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Ryanodine receptors (RyRs) are main regulators of intracellular Ca 2+ release and muscle contraction. The Y522S mutation of RyR1 causes central core disease, a weakening myopathy, and malignant hyperthermia, a sudden and potentially fatal response to anesthetics or heat. Y522 is in the core of the N-terminal subdomain C of RyR1 and the mechanism of how this mutation orchestrates malfunction is unpredictable for this 2-MDa ion channel, which has four identical subunits composed of 15 distinct cytoplasmic domains each. We expressed and purified the RyR1 rabbit homolog, Y523S, from HEK293 cells and reconstituted it in nanodiscs under closed and open states. The high-resolution cryogenic electron microscopic (cryo-EM) three-dimensional (3D) structures show that the phenyl ring of Tyr functions in a manner analogous to a “spacer” within an α-helical bundle. Mutation to the much smaller Ser alters the hydrophobic network within the bundle, triggering rearrangement of its α-helices with repercussions in the orientation of most cytoplasmic domains. Examining the mutation-induced readjustments exposed a series of connected α-helices acting as an ∼100 Å-long lever: One end protrudes toward the dihydropyridine receptor, its molecular activator (akin to an antenna), while the other end reaches the Ca 2+ activation site. The Y523S mutation elicits channel preactivation in the absence of any activator and full opening at 1.5 µM free Ca 2+ , increasing by ∼20-fold the potency of Ca 2+ to activate the channel compared with RyR1 wild type (WT). This study identified a preactivated pathological state of RyR1 and a long-range lever that may work as a molecular switch to open the channel.

To investigate the possible causative factors of central core disease(CCD), the clinical features of a neonatal case with CCD and five patients in the pedigree line were analyzed for RYR1 gene variant.Medical and family history inquiries and detailed clinical examinations were performed in the proband. High-throughput sequencing technology was applied to analyze the gene variant of the proband, and Sanger sequencing was applied to verify the pedigree distribution of the variant.The whole exon sequencing results showed that the proband has a missense variant of c. 14591AC (p.Tyr4864Ser) in the RYR1 gene which was unreported previously; Sanger sequencing results showed that the father, grandfather, the eldest aunt and second aunt of the proband all carried the same variant. The c.14591 AC variant of RYR1 gene was predicted to be a likely pathogenic (PM2+PM5+PP1+PP3) according to the American College of Medical Genetics and Genomics standards and guidelines.The RYR1 gene c.14591AC (p.Tyr4864Ser) variant may be the genetic cause of the pedigree and genetic testing helps to clarify the diagnosis. Identification of this variant has enriched the variant spectrum of the RYR1 gene.

Central Core Disease (CCD) is an inherited neuromuscular disorder characterized by the presence of cores in muscle biopsy. CCD is caused by mutations in the RYR1 gene. This gene encodes the ryanodine receptor 1, which is an intracellular calcium release channel from the sarcoplasmic reticulum to the cytosol in response to depolarization of the plasma membrane. Mutations in this gene are also associated with susceptibility to Malignant Hyperthermia (MHS). In this study, we evaluated 20 families with clinical and histological characteristics of CCD to identify primary mutations in patients, for diagnosis and genetic counseling of the families. We identified variants in the RYR1 gene in 19/20 families. The molecular pathogenicity was confirmed in 16 of them. Most of these variants (22/23) are missense and unique in the families. Two variants were recurrent in two different families. We identified six families with biallelic mutations, five compound heterozygotes with no consanguinity, and one homozygous, with consanguineous parents, resulting in 30% of cases with possible autosomal recessive inheritance. We identified seven novel variants, four of them classified as pathogenic. In one family, we identified two mutations in exon 102, segregating in cis, suggesting an additive effect of two mutations in the same allele. This work highlights the importance of using Next-Generation Sequencing technology for the molecular diagnosis of genetic diseases when a very large gene is involved, associated to a broad distribution of the mutations along it. These data also influence the prevention through adequate genetic counseling for the families and cautions against malignant hyperthermia susceptibility.

Central Core Disease (CCD) is a genetic neuromuscular disorder characterized by the presence of cores in muscle biopsy. The inheritance has been described as predominantly autosomal dominant (AD), and the disease may present as severe neonatal or mild adult forms. Here we report clinical and molecular data on a large cohort of Brazilian CCD patients, including a retrospective clinical analysis and molecular screening for RYR1 variants using Next-Generation Sequencing (NGS). We analyzed 27 patients from 19 unrelated families: four families (11 patients) with autosomal dominant inheritance (AD), two families (3 patients) with autosomal recessive (AR), and 13 sporadic cases. Biallelic RYR1 variants were found in six families (two AR and four sporadic cases) of the 14 molecularly analyzed families (~43%), suggesting a higher frequency of AR inheritance than expected. None of these cases presented a severe phenotype. Facial weakness was more common in biallelic than in monoallelic patients (p = 0.0043) and might be a marker for AR forms. NGS is highly effective for the identification of RYR1 variants in CCD patients, allowing the discovery of a higher proportion of AR cases with biallelic mutations. These data have important implications for the genetic counseling of the families.

Background The diagnosis of uncommon pediatric neuromuscular disease (NMD) is challenging due to genetic and phenotypic heterogeneity, yet is important to guide treatment, prognosis, and recurrence risk. Patients with diagnostically challenging presentations typically undergo extensive testing with variable molecular diagnostic yield. Given the advancement in next generation sequencing (NGS), we investigated the value of clinical whole exome sequencing (ES) in uncommon pediatric NMD. Methods A retrospective cohort study of 106 pediatric NMD patients with a combination of ES, chromosomal microarray (CMA), and candidate gene testing was completed at a large tertiary referral center. Results A molecular diagnosis was achieved in 37/79 (46%) patients with ES, 4/44 (9%) patients with CMA, and 15/74 (20%) patients with candidate gene testing. In 2/79 (3%) patients, a dual molecular diagnosis explaining the neuromuscular disease process was identified. A total of 42 patients (53%) who received ES remained without a molecular diagnosis at the conclusion of the study. Conclusions Due to NGS, molecular diagnostic yield of rare neurological diseases is at an all-time high. We show that ES has a higher diagnostic rate compared to other genetic tests in a complex pediatric neuromuscular disease cohort and should be considered early in the diagnostic journey for select NMD patients with challenging presentations in which a clinical diagnosis is not evident.

