prostaglandin-d2 and 15-hydroperoxy-5-8-11-13-eicosatetraenoic-acid

Capsaicin, a pungent ingredient of hot peppers, causes excitation of small sensory neurons, and thereby produces severe pain. A nonselective cation channel activated by capsaicin has been identified in sensory neurons and a cDNA encoding the channel has been cloned recently. However, an endogenous activator of the receptor has not yet been found. In this study, we show that several products of lipoxygenases directly activate the capsaicin-activated channel in isolated membrane patches of sensory neurons. Among them, 12- and 15-(S)-hydroperoxyeicosatetraenoic acids, 5- and 15-(S)-hydroxyeicosatetraenoic acids, and leukotriene B(4) possessed the highest potency. The eicosanoids also activated the cloned capsaicin receptor (VR1) expressed in HEK cells. Prostaglandins and unsaturated fatty acids failed to activate the channel. These results suggest a novel signaling mechanism underlying the pain sensory transduction.

Topics: Animals; Capsaicin; Cell Line; Cells, Cultured; Dinoprostone; Eicosanoids; Ganglia, Spinal; Humans; Hydroxyeicosatetraenoic Acids; Inflammation; Ion Channel Gating; Leukotriene B4; Leukotrienes; Ligands; Lipid Peroxides; Lipoxygenase; Molecular Structure; Neurons, Afferent; Prostaglandin D2; Prostaglandin H2; Prostaglandins H; Rats; Receptors, Drug; Structure-Activity Relationship

The oxidation of glutathione to a thiyl radical by prostaglandin H synthase was investigated. Ram seminal vesicle microsomes, in the presence of arachidonic acid, oxidized glutathione to its thiyl-free radical metabolite, which was detected by ESR using the spin trap 5,5-dimethyl-1-pyrroline-N-oxide. Oxidation of glutathione was dependent on arachidonic acid and inhibited by indomethacin. Peroxides also supported oxidation, indicating that the oxidation was by prostaglandin hydroperoxidase. Glutathione served as a reducingcofactor for the reduction of 15-hydroperoxy-5,8,11,13-eicosatetraenoic acid to 15-hydroxy-5,8,11,13-eicosatetraenoic acid at 1.5-2 times the nonenzymatic rate. Although purified prostaglandin H synthase in the presence of either H2O2 or 15-hydroperoxy-5,8,11,13-eicosatetraenoic acid oxidized glutathione to a thiyl radical, arachidonic acid did not support glutathione oxidation. Glutathione also inhibited cyclooxygenase activity as determined by measuring oxygen incorporation into arachidonic acid. Reverse-phase high pressure liquid chromatography analysis of the arachidonic acid metabolites indicated that the presence of glutathione in an incubation altered the metabolite profile. In the absence of the cofactor, the metabolites were PGD2, PGE2, and 15-hydroperoxy-PGE2 (where PG indicates prostaglandin), while in the presence of glutathione, the only metabolite was PGE2. These results indicate that glutathione not only serves as a cofactor for prostaglandin E isomerase but is also a reducing cofactor for prostaglandin H hydroperoxidase. Assuming that glutathione thiyl-free radical observed in the trapping experiments is involved in the enzymatic reduction of 15-hydroperoxy-5,8,11,13-eicosatetraenoic acid to 15-hydroxy-5,8,11,13-eicosatetraenoic acid, then a 1-electron donation from glutathione to prostaglandin hydroperoxidase is indicated.

Topics: Animals; Arachidonic Acid; Arachidonic Acids; Dinoprostone; Electron Spin Resonance Spectroscopy; Free Radicals; Glutathione; Hydroxyeicosatetraenoic Acids; Indomethacin; Leukotrienes; Lipid Peroxides; Male; Microsomes; Oxidation-Reduction; Peroxidases; Peroxides; Prostaglandin D2; Prostaglandin-Endoperoxide Synthases; Prostaglandins D; Prostaglandins E; Seminal Vesicles; Sheep