Excitation-contraction Coupling Proteins Research Articles

Sexual dimorphism in a mouse model of Friedreich’s ataxia with severe cardiomyopathy

Friedreich’s ataxia (FA) is an autosomal recessive disorder caused by reduced frataxin (FXN) expression in mitochondria, where the lethal component is cardiomyopathy. Using the conditional Fxnflox/null::MCK-Cre knock-out (Fxn-cKO) mouse model, we discovered significant sex differences in the progression towards heart failure, with Fxn-cKO males exhibiting a worse cardiac phenotype, low survival rate, kidney and reproductive organ deficiencies. These differences are likely due to a decline in testosterone in Fxn-cKO males. The decrease in testosterone was related to decreased expression of proteins involved in cholesterol transfer into the mitochondria: StAR and TSPO on the outer mitochondrial membrane, and the cholesterol side-chain cleavage enzyme P450scc and ferredoxin on the inner mitochondrial membrane. Expression of excitation-contraction coupling proteins (L-type calcium channel, RyR2, SERCA2, phospholamban and CaMKIIδ) was decreased significantly more in Fxn-cKO males. This is the first study that extensively investigates the sexual dimorphism in FA mouse model with cardiac calcium signaling impairment.

Speg interactions that regulate the stability of excitation-contraction coupling protein complexes in triads and dyads

Here we show that striated muscle preferentially expressed protein kinase α (Spegα) maintains cardiac function in hearts with Spegβ deficiency. Speg is required for stability of excitation-contraction coupling (ECC) complexes and interacts with esterase D (Esd), Cardiomyopathy-Associated Protein 5 (Cmya5), and Fibronectin Type III and SPRY Domain Containing 2 (Fsd2) in cardiac and skeletal muscle. Mice with a sequence encoding a V5/HA tag inserted into the first exon of the Speg gene (HA-Speg mice) display a >90% decrease in Spegβ but Spegα is expressed at ~50% of normal levels. Mice deficient in both Spegα and Speg β (Speg KO mice) develop a severe dilated cardiomyopathy and muscle weakness and atrophy, but HA-Speg mice display mild muscle weakness with no cardiac involvement. Spegα in HA-Speg mice suppresses Ca2+ leak, proteolytic cleavage of Jph2, and disruption of transverse tubules. Despite it’s low levels, HA-Spegβ immunoprecipitation identified Esd, Cmya5 and Fsd2 as Spegβ binding partners that localize to triads and dyads to stabilize ECC complexes. This study suggests that Spegα and Spegβ display functional redundancy, identifies Esd, Cmya5 and Fsd2 as components of both cardiac dyads and skeletal muscle triads and lays the groundwork for the identification of new therapeutic targets for centronuclear myopathy.

Dissociation of SH3 and cysteine-rich domain 3 and junctophilin 1 from dihydropyridine receptor in dystrophin-deficient muscles.

The disruption of excitation-contraction (EC) coupling and subsequent reduction in Ca2+ release from the sarcoplasmic reticulum (SR) have been shown to account for muscle weakness seen in patients with Duchenne muscular dystrophy (DMD). Here, we examined the mechanisms underlying EC uncoupling in skeletal muscles from mdx52 and DMD-null/NSG mice, animal models for DMD, focusing on the SH3 and cysteine-rich domain 3 (STAC3) and junctophilin 1 (JP1), which link the dihydropyridine receptor (DHPR) in the transverse tubule and the ryanodine receptor 1 in the SR. The isometric plantarflexion torque normalized to muscle weight of whole plantar flexor muscles was depressed in mdx52 and DMD-null/NSG mice compared with their control mice. This was accompanied by increased autolysis of calpain-1, decreased levels of STAC3 and JP1 content, and dissociation of STAC3 and JP1 from DHPR-α1s in gastrocnemius muscles. Moreover, in vitro mechanistic experiments demonstrated that STAC3 and JP1 underwent Ca2+-dependent proteolysis that was less pronounced in dystrophin-deficient muscles where calpastatin, the endogenous calpain inhibitor, was upregulated. Eccentric contractions further enhanced autolysis of calpain-1 and proteolysis of STAC3 and JP1 that were associated with severe torque depression in gastrocnemius muscles from DMD-null/NSG mice. These data suggest that Ca2+-dependent proteolysis of STAC3 and JP1 may be an essential factor causing muscle weakness due to EC coupling failure in dystrophin-deficient muscles.NEW & NOTEWORTHY The mechanisms underlying the disruption of excitation-contraction (EC) coupling in dystrophin-deficient muscles are not well understood. Here, using animal models for Duchenne muscular dystrophies (DMD), we show a Ca2+-dependent protease (calpain-1)-mediated proteolysis of SH3 and cysteine-rich domain 3 (STAC3) and junctophilin 1 (JP1), essential EC coupling proteins, in dystrophin-deficient muscle, and highlighting the dissociation of STAC3 and JP1 from dihydropyridine receptor as a causative factor in EC uncoupling of dystrophic muscles.

