In Vitro Contracture Test Research Articles

Predictive factors of the contracture test for diagnosing malignant hyperthermia in a Brazilian population sample: a retrospective observational study

IntroductionMalignant Hyperthermia (MH) is a pharmacogenetic, hereditary and autosomal dominant syndrome triggered by halogenates/succinylcholine. The In Vitro Contracture Test (IVCT) is the gold standard diagnostic test for MH, and it evaluates abnormal skeletal muscle reactions of susceptible individuals (earlier/greater contracture) when exposed to caffeine/halothane. MH susceptibility episodes and IVCT seem to be related to individual features. ObjectiveTo assess variables that correlate with IVCT in Brazilian patients referred for MH investigation due to a history of personal/family MH. MethodsWe examined IVCTs of 80 patients investigated for MH between 2004‒2019. We recorded clinical data (age, sex, presence of muscle weakness or myopathy with muscle biopsy showing cores, genetic evaluation, IVCT result) and IVCT features (initial and final maximum contraction, caffeine/halothane concentration triggering contracture of 0.2g, contracture at caffeine concentration of 2 and 32 mmoL and at 2% halothane, and contraction after 100 Hz stimulation). ResultsMean age of the sample was 35±13.3 years, and most of the subjects were female (n=43 or 54%) and MH susceptible (60%). Of the 20 subjects undergoing genetic investigation, 65% showed variants in RYR1/CACNA1S genes. We found no difference between the positive and negative IVCT groups regarding age, sex, number of probands, presence of muscle weakness or myopathy with muscle biopsy showing cores. Regression analysis revealed that the best predictors of positive IVCT were male sex (+12%), absence of muscle weakness (+20%), and personal MH background (+17%). ConclusionsPositive IVCT results have been correlated to male probands, in accordance with early publications. Furthermore, normal muscle strength has been confirmed as a significant predictor of positive IVCT while investigating suspected MH cases.

Anomalous Kv 7 channel activity in human malignant hyperthermia syndrome unmasks a key role for H2 S and persulfidation in skeletal muscle.

Human malignant hyperthermia (MH) syndrome is induced by volatile anaesthetics and involves increased levels of cystathionine β-synthase (CBS)-derived H2 S within skeletal muscle. This increase contributes to skeletal muscle hypercontractility. Kv 7 channels, expressed in skeletal muscle, may be a molecular target for H2 S. Here, we have investigated the role of Kv 7 channels in MH. Skeletal muscle biopsies were obtained from MH-susceptible (MHS) and MH-negative (MHN) patients. Immunohistochemistry, RT-PCR, Western blot, and in vitro contracture test (IVCT) were carried out. Development and characterization of primary human skeletal muscle cells (PHSKMC) and evaluation of cell membrane potential were also performed. The persulfidation state of Kv 7 channels and polysulfide levels were measured. Kv 7 channels were similarly expressed in MHN and MHS biopsies. The IVCT revealed an anomalous contractility of MHS biopsies following exposure to the Kv 7 channel opener retigabine. Incubation of negative biopsies with NaHS, prior to retigabine addition, led to an MHS-like positive response. MHS-derived PHSKMC challenged with retigabine showed a paradoxical depolarizing effect, compared with the canonical hyperpolarizing effect. CBS expression and activity were increased in MHS biopsies, resulting in a major polysulfide bioavailability. Persulfidation of Kv 7.4 channels was significantly higher in MHS than in MHN biopsies. In skeletal muscle of MHS patients, CBS-derived H2 S induced persulfidation of Kv 7 channels. This post-translational modification switches the hyperpolarizing activity into depolarizing. This mechanism can contribute to the pathological skeletal muscle hypercontractility typical of MH syndrome. This article is part of a themed section on Hydrogen Sulfide in Biology & Medicine. To view the other articles in this section visit http://onlinelibrary.wiley.com/doi/10.1111/bph.v177.4/issuetoc.

Masseter muscle rigidity and the role of DNA analysis to confirm malignant hyperthermia susceptibility.

