title: OX1 orexin/hypocretin receptor signaling through arachidonic acid and endocannabinoid release.
abstract: We showed previously that OX(1) orexin receptor stimulation produced a strong (3)H overflow response from [(3)H]arachidonic acid (AA)-labeled cells. Here we addressed this issue with a novel set of tools and methods, to distinguish the enzyme pathways responsible for this response. CHO-K1 cells heterologously expressing human OX(1) receptors were used as a model system. By using selective pharmacological inhibitors, we showed that, in orexin-A-stimulated cells, the AA-derived radioactivity was released as two distinct components, i.e., free AA and the endocannabinoid 2-arachidonoyl glycerol (2-AG). Two orexin-activated enzymatic cascades are responsible for this response: cytosolic phospholipase A(2) (cPLA(2)) and diacylglycerol lipase; the former cascade is responsible for part of the AA release, whereas the latter is responsible for all of the 2-AG release and part of the AA release. Essentially only diacylglycerol released by phospholipase C but not by phospholipase D was implicated as a substrate for 2-AG production, although both phospholipases were strongly activated. The 2-AG released acted as a potent paracrine messenger through cannabinoid CB(1) receptors in an artificial cell-cell communication assay that was developed. The cPLA(2) cascade, in contrast, was involved in the activation of orexin receptor-operated Ca(2+) influx. 2-AG was also released upon OX(1) receptor stimulation in recombinant HEK-293 and neuro-2a cells. The results directly show, for the first time, that orexin receptors are able to generate potent endocannabinoid signals in addition to arachidonic acid signals, which may explain the proposed orexin-cannabinoid interactions (e.g., in neurons).
authors.affiliation: Biochemistry and Cell Biology, Department of Veterinary Biosciences, University of Helsinki, Helsinki, Finland.
authors.forename: Pauli M
authors.initials: PM
authors.lastname: Turunen
authors.forename: Maria H
authors.initials: MH
authors.lastname: Jäntti
authors.forename: Jyrki P
authors.initials: JP
authors.lastname: Kukkonen
mesh_terms: D000818:Animals; D016718:Arachidonic Acid / Q000378:metabolism*; D001583:Benzoxazoles / Q000494:pharmacology; D016466:CHO Cells; D063385:Cannabinoid Receptor Modulators / Q000378:metabolism*; D006224:Cricetinae; D003412:Cricetulus; D063388:Endocannabinoids; D057809:HEK293 Cells; D006801:Humans; D009287:Naphthyridines; D064446:Orexin Receptors; D043562:Receptors, G-Protein-Coupled / Q000037:antagonists & inhibitors / Q000502:physiology*; D018013:Receptors, Neuropeptide / Q000037:antagonists & inhibitors / Q000502:physiology*; D015398:Signal Transduction / Q000187:drug effects / Q000502:physiology*; D014508:Urea / Q000031:analogs & derivatives / Q000494:pharmacology
chemical_list: C420062:1-(2-methylbenzoxazol-6-yl)-3-(1,5)naphthyridin-4-yl urea; D001583:Benzoxazoles; D063385:Cannabinoid Receptor Modulators; D063388:Endocannabinoids; D009287:Naphthyridines; D064446:Orexin Receptors; D043562:Receptors, G-Protein-Coupled; D018013:Receptors, Neuropeptide; D016718:Arachidonic Acid; D014508:Urea