title: Isoflurane selectively inhibits distal mitochondrial complex I in Caenorhabditis elegans.
abstract: BACKGROUND
  Complex I of the electron transport chain (ETC) is a possible target of volatile anesthetics (VAs). Complex I enzymatic activities are inhibited by VAs, and dysfunction of complex I can lead to hypersensitivity to VAs in worms and in people. Mutant analysis in Caenorhabditis (C.) elegans suggests that VAs may specifically interfere with complex I function at the binding site for its substrate ubiquinone. We hypothesized that isoflurane inhibits electron transport by competing with ubiquinone for binding to complex I.
  
  
  METHODS
  Wildtype and mutant C. elegans were used to study the effects of isoflurane on isolated mitochondria. Enzymatic activities of the ETC were assayed and dose-response curves determined using established techniques. Two-dimensional native gels of mitochondrial proteins were performed after exposure of mitochondria to isoflurane.
  
  
  RESULTS
  Complex I is the most sensitive component of the ETC to isoflurane inhibition; however, the proximal portion of complex I (the flavoprotein) is relatively insensitive to isoflurane. Isoflurane and quinone do not compete for a common binding site on complex I. The absolute rate of complex I enzymatic activity in vitro does not predict immobilization of the animal by isoflurane. Isoflurane had no measurable effect on stability of mitochondrial supercomplexes. Reduction of ubiquinone by complex I displayed positive cooperative kinetics not disrupted by isoflurane.
  
  
  CONCLUSIONS
  Isoflurane directly inhibits complex I at a site distal to the flavoprotein subcomplex. However, we have excluded our original hypothesis that isoflurane and ubiquinone compete for a common hydrophobic binding site on complex I. In addition, immobilization of the nematode by isoflurane is not due to limiting absolute amounts of complex I electron transport as measured in isolated mitochondria.
authors.affiliation: Department of Anesthesiology, University of Washington and Center for Developmental Therapeutics, Seattle Children's Research Institute, Seattle, WA 98101-1304, USA. philip.morgan@seattlechildrens.org
authors.forename: Ernst-Bernhard
authors.initials: EB
authors.lastname: Kayser
authors.forename: Wichit
authors.initials: W
authors.lastname: Suthammarak
authors.forename: Phil G
authors.initials: PG
authors.lastname: Morgan
authors.forename: Margaret M
authors.initials: MM
authors.lastname: Sedensky
mesh_terms: D018685:Anesthetics, Inhalation / Q000494:pharmacology*; D000818:Animals; D001665:Binding Sites; D017173:Caenorhabditis elegans; D029742:Caenorhabditis elegans Proteins / Q000378:metabolism; D045304:Cytochromes c / Q000378:metabolism; D004579:Electron Transport; D042967:Electron Transport Complex I / Q000378:metabolism*; D015180:Electrophoresis, Gel, Two-Dimensional / Q000379:methods; D004591:Electrophoresis, Polyacrylamide Gel; D006736:Horses; D057927:Hydrophobic and Hydrophilic Interactions; D007530:Isoflurane / Q000737:chemistry / Q000494:pharmacology*; D007700:Kinetics; D008928:Mitochondria / Q000378:metabolism; D009154:Mutation; D009245:NADH Dehydrogenase / Q000378:metabolism; D012995:Solubility; D014451:Ubiquinone / Q000737:chemistry; D014867:Water / Q000737:chemistry
chemical_list: D018685:Anesthetics, Inhalation; D029742:Caenorhabditis elegans Proteins; D014867:Water; D014451:Ubiquinone; D045304:Cytochromes c; D007530:Isoflurane; D009245:NADH Dehydrogenase; D042967:Electron Transport Complex I; C468009:GAS-1 protein, C elegans