA/Io-stimulated interleukin-2 was not mediated via CB1 or CB2, alternative targets of cannabinoids in T cells were examined. Specifically, it was demonstrated that cannabinoids elevated intracellular calcium concentration in resting splenocytes and that the cannabinol-induced elevation in intracellular calcium concentration was attenuated by treatment with both SR144528 and SR141716A. Interestingly, pretreatment of splenocytes with agents that elevate intracellular calcium concentration inhibited PMA/Io-stimulated interleukin-2 production, suggesting that an elevation in intracellular calcium concentration might be involved in the mechanism of interleukin-2 inhibition. These studies suggest that immune modulation produced by cannabinoids involves multiple mechanisms, which might be both cannabinoid receptor-dependent and -independent.
authors.affiliation: Department of Pharmacology and Toxicology, Michigan State University, East Lansing, MI 48824, USA.
authors.forename: Barbara L Faubert
authors.initials: BL
authors.lastname: Kaplan
authors.forename: Cheryl E
authors.initials: CE
authors.lastname: Rockwell
authors.forename: Norbert E
authors.initials: NE
authors.lastname: Kaminski
mesh_terms: D000818:Animals; D048588:Benzoxazines; D002118:Calcium / Q000378:metabolism; D002185:Cannabidiol / Q000494:pharmacology; D004347:Drug Interactions; D005260:Female; D007376:Interleukin-2 / Q000378:metabolism; D007962:Leukocytes / Q000187:drug effects / Q000378:metabolism / Q000502:physiology*; D051379:Mice; D009025:Morpholines / Q000494:pharmacology; D009281:Naphthalenes / Q000494:pharmacology; D037342:Pertussis Toxin / Q000494:pharmacology; D043882:Receptors, Cannabinoid; D011955:Receptors, Drug / Q000037:antagonists & inhibitors / Q000502:physiology*; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology
chemical_list: D048588:Benzoxazines; D007376:Interleukin-2; D009025:Morpholines; D009281:Naphthalenes; D043882:Receptors, Cannabinoid; D011955:Receptors, Drug; D002185:Cannabidiol; C070417:(3R)-((2,3-dihydro-5-methyl-3-((4-morpholinyl)methyl)pyrrolo-(1,2,3-de)-1,4-benzoxazin-6-yl)(1-naphthalenyl))methanone; D037342:Pertussis Toxin; D013755:Tetradecanoylphorbol Acetate; D002118:Calcium