Myasthenia gravis sera containing antiryanodine receptor antibodies inhibit binding of [3H]-ryanodine to sacroplasmic reticulum

Myasthenia gravis (MG) patients with thymoma often have antibodies against the calcium-release channel of the sarcoplasmic reticulum (SR) in striated muscle, the ryanodine receptor (RyR). RyR function can be tested in vitro by measuring the degree of [³H]-ryanodine binding to SR. In this study, sera from 9 out of 14 MG patients containing RyR antibodies inhibited [³H]-ryanodine binding to SR membranes from rat skeletal muscle. The 9 patients with antibodies inhibiting ryanodine binding had more severe MG than those with noninhibiting antibodies (P = 0.006). Sera from MG patients with acetylcholine receptor and titin muscle antibodies but no antibodies against RyR and blood-donor sera did not have an inhibiting effect in the [³H]-ryanodine binding assay. The results show that RyR antibodies in MG patients have high affinity for the RyR, and that the binding of antibodies probably affects calcium release from SR by locking the RyR ion channel in a closed position. © 1998 John Wiley & Sons, Inc. Muscle Nerve 21:329–335.     

Potassium and caffeine contractures of mouse muscles before and after fatiguing stimulation

To assess the impairment of muscle membrane excitation, excitation–contraction (E–C) coupling, and contractility during muscle fatigue, we monitored the contracture responses of resting and fatigued muscles on exposure to high potassium and caffeine. On exposure to 140 mmol/L potassium, mouse extensor digitorum longus (EDL) developed a contracture which was 15.7% of tetanic tension before fatigue and 31.7% after fatigue, while soleus developed 59.4% contracture before and 68.8% after fatigue. Potassium causes contractures by depolarizing the muscle fiber membrane. Hence, membrane excitation is reduced in fatigued EDL and soleus. On exposure to 32 mmol/L caffeine, the contracture was 7.1% in resting EDL, 8.5% in fatigued EDL, 50.1% in resting soleus, and 43.7% in fatigued soleus. On exposure to 1 mmol/L caffeine followed by rapid cooling, the contracture was 3.0% in resting EDL, 3.2% in fatigued EDL, 21.5% in resting soleus, and 10.3% in fatigued soleus. Caffeine causes contracture by releasing Ca^{+ +} from the sarcoplasmic reticulum. Our results indicate reduced E–C coupling attributable to reduced membrane excitation in fatigued EDL, and reduced contractility in fatigued soleus. © 1994 John Wiley & Sons, Inc.     

CaM kinase II in colonic smooth muscle contributes to dysmotility in murine DSS‐colitis

Background Altered calcium mobilization has been implicated in the development of colonic dysmotility in inflammatory bowel disease. The aim of this study was to investigate the mechanisms by which disrupted intracellular Ca²⁺ signalling contributes to the impaired contractility of colon circular smooth muscles. Methods Acute colitis was induced in C57Bl/6 mice with dextran sulphate sodium (DSS) in the drinking water for 5 days. Key Results Spontaneous and acetylcholine‐evoked contractions, caffeine‐evoked hyperpolarization, and SERCA2 and phospholamban expression were reduced compared with controls. Tetrodotoxin did not restore control levels of contractile activity. The amplitudes, but not the frequency, of intracellular Ca²⁺ waves were increased compared with controls. Caffeine abolished intracellular Ca²⁺ waves in control smooth muscle cells, but not in smooth muscle cells from DSS‐treated mice. CaM kinase II activity and cytosolic levels of HDAC4 were increased, and IκBα levels were decreased in distal colon smooth muscles from DSS‐treated mice. Conclusions & Inferences These results suggest that disruptions in intracellular Ca²⁺ mobilization due to down‐regulation of SERCA2 and phospholamban expression lead to increased CaM kinase II activity and cytosolic HDAC4 that may contribute to the dysmotility of colonic smooth muscles in colitis by enhancing NF‐κB activity.     

