Objective-To improve the accuracy of genotype prediction and guide genetic testing in patients with muscle channelopathies we applied and refined specialised electrophysiological exercise test parameters.Methods-We studied 56 genetically confirmed patients and 65 controls using needle electromyography, the long exercise test, and short exercise tests at room temperature, after cooling, and rewarming.Results-Concordant amplitude-and-area decrements were more reliable than amplitude-only measurements when interpreting patterns of change during the short exercise tests. Concordant amplitude-and-area pattern I and pattern II decrements of >20% were 100% specific for PMC and MC respectively. When decrements at room temperature and after cooling were <20%, a repeat short exercise test after rewarming was useful in patients with myotonia congenita. Area measurements and rewarming distinguished true temperature sensitivity from amplitude reduction due to cold-induced slowing of muscle fibre conduction. In patients with negative short exercise tests, symptomatic eye closure myotonia predicted sodium channel myotonia over myotonia congenita. Distinctive 'tornado-shaped' neuromyotonia-like discharges may be seen in patients with paramyotonia congenita. In the long exercise test, area decrements from pre-exercise baseline were more sensitive than amplitude decrements-from-maximum-CMAP in patients with Andersen-Tawil syndrome. Possible ethnic differences in the normative data of the long exercise test argue for the use of appropriate ethnically-matched controls.Interpretation-Concordant CMAP amplitude-and-area decrements of >20% allow more reliable interpretation of the short exercise tests and aid accurate DNA-based diagnosis. In patients

Objectives: To obtain minimum point prevalence rates for the skeletal muscle channelopathies and to evaluate the frequency distribution of mutations associated with these disorders.Methods: Analysis of demographic, clinical, electrophysiologic, and genetic data of all patients assessed at our national specialist channelopathy service. Only patients living in the United Kingdom with a genetically defined diagnosis of nondystrophic myotonia or periodic paralysis were eligible for the study. Prevalence rates were estimated for England, December 2011.Results: A total of 665 patients fulfilled the inclusion criteria, of which 593 were living in England, giving a minimum point prevalence of 1.12/100,000 (95% confidence interval [CI] 1.03-1.21). Disease-specific prevalence figures were as follows: myotonia congenita 0.52/100,000 (95% CI 0.46-0.59), paramyotonia congenita 0.17/100,000 (95% CI 0.13-0.20), sodium channel myotonias 0.06/100,000 (95% CI 0.04-0.08), hyperkalemic periodic paralysis 0.17/100,000 (95% CI 0.13-0.20), hypokalemic periodic paralysis 0.13/100,000 (95% CI 0.10-0.17), and Andersen-Tawil syndrome (ATS) 0.08/100,000 (95% CI 0.05-0.10). In the whole sample (665 patients), 15 out of 104 different CLCN1 mutations accounted for 60% of all patients with myotonia congenita, 11 out of 22 SCN4A mutations for 86% of paramyotonia congenita/sodium channel myotonia pedigrees, and 3 out of 17 KCNJ2 mutations for 42% of ATS pedigrees. Conclusion:We describe for the first time the overall prevalence of genetically defined skeletal muscle channelopathies in England. Despite the large variety of mutations observed in patients with nondystrophic myotonia and ATS, a limited number accounted for a large proportion of cases. Neurology â 2013;80:1472-1475 GLOSSARY ATS 5 Andersen-Tawil syndrome; CACNA1S 5 calcium channel, voltage-dependent, L type, a 1S subunit gene; CI 5 confidence interval; CLCN1 5 chloride channel, voltage-sensitive 1 gene; HyperPP 5 hyperkalemic periodic paralysis; HypoPP 5 hypokalemic periodic paralysis; KCNJ2 5 potassium inwardly rectifying channel, subfamily J, member 2 gene; MC 5 myotonia congenita; NDM 5 nondystrophic myotonias; PMC 5 paramyotonia congenita; PP 5 periodic paralyses; SCM 5 sodium channel myotonias; SCN4A 5 sodium channel, voltage-gated, type 4, a subunit gene.Nondystrophic myotonias (NDM) and periodic paralyses (PP) comprise a heterogeneous group of skeletal muscle disorders caused by mutations in genes encoding ion channels.e1 NDM include myotonia congenita (MC), paramyotonia congenita (PMC), and the sodium channel myotonias (SCM). PP encompass hypokalemic periodic paralysis (HypoPP), hyperkalemic periodic paralysis (HyperPP), and Andersen-Tawil syndrome (ATS). These are autosomal dominant disorders except for MC, which is inherited in either a dominant or recessive manner. e1 Although significant progress in the clinical, electrophysiologic, and genetic characterization of the skeletal muscle channelopathies has taken place in the past 2 decades, their actual prevalence rem...