Distinct features of calcium handling and β-adrenergic sensitivity in heart failure with preserved versus reduced ejection fraction.

Key points Heart failure (HF), the leading cause of death in developed countries, occurs in the setting of reduced (HFrEF) or preserved (HFpEF) ejection fraction. Unlike HFrEF, there are no effective treatments for HFpEF, which accounts for ∼50% of heart failure.Abnormal intracellular calcium dynamics in cardiomyocytes have major implications for contractility and rhythm, but compared to HFrEF, very little is known about calcium cycling in HFpEF.We used rat models of HFpEF and HFrEF to reveal distinct differences in intracellular calcium regulation and excitation‐contraction (EC) coupling.While HFrEF is characterized by defective EC coupling at baseline, HFpEF exhibits enhanced coupling fidelity, further aggravated by a reduction in β‐adrenergic sensitivity.These differences in EC coupling and β‐adrenergic sensitivity may help explain why therapies that work in HFrEF are ineffective in HFpEF. Heart failure with reduced or preserved ejection fraction (respectively, HFrEF and HFpEF) is the leading cause of death in developed countries. Although numerous therapies improve outcomes in HFrEF, there are no effective treatments for HFpEF. We studied phenotypically verified rat models of HFrEF and HFpEF to compare excitation‐contraction (EC) coupling and protein expression in these two forms of heart failure. Dahl salt‐sensitive rats were fed a high‐salt diet (8% NaCl) from 7 weeks of age to induce HFpEF. Impaired diastolic relaxation and preserved ejection fraction were confirmed in each animal echocardiographically, and clinical signs of heart failure were documented. To generate HFrEF, Sprague‐Dawley (SD) rats underwent permanent left anterior descending coronary artery ligation which, 8–10 weeks later, led to systolic dysfunction (verified echocardiographically) and clinical signs of heart failure. Calcium (Ca2+) transients were measured in isolated cardiomyocytes under field stimulation or patch clamp. Ultra‐high‐speed laser scanning confocal imaging captured Ca2+ sparks evoked by voltage steps. Western blotting and PCR were used to assay changes in EC coupling protein and RNA expression. Cardiomyocytes from rats with HFrEF exhibited impaired EC coupling, including decreased Ca2+ transient (CaT) amplitude and defective couplon recruitment, associated with transverse (t)‐tubule disruption. In stark contrast, HFpEF cardiomyocytes showed saturated EC coupling (increased I Ca, high probability of couplon recruitment with greater Ca2+ release synchrony, increased CaT) and preserved t‐tubule integrity. β‐Adrenergic stimulation of HFpEF myocytes with isoprenaline (isoproterenol) failed to elicit robust increases in I Ca or CaT and relaxation kinetics. Fundamental differences in EC coupling distinguish HFrEF from HFpEF.

Sustaining Circulating Regulatory T Cell Subset Contributes to the Therapeutic Effect of Paroxetine on Mice With Diabetic Cardiomyopathy.

