title: S-nitrosoglutathione inhibits alpha1-adrenergic receptor-mediated vasoconstriction and ligand binding in pulmonary artery.
abstract: Endogenous nitric oxide donor compounds (S-nitrosothiols) contribute to low vascular tone by both cGMP-dependent and -independent pathways. We have reported that S-nitrosoglutathione (GSNO) inhibits 5-hydroxytryptamine (5-HT)-mediated pulmonary vasoconstriction via a cGMP-independent mechanism likely involving S-nitrosylation of its G protein-coupled receptor (GPCR) system. Because catecholamines, like 5-HT, constrict lung vessels via a GPCR coupled to G(q), we hypothesized that S-nitrosothiols modify the alpha1-adrenergic GPCR system to inhibit pulmonary vasoconstriction by receptor agonists, e.g., phenylephrine (PE). Rat pulmonary artery rings were pretreated for 30 min with and without an S-nitrosothiol, either GSNO or S-nitrosocysteine (CSNO), and constricted with sequential concentrations of PE (10(-8)-10(-6) M). Effective cGMP-dependence was tested in rings pretreated with soluble guanylate cyclase inhibitors {either 1H-[1,2,4]oxadiazolo[4,3-a]quinoxalin-1-one (ODQ) or LY-83583} or G kinase inhibitor (KT-5823), and a thiol reductant [dithiothreitol (DTT)] was used to test reversibility of S-nitrosylation. Both S-nitrosothiols attenuated the PE dose response. The GSNO effect was not prevented by LY-83583, ODQ, or KT-5823, indicating cGMP independence. GSNO inhibition was reversed by DTT, consistent with S-nitrosylation or other GSNO-mediated cysteine modifications. In CSNO-treated lung protein, the alpha1-adrenergic receptor was shown to undergo S-nitrosylation in vitro using a biotin switch assay. Studies of alpha1-adrenergic receptor subtype expression and receptor density by saturation binding with 125I-HEAT showed that GSNO decreased alpha1-adrenergic receptor density but did not alter affinity for antagonist or agonist. These data demonstrate a novel cGMP-independent mechanism of reversible alpha1-adrenergic receptor inhibition by S-nitrosothiols.
authors.affiliation: Department of Pediatrics, University of Colorado Health Science Center, Denver, CO 80262, USA. eva.grayck@uchsc.edu
authors.forename: Eva
authors.initials: E
authors.lastname: Nozik-Grayck
authors.forename: Erin J
authors.initials: EJ
authors.lastname: Whalen
authors.forename: Jonathan S
authors.initials: JS
authors.lastname: Stamler
authors.forename: Timothy J
authors.initials: TJ
authors.lastname: McMahon
authors.forename: Pasquale
authors.initials: P
authors.lastname: Chitano
authors.forename: Claude A
authors.initials: CA
authors.lastname: Piantadosi
mesh_terms: D000316:Adrenergic alpha-Agonists / Q000494:pharmacology; D000818:Animals; D017869:Cyclic GMP-Dependent Protein Kinases / Q000037:antagonists & inhibitors; D004229:Dithiothreitol / Q000494:pharmacology; D004305:Dose-Response Relationship, Drug; D004791:Enzyme Inhibitors / Q000494:pharmacology; D006162:Guanylate Cyclase / Q000037:antagonists & inhibitors; D066298:In Vitro Techniques; D008024:Ligands; D008297:Male; D010656:Phenylephrine / Q000494:pharmacology; D011651:Pulmonary Artery / Q000187:drug effects / Q000378:metabolism / Q000502:physiology*; D051381:Rats; D017207:Rats, Sprague-Dawley; D018340:Receptors, Adrenergic, alpha-1 / Q000378:metabolism / Q000502:physiology*; D026422:S-Nitrosoglutathione / Q000009:adverse effects / Q000494:pharmacology*; D026403:S-Nitrosothiols / Q000494:pharmacology; D014661:Vasoconstriction / Q000187:drug effects* / Q000502:physiology*; D014662:Vasoconstrictor Agents / Q000494:pharmacology; D014665:Vasodilator Agents / Q000009:adverse effects / Q000494:pharmacology*
chemical_list: D000316:Adrenergic alpha-Agonists; D004791:Enzyme Inhibitors; D008024:Ligands; D018340:Receptors, Adrenergic, alpha-1; D026403:S-Nitrosothiols; D014662:Vasoconstrictor Agents; D014665:Vasodilator Agents; D010656:Phenylephrine; D026422:S-Nitrosoglutathione; D017869:Cyclic GMP-Dependent Protein Kinases; D006162:Guanylate Cyclase; D004229:Dithiothreitol