Abstract
Arachidonyl ethanolamine, which is commonly known as anandamide, was the first endogenous compound to be identified that binds to the cannabinoid receptors. Anandamide mimics many of the physiological effects of Δ9-tetrahydrocannabinol (Δ9-THC), including hypothermia, antinociception, immobility, catalepsy, and immune modulation. In the present studies, we show that anandamide caused a concentration-dependent inhibition of interleukin-2 in primary splenocytes. The CB1 and CB2 antagonists, SR141716A [N-(piperidin-1-yl)-5-(4-chlorophenyl)-1-(2,4-dichlorphenyl)-4-methyl-H-pyrazole-3 carboxyamidehydrochloride] and SR144528 [N-[(1S)-endo-1,3,3,-trimethylbicyclo[2,2,1]heptan-2-yl]-5-(4-chloro-3-methylphenyl)-1-(4-methylbenzyl)-pyrazole-3-carboxamide], when used in combination, did not antagonize the inhibition of interleukin-2 by anandamide. Additionally, neither UCM707 [N-(3-furanylmethyl)-5Z,8Z,11Z,14Z-eicosatetraenamide], the inhibitor of the putative anandamide membrane transporter (AMT), nor methyl arachidonoyl fluorophosphonate (MAFP), the inhibitor of fatty acid amidohydrolase (FAAH), were able to affect the inhibitory activity of anandamide upon interleukin-2. Interestingly, arachidonic acid caused a concentration-dependent inhibition of interleukin-2 secretion (IC50 = 10.3 μM), which was similar to that of structurally related anandamide (IC50 = 11.4 μM). The inhibition of interleukin-2 by anandamide and arachidonic acid was partially reversed by pretreatment with the nonspecific cyclooxygenase inhibitors, flurbiprofen and piroxicam. Moreover, NS398 [N-[2-(cyclohexyloxy)-4-nitrophenyl]-methanesulfonamide], a cyclooxygenase-2-specific inhibitor, also attenuated the inhibitory effects of anandamide and arachidonic acid upon interleukin-2 secretion. Finally, pretreatment with a peroxisome proliferator-activated receptor γ (PPARγ)-specific antagonist, T0070907 [2-chloro-5-nitro-N-4-pyridinyl-benzamide], partially antagonized anandamide-mediated suppression of IL-2 secretion. Collectively, the aforementioned studies suggest that inhibition of interleukin-2 secretion by anandamide is independent of CB1/CB2 and the AMT/FAAH system. Additionally, these studies also suggest that inhibition of interleukin-2 is mediated by a PPARγ, which is activated by a cyclooxygenase-2 metabolite of anandamide.
Footnotes
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This work was supported with funds from the National Institute on Drug Abuse Grant DA12740.
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doi:10.1124/jpet.104.065524.
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ABBREVIATIONS: LPS, lipopolysaccharide; AMT, anandamide membrane transporter; ANOVA, analysis of variance; CB1, cannabinoid receptor 1; CB2, cannabinoid receptor 2; ELISA, enzyme-linked immunosorbent assay; FAAH, fatty acid amidohydrolase; FR122047, 1-[[4,5-bis(4-methoxyphenyl)-2-thiazolyl]carbonyl]-4-methylpiperazine; IL, interleukin; MAFP, methyl arachidonyl fluorophosphonate; NFAT, nuclear factor of activated T cells; NS398, N-[2-(cyclohexyloxy)-4-nitrophenyl]-methanesulfonamide; PG, prostaglandin; 15d-PGJ2, 15-deoxy-prostaglandin J2; PMA, phorbol 12-myristate 13-acetate; PPAR, peroxisome proliferator-activated receptor; SC560, 5-(4-chlorophenyl)-1-(4-methoxyphenyl)3-3(trifluoromethyl)1H-pyrazole; SR141716A, N-(piperidin-1-yl)-5-(4-chlorophenyl)-1-(2,4-dichlorphenyl)-4-methyl-H-pyrazole-3 carboxyamidehydrochloride; SR144528, 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; T0070907, 2-chloro-5-nitro-N-4-pyridinyl-benzamide; UCM707, N-(3-furanylmethyl)-5Z,8Z,11Z,14Z-eicosatetraenamide; VR1, vanilloid receptor 1.
- Received January 27, 2004.
- Accepted July 29, 2004.
- The American Society for Pharmacology and Experimental Therapeutics
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