Contractile properties of slow and fast skeletal muscles from protease activated receptor‐1 null mice

Introduction: Protease‐activated receptors (PARs) may play a role in skeletal muscle development. We compared the contractile properties of slow‐twitch soleus muscles and fast‐twitch extensor digitorum longus (EDL) muscles from PAR‐1 null and littermate control mice. Methods: Contractile function was measured using a force transducer system. Fiber type proportions were determined using immunohistochemistry. Results: Soleus muscles from PAR‐1 null mice exhibited longer contraction times, a leftward shift in the force–stimulation frequency relationship, and decreased fatiguability compared with controls. PAR‐1 null soleus muscles also had increased type 1 and decreased type IIb/x fiber numbers compared with controls. In PAR‐1 null EDL muscles, no differences were found, except for a slower rate of fatigue compared with controls. Conclusions: The absence of PAR‐1 results in a slower skeletal muscle contractile phenotype, likely due to an increase in type I and a decrease in type IIb/x fiber numbers. Muscle Nerve 50: 991–998, 2014     

Ca2+ sensitizers: An emerging class of agents for counterbalancing weakness in skeletal muscle diseases?

Ca²⁺ ions are key regulators of skeletal muscle contraction. By binding to contractile proteins, they initiate a cascade of molecular events leading to cross-bridge formation and ultimately, muscle shortening and force production. The ability of contractile proteins to respond to Ca²⁺ attachment, also known as Ca²⁺ sensitivity, is often compromised in acquired and congenital skeletal muscle disorders. It constitutes, undoubtedly, a major physiological cause of weakness for patients. In this review, we discuss recent studies giving strong molecular and cellular evidence that pharmacological modulators of some of the contractile proteins, also termed Ca²⁺ sensitizers, are efficient agents to improve Ca²⁺ sensitivity and function in diseased skeletal muscle cells. In fact, they compensate for the impaired contractile proteins response to Ca²⁺ binding. Currently, such Ca²⁺ sensitizing compounds are successfully used for reducing problems in cardiac disorders. Therefore, in the future, under certain conditions, these agents may represent an emerging class of agents to enhance the quality of life of patients suffering from skeletal muscle weakness.     

Inflammatory response during slow‐ and fast‐twitch muscle regeneration

Introduction: Skeletal muscles are characterized by their unique ability to regenerate. Injury of a so‐called fast‐twitch muscle, extensor digitorum longus (EDL), results in efficient regeneration and reconstruction of the functional tissue. In contrast, slow‐twitch muscle (soleus) fails to properly reconstruct and develops fibrosis. This study focuses on soleus and EDL muscle regeneration and associated inflammation. Methods: We determined differences in the activity of neutrophils and M1 and M2 macrophages using flow cytometry and differences in the levels of proinflammatory cytokines using Western blotting and immunolocalization at different times after muscle injury. Results: Soleus muscle repair is accompanied by increased and prolonged inflammation, as compared to EDL. The proinflammatory cytokine profile is different in the soleus and ED muscles. Conclusions: Muscle repair efficiency differs by muscle fiber type. The inflammatory response affects the repair efficiency of slow‐ and fast‐twitch muscles. Muscle Nerve 55 : 400–409, 2017     

Regulation of isometric contraction in skeletal muscle

The purpose of this study was to determine whether differences in isometric twitch contraction times of skeletal muscles are related more closely to myosin adenosine triphosphatase (ATPase) activity or to the ability of the sarcotubules to accumulate Ca²⁺. The isometric contraction time was observed to be shorter in the crureus muscle than in the soleus muscle of the rabbit. These muscles were found to have similar myosin ATPase activities, but sarcotubules isolated from crureus had a faster rate of Ca²⁺ uptake than soleus sarcotubules. Furthermore, the yield of sarcotubules was 41% higher from crureus. Likewise, the isometric contraction time of rat soleus decreased during the first 3 weeks of life, but no change was observed in myosin ATPase activity. However, sarcotubular uptake of Ca²⁺ increased as did the yield of sarcotubules (44% higher). It is suggested that the differences in isometric contraction time between skeletal muscles are related more closely to the sarcotubular Ca²⁺ uptake than to the activity of myosin ATPase.     

Calmodulin antagonists induce platelet apoptosis

Calmodulin (CaM) antagonists induce apoptosis in various tumor models and inhibit tumor cell invasion and metastasis, thus some of which have been extensively used as anti-cancer agents. In platelets, CaM has been found to bind directly to the cytoplasmic domains of several platelet receptors. Incubation of platelets with CaM antagonists impairs the receptors-related platelet functions. However, it is still unknown whether CaM antagonists induce platelet apoptosis. Here we show that CaM antagonists N-(6-aminohexyl)-5-chloro-1-naphthalene sulfonamide (W7), tamoxifen (TMX), and trifluoperazine (TFP) induce apoptotic events in human platelets, including depolarization of mitochondrial inner transmembrane potential, caspase-3 activation, and phosphatidylserine exposure. CaM antagonists did not incur platelet activation as detected by P-selectin surface expression and PAC-1 binding. However, ADP-, botrocetin-, and α-thrombin-induced platelet aggregation, platelet adhesion and spreading on von Willebrand factor surface were significantly reduced in platelets pre-treated with CaM antagonists. Furthermore, cytosolic Ca²⁺ levels were obviously elevated by both W7 and TMX, and membrane-permeable Ca²⁺ chelator BAPTA-AM significantly reduced apoptotic events in platelets induced by W7. Therefore, these findings indicate that CaM antagonists induce platelet apoptosis. The elevation of the cytosolic Ca²⁺ levels may be involved in the regulation of CaM antagonists-induced platelet apoptosis.     

Calendar of events

Intramyofiber accumulation of β‐amyloid fragments (Aβ) is a pathologic hallmark of inclusion‐body myositis (IBM), a progressive skeletal muscle disorder. We investigated the temporal pattern of alterations in the resting cytoplasmic [Ca²⁺] ([Ca²⁺]_i) as well as the depolarization‐evoked Ca²⁺ release from the sarcoplasmic reticulum in skeletal muscle from transgenic mice expressing human βAPP (MCK‐βAPP). MCK‐βAPP mice show an age‐dependent increase in [Ca²⁺]_i along with a reduction in depolarization‐evoked Ca²⁺ release, which appear well before the other reported aspects of IBM, such as inclusion formation, inflammation, centralized nuclei, atrophy, and skeletal muscle weakness. In the young MCK‐βAPP animals the increase in resting [Ca²⁺]_i can be attributed largely to Ca²⁺ influx through nifedipine‐sensitive Ca²⁺ channels. In the adult MCK‐βAPP mice, in addition to the nifedipine‐sensitive pathway, there is also a substantial contribution by the intracellular compartments to the increase in [Ca²⁺]_i. These results suggest that β‐amyloid‐induced disuption of Ca²⁺ handling may represent an early event in the pathogenesis of IBM. Muscle Nerve, 2010     

Autoantibodies against TRPC3 and ryanodine receptor in myasthenia gravis

The transient receptor potential canonical type-3 (TRPC3, receptor- and store-operated Ca²⁺ influx channel) participates in skeletal muscle contraction; its functional interactions with ryanodine receptor-1 (RyR1) are independent of sarcoplasmic Ca²⁺ content and dihydropyridine receptor. In 25 generalized myasthenia gravis (MG), we detected antibodies against human TRPC3 peptide in 9 patients (8 with thymoma and one with hyperplastic thymus) and those against human RyR1 peptides in 16 patients (15 with thymoma and one with hyperplastic thymus). Both antibodies were found in patients with more severe myasthenia and could contribute to the contractile abnormalities in MG.     

Enhanced muscle shortening and impaired Ca2+ channel function in an acute septic myopathy model

