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Sample records for striated muscle sarcoplasmic

  1. Minor sarcoplasmic reticulum membrane components that modulate excitation–contraction coupling in striated muscles

    PubMed Central

    Treves, Susan; Vukcevic, Mirko; Maj, Marcin; Thurnheer, Raphael; Mosca, Barbara; Zorzato, Francesco

    2009-01-01

    In striated muscle, activation of contraction is initiated by membrane depolarisation caused by an action potential, which triggers the release of Ca2+ stored in the sarcoplasmic reticulum by a process called excitation–contraction coupling. Excitation–contraction coupling occurs via a highly sophisticated supramolecular signalling complex at the junction between the sarcoplasmic reticulum and the transverse tubules. It is generally accepted that the core components of the excitation–contraction coupling machinery are the dihydropyridine receptors, ryanodine receptors and calsequestrin, which serve as voltage sensor, Ca2+ release channel, and Ca2+ storage protein, respectively. Nevertheless, a number of additional proteins have been shown to be essential both for the structural formation of the machinery involved in excitation–contraction coupling and for its fine tuning. In this review we discuss the functional role of minor sarcoplasmic reticulum protein components. The definition of their roles in excitation–contraction coupling is important in order to understand how mutations in genes involved in Ca2+ signalling cause neuromuscular disorders. PMID:19403606

  2. Eudistomin D and penaresin derivatives as modulators of ryanodine receptor channels and sarcoplasmic reticulum Ca2+ ATPase in striated muscle.

    PubMed

    Diaz-Sylvester, Paula L; Porta, Maura; Juettner, Vanessa V; Lv, Yuanzhao; Fleischer, Sidney; Copello, Julio A

    2014-04-01

    Eudistomin D (EuD) and penaresin (Pen) derivatives are bioactive alkaloids from marine sponges found to induce Ca(2+) release from striated muscle sarcoplasmic reticulum (SR). Although these alkaloids are believed to affect ryanodine receptor (RyR) gating in a "caffeine-like" manner, no single-channel study confirmed this assumption. Here, EuD and MBED (9-methyl-7-bromoeudistomin D) were contrasted against caffeine on their ability to modulate the SR Ca(2+) loading/leak from cardiac and skeletal muscle SR microsomes as well as the function of RyRs in planar bilayers. The effects of these alkaloids on [(3)H]ryanodine binding and SR Ca(2+) ATPase (SERCA) activity were also tested. MBED (1-5 μM) fully mimicked maximal activating effects of caffeine (20 mM) on SR Ca(2+) leak. At the single-channel level, MBED mimicked the agonistic action of caffeine on cardiac RyR gating (i.e., stabilized long openings characteristic of "high-open-probability" mode). EuD was a partial agonist at the maximal doses tested. The tested Pen derivatives displayed mild to no agonism on RyRs, SR Ca(2+) leak, or [(3)H]ryanodine binding studies. Unlike caffeine, EuD and some Pen derivatives significantly inhibited SERCA at concentrations required to modulate RyRs. Instead, MBED's affinity for RyRs (EC50 ∼ 0.5 μM) was much larger than for SERCA (IC50 > 285 μM). In conclusion, MBED is a potent RyR agonist and, potentially, a better choice than caffeine for microsomal and cell studies due to its reported lack of effects on adenosine receptors and phosphodiesterases. As a high-affinity caffeine-like probe, MBED could also help identify the caffeine-binding site in RyRs. PMID:24423447