Malignant hyperthermia (MH) is an uncommon, autosomal dominant disorder of skeletal muscle, triggered by inhalational anaesthetics or depolarizing muscle relaxants. Masseter muscle rigidity (MMR) can be regarded as potentially a preceding sign for an MH reaction. Susceptibility to MH can be determined by the in vitro contracture test (IVCT) or DNA analysis where a familial variant is known. Our aims were to review patients with MMR, where IVCT and DNA analysis had been undertaken, to determine if DNA analysis could be used as an initial screening tool for MH susceptibility, and, by reviewing standard monitored variables (SMVs), to determine if any clinical characteristics could be used to differentiate between MMR patients who are MH susceptible (MHS) and those who are not. Patients with MMR were identified from the Palmerston North Hospital MH Reactions Database. IVCT and DNA analysis results were documented. DNA testing was performed retrospectively in the majority of patients as many patients had presented before DNA analysis was available. Forty-one patients were analysed. Fourteen were DNA positive/IVCT positive and six DNA positive only (48% in total), seven were IVCT positive/DNA negative and 14 were IVCT normal. Increased creatine kinase (>18,000 units/L) was consistent with MH susceptibility. Severity of MMR was not linked to MH susceptibility. This study confirmed that DNA analysis can be used as a first-line test for MH susceptibility in patients presenting with MMR (consistent with European MH Group recommendations). Creatine kinase was the only SMV that was significantly different between MHS and MH normal individuals.

Epigenetic responses to exertional heat stroke in mice: a potential link to long term Ca2+ dysregulation in skeletal muscle

Exposure to extreme environments can induce a long‐term epigenetic memory of stress that is stored as altered DNA‐methylation and/or histone posttranslational modifications. These ultimately affect gene and protein expression as well as cellular functions. Exertional heat stroke (EHS) is a life‐threatening stress, but whether it induces epigenetic changes is unknown. Previous studies have suggested long‐term complications of EHS that could be mediated by epigenetics, including greater vulnerability to disease, heat stress susceptibility and disordered muscle Ca2+ signaling; the latter characterized as a malignant‐hyperthermia‐type muscle phenotype, based on the in vitro contracture test (IVCT).PURPOSETo determine whether a single exposure to EHS using a preclinical mouse model induces significant changes in DNA methylation and altered skeletal muscle Ca2+ regulation.METHODSMice were either subjected to a standardized EHS protocol using a forced running wheel at 37.5°C or a matched exercise control trial (22.5 °C), N=4/group. The EHS mice achieved peak core temps of ~42.2 °C, accompanied by transient loss of consciousness. Mice were euthanized after 4 or 30 days. At day 4, cells of a monocyte lineage were isolated from bone marrow, and DNA was extracted and tested using bisulfite DNA sequencing. At 30 days, soleus muscles were excised (N=5/group) and tested for a Ca2+ response to caffeine and halothane (IVCT test).RESULTS>3,000 genes from monocytes were significantly hyper‐ or hypomethylated in EHS mice compared to matched controls. Some genes of physiological interest included Ryr1&2, Cacna1s&c (DHPR isoforms), Orai1 (store operated calcium channel), Camk2d, Pde4d (cAMP signaling), Atp2a1/2, Sod2, Nox4, Nos1&2 isoforms, Hsp72, a number of K+ channels, and isoforms of Mir669a‐1 (inhibitors of skeletal muscle myogenesis). The IVCT displayed an abnormal response in solei from EHS mice (3/5) vs. control (0/5) (p=0.031; adjusted Wald Chi‐Square test). Two mice showed an abnormal response to caffeine; one displayed abnormal responses to both caffeine and halothane.CONCLUSIONSThe data demonstrates that EHS induces striking alterations in the epigenome at 4 days. Most biological and behavioral responses return to normal by this time. DNA methylation profiles from accessible blood cell populations may prove to be a useful biomarker for severity of EHS exposures. The IVCT findings mirror the incidence in military EHS populations. Coupled with the epigenetic changes observed in Ca2+‐related genes, studying skeletal muscle Ca2+ regulation may prove a promising target for future discovery.Support or Funding InformationBK and Betty Stevens Endowment and DOD Grant W81XWH‐15‐2‐0038. Disclaimers: Research was conducted in compliance with the Animal Welfare Act, and all other Federal requirements. The views expressed are those of the authors and do not constitute endorsement by the U.S. Army.This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Cross-Influence of Halothane and Voltage on Intracellular Ca2+ in Isolated Muscle Fibers of Mice Expressing Human RyR1 Mutation Y522S

