Malignant hyperthermia in humans--standardization of contracture testing protocol

Malignant hyperthermia (MH) diagnostic biopsy centers across North America have not previously been standardized in regard to protocols and specific muscles. Recent standardization criteria prompted this study of the vastus and rectus abdominis muscles. This study evaluated changes in contracture tension after electrical stimulation of 271 bundles taken from the vastus (n = 16) and rectus abdominus (n = 19) muscle biopsies of normal individuals when exposed to tissue baths in the absence of and in the presence of caffeine (0.5, 1.0, 2.0, 4.0, 8.0, and 32.0 mM) alone, halothane (1% or 3%) alone, or the combination of halothane (1%) plus caffeine (0.25, 0.5, 1.0, 2.0, 4.0, and 32.0). Caffeine threshold concentration was that concentration of caffeine that produced a 7% increase in tension. Caffeine specific concentration (CSC) and halothane caffeine specific concentration (HCSC) were those concentrations of caffeine alone or of halothane plus caffeine that produced a 1 g increase in tension. The concentration of caffeine alone that increased the contracture tension by 7% averaged 6.7 +/- 0.3 mM for vastus, significantly greater than 4.1 +/- 0.2 mM for the rectus muscle biopsies. Caffeine specific concentration was significantly greater for vastus muscle (7.7 +/- 0.7 mM) than it was for rectus muscle (4.9 +/- 0.4 mM). Three percent halothane alone showed contractures in 3/41 vastus (all less than 0.5 g) and 18/54 rectus muscle bundles (8 greater than 0.5 g). Mean HCSC was statistically significantly greater for vastus muscle (1.9 +/- 0.2 mM) than for rectus muscle (1.2 +/- 0.2 mM).(ABSTRACT TRUNCATED AT 250 WORDS)     

A comparison of the caffeine halothane muscle contracture test with the molecular genetic diagnosis of malignant hyperthermia

Malignant hyperthermia (MH) is currently diagnosed by the caffeine-halothane contracture (CHC) test. In a previous study, this test was used to establish linkage between the human gene for MH susceptibility and the ryanodine receptor (RYR) gene. The current study extends the genetic linkage analysis to a large French-Canadian kindred. In this family, genetic linkage between RYR and MH genes was not demonstrable using the currently recommended limits of normal for the CHC test in the identification of MH-susceptible individuals. With CHC test threshold limits below those currently recommended, however, complete linkage between the RYR and MH genes was seen. Comparisons of CHC test results with genetic linkage studies will increase the diagnostic accuracy of both tests as well as generate new insights into the biology of MH.     

Abnormal calcium signalling and the caffeine–halothane contracture test

Background

The variable clinical presentation of malignant hyperthermia (MH), a disorder of calcium signalling, hinders its diagnosis and management. Diagnosis relies on the caffeine–halothane contracture test, measuring contraction forces upon exposure of muscle to caffeine or halothane (F_C and F_H, respectively). Patients with above-threshold F_C or F_H are diagnosed as MH susceptible. Many patients test positive to halothane only (termed ‘HH’). Our objective was to determine the characteristics of these HH patients, including their clinical symptoms and features of cytosolic Ca²⁺ signalling related to excitation–contraction coupling in myotubes.

In vitro contracture test for diagnosis of malignant hyperthermia following the protocol of the European MH Group: Results of testing patients surviving fulminant MH and unrelated low-risk subjects