  3. OBLIQUELY STRIATED MUSCLE

    PubMed Central

    Rosenbluth, Jack

    1967-01-01

    Segments of the obliquely striated body muscle of Ascaris were fixed at minimum body length after treatment with acetylcholine and at maximum body length after treatment with piperazine citrate and then studied by light and electron microscopy. Evidence was found for two mechanisms of length change: sliding of thin filaments with respect to thick filaments such as occurs in cross-striated muscle, and shearing of thick filaments with respect to each other such that the degree of their stagger increases with extension and decreases with shortening. The shearing mechanism could account for great extensibility in this muscle and in nonstriated muscles in general and could underlie other manifestations of "plasticity" as well. In addition, it is suggested that the contractile apparatus is attached to the endomysium in such a way that the sarcomeres can act either in series, as in cross-striated muscle, or individually. Since the sarcomeres are virtually longitudinal in orientation and are almost coextensive with the muscle fiber, it would, therefore, be possible for a single sarcomere contracting independently to develop tension effectively between widely separated points on the fiber surface, thus permitting very efficient maintenance of isometric tension. PMID:6040534

  4. Ophthalmic striated muscle neoplasms.

    PubMed

    Knowles, D M; Jakobiec, F A; Potter, G D; Jones, I S

    1976-01-01

    Rhabdomyosarcoma, the most common primary malignant childhood orbital tumor, is composed of neoplastic striated muscle cells (rhabdomyoblasts) in various stages of differentiation and in patterns suggestive of neoplastic analogs of normal muscle embryogenesis. Orbital rhabdomyosarcoma is most commonly seen in children and adolescents, the average age of onset of symptoms being 7.8 years. The tumor usually presents as a rapidly evolving exophthalmos, often associated with drooping of the upper eyelid. A mass is palapable in only 25% of cases, loss of central vision at the time of presentation is uncommon, and laboratory studies are often of little help in diagnosis. The best diagnostic aid is a high index of suspicion whenever one sees a rapidly progressive exophthalamos in a child. Orbital rhabdomyosarcoma is almost always of the embryonal type, believed to originate in the orbital soft tissues from undifferentiated pluripotential embryonic mesenchyme. In the past, orbital exenteration has been the primary therapy. Review of 162 literature cases of orbital rhabdomyosarcoma, generally treated by unassisted surgery, revealed that only 25% of the patients survived 3 or more years. Recently, it has been shown that radiation therapy, alone or combined with chemotherapy, can be successful. A multidisciplinary approach, utilizing surgery, radiation therapy and chemotherapy has also been advocated. Both approaches appear to offer greater survival than unassisted orbital exenteration. The possibility of primary radiation therapy is extremely promising; if it becomes increasingly effective, a mutilating surgical procedure may become obsolete.

  5. An invertebrate smooth muscle with striated muscle myosin filaments

    PubMed Central

    Sulbarán, Guidenn; Alamo, Lorenzo; Pinto, Antonio; Márquez, Gustavo; Méndez, Franklin; Padrón, Raúl; Craig, Roger

    2015-01-01

    Muscle tissues are classically divided into two major types, depending on the presence or absence of striations. In striated muscles, the actin filaments are anchored at Z-lines and the myosin and actin filaments are in register, whereas in smooth muscles, the actin filaments are attached to dense bodies and the myosin and actin filaments are out of register. The structure of the filaments in smooth muscles is also different from that in striated muscles. Here we have studied the structure of myosin filaments from the smooth muscles of the human parasite Schistosoma mansoni. We find, surprisingly, that they are indistinguishable from those in an arthropod striated muscle. This structural similarity is supported by sequence comparison between the schistosome myosin II heavy chain and known striated muscle myosins. In contrast, the actin filaments of schistosomes are similar to those of smooth muscles, lacking troponin-dependent regulation. We conclude that schistosome muscles are hybrids, containing striated muscle-like myosin filaments and smooth muscle-like actin filaments in a smooth muscle architecture. This surprising finding has broad significance for understanding how muscles are built and how they evolved, and challenges the paradigm that smooth and striated muscles always have distinctly different components. PMID:26443857