We studied the effect of halothane and its interference with membrane voltage in malignant hyperthermia (MH) susceptible muscle by using enzymatically isolated single fibers (m. interosseus) of knock-in mice with the RyR1 mutation Y524S (human MH mutation Y522S). To investigate anesthetic- and voltage-triggered changes of Ca2+ concentration, we measured fluorescence from fibers loaded with Fura-2-AM. Similar as in the diagnostic in-vitro contracture test (IVCT) for MH susceptibility, cells were exposed to different halothane-containing solutions using a vaporizer system. The percentage of halothane in the gas phase was determined using infrared photometry. The temporal change of the halothane concentration in the recording chamber was checked using gas chromatography. In Krebs-Ringer's solution at room temperature, a step from 0 to 0.5% halothane in the gas phase caused a clear increase in resting Ca2+ concentration in most of the investigated fibers of mutant mice but very little response in fibers of WT littermates. The Ca2+ response was phasic: after reaching a peak the Ca2+ signal decreased at a rate of about 10%/min indicating either reduced sensitivity to halothane or inhibition of Ca2+ release after activation. A likely explanation is Ca2+-induced inactivation of RyR1. In further experiments, we applied halothane to single fibers which were voltage-clamped using two intracellular microelectrodes. Depolarizing pulses to different membrane potentials from a holding potential of −80 mV revealed a strong shift in the voltage threshold for activation of Ca2+ release to more negative potentials. Correspondingly, hyperpolarizing steps led to a rapid partial recovery from the halothane-induced increase in basal Ca2+ concentration. These results demonstrate reciprocal effects of a volatile anesthetic drug and membrane voltage on Ca2+ release in MH-susceptible skeletal muscle.

Cystathionine β-synthase-derived hydrogen sulfide is involved in human malignant hyperthermia.

Hydrogen sulfide is an endogenous gasotransmitter and its mechanism of action involves activation of ATP-sensitive K(+) channels and phosphodiesterase inhibition. As both mechanisms are potentially involved in malignant hyperthermia (MH), in the present study we addressed the involvement of the L-cysteine/hydrogen sulfide pathway in MH. Skeletal muscle biopsies obtained from 25 MH-susceptible (MHS) and 56 MH-negative (MHN) individuals have been used to perform the invitro contracture test (IVCT). Quantitative real-time PCR (qPCR) and Western blotting studies have also been performed. Hydrogen sulfide levels are measured in both tissue samples and plasma. In MHS biopsies an increase in cystathionine β-synthase (CBS) occurs, as both mRNA and protein expression compared with MHN biopsies. Hydrogen sulfide biosynthesis is increased in MHS biopsies (0.128±0.12 compared with 0.943±0.13 nmol/mg of protein per min for MHN and MHS biopsies, respectively; P<0.01). Addition of sodium hydrosulfide (NaHS) to MHS samples evokes a response similar, in the IVCT, to that elicited by either caffeine or halothane. Incubation of MHN biopsies with NaHS, before caffeine or halothane challenge, switches an MHN to an MHS response. In conclusion we demonstrate the involvement of the L-cysteine/hydrogen sulfide pathway in MH, giving new insight into MH molecular mechanisms. This finding has potential implications for clinical care and could help to define less invasive diagnostic procedures.

Is there a link between exertional heat stroke and susceptibility to malignant hyperthermia?