Mutations in the RYR1 gene, encoding ryanodine receptor 1 (RyR1), are a well-known cause of Central Core Disease (CCD) and Multi-minicore Disease (MmD). We screened a cohort of 153 patients carrying an histopathological diagnosis of core myopathy (cores and minicores) for RYR1 mutation. At least one RYR1 mutation was identified in 69 of them and these patients were further studied. Clinical and histopathological features were collected. Clinical phenotype was highly heterogeneous ranging from asymptomatic or paucisymptomatic hyperCKemia to severe muscle weakness and skeletal deformity with loss of ambulation. Sixty-eight RYR1 mutations, generally missense, were identified, of which 16 were novel. The combined analysis of the clinical presentation, disease progression and the structural bioinformatic analyses of RYR1 allowed to associate some phenotypes to mutations in specific domains. In addition, this study highlighted the structural bioinformatics potential in the prediction of the pathogenicity of RYR1 mutations. Further improvement in the comprehension of genotype–phenotype relationship of core myopathies can be expected in the next future: the actual lack of the human RyR1 crystal structure paired with the presence of large intrinsically disordered regions in RyR1, and the frequent presence of more than one RYR1 mutation in core myopathy patients, require designing novel investigation strategies to completely address RyR1 mutation effect.

Core myopathies are clinically, pathologically, and genetically heterogeneous muscle diseases. Their onset and clinical severity are variable. Core myopathies are diagnosed by muscle biopsy showing focally reduced oxidative enzyme activity and can be pathologically divided into central core disease, multiminicore disease, dusty core disease, and core-rod myopathy. Although RYR1-related myopathy is the most common core myopathy, an increasing number of other causative genes have been reported, including SELENON, MYH2, MYH7, TTN, CCDC78, UNC45B, ACTN2, MEGF10, CFL2, KBTBD13, and TRIP4. Furthermore, the genes originally reported to cause nemaline myopathy, namely ACTA1, NEB, and TNNT1, have been recently associated with core-rod myopathy. Genetic analysis allows us to diagnose each core myopathy more accurately. In this review, we aim to provide up-to-date information about core myopathies.

Background Dermatomyositis is an inflammatory muscle disease caused by immune-mediated muscle injury, and central core disease (CCD) is a congenital myopathy associated with disturbed intracellular calcium homeostasis and excitation-contraction coupling. To date, CCD has not been reported to have autoantibodies or coexist with inflammatory myopathy. Case presentation Here, we described the case of a 25-year-old woman who had progressive proximal muscle weakness, myalgia, pruritic macular rash, skin ulcers, and calcinosis. Dermatomyositis was initially suspected based on the clinical symptoms accompanied by elevated muscle enzyme levels, electromyography abnormalities, and a positive antinuclear antibody test. However, the patient’s muscle biopsy revealed the characteristic findings of both dermatomyositis and CCD, suggesting that dermatomyositis occurred in this patient with previously asymptomatic CCD. The patient did not have any pathogenic gene mutations associated with congenital myopathy, including RYR1 and SEPN1 in targeted next-generation sequencing. She received high-dose glucocorticoid therapy and azathioprine with a significant improvement in muscle strength. Conclusions We present a case of rare coexistence of dermatomyositis and CCD. Clinicians should be aware that patients with CCD may have inflammatory myopathy that responds well to immunosuppressive therapy.

Recessive ryanodine receptor 1 (RYR1) mutations cause congenital myopathies including multiminicore disease (MmD), congenital fiber-type disproportion and centronuclear myopathy. We created a mouse model knocked-in for the Q1970fsX16+A4329D RYR1 mutations, which are isogenic with those identified in a severely affected child with MmD. During the first 20 weeks after birth the body weight and the spontaneous running distance of the mutant mice were 20% and 50% lower compared to wild-type littermates. Skeletal muscles from mutant mice contained ‘cores’ characterized by severe myofibrillar disorganization associated with misplacement of mitochondria. Furthermore, their muscles developed less force and had smaller electrically evoked calcium transients. Mutant RyR1 channels incorporated into lipid bilayers were less sensitive to calcium and caffeine, but no change in single-channel conductance was observed. Our results demonstrate that the phenotype of the RyR1Q1970fsX16+A4329D compound heterozygous mice recapitulates the clinical picture of multiminicore patients and provide evidence of the molecular mechanisms responsible for skeletal muscle defects.

Congenital myopathies represent a clinically and genetically heterogeneous group of early-onset neuromuscular diseases with characteristic, but not always specific, histopathological features, often presenting with stable and/or slowly progressive truncal and proximal weakness. It is often not possible to have a diagnosis on clinical ground alone. Additional extraocular, respiratory, distal involvement, scoliosis, and distal laxity may provide clues. The “core myopathies” collectively represent the most common form of congenital myopathies, and the name pathologically corresponds to histochemical appearance of focally reduced oxidative enzyme activity and myofibrillar changes on ultrastructural studies. Because of the clinical, pathological, and molecular overlaps, central core disease and multiminicore disease will be discussed together.

Typical central core disease (CCD) is characterized pathologically by the presence of a core and is accompanied by type 1 fiber uniformity. Congenital neuromuscular disease with uniform type 1 fiber (CNMDU1) is characterized pathologically by the presence of type 1 fiber uniformity but without the abnormal structural changes in muscle fibers. Interestingly, typical CCD and 40% of CNMDU1 cases are caused by the same mutations in RYR1, and thus CNMDU1 has been considered an early precursor to CCD. To better understand the nature of CNMDU1, we re-evaluated muscle biopsies from 16 patients with CNMDU1 using immunohistochemistry to RYR1, triadin and TOM20, and compared this to muscle biopsies from 36 typical CCD patients. In CCD, RYR1, and triadin were present in the core regions, while TOM20 was absent in the core regions. Interestingly, in 5 CNMDU1 cases with the RYR1 mutation, RYR1, and triadin were similarly present in core-like areas, while TOM20 was absent in the subsarcolemmal region. Furthermore, there was a correlation between the core position and the disease duration or progression-the older patients in more advanced stages had more centralized cores. Our results indicate that CNMDU1 due to RYR1 mutation is a de facto core myopathy.