  6. Titin: major myofibrillar components of striated muscle.

    PubMed Central

    Wang, K; McClure, J; Tu, A

    1979-01-01

    Electrophoretic analyses of protein components of striated muscle myofibril purified from various vertebrate and invertebrate species revealed that proteins much larger than myosin heavy chain are present in significant amounts. To define possible roles of these heretofore unidentified proteins, we purified a combination of two uncommonly large proteins, designated as titin, from chicken breast myofibrils. Chemical and immunological studies indicated that titin is distinct from myosin, actin, and filamin. Specific titin anti body crossreacts with similar protein in both skeletal and cardiac myofibrils of many vertebrate and invertebrate species. Immunofluorescent staining of glycerinated chicken breast myofibrils indicated that titin is present in M lines, Z lines, the junctions of A and I bands, and perhaps throughout the entire A bands. Similar staining studies of myofibrils from other species suggest that titinlike proteins may be organized in all myofibrils according to a common architectural plan. We conclude that titin is a structurally conserved myofibrillar component of vertebrate and invertebrate striated muscles. Images PMID:291034

  7. Independent evolution of striated muscles in cnidarians and bilaterians

    PubMed Central

    Steinmetz, Patrick R.H.; Kraus, Johanna E.M.; Larroux, Claire; U. Hammel, Jörg; Amon-Hassenzahl, Annette; Houliston, Evelyn; Wörheide, Gert; Nickel, Michael; Degnan, Bernard M.; Technau, Ulrich

    2012-01-01

    Striated muscles are present in bilaterian animals (e.g. vertebrates, insects, annelids) and some non-bilaterian eumetazoans (i.e. cnidarians and ctenophores). The striking ultrastructural similarity of striated muscles between these animal groups is thought to reflect a common evolutionary origin1, 2. Here we show that a muscle protein core set, including a Myosin type II Heavy Chain motor protein characteristic of striated muscles in vertebrates (MyHC-st), was already present in unicellular organisms before the origin of multicellular animals. Furthermore, myhc-st and myhc-non-muscle (myhc-nm) orthologues are expressed differentially in two sponges, compatible with the functional diversification of myhc paralogues before the origin of true muscles and the subsequent deployment of MyHC-st in fast-contracting smooth and striated muscle. Cnidarians and ctenophores possess myhc-st orthologues but lack crucial components of bilaterian striated muscles, such as troponin complex and titin genes, suggesting the convergent evolution of striated muscles. Consistently, jellyfish orthologues of a shared set of bilaterian z-disc proteins are not associated with striated muscles, but are instead expressed elsewhere or ubiquitously. The independent evolution of eumetazoan striated muscles through the addition of novel proteins to a pre-existing, ancestral contractile apparatus may serve as a paradigm for the evolution of complex animal cell types. PMID:22763458

  8. Subcellular distribution of potassium in striated muscles

    SciTech Connect

    Edelmann, L.

    1984-01-01

    Microanalytical experiments have been performed to answer the question whether the main cellular cation, K+, follows the water distribution in the striated muscle cell or whether K+ follows the distribution of negative fixed charges (beta- and gamma-carboxyl groups of aspartic and glutamic acid residues). Subcellular localization of K and/or of the K surrogates Rb, Cs, and Tl has been investigated by the following methods: Chemical precipitation of K with tetraphenylborate. Autoradiography of alkali-metals and Tl in air-dried and frozen-hydrated preparations. TEM visualization of electron dense Cs and Tl in sections of freeze-dried and plastic embedded muscle. X-ray microanalysis of air-dried myofibrils and muscle cryosections. The experiments consistently show that K, Rb, Cs, and Tl do not follow the water distribution but are mainly accumulated in the A band, especially in the marginal regions, and at Z lines. The same sites preferentially accumulate Cs or uranyl cations when sections of freeze-dried, embedded muscle are exposed to these electron microscopic stains. It is concluded that the detected uneven distribution of K, Rb, Cs, and Tl in muscle is neither a freeze-drying artifact nor an embedding artifact and may result from a weak ion binding to the beta- and gamma-carboxyl groups of cellular proteins.