G protein coupled receptor kinase 2 (GRK2) inhibitor, paroxetine, has been approved to ameliorate diabetic cardiomyopathy (DCM). GRK2 is also involved in regulating T cell functions; the potential modifications of paroxetine on the immune response to DCM is unclear.Methods and Results:DCM mouse was induced by high-fat diet (HFD) feeding. A remarkable reduction in the regulatory T (Treg) cell subset in DCM mouse was found by flow cytometry, with impaired cardiac function evaluated by echocardiography. The inhibited Treg differentiation was attributable to insulin chronic stimulation in a GRK2-PI3K-Akt signaling-dependent manner. The selective GRK2 inhibitor, paroxetine, rescued Treg differentiation in vitro and in vivo. Furthermore, heart function, as well as the activation of excitation-contraction coupling proteins such as phospholamban (PLB) and troponin I (TnI) was effectively promoted in paroxetine-treated DCM mice compared with vehicle-treated DCM mice. Blockade of FoxP3 expression sufficiently inhibited the proportion of Treg cells, abolished the protective effect of paroxetine on heart function as well as PLB and TnI activation in HFD-fed mice. Neither paroxetine nor carvedilol could effectively ameliorate the metabolic disorder of HFD mice. The impaired systolic heart function of DCM mice was effectively improved by paroxetine therapy, partially through restoring the population of circulating Treg cells by targeting the GRK2-PI3K-Akt pathway.

170 Subcellular Distribution of Epac 2 in Relation to Key Excitation-Contraction Coupling Proteins in Human Atrial Tissue

170 Subcellular Distribution of Epac 2 in Relation to Key Excitation-Contraction Coupling Proteins in Human Atrial Tissue

Abstract 520: Overexpression of a Non-muscle RBFOX2 Splice Isoform Induce Cardiac Arrhythmias in Myotonic Dystrophy

Myotonic dystrophy type 1 (DM1) is a dominantly inherited neuromuscular disease caused by a CTG repeat expansion in 3’-UTR of DMPK gene. DM1 affects multiple tissues, but cardiac dysfunctions are the second leading cause of death which often include conduction defects and arrhythmias. The best characterized pathogenic mechanism of DM1 is toxic gain-of-function of expanded CUG repeat (CUG exp ) RNA that accumulates to form ribonuclear foci causing sequestration of MBNL and overexpression of CELF1 family of splicing factors. However, loss of MBNL or gain of CELF1 activity does not explain the cardiac phenotypes observed in DM1. Here we report that steady-state protein levels of RBFOX2, a critical splicing regulator, are drastically upregulated in DM1 heart tissue. This is accompanied by aberrant skipping of a muscle-specific 43bp exon in RBFOX2 transcript, resulting in selective up-regulation of the non-muscle RBFOX2 splice isoform in adult cardiomyocytes. We demonstrate that expression of CUG exp RNA in DM1 affects RBFOX2 in two distinct ways: reduced expression of a subset of microRNAs causes de-repression of RBFOX2 protein production; and CELF1 overexpression promotes skipping of the muscle-specific exon of RBFOX2 . Remarkably, tet -inducible overexpression of the non-muscle RBFOX2 isoform, or CRISPR/ Cas9 mediated deletion of the muscle-specific RBFOX2 exon in the mouse heart results in prolonged PR and QRS intervals, slower conduction velocity and spontaneous cardiac arrhythmias that mirror human DM1 pathology. Integration of RBFOX2-CLIP with RNA-seq data from cardiomyocytes of mice expressing the non-muscle RBFOX2 isoform identified a core network of mRNA splicing defects in genes encoding sodium, potassium, and calcium ion channels and key excitation-contraction coupling proteins that are similarly misspliced in hearts of DM1 patients. Thus, we have uncovered a central role for RBFOX2 isoform switching in DM1 cardiac pathogenesis and identified the molecular mechanisms that may alter electrophysiological properties and thereby induce cardiac arrhythmias in the DM1 heart.

