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- Title
Phosphatidic Acid as the Biosynthetic Precursor of the Endocannabinoid 2-Arachidonoylglycerol in Intact Mouse Neuroblastoma Cells Stimulated with Ionomycin.
- Authors
Bisogno, Tiziana; Melck, Dominique; De Petrocellis, Luciano; Di Marzo, Vincenzo
- Abstract
In mouse neuroblastoma N18TG2 cells prelabeled with [3H]arachidonic acid ([3H]AA) the biosynthesis of 2-arachidonoylglycerol (2-AG) is induced by ionomycin in a fashion sensitive to an inhibitor of diacylglycerol (DAG) lipase, RHC 80267, but not to four different phospholipase C (PLC) blockers. Pulse experiments with [3H]AA showed that ionomycin stimulation leads to the sequential formation of [3H]phosphatidic acid ([3H]PA), [3H]DAG, and [3H]2-AG. [3H]2-AG biosynthesis in N18TG2 cells prelabeled with [3H]AA was counteracted by propranolol and N-ethylmaleimide, two inhibitors of the Mg2+/Ca2+-dependent brain PA phosphohydrolase. Pretreatment of cells with exogenous phospholipase D (PLD) led to a strong potentiation of ionomycin-induced [3H]2-AG formation. These data indicate that DAG precursors for 2-AG in intact N18TG2 cells are obtained from the hydrolysis of PA and not through the activation of PLC. The presence of 2% ethanol during ionomycin stimulation failed to elicit the synthesis of [3H]phosphatidylethanol and did not counteract the formation of [3H]PA, thus arguing against the activation of PLD by the Ca2+ ionophore. Selective inhibitors of secretory phospholipase A2 and the acyl-CoA acylase inhibitor thimerosal significantly reduced [3H]2-AG biosynthesis. The implications of these latter findings, and of the PA-dependent pathways of 2-AG formation described here, are discussed.
- Subjects
ARACHIDONIC acid; NEUROBLASTOMA
- Publication
Journal of Neurochemistry, 1999, Vol 72, Issue 5, p2113
- ISSN
0022-3042
- Publication type
Article
- DOI
10.1046/j.1471-4159.1999.0722113.x