Myopathies in critically ill patients are increasingly documented. Various animal models of chronic sepsis have been employed to investigate reduced membrane excitability or altered isometric contractility of skeletal muscle. In contrast, immediate changes occurring during acute sepsis are significantly under-characterised; L-type Ca²⁺ channel function or isotonic shortening are examples. We recorded slowly activating L-type Ca²⁺ currents (I _Ca) in voltage-clamped single intact mouse skeletal muscle fibres and tested the effects of acute challenge with serum fractions from critical illness myopathy patients (CIM). Using a high-speed camera system, we simultaneously recorded unloaded fibre shortening during isotonic contractions with unprecedented temporal resolution (~1,600 frames/s). Time courses of fibre lengths and shortening velocity were determined from automated imaging algorithms. CIM fractions acutely induced depression of I _Ca amplitudes with no shifts in I _Ca–V-relations. Voltage-dependent inactivation was unaltered and I _Ca activation and inactivation kinetics were prolonged compared to controls. Unexpectedly, maximum unloaded speed of shortening was slightly faster following CIM serum applications, suggesting a direct action of CIM serum on weak-binding-state cross-bridges. Our results are compatible with a model where CIM serum might acutely reduce a fraction of functional L-type Ca²⁺ channels and could account for reduced SR Ca²⁺ release and force production in CIM patients. Acute increase in isotonic shortening velocity might be an early diagnostic feature suitable for testing in clinical studies. The acute challenge model is also robust against atrophy or fibre type changes that ordinarily would have to be considered in chronic sepsis models.     

Recovery of rat skeletal muscles after partial denervation is enhanced by treatment with nifedipine

Following partial denervation of adult rat skeletal muscle intact axons sprout to reinnervate denervated muscle fibres and increase their territory. The extent of this increase is limited and may depend on the ability of axon terminals to form and maintain synaptic contacts with the denervated muscle fibres. Here we tested the possibility whether reducing Ca²⁺ entry into presynaptic nerve terminals through dihydropyridine sensitive channels may allow more nerve–muscle contacts to be formed and maintained. Hindlimb muscles of adult Wistar rats were partially denervated by removing a small segment of the L4 or L5 spinal nerve on one side. A nifedipine-containing silastic rubber strip was subsequently implanted close to the partially denervated soleus or extensor digitorum longus (EDL) muscles in some animals. In control experiments silastic strips which did not contain nifedipine were used. Several weeks later isometric contractions were recorded, to determine the effect of (a) partial denervation and (b) nifedipine treatment on force output and motor unit numbers. The tension produced by nifedipine treated partially denervated muscles was 82% and 79% of the unoperated contralateral value for soleus and EDL, respectively. This was significantly greater than in untreated muscles, which only produced 61% and 48%, respectively. Mean motor unit force was also significantly larger with nifedipine treatment. Histological analysis revealed that a significantly larger proportion of the total number of muscle fibres remained in nifedipine-treated partially denervated muscles (soleus, 90% and EDL, 101%) compared with untreated muscles (soleus, 51% and EDL, 66%). Thus the number of neuromuscular contacts was increased with nifedipine treatment.     

Vitamin D and/or calcium deficient diets may differentially affect muscle fiber neuromuscular junction innervation

Introduction: There is evidence that supports a role for Vitamin D (Vit. D) in muscle. The exact mechanism by which Vit. D deficiency impairs muscle strength and function is not clear. Methods: Three‐week‐old mice were fed diets with varied combinations of Vit. D and Ca²⁺ deficiency. Behavioral testing, genomic and protein analysis, and muscle histology were performed with a focus on neuromuscular junction (NMJ) ‐related genes. Results: Vit. D and Ca²⁺ deficient mice performed more poorly on given behavioral tasks than animals with Vit. D deficiency alone. Genomic and protein analysis of the soleus and tibialis anterior muscles revealed changes in several Vit. D metabolic, NMJ‐related, and protein chaperoning and refolding genes. Conclusions: These data suggest that detrimental effects of a Vit. D deficient or a Vit. D and Ca²⁺ deficient diet may be a result of differential alterations in the structure and function of the NMJ and a lack of a sustained stress response in muscles. Muscle Nerve 54 : 1120–1132, 2016     

Histamine resets the circadian clock in the suprachiasmatic nucleus through the H1R‐CaV1.3‐RyR pathway in the mouse

Histamine, a neurotransmitter/neuromodulator implicated in the control of arousal state, exerts a potent phase‐shifting effect on the circadian clock in the rodent suprachiasmatic nucleus (SCN). In this study, the mechanisms by which histamine resets the circadian clock in the mouse SCN were investigated. As a first step, Ca²⁺‐imaging techniques were used to demonstrate that histamine increases intracellular Ca²⁺ concentration ([Ca²⁺]_i) in acutely dissociated SCN neurons and that this increase is blocked by the H1 histamine receptor (H1R) antagonist pyrilamine, the removal of extracellular Ca²⁺ and the L‐type Ca²⁺ channel blocker nimodipine. The histamine‐induced Ca²⁺ transient is reduced, but not blocked, by application of the ryanodine receptor (RyR) blocker dantrolene. Immunohistochemical techniques indicated that Ca_V1.3 L‐type Ca²⁺ channels are expressed mainly in the somata of SCN cells along with the H1R, whereas Ca_V1.2 channels are located primarily in the processes. Finally, extracellular single‐unit recordings demonstrated that the histamine‐elicited phase delay of the circadian neural activity rhythm recorded from SCN slices is blocked by pyrilamine, nimodipine and the knockout of Ca_V1.3 channel. Again, application of dantrolene reduced but did not block the histamine‐induced phase delays. Collectively, these results indicate that, to reset the circadian clock, histamine increases [Ca²⁺]_i in SCN neurons by activating Ca_V1.3 channels through H1R, and secondarily by causing Ca²⁺‐induced Ca²⁺ release from RyR‐mediated internal stores.     

Calcium exchange hypothesis of skeletal muscle fatigue: A brief review

Skeletal muscle fatigue is often associated with diminished athletic performance and work productivity as well as increased susceptibility to injury. The exact cause of muscle fatigue probably involves a number of factors which influence force production in a manner dependent on muscle fiber type and activation pattern. However, a growing body of evidence implicates alterations in intracellular Ca²⁺ exchange as a major role in the fatigue process. These changes are thought to occur secondary to reductions in the rates of Ca²⁺ uptake and release by the sarcoplasmic reticulum (SR). This hypothesis is based on the finding that peak myoplasmic Ca²⁺ concentration ([Ca²⁺]_i) is reduced as force declines during fatigue. In addition, direct measurements of Ca²⁺ uptake and release show that fatiguing activity causes intrinsic alterations in the functional properties of the SR. We also propose that fatigue-induced alterations in Ca²⁺ exchange may be beneficial, reducing the rate of energy utilization by the muscle fiber and preventing irreversible damage to the cell. © 1995 John Wiley & Sons, Inc.     

Calmodulin activity regulates group I metabotropic glutamate receptor‐mediated signal transduction and synaptic depression

Group I metabotropic glutamate receptors (mGluR), including mGluR1 and mGluR 5 (mGluR1/5), are coupled to Gq and modulate activity‐dependent synaptic plasticity. Direct activation of mGluR1/5 causes protein translation‐dependent long‐term depression (LTD). Although it has been established that intracellular Ca²⁺ and the Gq‐regulated signaling molecules are required for mGluR1/5 LTD, whether and how Ca²⁺ regulates Gq signaling and upregulation of protein expression remain unknown. Through pharmacological inhibition, we tested the function of the Ca²⁺ sensor calmodulin (CaM) in intracellular signaling triggered by the activation of mGluR1/5. CaM inhibitor N‐[4‐aminobutyl]‐5‐chloro‐2‐naphthalenesulfonamide hydrochloride (W13) suppressed the mGluR1/5‐stimulated activation of extracellular signal‐regulated kinase 1/2 (ERK1/2) and p70‐S6 kinase 1 (S6K1) in hippocampal neurons. W13 also blocked the mGluR1/5 agonist‐induced synaptic depression in hippocampal slices and in anesthetized mice. Consistent with the function of CaM, inhibiting the downstream targets Ca²⁺/CaM‐dependent protein kinases (CaMK) blocked ERK1/2 and S6K1 activation. Furthermore, disruption of the CaM–CaMK–ERK1/2 signaling cascade suppressed the mGluR1/5‐stimulated upregulation of Arc expression. Altogether, our data suggest CaM as a new Gq signaling component for coupling Ca²⁺ and protein upregulation and regulating mGluR1/5‐mediated synaptic modification. © 2016 Wiley Periodicals, Inc.