Mutations in the RYR1 gene, encoding the skeletal muscle calcium channel RyR1, lead to congenital myopathies, through expression of a channel with abnormal permeability and/or in reduced amount, but the direct functional whole organism consequences of exclusive reduction in RyR1 amount have never been studied. We have developed and characterized a mouse model with inducible muscle specific RYR1 deletion. Tamoxifen-induced recombination in the RYR1 gene at adult age resulted in a progressive reduction in the protein amount reaching a stable level of 50% of the initial amount, and was associated with a progressive muscle weakness and atrophy. Measurement of calcium fluxes in isolated muscle fibers demonstrated a reduction in the amplitude of RyR1-related calcium release mirroring the reduction in the protein amount. Alterations in the muscle structure were observed, with fibers atrophy, abnormal mitochondria distribution and membrane remodeling. An increase in the expression level of many proteins was observed, as well as an inhibition of the autophagy process. This model demonstrates that RyR1 reduction is sufficient to recapitulate most features of Central Core Disease, and accordingly similar alterations were observed in muscle biopsies from Dusty Core Disease patients (a subtype of Central Core Disease), pointing to common pathophysiological mechanisms related to RyR1 reduction. Electronic supplementary material The online version of this article (10.1186/s40478-020-01068-4) contains supplementary material, which is available to authorized users.

The core myopathies are a group of congenital myopathies with variable clinical expression – ranging from early-onset skeletal-muscle weakness to later-onset disease of variable severity – that are identified by characteristic ‘core-like’ lesions in myofibers and the presence of hypothonia and slowly or rather non-progressive muscle weakness. The genetic causes are diverse; central core disease is most often caused by mutations in ryanodine receptor 1 (RYR1), whereas multi-minicore disease is linked to pathogenic variants of several genes, including selenoprotein N (SELENON), RYR1 and titin (TTN). Understanding the mechanisms that drive core development and muscle weakness remains challenging due to the diversity of the excitation-contraction coupling (ECC) proteins involved and the differential effects of mutations across proteins. Because of this, the use of representative models expressing a mature ECC apparatus is crucial. Animal models have facilitated the identification of disease progression mechanisms for some mutations and have provided evidence to help explain genotype-phenotype correlations. However, many unanswered questions remain about the common and divergent pathological mechanisms that drive disease progression, and these mechanisms need to be understood in order to identify therapeutic targets. Several new transgenic animals have been described recently, expanding the spectrum of core myopathy models, including mice with patient-specific mutations. Furthermore, recent developments in 3D tissue engineering are expected to enable the study of core myopathy disease progression and the effects of potential therapeutic interventions in the context of human cells. In this Review, we summarize the current landscape of core myopathy models, and assess the hurdles and opportunities of future modeling strategies., Summary: The core myopathies are neuromuscular disorders with no cure. In this Review, we outline our current understanding of pathomechanisms, the advantages and limitations of available models, and discuss emerging modeling technologies.

INTRODUCTION Congenital myopathies are muscle diseases characterized by specific histopathologic features, generalized hypotonia from birth, and perinatal complications, although some cases develop during childhood or, rarely, in adulthood. We undertook this study to characterize congenital myopathies among patients registered at our institution. METHODS Clinical, histopathologic, and genetic features were evaluated in 34 patients recruited for this study. RESULTS The majority of patients experienced a childhood onset, and no disease-related mortality was recorded during follow-up. Functional outcomes were no better for those with late-onset disease, indicating later disease progression can be significant. Nemaline myopathy was the most frequent pathology, followed by central core disease and centronuclear myopathy. Among the 18 (54.5%) genetically confirmed patients, NEB and RYR1 mutations were the most common, followed by DNM2 mutations. DISCUSSION This study shows features not previously reported and suggests that congenital myopathy should be considered an important issue among adult patients. Muscle Nerve 58: 235-244, 2018.

Background Central core disease and malignant hyperthermia are human disorders of skeletal muscle resulting from aberrant Ca2+ handling. Most malignant hyperthermia and central core disease cases are associated with amino acid changes in the type 1 ryanodine receptor (RyR1), the skeletal muscle Ca2+-release channel. Malignant hyperthermia exhibits a gain-of-function phenotype, and central core disease results from loss of channel function. For a variant to be classified as pathogenic, functional studies must demonstrate a correlation with the pathophysiology of malignant hyperthermia or central core disease. Objective We assessed the pathogenicity of four C-terminal variants of the ryanodine receptor using functional analysis. The variants were identified in families affected by either malignant hyperthermia or central core disease. Methods Four variants were introduced separately into human cDNA encoding the skeletal muscle ryanodine receptor. Following transient expression in HEK-293T cells, functional studies were carried out using calcium release assays in response to an agonist. Two previously characterized variants and wild-type skeletal muscle ryanodine receptor were used as controls. Results The p.Met4640Ile variant associated with central core disease showed no difference in calcium release compared to wild-type. The p.Val4849Ile variant associated with malignant hyperthermia was more sensitive to agonist than wild-type but did not reach statistical significance and two variants (p.Phe4857Ser and p.Asp4918Asn) associated with central core disease were completely inactive. Conclusions The p.Val4849Ile variant should be considered a risk factor for malignant hyperthermia, while the p.Phe4857Ser and p.Asp4918Asn variants should be classified as pathogenic for central core disease.

