This thesis focuses on bioactive lipids as (1) metabolites of the widely used antipyretic and analgesic drug acetaminophen and (2) activators of the ion channel TRPV1, an important downstream target for inflammatory mediators, in the phospholipase C (PLC)/TRPV1 signaling pathway. Evidence is presented for a fatty acid amide hydrolase (FAAH)-dependent fatty acid conjugation of p-aminophenol, a known acetaminophen metabolite, to form the potent TRPV1 activator AM404 in the central nervous system. We show that acetaminophen is able to reduce the content of prostanoids in brain and kidney, but this effect is independent of FAAH. While ibuprofen produces a similar reduction as acetaminophen of the PGE2 level in brain, only acetaminophen displays antinociceptive activity in animal tests of acute non-inflammatory pain, indicating that other mechanisms than central cyclooxygenase inhibition must be sought to explain the analgesic effect of acetaminophen in these tests. Knowledge of the mechanisms of action of acetaminophen may help to develop analogs with improved analgesic activity. Many TRP ion channels, including TRPV1, are regulated by PLC-coupled surface receptors. After PLC-dependent cleavage of phosphatidylinositol bisphosphate into diacylglycerol (DAG) and inositoltrisphosphate, diacylglycerol may be further metabolized to monoacyl-glycerols, such as 2-arachidonoylglycerol (2-AG) and 2-oleoylglycerol by the action of DAG lipase. 2-Arachidonoylglycerol and 1-AG activate both native TRPV1 on sensory nerve fibers in rodent mesenteric arteries and heterologously expressed rat and human TRPV1. The effects of 2-AG and the metabolically stable analog noladin ether were almost absent in TRPV1 gene-deficient mice. Stimulation of isolated rat dorsal root ganglia with the inflammatory mediators bradykinin and ATP led to an increase in the level of 2-AG, whereas the levels of anandamide and 2-oleoylglycerol were unaffected. Extensive metabolism of d8-2-AG and d5-1-AG, in contrast to d8-anandamide, was demonstrated in homogenates of rat mesenteric arteries. The monoacylglycerol lipase inhibitor MAFP prevented the metabolism of these lipids and enhanced the TRPV1-mediated vascular effects of 2-AG and 1-AG, but not anandamide. The existence of a regulated biosynthesis and enzymatic degradation of 2-AG and 1-AG in TRPV1-containing tissues is compatible with a physiological role for these monoacylglycerols as messengers in the PLC/TRPV1 signaling cascade. Bioactive lipids, with TRPV1 activity, can be formed via drug metabolism and after stimulation of PLC-coupled surface receptors.
Medical and Health Sciences, Cannabinoids, Anandamide, AM404, 2-Arachidonoylglycerol, Acetaminophen, Cyclooxygenase, Nocicieption, TRPV1
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Clinical Chemistry and Pharmacology, Dept of Laboratory Medicine