Organization of junctional sarcoplasmic reticulum proteins in skeletal muscle fibers.

The sarcoplasmic reticulum (SR) of striated muscles is specialized for releasing Ca(2+) following sarcolemma depolarization in order to activate muscle contraction. To this end, the SR forms a network of longitudinal tubules and cisternae that surrounds the myofibrils and, at the same time, participates to the assembly of the triadic junctional membrane complexes formed by the close apposition of one t-tubule, originated from the sarcolemma, and two SR terminal cisternae. Advancements in understanding the molecular basis of the SR structural organization have identified an interaction between sAnk1, a transmembrane protein located on the longitudinal SR (l-SR) tubules, and obscurin, a myofibrillar protein. The direct interaction between these two proteins results in molecular contacts that have the overall effect to stabilize the l-SR tubules along myofibrils in skeletal muscle fibers. Less known is the structural organization of the sites in the SR that are specialized for Ca(2+) release and are positioned at the junctional SR (j-SR), i.e. the region of the terminal cisternae that faces the t-tubule at triads. At the j-SR, several trans-membrane proteins like triadin, junctin, or intra-luminal SR proteins like calsequestrin, are assembled together with the ryanodine receptor, the SR Ca(2+) release channel, into a macromolecular complex specialized in releasing Ca(2+). At triads, the 12nm-wide gap between the t-tubule and the j-SR allows the ryanodine receptor on the j-SR to be functionally coupled with the voltage-gated L-type calcium channel on the t-tubule in order to allow the transduction of the voltage-induced signal into Ca(2+) release through the ryanodine receptor channels. The muscle-specific junctophilin isoforms (JPH1 and JPH2) are anchored to the j-SR with a trans-membrane segment present at the C-terminus and are capable to bind the sarcolemma with a series of phospholipid-binding motifs localized at the N-terminus. Accordingly, through this dual interaction, JPH1 and JPH2 are responsible for the assembly of the triadic junctional membrane complexes. Recent data indicate that junctophilins seem also to interact with other proteins of the excitation-contraction machinery, suggesting that they may contribute to hold excitation-contraction coupling proteins to the sites where the j-SR is being organized.

Protein S‐nitrosylation regulates Ca2+ handling and myofilament Ca2+ sensitivity in beta‐adrenergic signaling

ObjectiveTo assess the impact of increased rotein denitrosylation on beta‐adrenergic receptor (b‐AR)‐dependent signaling and to evaluate effects of augmented denitrosylation during nitrosative stress in cardiomyocytes.Methods and ResultsWe studied cardiomyocytes isolated from wild‐type (WT) and cardiomyocytespecific S‐nitrosoglutathione reductase (GSNOR) overexpressing transgenic (GSNOR‐CMTg) mice. Stimulation of the b‐AR with isoproterenol (ISO)increased sarcomere shortening in both genotypes, while systolic Ca2+ release and sarcoplasmic Ca2+ load only increased in WT. This effect was correlated with increased myofilament Ca2+ sensitivity in ISO‐treated GSNOR‐CMTg cells. Both genotypes showed similar increases in phosphorylation of phospholamban (PLN) and troponin I after b‐AR stimulation while S‐nitrosylation of PLN and troponin C was only increased in WT. Exposure of ISO‐stimulated WT cells to S‐nitrosocysteine severely depressed contractility and systolic Ca2+ release, associated with sarcoplasmic Ca2+ leak and increased ryanodine receptor (RyR2) S‐nitrosylation. GSNOR overexpression prevented RyR2 S‐nitrosylation and preserved contractile function during nitrosative stress.ConclusionThese results suggest that b‐AR‐induced S‐nitrosylation of excitation‐contraction coupling proteins is required for b‐AR‐induced augmentation of cardiomyocyte function.