Background: Determination of sensitivity and specificity of the in vitro contracture test (IVCT) for malignant hyperthermia (MH) susceptibility using the European MH Group (EMHG) protocol has been performed in some laboratories but only on a small sample from the combined EMHG. Thus, the purpose of the present study was to determine combined EMHG sensitivity and specificity of the test. Methods: Results of IVCT of patients with previous fulminant MH and normal, low-risk subjects (controls) were collected from 22 centresof the EMHG. IVCT was performed according to the EMHG protocol. Patients were included inthe study if the clinical crisis had a score of at least 50 points with the Clinical Grading Scale. Low-risk subjects were included provided they did not belong to a family with known MH susceptibility, they had not developed any signs of MH at previous anaesthetics, and they did not suffer from any neuromuscular disease. For inclusion of both MH patientsand low-risk subjects, at least 1 muscle bundle in the IVCT should have twitches of 10 mN(1 g) or more. For evaluation of individual tests, only muscle bundles with twitch heights of 10 mN (1 g) or more were used. Results: A total of 1502 probands had undergone IVCT because of a previous anaesthesia with symptoms and signs suggestive of MH. Of these, 119 had clinical scores of 50 and above. From these 119 MH-suspected patients and from 202 low-risk subjects, IVCT data were collected. Subsequently, 14 MH-suspected patients were excluded from further analysis for thefollowing reasons: In 3 patients, the suspected MH episode could be fully explained by diseases other than MH; in 11 MHS patients, IVCT was incomplete (n=l), data were lost (n=3), or none of the muscle bundles fulfilled twitch criteria (n=7). Of the remaining 105 MH-suspected patients, 89 were MHS, 10 MHEh, 5 MHEc, and one MHN. Thus, we observed a diagnostic sensitivity of the IVCT of 99.0% if the MHE group is considered susceptible(95% confidence interval 94.8–100.0%). Of the 202 low-risk subjects, 3 were MHS, 5 MHEh, 5 MHEc, and 189 MHN. This gives a specificity of the IVCT of 93.6% (95% confidence interval 89.2–96.5%). Conclusion: The IVCT for diagnosis of MH susceptibility in Europe has a high sensitivity and a satisfactory specificity.     

Comparison of the caffeine skinned fibre tension (csft) test with the caffeine-halothane contracture (chc) test in the diagnosis of malignant hyperthermia

We have compared and contrasted two diagnostic tests for Malignant Hyperthermia (MH) -the Caffeine-Halothane Contracture Test and the Caffeine Skinned Fibre Tension Test. Both tests show a strongly positive relationship both with the occurrence of MH reactions and with each other. The former test is more rapid and requires less skill. The latter test can be performed on much less muscle and permits storage of the muscle over prolonged periods of time.     

Sources of variability in halothane and caffeine contracture tests for susceptibility to malignant hyperthermia.

In vitro contracture tests for susceptibility to malignant hyperthermia (MH) were performed in 96 patients according to the protocol of the European MH Group. In addition, tests were performed with halothane 0.44 mmol l-1 and 0.66 mmol l-1, and caffeine 2 mmol l-1, each added as a single bolus dose to fresh specimens. For all tests the size of contractures were recorded, and for the diagnostic tests the halothane and caffeine threshold concentrations were determined (i.e. the minimal concentrations eliciting a contracture of 0.2 g). The caffeine specific concentration (CSC, i.e. the concentration increasing force 1.0 g) and the % increase with caffeine 2 mmol l-1 were calculated from the dose response curves. Various diagnostic criteria in use by the North American MH Group were applied, and diagnostic outcome compared with the result obtained by the protocol of the European MH Group. Thirty-five patients were susceptible to MH (MHS), 33 were non-susceptible (MHN), and 28 had equivocal results of the tests (MHE). Additional tests were made in 34 MHS, 32 MHN, and 26 MHE patients. Contractures elicited by bolus addition of halothane or caffeine were significantly larger than those observed following the same dose of drug added cumulatively (P less than 0.05). Contractures greater than or equal to 0.7 g following halothane 3% (bolus dose) were seen in 78% of MHS patients and 18% of MHN patients, A CSC less than 4 mmol l-1 was elicited in 86% of MHS and 30% of MHN patients, whereas an increase in force greater than or equal to 4% or greater than or equal to 7% was seen in 71% and 34% of MHS patients, respectively, and in none of the MHN patients. Using the criterion of greater than or equal to 0.7 g in the halothane test and greater than or equal to 4% increase in the caffeine test gave the best agreement between diagnostic outcome with the European and North American protocols: All 34 MHS patients (100%) were positive to one or more tests, but so were eight of 32 MHN patients (25%), giving an overall diagnostic agreement of 88%. We conclude that, in our laboratory, the results obtained with the two major protocols for investigation of MH susceptibility are not identical. Patients surviving fulminant MH, however, react abnormally to nearly all the tests. For validation and possibly further standardization of the tests each laboratory must investigate a large number of normal controls and as many patients surviving fulminant MH as possible.     