ObjectiveThe identification of a predisposition toward malignant hyperthermia (MH) as a risk factor for exertional heat stroke (EHS) remains a matter of debate. Such a predisposition indicates a causal role for MH susceptibility (MHS) after EHS in certain national recommendations and has led to the use of an in vitro contracture test (IVCT) to identify the MHS trait in selected or unselected EHS patients. The aim of this study was to determine whether the MHS trait is associated with EHS.MethodsEHS subjects in the French Armed Forces were routinely examined for MHS after experiencing an EHS episode. This retrospective study compared the features of IVCT-diagnosed MHS (iMHS) EHS subjects with those of MH-normal EHS patients and MH patients during the 2004–2010 period. MHS status was assessed using the European protocol.ResultsDuring the study period, 466 subjects (median age 25 years; 31 women) underwent MHS status investigation following an EHS episode. None of the subjects reported previous MH events. An IVCT was performed in 454 cases and was diagnostic of MHS in 45.6% of the study population, of MH susceptibility to halothane in 18.5%, of MH susceptibility to caffeine in 9.9%, and of MH susceptibility to halothane and caffeine in 17.2%. There were no differences in the clinical features, biological features or outcomes of iMHS EHS subjects compared with those of MH-normal or caffeine or halothane MHS subjects without known prior EHS episode. The recurrence rate was 12.7% and was not associated with MH status or any clinical or biological features. iMHS EHS patients exhibited a significantly less informative IVCT response than MH patients.ConclusionsThe unexpected high prevalence of the MHS trait after EHS suggested a latent disturbance of calcium homeostasis that accounted for the positive IVCT results. This study did not determine whether EHS patients have an increased risk of MH, and it could not determine whether MH susceptibility is a risk factor for EHS.

Analysis of malignant hyperthermia results in Padua and Siena

Background Malignant Hyperthermia is pharmacogenetic disease triggered by some anaesthetics such as halogenated gases and non depolarizing muscle relaxant. The diagnosis of malignant hyperthermia susceptibility may be done by 1) relevant clinical signs observed during the crisis, 2) the in vitro contracture test (IVCT) with caffeine-halothane according to the European and the American Malignant Hyperthermia Group protocols and/or 3) the search for mutations of RYR 1 and CACNA1S genes. The IVCT identified three categories of patients: a) MHS (malignant hyperthermia susceptible), b) MHE (malignant hyperthermia equivocal) and c) MHN (non susceptible). A gene mutation may be defined as diagnostic only if it can be defined as “causative mutations” capable to alter the protein function in a cell culture test. The patients carrying “no causative” mutations must carry out the IVCT to confirm a susceptibility condition. Usually, the first investigated patients, the probands, have had an abnormal reactions during trigger anaesthesia such as early signs (inappropriately elevated CO2, inappropriate tachycardia and/or arrhythmia and muscle signs like masseter spasm or generalized muscle rigidity) or late signs (rapid increase in core body temperature, high value of CK etc). If the anaesthesia report has been well done it is possible to attribute a Larach Clinical Grading Scale that classifying the reactions as” almost certain” (more than 50 scores) or very rare “almost never” with 0 scores. The susceptible probands may be also genetically studied and the relatives warned of the genetic risk of an autosomal dominant trait. The problems for the physicians may be relevant when the probands have a raw score range from 20 to 34 or more, and the IVCT results is negative (MHN). Although negative patients may be happy to result negative, they remain worried regarding anaesthesia own and of their relatives. The worry for General Anesthesia (GA) does not disappear with a negative test and it may be very difficult explaining that other possible causes may be involved in MH like reactions. In some MHN probands it may be easier to find an explanation, as it is possible an alternative diagnosis or because.... the not clear or wrong anaesthetic report but what about the others. In some of similar cases, we decide to perform a genetic analysis and in one patient MHN patient we found a “no causative mutation”. At this point the question is 1) can be assure the MHN patients regarding GA with trigger agents? 2) Are there other possible investigations to give more assurance to such patients? 3) genetic testing should be prescribed also in the MHN probands? We present the data of our MH laboratories highlighting the IVCT and genetic results obtained on our probands and the suggestions and information that we give to the MHN probands as well as some questions to the auditorium.