Triclosan Impairs Swimming Behavior and Alters Expression of Excitation-Contraction Coupling Proteins in Fathead Minnow (Pimephales promelas)

Triclosan (TCS), a high volume chemical widely used in consumer products, is a known aquatic contaminant found in fish inhabiting polluted watersheds. Mammalian studies have recently demonstrated that TCS disrupts signaling between the ryanodine receptor (RyR) and the dihydropyridine receptor (DHPR), two proteins essential for excitation-contraction (EC) coupling in striated muscle. We investigated the swimming behavior and expression of EC coupling proteins in larval fathead minnows (Pimephales promelas) exposed to TCS for up to 7 days. Concentrations as low as 75 μg L(-1) significantly altered fish swimming activity after 1 day; which was consistent after 7 days of exposure. The mRNA transcription and protein levels of RyR and DHPR (subunit CaV1.1) isoforms changed in a dose and time dependent manner. Crude muscle homogenates from exposed larvae did not display any apparent changes in receptor affinity toward known radioligands. In nonexposed crude muscle homogenates, TCS decreased the binding of [(3)H]PN20-110 to the DHPR and decreased the binding of [(3)H]-ryanodine to the RyR, demonstrating a direct impact at the receptor level. These results support TCS's impact on muscle function in vertebrates further exemplifying the need to re-evaluate the risks this pollutant poses to aquatic environments.

Shortening and intracellular Ca2+ in ventricular myocytes and expression of genes encoding cardiac muscle proteins in early onset type 2 diabetic Goto–Kakizaki rats

There has been a spectacular rise in the global prevalence of type 2 diabetes mellitus. Cardiovascular complications are the major cause of morbidity and mortality in diabetic patients. Contractile dysfunction, associated with disturbances in excitation-contraction coupling, has been widely demonstrated in the diabetic heart. The aim of this study was to investigate the pattern of cardiac muscle genes that are involved in the process of excitation-contraction coupling in the hearts of early onset (8-10 weeks of age) type 2 diabetic Goto-Kakizaki (GK) rats. Gene expression was assessed in ventricular muscle with real-time RT-PCR; shortening and intracellular Ca(2+) were measured in ventricular myocytes with video edge detection and fluorescence photometry, respectively. The general characteristics of the GK rats included elevated fasting and non-fasting blood glucose and blood glucose at 120 min following a glucose challenge. Expression of genes encoding cardiac muscle proteins (Myh6/7, Mybpc3, Myl1/3, Actc1, Tnni3, Tnn2, Tpm1/2/4 and Dbi) and intercellular proteins (Gja1/4/5/7, Dsp and Cav1/3) were unaltered in GK ventricle compared with control ventricle. The expression of genes encoding some membrane pumps and exchange proteins was unaltered (Atp1a1/2, Atp1b1 and Slc8a1), whilst others were either upregulated (Atp1a3, relative expression 2.61 ± 0.69 versus 0.84 ± 0.23) or downregulated (Slc9a1, 0.62 ± 0.07 versus 1.08 ± 0.08) in GK ventricle compared with control ventricle. The expression of genes encoding some calcium (Cacna1c/1g, Cacna2d1/2d2 and Cacnb1/b2), sodium (Scn5a) and potassium channels (Kcna3/5, Kcnj3/5/8/11/12, Kchip2, Kcnab1, Kcnb1, Kcnd1/2/3, Kcne1/4, Kcnq1, Kcng2, Kcnh2, Kcnk3 and Kcnn2) were unaltered, whilst others were either upregulated (Cacna1h, 0.95 ± 0.16 versus 0.47 ± 0.09; Scn1b, 1.84 ± 0.16 versus 1.11 ± 0.11; and Hcn2, 1.55 ± 0.15 versus 1.03 ± 0.08) or downregulated (Hcn4, 0.16 ± 0.03 versus 0.37 ± 0.08; Kcna2, 0.35 ± 0.03 versus 0.80 ± 0.11; Kcna4, 0.79 ± 0.25 versus 1.90 ± 0.26; and Kcnj2, 0.52 ± 0.07 versus 0.78 ± 0.08) in GK ventricle compared with control ventricle. The amplitude of ventricular myocyte shortening and the intracellular Ca(2+) transient were unaltered; however, the time-to-peak shortening was prolonged and time-to-half decay of the Ca(2+) transient was shortened in GK myocytes compared with control myocytes. The results of this study demonstrate changes in expression of genes encoding various excitation-contraction coupling proteins that are associated with disturbances in myocyte shortening and intracellular Ca(2+) transport.