Diagnosis of susceptibility to malignant hyperthermia using the in vitro contracture test

Though a malignant hyperthermia (MH) crisis is still a critical event during general anesthesia, recent developments in prophylaxis and treatment should help in avoiding fatal episodes. The best means to avoid MH episodes would be early recognition of MH susceptibility. Today the only reliable test to identify MH susceptibility is the in vitro contracture test. Thus, to diagnose MH susceptibility we performed this test on muscle biopsies from 26 individuals who: (1) had an event during general anesthesia that may have been indicative of MH (4 patients); (2) had a family member with a medical history of MH (20 patients); or (3) had unexplained elevated CK levels (1 patient). The criteria according to which patients were submitted to the testing are shown in detail in Table 1. We used the standardized version of the contracture test that has been proposed by the European Malignant Hyperpyrexia Group. Muscle biopsies (20-30 mm long, 8 mm diameter) were dissected into 8-10 small bundles (2-3 mm diameter) and tested within 3 h post-biopsy in four independent tissue baths with various concentrations of caffeine or halothane. According to the concentration of caffeine or halothane necessary to elicit contractures exceeding a predefined force threshold (20 mN), it was possible to classify the patients as MHS (MH-susceptible), MHE (equivocal), or MHN (negative). In addition to the in vitro test, clinical, laboratory, and neurophysiological data were collected from these patients and correlated with the individual test results (Table 2). Thirteen patients were classified as MHS, five were MHE, and seven patients MHN (Fig. 3).(ABSTRACT TRUNCATED AT 250 WORDS)     

Malignant hyperthermia (MH) diagnostics: a comparison between the halothane-caffeine-and the ryanodine-contracture-test results in MH susceptible, normal and control muscle

Recent studies demonstrated different contracture responses in muscle from malignant hyperthermia susceptible (MHS) compared to normal (MHN) individuals following exposure to the plant alkaloid ryanodine in-vitro. To confirm if ryanodine has a specific action in MHS muscle, the effect of a single concentration was investigated in skeletal muscle from MHS, MHN and control subjects using a new evaluation technique. In-vitro contracture test (IVCT) and MH diagnosis were performed according to the European Protocol in 86 patients sent to us for MH diagnostic testing and in 24 controls. Viable fresh muscle bundles were exposed to a single bolus of ryanodine 1.0 μM. Contracture onset time (OTp: defined as the time (min) from administration of ryanodine to the start of a contracture as measured by a contracture exceeding predrug baseline height), and the time to an increase of the baseline height to 10 mN above the predrug level (10Tp) were recorded. 29 patients were identified by IVCT to be MHS, 50 MHN, 7 MHE (equivocal) and 24 controls MHN. The indices from the ryanodine test separated all MHS (OTp: <16 min; 10Tp <27.4 min) from MHN (>18 and >27.7 min) and control subjects (>17.4 and >29 min). Values for MHE (equivocal) individuals ranged from 17.1 to 27.8 min for the OTp and from 32 to 49.2 min for the 10Tp. 5 patients with fulminant MH crises were included in the MHS group and showed the 95% confidence intervals (CI) of the median value ≤8.05min (OTp) and ≤13.35min (IOTP) for MHS. In contrast, CI of the median value for the control group were found to be ≥25.2min (OTp) and 43.15min (10Tp) for normal muscle. Thus the ryanodine test protocol showed markedly different contractures in MHS and MHN or control muscle. These results suggest that MHS muscle has a higher sensitivity to ryanodine. However, the protocol should be investigated for reproducibility and validation of thresholds by other laboratories. Ryanodine can help to improve MH diagnostic tests.     