Introduction The objective of this study was to obtain a 6‐month natural history of motor function performance in individuals with RYR1‐ related myopathy (RYR1‐RM) by using the Motor Function Measure‐32 (MFM‐32) and graded functional tests (GFT) while facilitating preparation for interventional trials. Methods In total, 34 participants completed the MFM‐32 and GFTs at baseline and 6‐month visits. Results Motor deficits according to MFM‐32 were primarily observed in the standing and transfers domain (D1; mean 71%). Among the GFTs, participants required the most time to ascend/descend stairs (>7.5 s). Functional movement, determined by GFT grades, was strongly correlated with MFM‐32 (D1; r ≥ 0.770, P < 0.001). Motor Function Measure‐32 and GFT scores did not reflect any change in performance between baseline and 6‐month visits. Discussion The MFM‐32 and GFTs detected motor impairment in RYR1‐RM, which remained stable over 6 months. Thus, these measures may be suitable for assessing change in motor function in response to therapeutic intervention. Muscle Nerve 60: 80–87, 2019

Malignant hyperthermia (MH) is a potentially lethal pharmacogenetic disorder. Numerous mutations of the ryanodine receptor 1 (RyR1) found in skeletal muscles have been identified to be causative for the susceptibility to the hypermetabolic status called MH crisis. This life-threatening state can be triggered by volatile anaesthetics, depolarising muscle relaxants and excessive body exercise in individuals carrying these mutations. Susceptibility to MH shows a certain overlap with the congenital myopathy central core disease (CCD), which is also mainly based on RyR1 mutations. An increased resting calcium concentration and a reduced sarcoplasmic reticulum (SR) calcium concentration were found in muscle cells expressing RyR1 and carrying so-called leaky channel mutations, leading to an inordinate calcium outflow from the SR and to store depletion. A reduced SR calcium concentration was found in some representatives of the so-called excitation contraction (EC)-uncoupling mutations, possibly caused by reduced induction of the sarcoplasmic/endoplasmic reticulum calcium-ATPase (SERCA) gene. Furthermore, the phenomenon of store operated calcium entry (SOCE) was discovered. It describes the activation of calcium channels in the plasma membrane after store depletion in the SR. This work investigated the influences of the lack of extracellular calcium on RyR1 mutants, preventing possible effects of SOCE. The Y524S (YS) mouse model served as a representative model for leaky channel mutations, the I4895T (IT) mouse model for EC-uncoupling mutations. I compared muscle cells of wild type and heterozygous mice in calcium free and calcium containing solutions, respectively. This study confirmed findings of a reduced maximal calcium concentration and a slowed calcium release in mice heterozygous for IT and gave evidence for a similar behaviour in heterozygous YS mice. Moreover, resting calcium concentration was decreased in calcium free solution compared to calcium containing solution in the YS mouse model. Looking at calcium removal, it appeared enhanced in IT mutants in calcium free solution. On the contrary, YS mutants showed a significantly slower calcium removal than their wild type littermates. Regarding the evaluation of single-pulse induced calcium transients, the stretched exponential function was introduced as an alternative method to describe and to compare calcium removal by simple exponential models.

Myopathies due to mutations in the skeletal muscle ryanodine receptor (RYR1) gene are amongst the most common non-dystrophic neuromuscular disorders and have been associated with both dominant and recessive inheritance. Several cases with apparently de novo dominant inheritance have been reported. Here we report two siblings with features of Central Core Disease (CCD) born to unaffected parents. Genetic testing revealed a heterozygous dominant RYR1 c.14582G>A (p. Arg4861His) mutation previously identified in other CCD pedigrees. The variant was absent in blood from the asymptomatic mother but detected at low but variable levels in blood- and saliva-derived DNA from the unaffected father, suggesting that this mutation has arisen as a paternal post-zygotic de novo event. These findings suggest that parental mosaicism should be considered in RYR1-related myopathies, and may provide one possible explanation for the marked intergenerational variability seen in some RYR1 pedigrees.

Mutations in the skeletal muscle ryanodine receptor (RyR1) cause malignant hyperthermia (MH) and central core disease (CCD), whereas mutations in the cardiac ryanodine receptor (RyR2) lead to catecholaminergic polymorphic ventricular tachycardia (CPVT). Most disease-associated RyR1 and RyR2 mutations are located in the N-terminal, central, and C-terminal regions of the corresponding ryanodine receptor (RyR) isoform. An increasing body of evidence demonstrates that CPVT-associated RyR2 mutations enhance the propensity for spontaneous Ca2+ release during store Ca2+ overload, a process known as store overload-induced Ca2+ release (SOICR). Considering the similar locations of disease-associated RyR1 and RyR2 mutations in the RyR structure, we hypothesize that like CPVT-associated RyR2 mutations, MH/CCD-associated RyR1 mutations also enhance SOICR. To test this hypothesis, we determined the impact on SOICR of 12 MH/CCD-associated RyR1 mutations E2347-del, R2163H, G2434R, R2435L, R2435H, and R2454H located in the central region, and Y4796C, T4826I, L4838V, A4940T, G4943V, and P4973L located in the C-terminal region of the channel. We found that all these RyR1 mutations reduced the threshold for SOICR. Dantrolene, an acute treatment for MH, suppressed SOICR in HEK293 cells expressing the RyR1 mutants R164C, Y523S, R2136H, R2435H, and Y4796C. Interestingly, carvedilol, a commonly used β-blocker that suppresses RyR2-mediated SOICR, also inhibits SOICR in these RyR1 mutant HEK293 cells. Therefore, these results indicate that a reduced SOICR threshold is a common defect of MH/CCD-associated RyR1 mutations, and that carvedilol, like dantrolene, can suppress RyR1-mediated SOICR. Clinical studies of the effectiveness of carvedilol as a long-term treatment for MH/CCD or other RyR1-associated disorders may be warranted.

