Occurrence and postmortem generation of anandamide and other long‐chain N‐acylethanolamines in mammalian brain

Abstract

Long-chain N-acylethanolamines (NAEs) were assayed in pig, sheep and cow brain by gas chromatography/mass spectrometry of their tert.-butyldimethylsilyl derivatives in the presence of deuterium-labeled internal standards. Immediately after death, total NAEs ranged from about 2.7/tglg wet weight (sheep, cow) to 6.5 /tg/g wet weight (pig) and consisted almost exclusively (99%) of saturated and monounsaturated species. Anandamide (20:4n-6 NAE) comprised about 1% of total NAE in pig and cow brain, but was absent in freshly extracted sheep brain. When pig brain was analysed after 0.5, 1, 3, 4 and 23 h at ambient temperature, NAE levels were found to increase substantially over the entire time period with 20:4n-6 NAE formation exhibiting a time lag compared to that of saturated and monounsaturated NAEs.Key words: N-Acylethanolamine; Anandamide; Arachidonic acid; Pig brain; Sheep brain; Cow brain dog heart [20,21] and have observed that N-acylethanolamine phospholipids are generated in young rat brain within 15 min postmortem [19]. In mammalian cells, N-acylethanolamine phospholipids are produced by acyl transfer from the sn-1 position of various glycerophospholipids to the free amino group of ethanolamine phospholipids [22][23][24], a reaction that does not involve ester hydrolysis [25]. They are metabolized to N-acylethanolamines and phosphatidic acid by phosphodiesterase (phospholipase D) activity [26,27]. This pathway generates primarily saturated and monounsaturated, as well as some diunsaturated [28], NAEs and these are the only ones for which quantitative information is available [29].Here, we report the quantitative assay of NAEs present in pig, sheep and cow brain at different periods postmortem using gas chromatography-mass spectrometry of their tert.-butyldimethylsilyl derivatives in the presence of deuterated analogs as internal standards.

Changes in anandamide levels in mouse uterus are associated with uterine receptivity for embryo implantation

“…Individual brains were homogenized in chloroform and the extracts were centrifuged. A mixture of internal standards was added during homogenization (21). The standards were as follows: N-acyl-1,1,2,2,-2 H-ethanolamines with acyl chain lengths of 16:0, 17:0, 18:0, 18:1n-9, 18:2n-6, and 20:4n-6 (d4 NAEs, 1 g each), and N-hept adecanoyl-1,2-dihept adecanoylphosphatidylethanolamine (17:0 NAPE, 2.7 nmol).…”

Biosynthesis and turnover of anandamide and other N‐acylethanolamines in peritoneal macrophages

“…Unlabeled ethanolamine and calcium ionophore A23187 were from Sigma (St. Louis, MO, USA). [1,[2][3][4][5][6][7][8][9][10][11][12][13][14] C]Ethanolamine (speci¢c activity, 2.5 mCi/ mmol) was from NEN Life Science Products (USA). L-Glutamine and all other chemicals used in the incubation experiments were analytical grade from Wako Pure Chemical Industries (Osaka, Japan).…”

“…In mammalian tissues, they accumulate post mortem [2] and under pathological conditions involving degenerative membrane changes [1]. It appears that under such conditions a loss of calcium homeostasis results in the activation of a transacylase that catalyzes the direct transfer of O-acyl groups from the sn-1 position of glycerophospholipids to the amino group of ethanolamine phospholipids [3].…”

Regional distribution and effects of postmortal delay on endocannabinoid content of the human brain

“…The concentration of AEA in the mammalian brains increases rapidly after death (Schmid et al, 1995;Kempe et al, 1996;Bisogno et al, 1999;Patel et al, 2004). A seven-fold increase in AEA levels has been measured in the rat brain 24 h after death (Maccarrone et al, 2001).…”

“…Studies in rodents have documented postmortal increases in brain levels of endocannabinoids (Schmid et al, 1995;Felder et al, 1996;Kempe et al, 1996;Sugiura et al, 2001;Patel et al, 2004). In the human brain, postmortal changes of cannabinoid receptors have been reported (Mato and Pazos, 2004), but similar changes in AEA and 2-AG have not yet been explored.…”