Effects of carnosine on contractile apparatus Ca2+sensitivity and sarcoplasmic reticulum Ca2+release in human skeletal muscle fibers

There is considerable interest in potential ergogenic and therapeutic effects of increasing skeletal muscle carnosine content, although its effects on excitation-contraction (EC) coupling in human muscle have not been defined. Consequently, we sought to characterize what effects carnosine, at levels attained by supplementation, has on human muscle fiber function, using a preparation with all key EC coupling proteins in their in situ positions. Fiber segments, obtained from vastus lateralis muscle of human subjects by needle biopsy, were mechanically skinned, and their Ca(2+) release and contractile apparatus properties were characterized. Ca(2+) sensitivity of the contractile apparatus was significantly increased by 8 and 16 mM carnosine (increase in pCa(50) of 0.073 ± 0.007 and 0.116 ± 0.006 pCa units, respectively, in six type I fibers, and 0.063 ± 0.018 and 0.103 ± 0.013 pCa units, respectively, in five type II fibers). Caffeine-induced force responses were potentiated by 8 mM carnosine in both type I and II fibers, with the potentiation in type II fibers being entirely explicable by the increase in Ca(2+) sensitivity of the contractile apparatus caused by carnosine. However, the potentiation of caffeine-induced responses caused by carnosine in type I fibers was beyond that expected from the associated increase in Ca(2+) sensitivity of the contractile apparatus and suggestive of increased Ca(2+)-induced Ca(2+) release. Thus increasing muscle carnosine content likely confers benefits to muscle performance in both fiber types by increasing the Ca(2+) sensitivity of the contractile apparatus and possibly also by aiding Ca(2+) release in type I fibers, helping to lessen or slow the decline in muscle performance during fatiguing stimulation.

Expression and regulation of excitation-contraction coupling proteins in aging skeletal muscle.

Functional and structural decline of the neuromuscular system is a recognized cause of decreased strength, impaired performance of daily living activities, and loss of independence in the elderly. However, in mammals, including humans, age-related loss of strength is greater than loss of muscle mass, so the underlying mechanisms remain only partially understood. This review focuses on the mechanisms underlying impaired skeletal muscle function with aging, including external calcium-dependent skeletal muscle contraction; increased voltage-sensitive calcium channel Cav1.1 β1asubunit and junctional face protein JP-45 and decreased Cav1.1 (α1) expression, and the potential role of these and other recently discovered molecules of the muscle T-tubule/sarcoplasmic reticulum junction in excitation-contraction uncoupling. We also examined neural influences and trophic factors, particularly insulin-like growth factor-I (IGF-1). Better insight into the triad proteins' involvement in muscle ECC and nerve/muscle interactions and regulation will lead to more rational interventions to delay or prevent muscle weakness with aging. The focus of this review is on the proteins mediating excitation-contraction coupling (ECC) and their expression and regulation in humans and rodent models of skeletal muscle functional decline with aging. Age-dependent changes in proteins other than those related to ECC, muscle composition, clinical assessment and interventions, have been extensively reviewed recently [1-3].