Central core disease (CCD) is a congenital myopathy linked to mutations in the ryanodine receptor type 1 (RYR1), the sarcoplasmic reticulum Ca2+ release channel of skeletal muscle. CCD is characterized by formation of amorphous cores within muscle fibers, lacking mitochondrial activity. In skeletal muscle of RYR1Y522S/WT knock-in mice, carrying a human mutation in RYR1 linked to malignant hyperthermia (MH) with cores, oxidative stress is elevated and fibers present severe mitochondrial damage and cores. We treated RYR1Y522S/WT mice with N-acetylcysteine (NAC), an antioxidant provided ad libitum in drinking water for either 2 or 6 months. Our results show that 2 months of NAC treatment starting at 2 months of age, when mitochondrial and fiber damage was still minimal, (i) reduce formation of unstructured and contracture cores, (ii) improve muscle function, and (iii) decrease mitochondrial damage. The beneficial effect of NAC treatment is also evident following 6 months of treatment starting at 4 months of age, when structural damage was at an advanced stage. NAC exerts its protective effect likely by lowering oxidative stress, as supported by the reduction of 3-NT and SOD2 levels. This work suggests that NAC administration is beneficial to prevent mitochondrial damage and formation of cores and improve muscle function in RYR1Y522S/WT mice.

Background Pathogenic variants in ryanodine receptor type 1 (RYR1) gene are an important cause of congenital myopathy. The clinical, histopathologic and genetic spectrum is wide. Objective Review a group of the patients diagnosed with ryanodinopathy in a tertiary centre from North Portugal, as an attempt to define some phenotypical patterns that may help guiding future diagnosis. Methods Patients were identified from the database of the reference centre for Neuromuscular Disorders in North Portugal. Their data (clinical, histological and genetic) was retrospectively accessed. Results Seventeen RYR1-related patients (including 4 familial cases) were identified. They were divided in groups according to three distinctive clinical characteristics: extraocular muscle (EOM) weakness (N = 6), disproportionate axial muscle weakness (N = 2) and joint laxity (N = 5). The fourth phenotype includes patients with mild tetraparesis and no distinctive clinical features (N = 4). Four different histopathological patterns were found: centronuclear (N = 5), central core (N = 4), type 1 fibres predominance (N = 4) and congenital fibre type disproportion (N = 1) myopathies. Each index case, except two patients, had a different RYR1 variant. Four new genetic variants were identified. All centronuclear myopathies were associated with autosomal recessive inheritance and EOM weakness. All central core myopathies were caused by pathogenic variants in hotspot 3 with autosomal dominant inheritance. Three genetic variants were reported to be associated to malignant hyperthermia susceptibility. Conclusions Distinctive clinical features were recognized as diagnostically relevant: extraocular muscle weakness (and centronuclear pattern on muscle biopsy), severe axial weakness disproportionate to the ambulatory state and mild tetraparesis associated with (proximal) joint laxity. There was a striking genetic heterogeneity, including four new RYR1 variants.

Central core disease (CCD) is a genetically heterogeneous congenital myopathy, and ryanodine receptor 1 ( RYR1 , gene ID6261) is the only pathogenicity gene until now. Data on mutation characteristics of RYR1 in the Chinese CCD population are scarce. This study searched for mutations in the C-terminal-encoding domain of RYR1 in six Chinese patients with CCD, and identified five missense mutations (N4807F, R4861H, R4893P, G4897D, and I4898T). Among them, N4807F, G4897D were novel while R4861H, R4893P, and I4898T were previously reported. All missense mutations were highly conserved across the species of human, mouse, rabbit, fish, and pig. This study found that mutations could be identified in about 85% CCD patients, even if only the C-terminal-encoding region of RYR1 was screened. Many mutations clustered in exons 100–102.

Summary The core myopathies are a subset of myopathies that present in infancy with hypotonia and muscle weakness. They were formerly considered a rare type of congenital myopathy but are now recognized as being more prevalent. Due to their genetic linkage to mutations in the ryanodine receptor gene (RYR1), core myopathies (in particular, central core disease) carry a high risk of malignant hyperthermia susceptibility. In this review article, we describe the phenotypical, genetic, and histopathological characteristics of core myopathies and further describe the currently understood nature of their risk of malignant hyperthermia. We also review the level of suspicion a clinician should exhibit with a child who has a possible core myopathy or other congenital myopathy presenting for an anesthetic prior to a definitive genetic analysis. For this review article, we performed literature searches using the key words anesthesiology, core myopathies, pediatric neurology, malignant hyperthermia, genetics, ryanodine receptor, and molecular biology. We also relied on literature accumulated by the two authors, who served as hotline consultants for the Malignant Hyperthermia Hotline of the Malignant Hyperthermia Association of the United States (MHAUS) for the past 12 years.

Central core disease (CCD) is a dominantly inherited congenital myopathy. CCD is also associated with muscular and skeletal abnormalities such as abnormal curvature of the spine (scoliosis), hip dislocation, and joint deformities. CCD and malignant hyperthermia (MH) are both associated with mutations in the ryanodine receptor on chromosome 19q13.1. An 11-year-old boy with CCD complicated with severe scoliosis was scheduled for spinal fusion surgery under general anesthesia. Furthermore, he had trismus caused by temporomandibular contracture. He was considered as MH susceptible. Anesthesia was managed with remifentanil and propofol without using muscle relaxtants and volatile anesthetics. We could intubate the trachea with Airtraq laryngoscope without any complications. The perioperative course was uneventful.

Central core disease is a rare inherited neuromuscular disorder caused by mutations in ryanodine receptor type 1 gene. The clinical phenotype of the disease is highly variable. We report a Chinese pedigree with central core disease confirmed by the gene sequencing. All 3 patients in the family presented with mild proximal limb weakness. The serum level of creatine kinase was normal, and electromyography suggested myogenic changes. The histologic analysis of muscle biopsy showed identical central core lesions in almost all of the muscle fibers in the index case. Exon 90-106 in the C-terminal domain of the ryanodine receptor type 1 gene was amplified using polymerase chain reaction. One heterozygous missense mutation G14678A (Arg4893Gln) in exon 102 was identified in all 3 patients. This is the first report of a familial case of central core disease confirmed by molecular study in mainland China.

