title: Effects of mitochondrial inhibitors and uncouplers on hypoxic vasoconstriction in rabbit lungs.
abstract: Hypoxic pulmonary vasoconstriction (HPV) matches lung perfusion to ventilation for optimizing pulmonary gas exchange; however, the underlying mechanism has not yet been fully elucidated. Lung nitric oxide (NO) generation appears to be involved in this process. Recently, mitochondria have been proposed as oxygen sensors, with HPV signaling via a hypoxia-induced increase in the generation of reactive oxygen species derived from mitochondrial complex III and escaping through an anion channel into the cytoplasm. In addition, complex II has been suggested to be specifically involved in hypoxia-dependent generation of reactive oxygen species in the lung. We investigated the effects of several mitochondrial inhibitors and uncouplers on the strength of HPV, and asked for their capacity to mimic HPV during normoxia in isolated buffer-perfused rabbit lungs. Specificity of the agents for HPV was tested by comparison of their effects on non-hypoxia-induced vasoconstriction, elicited by the thromboxane mimetic U46619. Interference with NO metabolism was determined by performing parallel studies with blocked lung NO generation and by measurement of exhaled NO. Rotenone, 3-nitroproprionic acid, and myxothiazol dose-dependently inhibited HPV without being mimics of HPV during normoxia. The inhibitory effect of these agents was only partly specific for HPV by comparison with U46619-induced vasoconstriction. During pre-blocked lung NO synthesis, the selectivity for HPV inhibition was increased for rotenone, but largely lost for myxothiazol. 2-tenoyltrifluoroacetone resulted in an unspecific inhibition of HPV as compared with U46619-induced vasoconstriction. 1-methyl-4-phenylpyridinium iodide and 2-heptyl-4-hydroxyquinoline-N-oxide specifically suppressed HPV and increased normoxic vascular tone. Antimycin A suppressed HPV, an effect being specific in lungs with intact NO synthesis and only partly specific while blocking NO. However, this agent did not mimic HPV during normoxia, as may be expected for interference with the mitochondrial electron transport downstream in complex III. The uncouplers 2,4-dinitrophenol (DNP, 10-200 microM) and carbonyl cyanide 4-(trifluoromethoxy)phenylhydrazone (FCCP, 1-3 microM) induced sustained vasoconstriction during normoxia, with enhancement of HPV by DNP at low and suppression of HPV for both agents at high concentrations. The anion channel blocker 4,4'-diisothiocyanatostilbene-2,2'-disulfonic acid inhibited HPV and U46619-induced vasoconstriction with identical dose-response curves. These findings suggest that mitochondria are in some manner involved in the regulation of HPV in intact rabbit lungs. The hypothesis that enhanced superoxide leak at complex III of mitochondria represents the underlying mechanism of acute HPV is supported by the rotenone and 2-heptyl-4-hydroxyquinoline-N-oxide data, but partly contradicted by the findings with 1-methyl-4-phenylpyridinium iodide, antimycin A, DNP, and FCCP. Further studies are mandatory to clarify the link between mitochondrial respiratory chain and hypoxic pulmonary vasoconstriction.
authors.affiliation: Department of Internal Medicine, Justus-Liebig-University Giessen, Klinikstrasse 36, 35392 Giessen, Germany. Norbert.Weissmann@innere.med.uni-giessen.de
authors.forename: Norbert
authors.initials: N
authors.lastname: Weissmann
authors.forename: Nadine
authors.initials: N
authors.lastname: Ebert
authors.forename: Marit
authors.initials: M
authors.lastname: Ahrens
authors.forename: Hossein A
authors.initials: HA
authors.lastname: Ghofrani
authors.forename: Ralph T
authors.initials: RT
authors.lastname: Schermuly
authors.forename: Jörg
authors.initials: J
authors.lastname: Hänze
authors.forename: Ludger
authors.initials: L
authors.lastname: Fink
authors.forename: Frank
authors.initials: F
authors.lastname: Rose
authors.forename: Jörg
authors.initials: J
authors.lastname: Conzen
authors.forename: Werner
authors.initials: W
authors.lastname: Seeger
authors.forename: Friedrich
authors.initials: F
authors.lastname: Grimminger
mesh_terms: D017878:4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid / Q000494:pharmacology; D000818:Animals; D000935:Antifungal Agents / Q000494:pharmacology; D000968:Antimycin A / Q000494:pharmacology; D004305:Dose-Response Relationship, Drug; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D000860:Hypoxia / Q000378:metabolism*; D066298:In Vitro Techniques; D008168:Lung / Q000166:cytology / Q000502:physiology*; D008689:Methacrylates; D008928:Mitochondria / Q000187:drug effects* / Q000378:metabolism*; D009569:Nitric Oxide / Q000378:metabolism; D011817:Rabbits; D012402:Rotenone / Q000494:pharmacology; D013844:Thiazoles / Q000494:pharmacology; D014475:Uncoupling Agents / Q000494:pharmacology*; D014661:Vasoconstriction / Q000502:physiology*; D019323:omega-N-Methylarginine / Q000494:pharmacology
chemical_list: D000935:Antifungal Agents; D004791:Enzyme Inhibitors; D008689:Methacrylates; D013844:Thiazoles; D014475:Uncoupling Agents; D012402:Rotenone; D019323:omega-N-Methylarginine; D009569:Nitric Oxide; D000968:Antimycin A; C030517:myxothiazol; D017878:4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid