A novel mutation of the CLCN1 gene associated with myotonia hereditaria in an Australian cattle dog

BACKGROUND: Heritable myotonia is a genetic muscle disorder characterized by slow relaxation of skeletal muscles. The main clinical signs are skeletal muscle stiffness, especially after vigorous contraction, and muscle hypertrophy. Muscle stiffness may be enhanced by inactivity, and often is relieved by exercise. Myotonia can be inherited in an autosomal dominant or recessive manner (Thomsen- or Becker-type myotonia, respectively). In mice, goats, Miniature Schnauzer dogs, and most affected humans, the disorder is caused by mutations in CLCN1, which encodes the skeletal muscle voltage-gated chloride channel, Cl1C-1. HYPOTHESIS: We hypothesized that an Australian Cattle Dog with generalized muscle stiffness and hypertrophy examined at the Ontario Veterinary College would have a mutation in the CLCN1 gene. ANIMALS: A pure-bred Australian Cattle Dog from Ontario, Canada, was used. METHODS: Based on clinical signs and electromyographic test results, a diagnosis of myotonia hereditaria was made, and a muscle biopsy was collected for genetic analysis. RESULTS: Sequence data obtained from the affected dog confirmed that it was homozygous for a single base insertion in the CLCN1 coding sequence. This mutation would result in a truncated ClC-1 protein being expressed, which, based on molecular evidence from other studies, would result in functionally compromised chloride conduction in the skeletal muscles of the animal. CONCLUSIONS AND CLINICAL IMPORTANCE: To the authors' knowledge, this report describes the Ist case of myotonia in an Australian Cattle Dog and represents the 1st non-Schnauzer canine myotonia to be genetically characterized. In addition, we developed a polymerase chain reaction-based genetic screen to detect heterozygotes with this mutation in the at-large Australian Cattle Dog population.     

Effect of phenytoin on the clinical signs and in vitro muscle twitch characteristics in horses with chronic intermittent rhabdomyolysis and myotonia

In vitro twitch characteristics of the semimembranosus muscle were evaluated in 9 clinically normal horses, in 15 horses with chronic intermittent rhabdomyolysis (CIR) and in 2 horses with myotonia. Effects of phenytoin on in vitro muscle twitch and clinical signs of CIR and myotonia were evaluated in these same horses. Times to 90% relaxation were prolonged in the horses with CIR (mean +/- SEM, 186 +/- 5.9 ms) and in 2 horses with myotonia (197 and 177 ms) compared with those in clinically normal horses (mean +/- SEM, 146 +/- 2.1 ms). Horses with CIR also had significantly (P less than 0.05) longer times to 50% relaxation, compared with clinically normal horses. In the group of horses with CIR, Standardbreds had significantly (P less than 0.05) longer times to 90% and 50% relaxation, compared with Thoroughbreds. Times to 100% peak tension did not differ among the groups. Administration of phenytoin directly into a muscle preparation bath solution had no effect on muscle twitch properties. After the initial muscle biopsy, phenytoin was administered orally for 7 to 10 days to 4 horses with CIR, 2 myotonic horses, and 2 clinically normal horses before repeat biopsy from the same site in the contralateral semimembranosus muscle. Times to 90% relaxation decreased from 197 and 177 ms to 144 and 126 ms, respectively, in the 2 myotonic horses, from a mean of 192 (+/- 9) ms to 170 (+/- 9) ms in the 4 horses with CIR and remained unchanged (154 and 140 ms before vs 155 and 139 ms after treatment) in the 2 clinically normal horses.(ABSTRACT TRUNCATED AT 250 WORDS)     

Myotonic dystrophy-like disease in a dog

A mature female Rhodesian Ridgeback was determined to have a progressive, degenerative myopathy associated with myotonia, dysphagia, and marked muscle wasting. Clinical findings revealed a diffuse muscular disease with percussion dimpling, dysphagia, and creatine kinase elevation. A paroxysmal atrial tachycardia was found. Electromyography revealed a diffuse myopathy with high-frequency bizarre waves, myotonic discharges especially in the masticatory, laryngeal, and pharyngeal muscles. A few positive sharp waves were found in some of the appendicular muscles. Histopathologic and histochemical stains on skeletal muscle biopsy specimens demonstrated moderate fiber-size variation, myofiber architectural changes, muscle-fiber splitting, focal necrosis and phagocytosis, high percentage of internal nuclei, and atrophy of type-2 muscle fibers. A review of myotonic myopathies in the dog is presented. The clinical, electrophysiologic, and histochemical findings are similar to those for myotonic muscular dystrophy in man.     

Hereditary myotonia in the Chow Chow

Four related Chow Chow puppies which were presented because of stiffness in their movements were shown to have myotonia similar to myotonia congenita of man. Electromyography revealed characteristic myotonic discharges. Repetitive nerve stimulation studies showed a marked fade in the compound muscle action potential (CMAP) which was most apparent at stimulation rates of 10 per sec or higher. If stimulation was continued at a rate of 10 per sec the CMAP returned to normal or sometimes greater than normal following several seconds of the reduced response. The decremental response could be exaggerated by cooling the muscle prior to repetitive nerve stimulation. A therapeutic trial to assess the efficacy of the commonly used membrane stabilizing agents, quinidine, procainamide and phenytoin, indicated that all three drugs were beneficial in the treatment of myotonia but procainamide produced the best response. Hypocholesterolaemia was documented in one case. The possibility of a multisystem membrane defect in hereditary myotonia associated with low serum cholesterol was considered.     

A regional curare test for evaluation of myotonia in dogs

This paper describes a regional curare test performed in a dog suffering from myotonia and hyperadrenocorticism. The test prevented innervation to the pronator teres muscle of the left limb and allowed electrical activity to be observed in the muscle without the need for a general neuromuscular blockade; various types of spontaneous activity other than myotonic discharges were also observed. Details of the method are given and its usefulness in the evaluation of myopathies is discussed.     

Pinpointing the candidate region for muscular dystrophy in chickens with an abnormal muscle gene

Muscular dystrophies, a group of inherited diseases with the progressive weakness and degeneration of skeletal muscle, contain genetically variable diseases. Though chicken muscular dystrophy with abnormal muscle (AM) has long been known, the gene responsible has not yet been identified. In this study, a resource family for AM was established with 487 F2 individuals and 22 gene markers, including microsatellite and insertion–deletion markers, were developed. The haplotypes were analyzed with these markers for the candidate region of GGA2q described in a previous study. The candidate region was successfully narrowed down to approximately 1Mbp. The region included seven functional genes predicted as the most likely AM candidates.     

Congenital myotonia in related kittens

Four closely related domestic shorthair kittens were investigated following the detection of abnormalities in their gait, difficulty opening their mouths and muscle hypertrophy. They walked with a stiff, stilted gait, with the stiffness reducing during exercise. Startling of the kittens resulted in hyperextension of the limbs and falling to lateral recumbency, or spasm of the orbicularis oculi muscle, prolonged prolapse of the nictitating membranes and flattening of the ears. One kitten was intermittently dysphonic. Endotracheal intubation of the anaesthetised kittens was difficult due to an inability to open the mouth to a wide angle, and narrowing of the glottis due to muscle spasm. A diagnosis of congenital myotonia was made based on the clinical signs, the kittens' ages, typical myotonic discharges on electromyography, and the histopathological and histochemical findings in muscle. This is the first report of congenital myotonia in this species.     

Myotonia in a chow chow

Congenital myotonia was diagnosed in an inbred Chow Chow pup with severe muscle stiffness that regressed with exercise. Voluntary movement, percussion, or needle insertion caused sustained contraction of the muscles involved. Electromyographic recordings from several muscles contained myotonic discharges. Creatine kinase activity was mildly increased. Slight myofiber hypertrophy and a few atrophic fibers were seen in muscle biopsy specimens. Treatment of the pup with procainamide caused a marked decrease in clinical signs. Myotonia congenita in the Chow Chow appears to be inherited as an autosomal recessive trait. This condition can be diagnosed on the basis of clinical signs. Satisfactory clinical management of myotonia congenita can be achieved with procainamide.     

Dental and craniofacial findings in eight miniature schnauzer dogs affected by myotonia congenita: preliminary results

Myotonia is a clinical sign characterized by the delay of skeletal muscle relaxation following the cessation of a voluntary activity or the termination of an electrical or mechanical stimulus. Recently, Miniature Schnauzers with myotonia congenita associated with defective chloride ion conductance across the skeletal muscle membrane were identified. Congenital myotonia in these dogs appears to follow an autosomal recessive mode of inheritance. Craniofacial and dental findings of eight Miniature Schnauzer dogs with myotonia congenita are described in the present paper. These findings include: delayed dental eruption of both deciduous and permanent dentition: persistent deciduous dentition; unerupted or partially erupted permanent teeth: crowding and rotation of premolar and or incisor teeth: missing teeth: increased interproximal space between the maxillary fourth premolar and first molar teeth: decreased interproximal space between the maxillary canine and lateral incisor teeth: inability to fully close the mouth due to malocclusion: distoclusion: and, decreased mandibular range of motion. A long narrow skull with a flattened zygomatic arch and greater mandibular body curvature were also consistent findings in the affected dogs. The small number of dogs studied prevents conclusive statements about the origin of these abnormalities, however it is interesting that only 1 of 45 unaffected Miniature Schnauzer dogs showed similar traits.     

Histologic margins and the residual tumour classification scheme: Is it time to use a validated scheme in human oncology to standardise margin assessment in veterinary oncology?

There is no consensus on the definition of a complete histologic excision in veterinary oncology; many definitions have been used in various studies, but these have been arbitrarily selected with no apparent justification. The residual tumour classification scheme, where a complete histologic excision is defined as a histologic tumour‐free margin >0 mm, has been used for >40 years in human oncology by all of the major clinical staging organizations and is considered highly prognostic for the vast majority of malignant tumours in people. Because of the widespread use of the residual tumour classification scheme both clinically and in research studies, this standardized approach permits better communication between clinicians, an evidence‐based decision‐making process for adjuvant treatment options following surgical resection, minimizes exposing patients to unnecessary adjuvant treatments and a better ability to compare local tumour control for specific tumours between different studies. The adoption of the residual tumour classification scheme in veterinary oncology would likely achieve similar outcomes and minimize the prevalent confusion within the veterinary community, amongst both general practitioners and specialists, regarding the definition of what constitutes a complete histologic excision.     

Dysferlin and animal models for dysferlinopathy

Dysferlin (DYSF) is involved in the membrane-repair process, in the intracellular vesicle system and in T-tubule development in skeletal muscle. It interacts with mitsugumin 53, annexins, caveolin-3, AHNAK, affixin, S100A10, calpain-3, tubulin and dihydropyridine receptor. Limb-girdle muscular dystrophy 2B (LGMD2B) and Miyoshi myopathy (MM) are muscular dystrophies associated with recessively inherited mutations in the DYSF gene. The diseases are characterized by weakness and muscle atrophy that progress slowly and symmetrically in the proximal muscles of the limb girdles. LGMD2B and MM, which are collectively termed "dysferlinopathy", both lead to abnormalities in vesicle traffic and membrane repair at the plasma membrane in muscle fibers. SJL/J (SJL) and A/J mice are naturally occurring animal models for dysferlinopathy. Since there has been no an approach to therapy for dysferlinopathy, the immediate development of a therapeutic method for this genetic disorder is desirable. The murine models are useful in verification experiments for new therapies and they are valuable tools for identifying factors that accelerate dystrophic changes in skeletal muscle. It could be possible that the genetic or immunological background in SJL or A/J mice could modify muscle damage in experiments involving these models, because SJL and A/J mice show differences in the progress and prevalent sites of skeletal muscle lesions as well as in the gene-expression profiles of their skeletal muscle. In this review, we provide up-to-date information on the function of dysferlin, the development of possible therapies for muscle dystrophies (including dysferlinopathy) and the detection of new therapeutic targets for dysferlinopathy by means of experiments using animal models for dysferlinopathy.     

Myotonia associated with hyper-adrenocorticism in two dogs

Two dogs developed a disabling gait abnormality characterised by stiffness. The abnormality was consistent with a diagnosis of myotonia secondary to hyper-adrenocorticism. The first dog had iatrogenic hyperadrenocorticism, and its signs improved substantially after corticosteroid administration was gradually withdrawn. The second had pituitary-dependent hyperadrenocorticism, but myotonic signs progressed despite effective mitotane therapy. Procainamide administration reduced the myotonic stiffness in the second case.     

Myotonia and disorders of altered muscle cell membrane excitability.

Altered excitability of the skeletal muscle membrane (sarcolemma) can result in clinical signs of muscle dysfunction. Hyperexcitability of the sarcolemma results in myotonia, and hypoexcitability results in paresis or paralysis. Our understanding of the physiologic and molecular bases of disorders of sarcolemmal excitability is rapidly increasing as techniques for evaluation are improved. This article reviews muscle excitability disorders in dogs and cats and their pathogenesis.     

Histochemical changes in skeletal muscles of four male horses with neuromuscular disease

Skeletal muscle biopsy specimens were taken from 4 male horses with neuromuscular disease such as myotonia congenita, chronic myositis, exertional rhabdomyolysis, and shivers. Histologic and histochemical techniques were used to evaluate skeletal muscle morphologic features and fiber-type population, size, and area, as well as muscle enzyme activities (acid phosphatase, alkaline phosphatase, and esterase). A histochemical and histologic profile were described for each muscle biopsy specimen.     

Foetal development of skeletal muscle in bovines as a function of maternal nutrition,foetal sex and gestational age

To determine the effects of maternal nutrition on modifications of foetal development of the skeletal muscle and possible increase in the potential of skeletal muscle growth in cattle, gestating cows were either fed 190% NRC recommendations (overnourished; ON) or 100% NRC recommendation (control; CO). Interaction between maternal nutrition (MN) and the foetal sex (FS) was also investigated. Foetuses were necropsied at four different time points throughout gestation (139, 199, 241 and 268 days of gestation) to assess the mRNA expression of myogenic, adipogenic and fibrogenic markers in skeletal muscle. Phenotypic indicators of the development of skeletal muscle fibres, intramuscular lipogenesis and collagen development were also evaluated. Modifications in mRNA expression of skeletal muscle of foetuses were observed in function of MN and FS despite the lack of effect of MN and FS on foetal weight at necropsy. Maternal ON increased the mRNA expression of the myogenic marker Cadherin‐associated protein, beta 1 (CTNNB1) and adipogenic markers Peroxissome proliferator‐activated receptor gamma (PPARG) and Zinc finger protein 423 (ZNF423) at midgestation. However, no differences on foetal skeletal muscle development were observed between treatments at late gestation indicating that a compensatory development may have occurred on CO foetuses making the effect of MN on skeletal muscle development not significant at late gestation. Moreover, our data have shown an evidence of sexual dimorphism during foetal stage with a greater skeletal muscle development in male than in female foetuses. In conclusion, providing a higher nutritional level to pregnant cows changes the trajectory of the development of skeletal muscle during midgestation, but apparently does not change the potential of post‐natal growth of muscle mass of the offspring, as no differences in skeletal muscle development were observed in late gestation.     

Ex vivo bupivacaine treatment results in increased adipogenesis of skeletal muscle cells in the rat

Intramuscular adipose tissue (IMAT) is observed in some skeletal muscle pathologies. IMAT is implicated not only in the disorders of muscle contraction, but also of metabolism and insulin sensitivity due to its nature as a secretary organ. Several studies indicate the presence of cells with adipogenic potential in skeletal muscle. However, the mechanism of fate specification that triggers these cells to enter an adipogenic program in vivo remains to be solved. In the present study, we examined whether activation of the adipogenic program of muscle‐resident cells precedes their proliferation upon muscle injury. For this purpose, muscle injury was induced by injecting bupivacaine (BPVC) to excised skeletal muscle ex vivo. Cells isolated from ex vivo BPVC‐treated muscle exhibited higher adipogenic potential than those from saline‐treated muscle. Pre‐plating exposure of skeletal muscle cells to basic fibroblast growth factor (bFGF) mimicked the effect of ex vivo BPVC‐treatment, suggesting that bFGF released from extracellular matrix in response to muscle injury activates their adipogenic program. Interestingly, the number of myotubes were significantly reduced in the culture from BPVC‐treated muscle, suggesting that adipocytes negatively regulate myogenesis.     

Evaluation of body condition and weight loss in dogs presented to a veterinary oncology service

Cancer cachexia is a well-recognized syndrome in human patients that is characterized by progressive involuntary weight loss. The prevalence of this syndrome in veterinary cancer patients is unknown. This study's objective was to investigate the occurrence of weight loss and cachexia, as characterized by body condition scoring, in dogs presented to a veterinary oncology service. Information collected on 100 dogs included signalment, diagnosis, weight at time of diagnosis, and, when available, weight from a time approximately 12 months before diagnosis. Body condition was assessed by using a 9-point system based on body silhouette and palpation of adipose tissue (4-5 = optimal, 1 = extreme cachexia, 9 = extreme obesity). Muscle wasting was scored based on palpation of skeletal muscle (3 = no wasting, 2 = mild, 1 = moderate, 0 = severe). Only 4% of the dogs exhibited cachexia as defined by a body condition score < or = 3, whereas 29% were classified as markedly overweight (> or = 7). Fifteen percent had evidence of clinically relevant muscle wasting (< or = 1). Body weight from a time before the diagnosis of cancer was available for 64 dogs. At the time of diagnosis, 31% had maintained or gained weight, 31% had lost up to 5%, 14% had lost between 5 and 10%, and 23% had lost >10% of body weight. Overall, the percentage of dogs with signs indicating a decline in nutritional status was less than what has been reported for human cancer patients. Future studies should investigate the extent to which weight loss occurs in canine patients on an appropriate plane of nutrition as well as to establish whether an association exists between poor nutritional status and outcome in canine cancer patients.     

Myoclonus and hypersensitivity of the hind limbs and tail with urinary retention following neuraxial administration of morphine in a cat

An adult female domestic shorthair cat developed myoclonus, muscle rigidity, and hypersensitivity of the hind limbs and tail with urinary retention following neuraxial administration of morphine. Myoclonic contractions resolved following treatment with midazolam and a urinary catheter was placed until normal micturition returned. The cat was clinically normal 36 hours after neuraxial morphine injection. The cat underwent a second surgery without neuraxial morphine and control of postoperative pain required more intervention.Key clinical message:Neuraxial morphine may cause myoclonus and urinary retention. The response to pharmacological treatment of myoclonus is varied, but a benzodiazepine drug may be effective.