The type 1 ryanodine receptor (RyR1) is expressed widely in the brain, with high levels in the cerebellum, hippocampus, and hypothalamus. We have shown that L-type Ca 2+ channels in terminals of hypothalamic magnocellular neurons are coupled to RyRs, as they are in skeletal muscle, allowing voltage-induced Ca 2+ release (VICaR) from internal Ca 2+ stores without Ca 2+ influx. Here we demonstrate that RyR1 plays a role in VICaR in nerve terminals. Furthermore, in heterozygotes from the Ryr1 I4895T/WT (IT/+) mouse line, carrying a knock-in mutation corresponding to one that causes a severe form of human central core disease, VICaR is absent, demonstrating that type 1 RyR mediates VICaR and that these mice have a neuronal phenotype. The absence of VICaR was shown in two ways: first, depolarization in the absence of Ca 2+ influx elicited Ca 2+ syntillas ( scintilla , spark, in a nerve terminal, a SYNaptic structure) in WT, but not in mutant terminals; second, in the presence of extracellular Ca 2+ , IT/+ terminals showed a twofold decrease in global Ca 2+ transients, with no change in plasmalemmal Ca 2+ current. From these studies we draw two conclusions: ( i ) RyR1 plays a role in VICaR in hypothalamic nerve terminals; and ( ii ) a neuronal alteration accompanies the myopathy in IT/+ mice, and, possibly in humans carrying the corresponding RyR1 mutation.

The core myopathies, Central Core Disease and Multiminicore Disease, are heterogeneous congenital myopathies with the common defining histopathological feature of focally reduced oxidative enzyme activity (central cores, multiminicores). Mutations in the gene encoding for the skeletal muscle ryanodine (RyR1) receptor are the most common cause. Mutations in the selenoprotein N (SEPN1) gene cause a less common variant. Pathogenic mechanisms underlying dominant RYR1 mutations have been extensively characterized, whereas those associated with recessive RYR1 and SEPN1 mutations are emerging. Identifying a specific genetic defect from the histopathological diagnosis of a core myopathy is complex and ought to be informed by a combined appraisal of histopathological, clinical, and, increasingly, muscle magnetic resonance imaging data. The present review aims at giving an overview of the main genetic and clinicopathological findings, with a major emphasis on features likely to inform the diagnostic process, as well as current treatments and perspectives for future research.

Introduction: Ryanodine receptor gene (RYR1) mutations have been associated with central core disease (CCD), multiminicore/minicore/multicore disease (MmD), and susceptibility to malignant hyperthermia (MH). Methods: Pa- tients with muscle symptoms in adulthood, who had features compatible with CCD/MmD, underwent clinical, histological, and genetic (RYR1 and SEPN1 genes) evaluations. Published cases of CCD and MmD with adult onset were also reviewed. Results: Eight patients fulfilled the criteria for further analysis. Five RYR1 mutations, 4 of them unreported, were detected in 3 patients. Compound heterozygosity was proven in 1 case. Con- clusions: To our knowledge, this is the only report of adult onset associated with recessive RYR1 mutations and central core/multiminicores on muscle biopsy. Although adult patients with CCD, MmD, and minimally symptomatic MH with abnormal muscle biopsy findings usually have a mild clinical course, dif- ferential diagnosis and carrier screening is crucial for prevention of potentially life-threatening reactions to general anesthesia. Muscle Nerve 44: 102-108, 2011

PURPOSE OF REVIEW: Anaesthesia for a child with a muscle disease is always challenging because there is a risk of malignant hyperthermia, rhabdomyolysis or hypermetabolic reaction if a halogenated agent is used. Ongoing progress in genetics helps in clarifying the link between malignant hyperthermia (a calcium channelopathy) and muscle diseases. RECENT FINDINGS: We provide a summary of the most recent clinical, pathophysiological and genetic information on those risks when the diagnosis is known or suspected preoperatively. Some simple clues are also given to help make a decision in the presence of an infant or child with hypotonia or motor delay but no diagnosis. SUMMARY: Only a few muscle diseases are really associated with a risk of malignant hyperthermia. The risk of rhabdomyolysis is more difficult to clarify and a multicentric database would be useful to evaluate the risk/benefit ratio of all anaesthetic drugs in patients with muscle diseases.

Ryanodine receptors (RyR) are intracellular Ca2+-permeable channels that provide the sarcoplasmic reticulum Ca2+ release required for skeletal and cardiac muscle contractions. RyR1 underlies skeletal muscle contraction, and RyR2 fulfills this role in cardiac muscle. Over the past 20 years, numerous mutations in both RyR isoforms have been identified and linked to skeletal and cardiac diseases. Malignant hyperthermia, central core disease, and catecholaminergic polymorphic ventricular tachycardia have been genetically linked to mutations in either RyR1 or RyR2. Thus, RyR channelopathies are both of interest because they cause significant human diseases and provide model systems that can be studied to elucidate important structure–function relationships of these ion channels.

To summarize advances in our understanding of the clinical phenotypes, genetics, and molecular pathophysiology of the periodic paralyses, the nondystrophic myotonias, and other muscle channelopathies.The number of pathogenic mutations causing periodic paralysis, nondystrophic myotonias, and ryanodinopathies continues to grow with the advent of exon hierarchy analysis strategies for genetic screening and better understanding and recognition of disease phenotypes. Recent studies have expanded and clarified the role of gating pore current in channelopathy pathogenesis. It has been shown that the gating pore current can account for the molecular and phenotypic diseases observed in the muscle sodium channelopathies, and, given that homologous residues are affected in mutations of calcium channels, it is possible that pore leak represents a pathomechanism applicable to many channel diseases. Improvements in treatment of the muscle channelopathies are on the horizon. A randomized controlled trial has been initiated for the study of mexiletine in nondystrophic myotonias. The class IC antiarrhythmia drug flecainide has been shown to depress ventricular ectopy and improve exercise capacity in patients with Andersen-Tawil syndrome.Recent studies have expanded our understanding of gating pore current as a disease-causing mechanism in the muscle channelopathies and have allowed new correlations to be drawn between disease genotype and phenotype.

The dynamic actin cytoskeleton, consisting of six actin isoforms in mammals and a variety of actin binding proteins is essential for all developmental processes and for the viability of the adult organism. Actin isoform specific functions have been proposed for muscle contraction, cell migration, endo- and exocytosis and maintaining cell shape. However, these specific functions for each of the actin isoforms during development are not well understood. Based on transgenic mouse models, we will discuss the expression patterns of the six conventional actin isoforms in mammals during development and adult life. Ablation of actin genes usually leads to lethality and affects expression of other actin isoforms at the cell or tissue level. A good knowledge of their expression and functions will contribute to fully understand severe phenotypes or diseases caused by mutations in actin isoforms.

In this article, we analyze myopathies with cores, for which an association to malignant hyperthermia (MH) has been suggested. We discuss the clinical features, the underlying genetic defects, subsequent effects on cellular calcium metabolism, and in vitro muscle responses to MH triggers. We describe in detail central core disease, multiminicore disease, and nemaline rod myopathy. We categorize the diseases according to the affected proteins and discuss the risk for MH, which is high or theoretically possible when the calcium-conducting proteins are affected.

Muscle contraction and relaxation is regulated by transient elevations of myoplasmic Ca 2+ . Ca 2+ is released from stores in the lumen of the sarco(endo)plasmic reticulum (SER) to initiate formation of the Ca 2+ transient by activation of a class of Ca 2+ release channels referred to as ryanodine receptors (RyRs) and is pumped back into the SER lumen by Ca 2+ -ATPases (SERCAs) to terminate the Ca 2+ transient. Mutations in the type 1 ryanodine receptor gene, RYR1 , are associated with 2 skeletal muscle disorders, malignant hyperthermia (MH), and central core disease (CCD). The evaluation of proposed mechanisms by which RyR1 mutations cause MH and CCD is hindered by the lack of high-resolution structural information. Here, we report the crystal structure of the N-terminal 210 residues of RyR1 (RyR NTD ) at 2.5 Å. The RyR NTD structure is similar to that of the suppressor domain of type 1 inositol 1,4,5-trisphosphate receptor (IP 3 Rsup), but lacks most of the long helix-turn-helix segment of the “arm” domain in IP 3 Rsup. The N-terminal β-trefoil fold, found in both RyR and IP 3 R, is likely to play a critical role in regulatory mechanisms in this channel family. A disease-associated mutation “hot spot” loop was identified between strands 8 and 9 in a highly basic region of RyR1. Biophysical studies showed that 3 MH-associated mutations (C36R, R164C, and R178C) do not adversely affect the global stability or fold of RyR NTD , supporting previously described mechanisms whereby mutations perturb protein–protein interactions.

Mutations in the skeletal muscle ryanodine receptor (RYR1) gene have been associated with a wide range of phenotypes including the malignant hyperthermia (MH) susceptibility trait, Central Core Disease (CCD) and other congenital myopathies characterized by early onset and predominant proximal weakness. We report a patient presenting at 77 years with a predominant axial myopathy associated with prominent involvement of spine extensors, confirmed on MRI and muscle biopsy, compatible with a core myopathy. RYR1 mutational analysis revealed a novel heterozygous missense mutation (c.119G>T; p.Gly40Val) affecting the RYR1 N-terminus, previously predominantly associated with MH susceptibility. This case expands the spectrum of RYR1-related phenotypes and suggests that MH-related RYR1 mutations may give rise to overt neuromuscular symptoms later in life, with clinical features not typically found in CCD due to C-terminal hotspot mutations. Late-onset congenital myopathies may be under-recognised and diagnosis requires a high degree of clinical suspicion.

Summary We describe the anaesthetic management of a 21-year-old lady with central core disease for elective Caesarean section. Central core disease is characterised by muscle weakness, skeletal deformities and susceptibility to malignant hyperthermia. Total intravenous anaesthesia was used because of the combination of potential malignant hyperthermia, severe kyphoscoliosis and extensive spinal scarring. The authors believe there is no previous report of propofol and remifentanil being used in these circumstances. A short review of central core disease and its anaesthetic implications is provided.

We report the case of a 7-year-old boy with thoracolumbar scoliosis and central core disease who had a history of malignant hyperthermia. He had scoliosis with Cobb's angle deteriorating to 67 degrees (thoracic) and 59 degrees (lumbar). A provocation test of general anesthesia was performed to confirm no hyperthermic reaction. Then, he underwent surgical correction by a posterior approach. The thoracic curve was reduced to 38 degrees and the lumbar curve to 42 degrees . He has been followed up for 2 years without any complications. This is the first report with a detailed description of perioperative management on surgical treatment of scoliosis associated with central core disease.

Central core disease (CCD) and malignant hyperthermia (MH) are skeletal muscle disorders that are linked to mutations in the gene that encodes the type 1 ryanodine receptor (RYR1). The RYR1 ion channel plays a central role in excitation-contraction (EC) coupling by releasing Ca(2+) from an internal store. Pathogenic CCD mutations in RYR1 result in changes in the magnitude of Ca(2+) release during EC coupling. CCD has recently been linked to two novel deletions (c.12640_12648delCGCCAGTTC [p.Arg4214_Phe4216del] and c.14779_14784delGTCATC [p.Val4927_Ile4928del]) in the C-terminal region of RYR1. To determine the phenotypic consequences of these mutations and extend our understanding of the pathogenic mechanisms that underlie CCD, we determined functional effects on Ca(2+) release channel activity of analogous deletions (p.Arg4215_Phe4217del and p.Val4926_Ile4927del) engineered into rabbit RYR1 following expression in RYR1-null (dyspedic) myotubes and HEK293 cells. In addition, we assessed effects of the p.Arg4214 Phe4216del mutation on RYR1 function in lymphoblastoid cells obtained from CCD patients heterozygous for the mutation. Here we report that both deletions significantly reduce Ca(2+) release following RYR1 activation, but by different mechanisms. While the p.Arg4214_Phe4216del deletion promotes Ca(2+) depletion from intracellular stores by exhibiting a classic "leaky channel" behavior, the p.Val4927_Ile4928del deletion reduces Ca(2+) release by disrupting Ca(2+) gating and eliminating Ca(2+) permeation through the open channel.