Chronic use of low dose simvastatin does not alter the IVCT in normal rats

Background Statins are currently the preferred agent for the treatment of hypercholesterolemia. However, myalgia, increased serum levels of creatinekinase (CK) and even rhabdomyolysis may occur, characterizing the cholesterol lowering agents myopathy (CLAM), with a multifactorial aetiology. In statin monotherapy, the incidence of CLAM is 0.1 to 0.5 % and is dose related. RYR1 (ryanodine 1) gene alteration was observed in the patients with CLAM, an animal model of MH (malignant hyperthermia) showed hyper metabolism after simvastatin, simvastatin induced in vitro contractures in muscle of animal model and susceptibility to MH by IVCT (in vitro contracture test) was diagnosed in patients with CLAM. On the other way, postoperative rhabdomyolysis occurred in patients treated with cholesterol-lowering drugs, leading to the suggestion of suspend these medications prior to surgery or do not use succinylcholine in patients taking these drugs . But this stance is challenged by other groups. Thus, this study examined, in rats chronically exposed to simvastatin dose previously associated with muscle alteration, the outcome of the IVCT.

Update of the quality assurance (QA) results of halothane and caffeine analysis of all current participating malignant hyperthermia units

Background The provision of a Quality Assurance Scheme for members of the European Malignant Hyperthermia Group (EMHG) is to ensure that the concentrations of halothane and caffeine in the tissue bath are as stipulated in the in vitro contracture test (IVCT) protocol. Confirmation of the correct concentrations in the tissue bath is an essential part of the IVCT protocol. The focus of this presentation will be to provide an update of the QA results from the analysis of both halothane and caffeine from all of the participating units over a period of three years. The aims are to show

Abstracts of the 33rd Annual Meeting of the European Malignant Hyperthermia Group (EMHG), May15-17, 2014, Würzburg, Germany.

Background Malignant hyperthermia (MH) is a complication of anaesthesia appearing as an acute potentially lethal hypermetabolic state in people carrying a genetic anomaly expressed in the skeletal muscle. MH susceptibility can be diagnosed by in vitro contracture test (IVCT) with halothane and caffeine requiring muscular biopsy, or by looking for the MH mutations directly in DNA extracted from the blood. Studies showed an influence of the type of mutation (genotype) on the various expressions (phenotypes) of the MH susceptibility. The aims of this study are to look for any correlation between the presence of a MH mutation and 1) the severity of the anesthetic complication and 2) the force of the contractures observed in the IVCT.

A case of prolonged exertional rhabdomyolysis in a MHS individual

Case report A 42-year old healthy Caucasian male contacted our malignant hyperthermia (MH) hotline due to cramps and muscle pain one week after moderate exercise. He was already diagnosed MHS in 1988 by in vitro contracture test (IVCT), after a suspected MH episode during general anaesthesia. At present, he reported severe muscle pain and cramps in both upper legs. The discomfort has started after moderate endurance training with mild muscle soreness for 2 days. On the 3 day, pain increased, accompanied by severe muscle cramps in both upper legs. Symptoms were worsened by cold temperature. Due to pain, muscle weakness and swelling in both upper legs, he had sought medical attention on day 5 in a surgical outpatient clinic, where an ultrasound examination showed a “homogenous increase in muscle density of both quadriceps muscles”. No laboratory tests except d-dimer were performed and after exclusion of an acute thromboembolic event, the patient was dismissed. Due to ongoing pain, he contacted our MH-Hotline 7 days after the exercise. He still felt sick and reported darker urine during the past 3 days. Body temperature was not yet measured. His general practitioner had already taken a blood examination one hour before the telephone call. One day later, laboratory analysis was available and showed a CPK of 39,628 U/l and myoglobin of 2,863 ng/ml, consistent with rhabdomyolysis. Also liver enzymes were elevated, but kidney function and potassium were normal. The patient reported a body temperature of 37.5°C in the morning. Due to the high CPK levels 7 days after exercise (normally, peak levels are expected within 48 to 96 h), we recommended an immediate admission to our unit and decided to start with dantrolene 2.5 mg/kg IV. After the initial dose of dantrolene he was admitted to an intermediate care unit for the first 24h. Dantrolene was not continued since he was clinically stable. No clinical signs of a compartment syndrome could be detected. Pain continuously decreased and laboratory parameters slowly returned to normal within the next 2 weeks except of a CPK of 330 U/l. Body temperature and kidney function remained normal during the whole period. Any other reason for rhabdomyolysis than MHS diagnosis, i.e. viral or bacterial infection, hypothyreosis, medication, drugs or dietary supplements could be ruled out. A MRI examination showed an inhomogenous enhancement of contrast medium and edema of both quadriceps muscles. The patient was dismissed from hospital 3 days later, additional genetic testing has been planned. Unfortunately, previous CPK levels of the patient were unknown and in 1988, when IVCT was performed, histology was not regularly done.

