title: Sphingosylphosphorylcholine potentiates vasoreactivity and voltage-gated Ca2+ entry via NOX1 and reactive oxygen species.
abstract: OBJECTIVE
  Sphingosylphosphorylcholine (SPC) elicits vasoconstriction at micromolar concentrations. At lower concentrations (≤1 µmol/L), however, it does not constrict intrapulmonary arteries (IPAs), but strongly potentiates vasoreactivity. Our aim was to determine whether this also occurs in a systemic artery and to delineate the signalling pathway.
  
  
  RESULTS
  Rat mesenteric arteries and IPAs mounted on a myograph were challenged with ∼25 mmol/L [K+] to induce a small vasoconstriction. SPC (1 µmol/L) dramatically potentiated this constriction in all arteries by ∼400%. The potentiation was greatly suppressed or abolished by inhibition of phospholipase C (PLC; U73122), PKCε (inhibitory peptide), Src (PP2), and NADPH oxidase (VAS2870), and also by Tempol (superoxide scavenger), but not by inhibition of Rho kinase (Y27632). Potentiation was lost in mesenteric arteries from p47(phox-/-), but not NOX2(-/-), mice. The intracellular superoxide generator LY83583 mimicked the effect of SPC. SPC elevated reactive oxygen species (ROS) in vascular smooth muscle cells, and this was blocked by PP2, VAS2870, and siRNA knockdown of PKCε. SPC (1 µmol/L) significantly reduced the EC50 for U46619-induced vasoconstriction, an action ablated by Tempol. In patch-clamped mesenteric artery cells, SPC (200 nmol/L) enhanced Ba2+ current through L-type Ca2+ channels, an action abolished by Tempol but mimicked by LY83583.
  
  
  CONCLUSIONS
  Our results suggest that low concentrations of SPC activate a PLC-coupled and NOX1-mediated increase in ROS, with consequent enhancement of voltage-gated Ca2+ entry and thus vasoreactivity. We speculate that this pathway is not specific for SPC, but may also contribute to vasoconstriction elicited by other G-protein coupled receptor and PLC-coupled agonists.
authors: Division of Asthma, Allergy, and Lung Biology, King's College London, 5th Floor Tower Wing, Guy's Campus, London SE1 9RT, UK.
authors: Yasin
authors: Y
authors: Shaifta
authors: Division of Asthma, Allergy, and Lung Biology, King's College London, 5th Floor Tower Wing, Guy's Campus, London SE1 9RT, UK.
authors: Vladimir A
authors: VA
authors: Snetkov
authors: Division of Asthma, Allergy, and Lung Biology, King's College London, 5th Floor Tower Wing, Guy's Campus, London SE1 9RT, UK.
authors: Jesus
authors: J
authors: Prieto-Lloret
authors: Division of Asthma, Allergy, and Lung Biology, King's College London, 5th Floor Tower Wing, Guy's Campus, London SE1 9RT, UK.
authors: Greg A
authors: GA
authors: Knock
authors: Department of Pharmacy and Pharmacology, University of Bath, Bath, UK.
authors: Sergey V
authors: SV
authors: Smirnov
authors: Division of Asthma, Allergy, and Lung Biology, King's College London, 5th Floor Tower Wing, Guy's Campus, London SE1 9RT, UK.
authors: Philip I
authors: PI
authors: Aaronson
authors: Division of Asthma, Allergy, and Lung Biology, King's College London, 5th Floor Tower Wing, Guy's Campus, London SE1 9RT, UK jeremy.ward@kcl.ac.uk.
authors: Jeremy P T
authors: JP
authors: Ward
mesh_terms: D000818:Animals; D015220:Calcium Channels / Q000187:drug effects* / Q000502:physiology; D003497:Cyclic N-Oxides / Q000494:pharmacology; D004305:Dose-Response Relationship, Drug; D008562:Membrane Glycoproteins / Q000172:deficiency / Q000235:genetics / Q000502:physiology; D008638:Mesenteric Arteries / Q000187:drug effects / Q000502:physiology*; D051379:Mice; D018345:Mice, Knockout; D023421:Models, Animal; D009247:NADH, NADPH Oxidoreductases / Q000502:physiology*; D000074624:NADPH Oxidase 1; D000074662:NADPH Oxidase 2; D019255:NADPH Oxidases / Q000172:deficiency / Q000235:genetics / Q000494:pharmacology / Q000502:physiology; D010767:Phosphorylcholine / Q000031:analogs & derivatives* / Q000494:pharmacology; D051744:Protein Kinase C-epsilon / Q000494:pharmacology; D011651:Pulmonary Artery / Q000187:drug effects / Q000502:physiology*; D017382:Reactive Oxygen Species / Q000378:metabolism*; D015398:Signal Transduction / Q000187:drug effects / Q000502:physiology; D013110:Sphingosine / Q000031:analogs & derivatives* / Q000494:pharmacology; D013113:Spin Labels; D010738:Type C Phospholipases / Q000494:pharmacology; D014661:Vasoconstriction / Q000187:drug effects* / Q000502:physiology
chemical_list: D015220:Calcium Channels; D003497:Cyclic N-Oxides; D008562:Membrane Glycoproteins; D017382:Reactive Oxygen Species; D013113:Spin Labels; C005356:sphingosine phosphorylcholine; D010767:Phosphorylcholine; D009247:NADH, NADPH Oxidoreductases; C488842:Cybb protein, mouse; D000074624:NADPH Oxidase 1; D000074662:NADPH Oxidase 2; D019255:NADPH Oxidases; C000614711:NOX1 protein, mouse; C000614712:NOX1 protein, rat; C060505:neutrophil cytosolic factor 1; D051744:Protein Kinase C-epsilon; D010738:Type C Phospholipases; D013110:Sphingosine; C001803:tempol
keywords: L-type Ca2+ channels; NADPH oxidase; Protein kinase C epsilon; Reactive oxygen species; Vascular smooth muscle