title: Orexin-stimulated MAP kinase cascades are activated through multiple G-protein signalling pathways in human H295R adrenocortical cells: diverse roles for orexins A and B.
abstract: Orexins A and B (ORA and ORB) are neuropeptide hormones found throughout the central nervous system and periphery. They are required for a host of physiological processes including mitogen-activated protein kinase (MAPK) regulation, steroidogenesis, appetite control and energy regulation. While some signalling mechanisms have been proposed for individual recombinant orexin receptors in generic mammalian cell types, it is clear that the peripheral effects of orexin are spatially and temporally complex. This study dissects the different G-protein signalling and MAPK pathways activated in a pluripotent human adrenal H295R cell line capable of all the physiological steps involved in steroidogenesis. Both extracellular receptor kinase 1/2 (ERK1/2) and p38 were phosphorylated rapidly with a subsequent decline, in a time- and dose-dependent manner, in response to both ORA and ORB. Conversely, there was little or no direct activation of the ERK5 or JNK pathway. Analysis using signalling and MAPK inhibitors as well as receptor-specific antagonists determined the precise mediators of the orexin response in these cells. Both ERK1/2 and p38 activation were predominantly G(q)- and to a lesser extent G(s)-mediated; p38 activation even had a small G(i)-component. Effects were broadly comparable for both orexin sub-types ORA and ORB and although most of the effects were transmitted through the orexin receptor-1 subtype, we did observe a role for orexin receptor-2-mediated activation of both ERK1/2 and p38. Cortisol secretion also differed in response to ORA and ORB. These data suggest multiple roles for orexin-mediated MAPK activation in an adrenal cell-line, this complexity may help to explain the diverse biological actions of orexins with wide-ranging consequences for our understanding of the mechanisms initiated by these steroidogenic molecules.
authors.affiliation: Warwick Medical School, Warwick University, Gibbet Hill Road, Coventry, UK.
authors.forename: Manjunath
authors.initials: M
authors.lastname: Ramanjaneya
authors.forename: Alex C
authors.initials: AC
authors.lastname: Conner
authors.forename: Jing
authors.initials: J
authors.lastname: Chen
authors.forename: Prashanth
authors.initials: P
authors.lastname: Kumar
authors.forename: James E P
authors.initials: JE
authors.lastname: Brown
authors.forename: Olaf
authors.initials: O
authors.lastname: Jöhren
authors.forename: Hendrik
authors.initials: H
authors.lastname: Lehnert
authors.forename: Peter R
authors.initials: PR
authors.lastname: Stanfield
authors.forename: Harpal S
authors.initials: HS
authors.lastname: Randeva
mesh_terms: D000302:Adrenal Cortex / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D001583:Benzoxazoles / Q000008:administration & dosage; D015153:Blotting, Western; D002460:Cell Line; D004305:Dose-Response Relationship, Drug; D004789:Enzyme Activation / Q000187:drug effects; D019204:GTP-Binding Proteins / Q000378:metabolism*; D006801:Humans; D006854:Hydrocortisone / Q000378:metabolism; D047908:Intracellular Signaling Peptides and Proteins / Q000008:administration & dosage / Q000494:pharmacology*; D020928:Mitogen-Activated Protein Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism*; D009287:Naphthyridines; D009479:Neuropeptides / Q000008:administration & dosage / Q000494:pharmacology*; D064446:Orexin Receptors; D000068797:Orexins; D020033:Protein Isoforms / Q000008:administration & dosage / Q000494:pharmacology; D043562:Receptors, G-Protein-Coupled / Q000037:antagonists & inhibitors; D018013:Receptors, Neuropeptide / Q000037:antagonists & inhibitors; D015398:Signal Transduction / Q000502:physiology*; D014508:Urea / Q000008:administration & dosage / Q000031:analogs & derivatives; D048051:p38 Mitogen-Activated Protein Kinases / Q000378:metabolism
chemical_list: C420062:1-(2-methylbenzoxazol-6-yl)-3-(1,5)naphthyridin-4-yl urea; D001583:Benzoxazoles; D047908:Intracellular Signaling Peptides and Proteins; D009287:Naphthyridines; D009479:Neuropeptides; D064446:Orexin Receptors; D000068797:Orexins; D020033:Protein Isoforms; D043562:Receptors, G-Protein-Coupled; D018013:Receptors, Neuropeptide; D014508:Urea; D020928:Mitogen-Activated Protein Kinases; D048051:p38 Mitogen-Activated Protein Kinases; D019204:GTP-Binding Proteins; D006854:Hydrocortisone