Malignant hyperthermia and beyond - a virtual BioBank

• A more detailed analysis of in vitro contracture test (IVCT) data to better define phenotype for research purposes, and to assess the effects of specific mutations on the range of phenotypes. We have previously conducted an analysis of the effects of a number of ryanodine receptor type 1 (RYR1) mutations on IVCT and MH reaction phenotypes. • We have previously published preliminary work considering the possibility of genetic modifiers, and this is a topical area with the role of common versus rare variants as contributors to complex disorders being hotly debated. It will be possible to initiate small projects on the effects of previously identified single nucleotide variants and SNV-SNV interactions on phenotype. • To analyse observations such as discordancy in greater depth. • To assess the statistical power for large-scale studies such as a Genome Wide Association Scan.

Targeted exon capture and NGS to investigate an undefined myopathy reveal RYR1 variants

Background The family under investigation consists of parents and two daughters, one being the proband. The mother and the proband have elongated facial features. The father and second daughter appear normal. The older daughter presented for elective tonsillectomy aged 8 years. She had severe masseter spasm after suxamethonium. The rest of the procedure was carried out under total intravenous anaesthesia. No blood gas analysis could be done, but a creatine kinase next day was significantly elevated (2934). This led to study of both parents. There was no family history of malignant hyperthermia but an undefined myopathy was suspected in mother and daughter. Both mother and father were diagnosed malignant hyperthermia (MH) susceptible by in vitro contracture test (IVCT). This prompted a DNA analysis for variants associated with MH.

Gene expression in the context of malignant hyperthermia status and ageing

Background This study aims to investigate gene expression in skeletal muscle in relation to age and MH status, focussing on particular genes of interest. Expression patterns have been analysed to address the question of whether major causative malignant hyperthermia (MH) mutations influence skeletal muscle ageing. In this context we have examined gene expression in MH susceptible, both with (MHS+) and without mutation (MHS-), and MH normal (MHN) material from in vitro contracture test (IVCT) muscle biopsies.

RYR1 mutation screening 1992 – 2014: a genetic report on 22 years from the Würzburg MH unit

Materials and methods In 1992, we began to screen MHS individual by PCR and restriction enzyme digestion for the few then known RYR1 mutations. At around 1995, we switched to Sanger sequencing of selected exons (mutation hot-spots) and for a number of years, we analysed five exons in MHS (exons 17, 39, 40, 45, 46) and four in myopathic subjects (exons 95, 100, 101, 102). From 2002 on, we also included whole-gene sequencing of all 106 exons by Sanger or NGS technologies (starting from 2011). In total, we have screened 1.029 unrelated index cases. The great majority of malignant hyperthermia (MH) individuals had a positive in vitro contracture test (IVCT) result (MHS or MH equivocal (MHE)) or a likely clinical MH episode. Patients with congenital myopathies were referred on the basis of their clinical presentation.

Relation between the intensity of the answer to the tests of in vitro contracture test (IVCT) and 31P magnetic resonance spectroscopy realized in 2008–2010 and the clinical, biological data in the military population having presented a heat stroke

Relation between the intensity of the answer to the tests of in vitro contracture test (IVCT) and 31P magnetic resonance spectroscopy realized in 2008–2010 and the clinical, biological data in the military population having presented a heat stroke

