Identification of biosynthetic precursors for the endocannabinoid anandamide in the rat brain

Abstract

Anandamide is an endogenous signaling lipid that binds to and activates cannabinoid receptors in the brain and peripheral tissues. The endogenous precursors of ananda­mide, N-arachidonoyl phosphatidylethanolamines (NArPEs), are a family of complex glycerophospholipids that derive from the exchange reaction of an arachidonoyl group between the sn-1 position of phosphatidylcholine and the primary amine of phosphatidylethanolamine catalyzed by N-acyl transferase activity. A precise characterization of the molec­ular composition of NArPE species generating anandamide has not yet been reported. In the present study, using liquid chromatography coupled to electrospray ionization ion-trap mass spectrometry, we identified the major endogenous NArPE species, which mainly contained sn-1 alkenyl groups (C16:0, C18:0, C18:1) and monounsaturated (C18:1) or polyunsaturated (C20:4, C22:4, C22:6) acyl groups at the sn-2 position of the glycerol backbone. Using rat brain particulate fractions, we observed a calcium-dependent increase in both NArPEs and anandamide formation after incubation at 37°C for 30 min. Furthermore, a targeted lipidomic analysis showed that Ca2+ specifically stimulated the formation of PUFA-containing NArPE species. These results reveal a previously unrecognized preference of brain N-acyl transferase activity for polyunsaturated NArPE and provide new insights on the physiological regulation of anandamide biosynthesis.—Astarita, G., F. Ahmed, and D. Piomelli. Identification of biosynthetic precursors for the endocannabinoid anandamide in the rat brain. J. Lipid Res. 2008. 49: 48–57.

Keywords

• fatty acid ethanolamine
• arachidonoyl-ethanolamide
• N-arachidonoyl phosphatidylethanolamines
• phosphatidylethanolamine
• N-acyl phosphatidylethanolamines
• N-acyl transferase