title: Antioxidants protect calsequestrin-1 knockout mice from halothane- and heat-induced sudden death.
abstract: BACKGROUND
  Mice lacking calsequestrin-1 (CASQ1-null), a Ca-binding protein that modulates the activity of Ca release in the skeletal muscle, exhibit lethal hypermetabolic episodes that resemble malignant hyperthermia in humans when exposed to halothane or heat stress.
  
  
  METHODS
  Because oxidative species may play a critical role in malignant hyperthermia crises, we treated CASQ1-null mice with two antioxidants, N-acetylcysteine (NAC, Sigma-Aldrich, Italy; provided ad libitum in drinking water) and (±)-6-hydroxy-2,5,7,8-tetramethylchromane-2-carboxylic acid (Trolox, Sigma-Aldrich; administered by intraperitoneal injection), before exposure to halothane (2%, 1 h) or heat (41°C, 1 h).
  
  
  RESULTS
  NAC and Trolox significantly protected CASQ1-null mice from lethal episodes, with mortality being 79% (n = 14), 25% (n = 16), and 20% (n = 5) during halothane exposure and 86% (n = 21), 29% (n = 21), and 33% (n = 6) during heat stress in untreated, NAC-treated, and Trolox-treated mice, respectively. During heat challenge, an increase in core temperature in CASQ1-null mice (42.3° ± 0.1°C, n=10) was significantly reduced by both NAC and Trolox (40.6° ± 0.3°C, n = 6 and 40.5° ± 0.2°C, n = 6). NAC treatment of CASQ1-null muscles/mice normalized caffeine sensitivity during in vitro contracture tests, Ca transients in single fibers, and significantly reduced the percentage of fibers undergoing rhabdomyolysis (37.6 ± 2.5%, 38/101 fibers in 3 mice; 11.6 ± 1.1%, 21/186 fibers in 5 mice). The protective effect of antioxidant treatment likely resulted from mitigation of oxidative stress, because NAC reduced mitochondrial superoxide production, superoxide dismutase type-1 expression, and 3-nitrotyrosine expression, and increased both reduced glutathione and reduced glutathione/oxidized glutathione ratio.
  
  
  CONCLUSIONS
  These studies provide a deeper understanding of the mechanisms that underlie hyperthermic crises in CASQ1-deficient muscle and demonstrate that antioxidant pretreatment may prevent them.
authors.affiliation: From the Center for Research on Ageing and Department of Neuroscience, Imaging and Clinical Sciences, University G. d'Annunzio of Chieti, Chieti, Italy (A.M., C.P., L.P., A.D.M., F.P.); Department of Biomedical Sciences, University of Padova, Padova, Italy (M.C., C.R.); and Department of Pharmacology and Physiology, University of Rochester Medical Center, Rochester, New York (L.W.-L., R.T.D.).
authors.forename: Antonio
authors.initials: A
authors.lastname: Michelucci
authors.forename: Cecilia
authors.initials: C
authors.lastname: Paolini
authors.forename: Marta
authors.initials: M
authors.lastname: Canato
authors.forename: Lan
authors.initials: L
authors.lastname: Wei-Lapierre
authors.forename: Laura
authors.initials: L
authors.lastname: Pietrangelo
authors.forename: Alessandro
authors.initials: A
authors.lastname: De Marco
authors.forename: Carlo
authors.initials: C
authors.lastname: Reggiani
authors.forename: Robert T
authors.initials: RT
authors.lastname: Dirksen
authors.forename: Feliciano
authors.initials: F
authors.lastname: Protasi
mesh_terms: D018685:Anesthetics, Inhalation / Q000633:toxicity*; D000818:Animals; D000975:Antioxidants / Q000627:therapeutic use*; D002135:Calcium-Binding Proteins / Q000172:deficiency*; D002155:Calsequestrin; D003645:Death, Sudden / Q000517:prevention & control*; D006221:Halothane / Q000633:toxicity*; D006358:Hot Temperature / Q000009:adverse effects*; D008297:Male; D051379:Mice; D018345:Mice, Knockout
chemical_list: D018685:Anesthetics, Inhalation; D000975:Antioxidants; D002135:Calcium-Binding Proteins; D002155:Calsequestrin; C503130:Casq1 protein, mouse; D006221:Halothane