Evaluation of suspected malignant hyperthermia events during anesthesia

BackgroundMalignant hyperthermia (MH), a metabolic myopathy triggered by volatile anesthetics and depolarizing muscle relaxants, is a potentially lethal complication of general anesthesia in susceptible patients. The implementation of modern inhalation anesthetics that research indicates as less potent trigger substances and the recommended limitations of succinylcholine use, suggests there may be considerable decline of fulminant MH cases. In the presented study, the authors analyzed suspected MH episodes during general anesthesia of patients that were referred to the Wuerzburg MH unit between 2007 and 2011, assuming that MH is still a relevant anesthetic problem in our days.MethodsWith approval of the local ethics committee data of patients that underwent muscle biopsy and in vitro contracture test (IVCT) between 2007 and 2011 were analyzed. Only patients with a history of suspected MH crisis were included in the study. The incidents were evaluated retrospectively using anesthetic documentation and medical records.ResultsBetween 2007 and 2011 a total of 124 patients were tested. 19 of them were referred because of suspected MH events; 7 patients were diagnosed MH-susceptible, 4 MH-equivocal and 8 MH-non-susceptible by IVCT. In a majority of cases masseter spasm after succinylcholine had been the primary symptom. Cardiac arrhythmias and hypercapnia frequently occurred early in the course of events. Interestingly, dantrolene treatment was initiated in a few cases only.ConclusionsMH is still an important anesthetic complication. Every anesthetist must be aware of this life-threatening syndrome at any time. The rapid onset of adequate therapy is crucial to avoid major harm and possibly lethal outcome. Dantrolene must be readily available wherever MH triggering agents are used for anesthesia.

N-Acetylcysteine, a Potent Anti-Oxidant, Rescues the Malignant Hyperthermia and Environmental Heat Stroke Phenotype of Calsequestrin-1 Knockout Mice

Calsequestrin type 1 (CASQ1) is both the major Ca2+ binding protein in the sarcoplasmic reticulum (SR) in skeletal muscle and a direct modulator of type 1 ryanodine receptor (RYR1) Ca2+-release channel activity. Mice lacking CASQ1 (CASQ1-null) exhibit increased susceptibility for Malignant Hyperthermia (MH) and Environmental Heat Stroke (EHS), life-threatening disorders triggered by volatile anaesthetics and high temperature, respectively. Excessive oxidative stress during MH/EHS crises are proposed to result in increased RYR1 nitrosylation, and subsequently, enhanced SR Ca2+ leak that leads to hypercontractures and rhabdomyolysis. Here, we treated CASQ1-null mice for 2 months (from 2-to-4 months of age) with N-acetylcysteine (NAC, a potent anti-oxidant) provided ad libitum in their drinking water (1% w/v). NAC treatment significantly protected CASQ1-null mice from lethal episodes induced by both halothane (2%, 1h at 32°C) and heat challenge (41°C, 1h) with the rate of mortality being 79 vs 25% and 86 vs 30% in control vs NAC-treated mice, respectively. This protection resulted from several factors including: a) reduced oxidative stress measured as both GSH/GSSG ratio and the frequency of mitochondrial superoxide flash activity (P<0.05); b) decreased maximum core temperature during heat challenge (from 42.1 to 40.8°C); and c) reduced number of fibers undergoingrhabdomyolysis (from 37.6 to 11.6 %). Furthermore, in-vitro contracture test (IVCT) showed that the threshold for caffeine-induced contracture is back to normal values. These results support the hypothesis that excessive production of reactive oxygen species plays a critical role in the onset of MH/EHS crises. Therefore, anti-oxidants may be useful for the treatment and prevention of oxidative stress related skeletal muscle disorders.

A family with discordance between malignant hyperthermia susceptibility and rippling muscle disease

Rippling muscle disease (RMD) is a disorder that affects striated muscle and involves disturbances in calcium homeostasis. Malignant hyperthermia susceptibility (MHS) is a potentially lethal disorder, characterized by extreme hypermetabolism and muscle rigidity/rhabdomyolysis during anesthesia with potent inhalational agents, in otherwise healthy individuals. The aim of this report was to search for a correlation between RMD and MHS in members of a family in which both disorders were present. Ten members of a large Swedish family segregating RMD were tested for MHS prior to establishing an RMD diagnosis. Results from diagnostic RMD investigations and anesthesia outcomes were collected and cross-referenced to evaluate whether phenotype variations could be predicted by in vitro contracture test (IVCT) results suggestive of MHS. No correlation was found between individual RMD phenotypes and the IVCT results. There were no recorded adverse reactions to anesthesia, and RMD and MHS did not co-segregate. We conclude that RMD patients should not, on the basis of our present knowledge, be classified as having MHS; however, an increased surveillance for MH reactions is recommended in these patients.