sq-23377 and palmidrol

Endocannabinoids and related N-acylethanolamines (NAEs) are involved in regulation of gut function, but relatively little is known as to whether inflammatory cytokines such as IFNγ affect their levels. We have investigated this in vitro using cultures of T84 colon cancer cells.. T84 cells, when cultured in monolayers, differentiate to form adult colonic crypt-like cells with excellent permeability barrier properties. The integrity of the permeability barrier in these monolayers was measured using transepithelial electrical resistance (TEER). NAE levels were determined by ultra-performance liquid chromatography-tandem mass spectrometric analysis. Expression of the enzymes involved in NAE and 2-arachidonoylglycerol (2-AG) turnover were assessed with qPCR.. IFNγ treatment for 8 or 24 h increased levels of both endocannabinoids (anandamide and 2-AG) and the related NAEs. The treatment did not affect the rate of hydrolysis of either anandamide or palmitoylethanolamide by intact cells, and in both cases, fatty acid amide hydrolase (FAAH) rather than NAE-hydrolysing acid amidase (NAAA) was mainly responsible for the hydrolysis of these NAEs. IFNγ treatment reduced the TEER of the cells in a manner that was not prevented by inhibition of either FAAH or NAAA but was partially reversed by apical administration of the NAE palmitoylethanolamide.. IFNγ treatment mobilized endocannabinoid and related NAE levels in T84 cells. However, blockade of anandamide or NAE hydrolysis was insufficient to negate the deleterious effects of this cytokine upon the permeability barrier of the cell monolayers.

Topics: Amides; Arachidonic Acids; Cell Culture Techniques; Cell Line, Tumor; Chromatography, High Pressure Liquid; Colonic Neoplasms; Endocannabinoids; Ethanolamines; Glycerides; Humans; Interferon-gamma; Ionomycin; Palmitic Acids; Polyunsaturated Alkamides

Understanding the mechanisms involved in the biogenesis of N-arachidonoylethanolamine (anandamide) and N-palmitoylethanolamine is important in view of the possible role of these lipids as endogenous cannabinoid substances. Anandamide (which activates cannabinoid CB1 receptors) and N-palmitoylethanolamine (which activates a CB2-like receptor subtype in mast cells) may both derive from cleavage of precursor phospholipid, N-acylphosphatidylethanolamine (NAPE), catalyzed by Ca(2+)-activated D-type phosphodiesterase activity. We report here that the de novo biosynthesis of NAPE is enhanced in a Ca(2+)-dependent manner when rat cortical neurons are stimulated with the Ca(2+)-ionophore ionomycin or with membrane-depolarizing agents such as veratridine and kainate. This reaction is likely to be mediated by a neuronal N-acyltransferase activity, which catalyzes the transfer of an acyl group from phosphatidylcholine to the ethanolamine moiety of phosphatidylethanolamine. In addition, we show that Ca2+-dependent NAPE biosynthesis is potentiated by agents that increase cAMP levels, including forskolin and vasoactive intestinal peptide. Our results thus indicate that NAPE levels in cortical neurons are controlled by Ca2+ ions and cAMP. Such regulatory effect may participate in maintaining a supply of cannabimimetic N-acylethanolamines during synaptic activity, and prime target neurons for release of these bioactive lipids.

Topics: Amides; Animals; Anti-Inflammatory Agents, Non-Steroidal; Arachidonic Acids; Arylamine N-Acetyltransferase; Astrocytes; Calcium; Calcium Channel Blockers; Calmodulin; Cannabinoids; Carbachol; Cyclic AMP; Endocannabinoids; Enzyme Inhibitors; Ethanolamine; Ethanolamines; Imidazoles; Ionomycin; Ionophores; Neurons; Nicotinic Agonists; Palmitic Acids; Phosphatidylethanolamines; Polyunsaturated Alkamides; Rats; Sodium Channel Agonists; Tritium; Vasoactive Intestinal Peptide; Veratridine