Malignant hyperthermia is a genetic defect of uncontrolled hypermetabolic skeletal muscle response to anesthetic triggering drugs. Some congenital myopathies are regarded as risk increasing factors. The use of volatile anaesthetics or suxamethonium (succinylcholine) in patients who are predisposed to malignant hyperthermia leads to an increase in Ca2+ release from sarcoplasmic reticulum, which in turn causes a set of biochemical and clinical symptoms, which can be a cause of death, if dantrolene is not administered adequately. The aim of the study was to draw attention to the problem of malignant hyperthermia, which is hardly ever described in Polish literature, and requires the necessity of intensifying the cooperation between the dentist and specialists from other medical fields. The origin of the article was a case of congenital myopathy with recognized malignant hyperthermia in an 18-year-old patient, in whom surgical extraction of teeth was indicated. The course of diagnostics and treatment showed once more that contemporary medicine is in need of holistic approach, and in consequence, promising and effective cooperation of many specialists.

Ryanodine receptor 1 (RYR1), myosin heavy chain 7 (MYH7), and selenoprotein N1 (SEPN1) mutations are associated with core myopathies. RYR1 mutations cause most cases of central core disease (CCD).We screened 8 Chinese patients with clinicopathological diagnosis of CCD. Genetic analysis was carried out by targeted next generation sequencing (NGS) to identify causative genes. Variants were assessed for pathogenicity using bioinformatic approaches, and NGS results were confirmed by Sanger sequencing.One novel (p.L4578V) and heterozygous missense mutations in RYR1 were identified in 7 patients. Two patients carried a novel mutation, 1 had p.M4640R, 3 had p.R4861H, and 1 had p.R4861C. All patients had mild to moderate severity phenotypes. Histopathological findings demonstrated central cores and type I fiber predominance.NGS is an efficient strategy to identify variants in RYR1 in CCD. However, genetic results revealed by NGS must be combined with clinicopathologic features to validate the diagnosis. Muscle Nerve, 2016 Muscle Nerve 54: 432-438, 2016.

Congenital lumbar hernias are rare malformations caused by defects in the development of the posterior abdominal wall. A known association exists with lumbocostovertebral syndrome; however other associated anomalies, including one case with arthrogryposis, have been previously reported. We present an infant girl with bilateral congenital lumbar hernias, multiple joint contractures, decreased muscle bulk and symptoms of malignant hyperthermia. Molecular testing revealed an R4861C mutation in the ryanodine receptor 1 (RYR1) gene, known to be associated with central core disease. This is the first reported case of the co-occurrence of congenital lumbar hernias and central core disease. We hypothesize that ryanodine receptor 1 mutations may interrupt muscle differentiation and development. Further, this case suggests an expansion of the ryanodine receptor 1-related myopathy phenotype to include congenital lumbar hernias.

The type 1 ryanodine receptor (RyR1) is a Ca2+ release channel in the sarcoplasmic reticulum of skeletal muscle and is mutated in several diseases, including malignant hyperthermia (MH) and central core disease (CCD). Most MH and CCD mutations cause accelerated Ca2+ release, resulting in abnormal Ca2+ homeostasis in skeletal muscle. However, how specific mutations affect the channel to produce different phenotypes is not well understood. In this study, we have investigated 11 mutations at 7 different positions in the amino (N)-terminal region of RyR1 (9 MH and 2 MH/CCD mutations) using a heterologous expression system in HEK293 cells. In live-cell Ca2+ imaging at room temperature (~25 °C), cells expressing mutant channels exhibited alterations in Ca2+ homeostasis, i.e., an enhanced sensitivity to caffeine, a depletion of Ca2+ in the ER and an increase in resting cytoplasmic Ca2+. RyR1 channel activity was quantitatively evaluated by [3H]ryanodine binding and three parameters (sensitivity to activating Ca2+, sensitivity to inactivating Ca2+ and attainable maximum activity, i.e., gain) were obtained by fitting analysis. The mutations increased the gain and the sensitivity to activating Ca2+ in a site-specific manner. The gain was consistently higher in both MH and MH/CCD mutations. Sensitivity to activating Ca2+ was markedly enhanced in MH/CCD mutations. The channel activity estimated from the three parameters provides a reasonable explanation to the pathological phenotype assessed by Ca2+ homeostasis. These properties were also observed at higher temperatures (~37 °C). Our data suggest that divergent activity profiles may cause varied disease phenotypes by specific mutations. This approach should be useful for diagnosis and treatment of diseases with mutations in RyR1.

Objective: To characterize the muscle involvement of patients with central core disease (CCD) caused by mutations in the ryanodine receptor 1 gene ( RYR1 ) and to compare these findings with those from patients with core myopathies unlinked to the RYR1 gene. Methods: We performed a systematic muscular imaging assessment in 11 patients with an RYR1 gene mutation and compared these findings with those of 5 patients from two unrelated families with autosomal dominant core myopathies not linked to RYR1 , ACTA1 , or MYH7 gene loci. Results: All patients with RYR1 CCD had a characteristic pattern with predominant involvement of the gluteus maximus, adductor magnus, sartorius, vastus intermediolateralis, soleus, and lateral gastrocnemius muscles. In contrast, muscle CT in the first family not linked to RYR1 showed predominant affection of the gluteus minimus and hamstring muscles, whereas the second family presented with predominant involvement of the gluteus minimus, vastus intermediolateralis, tibialis anterior, and medial gastrocnemius muscles. In addition to muscle imaging data, we present detailed information on the clinical and pathologic findings of these novel phenotypes of core myopathies not linked to RYR1 . Conclusions: Our data suggest genetic heterogeneity in autosomal dominant core myopathies and the existence of additional unidentified genes.