These studies provide Class I evidence that DCP significantly reduces attack frequency in HOP but lacked the precision to support either efficacy or lack of efficacy of DCP in HYP.

The nondystrophic myotonias and primary periodic paralyses are an important group of genetic muscle diseases characterized by dysfunction of ion channels that regulate membrane excitability. Clinical manifestations vary and include myotonia, hyperkalemic and hypokalemic periodic paralysis, progressive myopathy, and cardiac arrhythmias. The severity of myotonia ranges from severe neonatal presentation causing respiratory compromise through to mild later-onset disease. It remains unclear why the frequency of attacks of paralysis varies greatly or why many patients develop a severe permanent fixed myopathy. Recent detailed characterizations of human genetic mutations in voltage-gated muscle sodium (gene: SCN4A), chloride (gene: CLCN1), calcium (gene: CACNA1S), and inward rectifier potassium (genes: KCNJ2, KCNJ18) channels have resulted in new insights into disease mechanisms, clinical phenotypic variation, and therapeutic options.

Objective: To assess whether exon deletions or duplications in CLCN1 are associated with recessive myotonia congenita (MC). Methods:We performed detailed clinical and electrophysiologic characterization in 60 patients with phenotypes consistent with MC. DNA sequencing of CLCN1 followed by multiplex ligation-dependent probe amplification to screen for exon copy number variation was undertaken in all patients. Results:Exon deletions or duplications in CLCN1 were identified in 6% of patients with MC. Half had heterozygous exonic rearrangements. The other 2 patients (50%), with severe disabling infantile onset myotonia, were identified with both a homozygous mutation, Pro744Thr, which functional electrophysiology studies suggested was nonpathogenic, and a triplication/homozygous duplication involving exons 8-14, suggesting an explanation for the severe phenotype. Conclusions:These data indicate that copy number variation in CLCN1 may be an important cause of recessive MC. Our observations suggest that it is important to check for exon deletions and duplications as part of the genetic analysis of patients with recessive MC, especially in patients in whom sequencing identifies no mutations or only a single recessive mutation. These results also indicate that additional, as yet unidentified, genetic mechanisms account for cases not currently explained by either CLCN1 point mutations or exonic deletions or duplications. Neurology Myotonia congenita (MC) is the most common skeletal muscle channelopathy, caused by mutations in the chloride channel gene, CLCN1. It can cause severe muscle stiffness after voluntary muscle contraction (myotonia) that warms-up on repeated contraction. Dominant MC has an early age of onset and primarily affects the upper limbs. Recessive MC predominantly affects the lower limbs, causing significant muscle hypertrophy and transient weakness on initiation of movement. More than 120 recessive and dominant acting CLCN1 mutations have been described. 1 These include missense and nonsense mutations, insertions, and small deletions, but to date no whole exon deletions or duplications have been described in the literature.An important unexplained diagnostic issue in MC is the occurrence of patients with recessive pedigrees but only a single loss of function mutation identified despite sequencing of all CLCN1 coding exons.2 This often makes genetic counseling difficult. In such patients, it is likely that other genetic mechanisms account for the recessive inheritance. Based upon the Human Gene Mutation database, large-scale deletions or duplications account for 7%-10% of reported mutations in the human genome. 3 We postulated that exon deletions or duplications

High throughput DNA sequencing is increasingly employed to diagnose single gene neurological and neuromuscular disorders. Large volumes of data present new challenges in data interpretation and its useful translation into clinical and genetic counselling for families. Even when a plausible gene is identified with confidence, interpretation of the clinical significance and inheritance pattern of variants can be challenging. We report our approach to evaluating variants in the skeletal muscle chloride channel ClC-1 identified in 223 probands with myotonia congenita (MC) as an example of these challenges. Sequencing of CLCN1, the gene that encodes CLC-1, is central to the diagnosis of MC. However, interpreting the pathogenicity and inheritance pattern of novel variants is notoriously difficult as both dominant and recessive mutations are reported throughout the channel sequence, ClC-1 structure-function is poorly understood and significant intra- and interfamilial variability in phenotype is reported. Heterologous expression systems to study functional consequences of CIC-1 variants are widely reported to aid the assessment of pathogenicity and inheritance pattern. However, heterogeneity of reported analyses does not allow for the systematic correlation of available functional and genetic data. We report the systematic evaluation of 95 CIC-1 variants in 223 probands, the largest reported patient cohort, in which we apply standardised functional analyses and correlate this with clinical assessment and inheritance pattern. Such correlation is important to determine if functional data improves the accuracy of variant interpretation and likely mode of inheritance. Our data provide an evidence-based approach that functional characterisation of ClC-1 variants improves clinical interpretation of their pathogenicity and inheritance pattern and serve as reference for 34 previously unreported and 28 previously uncharacterised CLCN1 variants. In addition, we identify novel pathogenic mechanisms and find that variants that alter voltage dependence of activation cluster in the first half of the transmembrane domains and variants that yield no currents cluster in the second half of the transmembrane domain. None of the variants in the intracellular domains were associated with dominant functional features or dominant inheritance pattern of MC. Our data help provide an initial estimate of the anticipated inheritance pattern based on the location of a novel variant and shows that systematic functional characterisation can significantly refine the assessment of risk of an associated inheritance pattern and consequently the clinical and genetic counselling.