Effect of temperature,time and fascicle size on the caffeine contracture test

The caffeine contracture test is the most commonly used method of diagnosing malignant hyperthermia. We have examined some factors which may influence the results of this test. These have included the temperature of the bathing solution, the size of the muscle fascicles, and the combined effect of the passage of time and prior equilibration with caffeine or with caffeine plus halothane. For both malignant hyperthermic susceptible (MHS) and normal fascicles, caffeine contractures were greater at 37° C than at 22° C, while halothane and caffeine plus halothane contractures were similar at 37° C and at 22° C. Good differentiation between the normal and the MHS fascicles were observed at both temperatures although the discrimination was slightly, although not always significantly, better at 22° C. The weight, length or diameter of the fascicles had little or no effect on the height of the caffeine or the caffeine plus halothane contractures. We compared caffeine plus halothane contractures exhibited by newly prepared muscle fascicles with caffeine plus halothane contractures manifested by fascicles which had already been equilibrated with either caffeine alone or with caffeine and halothane for at least one hour. Differences in contracture heights among the techniques were small and often not significant, particularly at 37° C. The greatest discrimination between the MHS and the normal muscle fascicles was provided by determining the caffeine plus halothane contracture curves at 22° C, using muscle fascicles which had previously been equilibrated with incremental doses of caffeine in the absence of halothane. Slightly less accurate but still reasonably satisfactory results were also obtained at 37° C using muscle strips which were either freshly prepared or which had prior exposure to caffeine or to caffeine in combination with halothane and, at 22° C, using either newly prepared muscle or muscle which had already been equilibrated with caffeine plus halothane. The widest differentiation between the MHS and the normal muscle was given by the caffeine plus halothane contractures and the least by the halothane contractures. MHS patients whose muscle fascicles exhibited greater than normal caffeine plus halothane contractures but normal caffeine contractures and normal halothane contractures had had the most mild clinical reactions. On the other hand MHS patients whose muscle fascicles demonstrated halothane contractures, caffeine contractures and caffeine plus halothane contractures which were all greater than normal had had the most severe reactions. It is concluded, therefore, that if the amount of muscle available is very small, a satisfactory caffeine contracture test can still be performed by doing the various parts of the test sequentially on the same fascicle. If the time available for performing the test is limited, then the several parts of the test should rather be performed simultaneously on separate muscle fascicles.     

The in-vitro caffeine contracture test: Influence of the muscle histochemical profile on test results

In vitro caffeine contracture tests were carried out on whole rat muscle composed primarily of either histochemical type I or type II fibers. Muscles composed primarily of type I fibers developed contractures at lower concentrations of caffeine and had lower caffeine specific concentrations than muscles composed primarily of type II fibers. These findings indicate that the histochemical profile of a muscle can influence the results of the in-vitro caffeine contracture test.     

Between-center variability of results of the in vitro contracture test for malignant hyperthermia susceptibility

The in vitro contracture test (IVCT) remains the standard test for the diagnosis of malignant hyperthermia (MH) susceptibility. The aim of this study was to investigate whether results of the IVCT varied between two diagnostic centers. The study took place at the national MH centers in Denmark and Sweden. Forty-three patients investigated for MH gave informed consent to have four extra muscle specimens excised. These were sent to the other center and immediately used for a parallel IVCT, according to the protocol of the European MH Group. Results of the IVCTs performed in the two centers on muscle samples from the same patients were compared. Each patient was assigned a diagnosis according to the result obtained in the "mother-center." Identical diagnostic results were obtained for 56% of the patients. The differing diagnostic outcomes were almost exclusively seen in cases with contractures of <5 mN (0.5 g) and abnormal results in only one or two muscle strips. We suggest different criteria for the interpretation of results for clinical and scientific purposes. The clinical criteria should remain unchanged. The scientific designation of susceptibility should be used in cases with contractures of > or =5 mN and abnormal results in at least 75% of the tested muscle strips. Implications: The diagnostic outcomes of tests for malignant hyperthermia susceptibility were compared between two laboratories by using muscle tissue from the same patients. Identical outcomes were found for 56% of the patients. Almost all diverging outcomes were seen in cases with a few small contractures near the cutoff limit. Different diagnostic criteria for clinical and scientific purposes are suggested.     

Prediction of malignant hyperthermia susceptibility in low-risk subjects. An epidemiologic investigation of caffeine halothane contracture responses. The North American Malignant Hyperthermia Registry.