Changes in the Organization of Excitation-Contraction Coupling Structures in Failing Human Heart

BackgroundThe cardiac myocyte t-tubular system ensures rapid, uniform cell activation and several experimental lines of evidence suggest changes in the t-tubular system and associated excitation-contraction coupling proteins may occur in heart failure.Methods and ResultsThe organization of t-tubules, L-type calcium channels (DHPRs), ryanodine receptors (RyRs) and contractile machinery were examined in fixed ventricular tissue samples from both normal and failing hearts (idiopathic (non-ischemic) dilated cardiomyopathy) using high resolution fluorescent imaging. Wheat germ agglutinin (WGA), Na-Ca exchanger, DHPR and caveolin-3 labels revealed a shift from a predominantly transverse orientation to oblique and axial directions in failing myocytes. In failure, dilation of peripheral t-tubules occurred and a change in the extent of protein glycosylation was evident. There was no change in the fractional area occupied by myofilaments (labeled with phalloidin) but there was a small reduction in the number of RyR clusters per unit area. The general relationship between DHPRs and RyR was not changed and RyR labeling overlapped with 51±3% of DHPR labeling in normal hearts. In longitudinal (but not transverse) sections there was an ∼30% reduction in the degree of colocalization between DHPRs and RyRs as measured by Pearson's correlation coefficient in failing hearts.ConclusionsThe results show that extensive remodelling of the t-tubular network and associated excitation-contraction coupling proteins occurs in failing human heart. These changes may contribute to abnormal calcium handling in heart failure. The general organization of the t-system and changes observed in failure samples have subtle differences to some animal models although the general direction of changes are generally similar.

S100A1 promotes action potential-initiated calcium release flux and force production in skeletal muscle

The role of S100A1 in skeletal muscle is just beginning to be elucidated. We have previously shown that skeletal muscle fibers from S100A1 knockout (KO) mice exhibit decreased action potential (AP)-evoked Ca(2+) transients, and that S100A1 binds competitively with calmodulin to a canonical S100 binding sequence within the calmodulin-binding domain of the skeletal muscle ryanodine receptor. Using voltage clamped fibers, we found that Ca(2+) release was suppressed at all test membrane potentials in S100A1(-/-) fibers. Here we examine the role of S100A1 during physiological AP-induced muscle activity, using an integrative approach spanning AP propagation to muscle force production. With the voltage-sensitive indicator di-8-aminonaphthylethenylpyridinium, we first demonstrate that the AP waveform is not altered in flexor digitorum brevis muscle fibers isolated from S100A1 KO mice. We then use a model for myoplasmic Ca(2+) binding and transport processes to calculate sarcoplasmic reticulum Ca(2+) release flux initiated by APs and demonstrate decreased release flux and greater inactivation of flux in KO fibers. Using in vivo stimulation of tibialis anterior muscles in anesthetized mice, we show that the maximal isometric force response to twitch and tetanic stimulation is decreased in S100A1(-/-) muscles. KO muscles also fatigue more rapidly upon repetitive stimulation than those of wild-type counterparts. We additionally show that fiber diameter, type, and expression of key excitation-contraction coupling proteins are unchanged in S100A1 KO muscle. We conclude that the absence of S100A1 suppresses physiological AP-induced Ca(2+) release flux, resulting in impaired contractile activation and force production in skeletal muscle.

Protein Aggregates and Novel Presenilin Gene Variants in Idiopathic Dilated Cardiomyopathy

Heart failure is a debilitating condition resulting in severe disability and death. In a subset of cases, clustered as idiopathic dilated cardiomyopathy (iDCM), the origin of heart failure is unknown. In the brain of patients with dementia, proteinaceous aggregates and abnormal oligomeric assemblies of beta-amyloid impair cell function and lead to cell death. We have similarly characterized fibrillar and oligomeric assemblies in the hearts of iDCM patients, pointing to abnormal protein aggregation as a determinant of iDCM. We also showed that oligomers alter myocyte Ca(2+) homeostasis. Additionally, we have identified 2 new sequence variants in the presenilin-1 (PSEN1) gene promoter leading to reduced gene and protein expression. We also show that presenilin-1 coimmunoprecipitates with SERCA2a. On the basis of these findings, we propose that 2 mechanisms may link protein aggregation and cardiac function: oligomer-induced changes on Ca(2+) handling and a direct effect of PSEN1 sequence variants on excitation-contraction coupling protein function.