Objective: The objective of this study was to validate the immunohistochemical assay for the diagnosis of nondystrophic myotonia and to provide full clarification of clinical disease to patients in whom basic genetic testing has failed to do so.Methods: An immunohistochemical assay of sarcolemmal chloride channel abundance using 2 different ClC1-specific antibodies.Results: This method led to the identification of new mutations, to the reclassification of W118G in CLCN1 as a moderately pathogenic mutation, and to confirmation of recessive (Becker) myotonia congenita in cases when only one recessive CLCN1 mutation had been identified by genetic testing. Conclusions:We have developed a robust immunohistochemical assay that can detect loss of sarcolemmal ClC-1 protein on muscle sections. This in combination with gene sequencing is a powerful approach to achieving a final diagnosis of nondystrophic myotonia. Neurology â 2012;79:2194-2200 GLOSSARY DM1 5 myotonic dystrophy type 1; DM2 5 myotonic dystrophy type 2; GFP 5 green fluorescent protein; HEK 5 human embryonic kidney; NDM 5 nondystrophic myotonia.We have previously reported an increased frequency of coexisting recessive CLCN1 mutations in patients with currently diagnosed myotonic dystrophy type 2 (DM2), 1 because patients with DM2 heterozygous for a recessive CLCN1 mutation have more pronounced myotonia.1 With the aim of showing this modifying effect of the cosegregating CLCN1 mutations on the protein level, we developed an immunohistochemical assay for ClC-1 protein expression. The method proved to be efficient in the molecular diagnostic clarification of nondystrophic myotonias (NDMs) caused by mutations in CLCN1 and SCN4A genes.Autosomal recessive Becker (OMIM #255700) and dominant Thomsen (OMIM #160800) congenital myotonia are NDMs caused by mutations in CLCN1 on chromosome 7q35.2 More than 100 different CLCN1 mutations have been identified.3 Some CLCN1 mutations are clearly more common than others. R894X (c.2680C.T) has an estimated carrier frequency of about 1% in the European population. In the Finnish population the mutation F413C (c.1238T.G) is almost as frequent, at least in northern Finland. 4 However, these 2 mutations explain only about half of the congenital myotonias in the studied population, and many myotonia patients remain with just one mutation identified when screening for these 2 common mutations.In this study, we focused on the validation of an immunohistochemical assay for the diagnosis of NDM. With this method combined with molecular genetics, we were able to clarify all undetermined myotonia patients, identify new recessive mutations, and verify normal protein expression with dominant CLCN1 mutations.

In a probe-type formation test, because of the geometry of the wellbore and the sealing effect of mudcake, the flow pattern is not perfectly spherical. To account for the deviation from spherical flow, several geometric correction factors were proposed for different analysis techniques (Steward and Wittmann 1979;Wilkinson and Hammond 1990;Dussan and Sharma 1992;Goode and Thambynayagam 1992;Proett and Chin 1996). A geometric factor is used in formation rate analysis (FRA) (Kasap et al. 1999), a technique used in analyzing a probe test to estimate formation pressure and permeability. Like other geometric correction factors, the geometric factor is a strong function of permeability anisotropy that is generally unknown before a test. When analyzing the test, we would logically assume an isotropic formation and use the corresponding isotropic geometric factor. Consequently, the FRA-estimated permeability does not represent the true spherical permeability. In contrast, the spherical permeability can be estimated from buildup analysis without prior knowledge of permeability anisotropy. Therefore, there is a discrepancy between the permeability estimates from the two analysis methods. In addition, if considered separately, neither FRA nor buildup analysis can decompose the estimated permeability into its horizontal and vertical components.This paper presents the derived numerical values of several geometric factors. Using these factors, we show that the discrepancy between the permeabilities estimated from FRA and from the conventional buildup analysis is attributable to the permeabilityanisotropy effect. A correct geometric-factor value must be used to estimate permeability correctly. On the basis of the permeabilityanisotropy effect, we present the procedures to estimate horizontal and vertical permeabilities by combining FRA permeability and buildup permeability or by history matching. These procedures are verified with a simulated probe test. Analysis of three actual tests is presented.