Recently we have shown that treatment of rat C6 glioma cells with the membrane cholesterol depletor and raft disruptor methyl-β-cyclodextrin (MCD) doubles the binding of anandamide (AEA) to type-1 cannabinoid receptors (CB1R), followed by CB1R-dependent signaling via adenylate cyclase (AC) and p42/p44 mitogen activated protein kinase (MAPK) activity (Bari, M., Battista, N., Fezza, F., Finazzi-Agrò, A. and Maccarrone, M. (2005) J. Biol. Chem. 280, 12212-12220). Here, we investigated whether also type-2 cannabinoid receptors (CB2R), widely expressed inimmune cells, are modulated by MCD. We show that treatment of human DAUDI leukemia cells with MCD does not affect AEA binding to CB2R. The activation of CB2R by AEA triggers a similar [35S]GTPγS binding in MCD-treated and controlcells, and thus a similar effect on AC and MAPK activity, and on MAPK-dependent protection against apoptosis. The other AEA-binding receptor TRPV1, the AEA synthetase NAPE-PLD and the AEA hydrolase FAAH were not affected by MCD,whereas the activity of the AEA membrane transporter AMT was reduced to ~55% of the controls. Furthermore, neither diacylglycerol lipase nor monoacylglycerol lipase, which respectively synthesize and degrade 2-arachidonoylglycerol, were affected by MCD, whereas the transport of 2-arachidonoylglycerol was reduced to ~50%. Instead, membrane cholesterol enrichment almost doubled the uptake of 2-arachidonoylglycerol and AEA. Transient expression of CB1R and CB2R in human immune cells ruled out that the different effect of raft disruption on the two receptor subtypes might be due to the gross cellular environment. Altogether, the present data demonstrate that lipid rafts control the activity of CB1R, but not that of CB2R, and endocannabinoid transport across the plasma membranes.[...]
LIPID RAFTS, CB2 RECEPTOR SIGNALING AND METABOLISM OF 2-ARACHIDONOYL-GLYCEROL IN HUMAN IMMUNE CELLS
ODDI, Sergio;
2006-01-01
Abstract
Recently we have shown that treatment of rat C6 glioma cells with the membrane cholesterol depletor and raft disruptor methyl-β-cyclodextrin (MCD) doubles the binding of anandamide (AEA) to type-1 cannabinoid receptors (CB1R), followed by CB1R-dependent signaling via adenylate cyclase (AC) and p42/p44 mitogen activated protein kinase (MAPK) activity (Bari, M., Battista, N., Fezza, F., Finazzi-Agrò, A. and Maccarrone, M. (2005) J. Biol. Chem. 280, 12212-12220). Here, we investigated whether also type-2 cannabinoid receptors (CB2R), widely expressed inimmune cells, are modulated by MCD. We show that treatment of human DAUDI leukemia cells with MCD does not affect AEA binding to CB2R. The activation of CB2R by AEA triggers a similar [35S]GTPγS binding in MCD-treated and controlcells, and thus a similar effect on AC and MAPK activity, and on MAPK-dependent protection against apoptosis. The other AEA-binding receptor TRPV1, the AEA synthetase NAPE-PLD and the AEA hydrolase FAAH were not affected by MCD,whereas the activity of the AEA membrane transporter AMT was reduced to ~55% of the controls. Furthermore, neither diacylglycerol lipase nor monoacylglycerol lipase, which respectively synthesize and degrade 2-arachidonoylglycerol, were affected by MCD, whereas the transport of 2-arachidonoylglycerol was reduced to ~50%. Instead, membrane cholesterol enrichment almost doubled the uptake of 2-arachidonoylglycerol and AEA. Transient expression of CB1R and CB2R in human immune cells ruled out that the different effect of raft disruption on the two receptor subtypes might be due to the gross cellular environment. Altogether, the present data demonstrate that lipid rafts control the activity of CB1R, but not that of CB2R, and endocannabinoid transport across the plasma membranes.[...]I documenti in IRIS sono protetti da copyright e tutti i diritti sono riservati, salvo diversa indicazione.