The most commonly used laboratory test for predicting malignant hyperthermia susceptibility is the caffeine halothane contracture test. However, the specificity and sensitivity of proposed North American diagnostic guidelines for this test have never been evaluated in a large, human study population. Therefore, the authors conducted a multiinstitutional, prospective study of skeletal muscle contracture responses in a subject population at low risk for malignant hyperthermia susceptibility to help determine the specificity of the proposed guidelines. Subjects were selected arbitrarily from a population of patients undergoing surgery unrelated to performance of a diagnostic muscle biopsy. Subjects were admitted to this study and were presumed nonsusceptible if there was no evidence of any of the following malignant hyperthermia risk factors: prior abnormal response to triggering anesthetic agents, myopathy, or family history of malignant hyperthermia susceptibility. The authors suggested rejection of the proposed diagnostic guidelines if an 85% specificity estimate among subjects could not be obtained. The authors analyzed the responses of 1,022 muscle fascicles, derived from 176 subjects, to the following: 1) separate administration of 3% halothane or incremental caffeine concentrations, or 2) the joint administration of 1% halothane and incremental caffeine concentrations. The following contracture results were obtained. First, for individual fascicles, 9.2% exceeded a greater than 0.7 g threshold for 3% halothane, 15.2% exceeded a greater than or equal to 0.2 g threshold for 2 mM caffeine, 32.4% exceeded a 1-g increase for less than 4 mM caffeine, 2.6% had a greater than 7% maximal increase in tension at 2 mM caffeine, and 63.5% had a "halothane caffeine-specific concentration" at less than or equal to 1 mM caffeine. Second, the percentages of subjects with 1 or more fascicles exceeding the proposed threshold were as follows: 45.8% for the four-component, 28.8% for the three-component, and 32.7% for the two-component contracture test. Third, the percentages of subjects with 1 or more fascicles exceeding the proposed threshold for both halothane and caffeine were as follows: 9.5% for 3% halothane and 2 mM caffeine, 2.0% for 3% halothane and 7% maximal increase in tension at 2 mM caffeine, and 11.0% for 1% halothane and 2 mM caffeine. Fourth, center-to-center differences were the major source of variation in the rate that subjects exceeded proposed thresholds. These data demonstrate that proposed diagnostic guidelines must be modified to improve specificity estimates before adoption by diagnostic centers.(ABSTRACT TRUNCATED AT 400 WORDS)     

No relationship between fiber type and halothane contracture test results in malignant hyperthermia

Previous studies in cat, rat, and swine have implicated fiber type as influencing the halothane and caffeine contracture test used to diagnose malignant hyperthermia (MH). The authors performed fiber type analysis using myosin ATPase stains on 31 fascicles of skeletal muscle from nine patients following contracture testing. There was no significant difference in fiber type composition between fascicles from MH negative (n = 5) and MH positive (n = 4) patients. Furthermore, examining each of the 31 fascicles, the authors found no correlation (P greater than .05) of contracture magnitude with percentage of either Type I or Type II fibers using the Pearson Product-Moment correlation calculation. The authors conclude that fiber type composition does not influence contracture test results in human biopsies.     

Pulmonary emboli in sclerotherapy for peripheral vascular malformations under general anesthesia; a report of two cases

Sclerotherapy with absolute ethanol and/or polidocanol is a well-established therapeutic modality for the treatment of peripheral vascular malformations, although systemic complications such as hemoglobinuria and pulmonary embolism could occur. We report two cases of pulmonary embolism associated with sclerotherapy for peripheral vascular malformations. Two patients, a 17-year-old man and a 17-year-old woman, undergoing absolute ethanol sclerotherapy for vascular malformations of the leg developed pulmonary embolism after injection of ethanol. Pulmonary embolism, suspected by the clinical symptoms such as hypoxia and hypocapnia, was confirmed by the pulmonary scintigraphy showing minimal pulmonary defects. Hemoglobinuria was also observed with injection of ethanol. Patients recovered rapidly with heparin and urokinase therapy. The review of perioperative complications with sclerotherapy for peripheral vascular malformations in our institution for past four years revealed that complications were observed in 18 out of 88 patients (20.5%), and in 32 out of 183 cases (17.5%). Major complications were hemoglobinuria, pulmonary embolism, shivering and delayed emergence from general anesthesia. We conclude that sclerotherapy for vascular malformations under general anesthesia is a risky procedure and this must be carefully managed with keen monitoring of Spo2 and Etco2.