Effect of age on cardiac excitation–contraction coupling

1. Cardiovascular diseases most commonly occur in the elderly and are a frequent cause of disability or death. However, the effect of age itself on cardiac function is not well understood. 2. Studies in both human and animal hearts indicate that contractile function is unaffected by age while at rest. However, the ability to increase cardiac contractile force during strenuous activities, such as exercise, declines with age. 3. Similar findings have been observed in individual ventricular myocytes isolated from aged hearts. When myocytes are stimulated with beta-adrenoceptor agonists or rapid pacing frequencies, aged cells show a much smaller increase in peak contractions and Ca(2+) transients than young adult cells. In addition, contractions and Ca(2+) transients are prolonged in aged cells compared with younger cells under these conditions. 4. These observations suggest that the age-related decline in cardiac contractile function originates at the cellular level and may reflect modifications in processes involved in excitation-contraction (EC) coupling. 5. Biochemical studies have shown that there are age-related modifications in the expression, regulation and function of a number of proteins essential to EC coupling in the heart. 6. Functional studies indicate that these changes in EC coupling proteins disrupt Ca(2+) homeostasis and contribute to decrease in peak contraction and prolongation of contraction duration observed in myocytes from aged hearts. 7. The present review describes modifications in cardiac contractile function that occur in the ageing heart and evaluates underlying alterations in the EC coupling pathway that may be responsible for this decline in contractile function in ageing.

Cardiac contractility modulation electrical signals normalize expression of the excitation-contraction coupling proteins in left ventricular myocardium of dogs with heart failure

Cardiac contractility modulation electrical signals normalize expression of the excitation-contraction coupling proteins in left ventricular myocardium of dogs with heart failure

Cardiac Contractility Modulation Electrical Signals Normalize Expression of the Excitation-Contraction Coupling Proteins Presenillin-2 and Sorcin in Left Ventricular Myocardium of Dogs with Chronic Heart Failure

Background: In the normal heart, during membrane depolarization, presenillin-2 (PS2) activates the sarcoplasmic reticulum (SR) ryanodine Ca2+ release channel (RyR2) to release large amounts of Ca2+ from the SR. At the end of systole (high Ca2+), sorcin (SOR) interacts with RyR2 to stop the SR Ca2+ release. In heart failure (HF), the RyR2 is “leaky” causing continuous release of Ca2+ into the cytosol and leading to Ca2+ overload as well as to depletion of calcium from the SR a maladaptation deemed detrimental in HF.

Isoform switching in myofibrillar and excitation-contraction coupling proteins contributes to diminished contractile function in regenerating rat soleus muscle

Postnatal development of skeletal muscle occurs through the progressive transformation of diverse biochemical, metabolic, morphological, and functional characteristics from the embryonic to the adult phenotype. Since muscle regeneration recapitulates postnatal development of muscle fiber, it offers an appropriate experimental model to investigate the existing relationships between diverse muscle functions and the expression of key protein isoforms, particularly at the single-fiber level. This study was carried out in regenerating soleus muscle 14 days after injury. At this intermediate stage, the regenerating muscle exhibited a recovery of mass greater than its force generation capacity. The lower specific tension of regenerating muscle suggested intrinsic defective excitation-contraction coupling and/or contractility processes. The presence of developmental isoforms of both the voltage-gated Ca(2+) channel (alpha(1)C) and of ryanodine receptor 3, paralleled by an abnormal caffeine contracture development, confirms the immature excitation-contraction coupling of the regenerating muscle. The defective Ca(2+) handling could also be confirmed by the lower sarcoplasmic reticulum caffeine sensitivity of regenerating single fibers. Also, regenerating single fibers revealed a lower maximal specific tension, which was associated with the residual presence of embryonic myosin heavy chains. Moreover, the fibers showed a reduced Ca(2+) sensitivity of myofibrillar proteins, particularly those simultaneously expressing the slow and fast isoforms of troponin C. The present results indicate that the expression of developmental proteins determines the incomplete functional recovery of regenerating soleus.