title: Activation of vanilloid receptor 1 by resiniferatoxin mobilizes calcium from inositol 1,4,5-trisphosphate-sensitive stores.
abstract: 1 Capsaicin and resiniferatoxin (RTX) stimulate Ca2+ influx by activating vanilloid receptor 1 (VR1), a ligand-gated Ca2+ channel on sensory neurones. We investigated whether VR1 activation could also trigger Ca2+ mobilization from intracellular Ca2+ stores. 2 Human VR1-transfected HEK293 cells (hVR1-HEK293) were loaded with Fluo-3 or a mixture of Fluo-4 and Fura Red and imaged on a fluorometric imaging plate reader (FLIPR) and confocal microscope respectively. 3 In Ca2+ -free media, RTX caused a transient elevation in intracellular free Ca2+ concentration in hVR1-HEK293 cells (pEC(50) 6.45+/-0.05) but not in wild type cells. Capsaicin (100 microM) did not cause Ca2+ mobilization under these conditions. 4 RTX-mediated Ca2+ mobilization was inhibited by the VR1 receptor antagonist capsazepine (pIC(50) 5.84+/-0.04), the Ca2+ pump inhibitor thapsigargin (pIC(50) 7.77+/-0.04), the phospholipase C inhibitor U-73122 (pIC(50) 5.35+/-0.05) and by depletion of inositol 1,4,5-trisphosphate-sensitive Ca2+ stores by pretreatment with the acetylcholine-receptor agonist carbachol (20 microM, 2 min). These data suggest that RTX causes Ca2+ mobilization from inositol 1,4,5-trisphosphate-sensitive Ca2+ stores in hVR1-HEK293 cells. 5 In the presence of extracellular Ca2+, both capsaicin-mediated and RTX-mediated Ca2+ rises were attenuated by U-73122 (10 microM, 30 min) and thapsigargin (1 microM, 30 min). We conclude that VR1 is able to couple to Ca2+ mobilization by a Ca2+ dependent mechanism, mediated by capsaicin and RTX, and a Ca2+ independent mechanism mediated by RTX alone.
authors.affiliation: Neurology Centre of Excellence for Drug Discovery, GlaxoSmithKline, New Frontiers Science Park, Third Avenue, Harlow, Essex CM19 5AW, UK. Ian_C_Marshall@gsk.com
authors.forename: Ian C B
authors.initials: IC
authors.lastname: Marshall
authors.forename: Davina E
authors.initials: DE
authors.lastname: Owen
authors.forename: Tim V
authors.initials: TV
authors.lastname: Cripps
authors.forename: John B
authors.initials: JB
authors.lastname: Davis
authors.forename: Shaun
authors.initials: S
authors.lastname: McNulty
authors.forename: Darren
authors.initials: D
authors.lastname: Smart
mesh_terms: D002118:Calcium / Q000378:metabolism* / Q000502:physiology; D020013:Calcium Signaling / Q000187:drug effects / Q000502:physiology; D002460:Cell Line; D004224:Diterpenes / Q000494:pharmacology*; D004305:Dose-Response Relationship, Drug; D006801:Humans; D015544:Inositol 1,4,5-Trisphosphate / Q000378:metabolism*; D011955:Receptors, Drug / Q000819:agonists / Q000037:antagonists & inhibitors / Q000378:metabolism*; D050916:TRPV Cation Channels
chemical_list: D004224:Diterpenes; D011955:Receptors, Drug; D050916:TRPV Cation Channels; C494484:TRPV1 receptor; D015544:Inositol 1,4,5-Trisphosphate; C024353:resiniferatoxin; D002118:Calcium