  9. Poorly Understood Aspects of Striated Muscle Contraction

    PubMed Central

    Månsson, Alf

    2015-01-01

    Muscle contraction results from cyclic interactions between the contractile proteins myosin and actin, driven by the turnover of adenosine triphosphate (ATP). Despite intense studies, several molecular events in the contraction process are poorly understood, including the relationship between force-generation and phosphate-release in the ATP-turnover. Different aspects of the force-generating transition are reflected in the changes in tension development by muscle cells, myofibrils and single molecules upon changes in temperature, altered phosphate concentration, or length perturbations. It has been notoriously difficult to explain all these events within a given theoretical framework and to unequivocally correlate observed events with the atomic structures of the myosin motor. Other incompletely understood issues include the role of the two heads of myosin II and structural changes in the actin filaments as well as the importance of the three-dimensional order. We here review these issues in relation to controversies regarding basic physiological properties of striated muscle. We also briefly consider actomyosin mutation effects in cardiac and skeletal muscle function and the possibility to treat these defects by drugs. PMID:25961006

  10. Neurohypophyseal Hormones: Novel Actors of Striated Muscle Development and Homeostasis

    PubMed Central

    Costa, Alessandra; Rossi, Eleonora; Scicchitano, Bianca Maria; Coletti, Dario; Moresi, Viviana

    2014-01-01

    Since the 1980’s, novel functional roles of the neurohypophyseal hormones vasopressin and oxytocin have emerged. Several studies have investigated the effects of these two neurohormones on striated muscle tissues, both in vitro and in vivo. The effects of vasopressin on skeletal myogenic cells, developing muscle and muscle homeostasis have been documented. Oxytocin appears to have a greater influence on cardiomyocite differentiation and heart homeostasis. This review summarizes the studies on these novel roles of the two neurohypophyseal hormones, and open the possibility of new therapeutic approaches for diseases affecting striated muscle. PMID:26913138

  11. Dynamic Regulation of Sarcomeric Actin Filaments in Striated Muscle

    PubMed Central

    Ono, Shoichiro

    2010-01-01

    In striated muscle, the actin cytoskeleton is differentiated into myofibrils. Actin and myosin filaments are organized in sarcomeres and specialized for producing contractile forces. Regular arrangement of actin filaments with uniform length and polarity is critical for the contractile function. However, the mechanisms of assembly and maintenance of sarcomeric actin filaments in striated muscle are not completely understood. Live imaging of actin in striated muscle has revealed that actin subunits within sarcomeric actin filaments are dynamically exchanged without altering overall sarcomeric structures. A number of regulators for actin dynamics have been identified, and malfunction of these regulators often result in disorganization of myofibril structures or muscle diseases. Therefore, proper regulation of actin dynamics in striated muscle is critical for assembly and maintenance of functional myofibrils. Recent studies have suggested that both enhancers of actin dynamics and stabilizers of actin filaments are important for sarcomeric actin organization. Further investigation of the regulatory mechanism of actin dynamics in striated muscle should be a key to understanding how myofibrils develop and operate. © 2010 Wiley-Liss, Inc. PMID:20737540

  12. The anatomy of the sarcoplasmic reticulum in vertebrate skeletal muscle: its implications for excitation contraction coupling.

    PubMed

    Sommer, J R

    1982-01-01

    The sarcoplasmic reticulum in situ is an intricate tubular network that surrounds the contractile material in striated muscle cells. Its topographical relationship to other intracellular components, especially the myofibrils, is rather rigidly maintained by a cytoskeleton which enmeshes Z line material and sarcoplasmic reticulum and, ultimately, is anchored at the plasmalemma. As a result, the two main components of the sarcoplasmic reticulum, the junctional SR and the free SR, retain their typical location in the A band region and in the I band region, respectively. The junctional SR, which is thought to be the site for calcium storage and release for contraction, is, thus, always well within one micron of the regulatory proteins associated with the actin filaments. The junctional SR, a synonym for terminal cisterna applying to both skeletal and cardiac muscle, is generally held to be involved in the translation of the action potential into calcium release, mainly because of the close topographic apposition between the junctional SR and the plasmalemma, especially in skeletal muscle. This attractive structure-function correlation is challenged by the observation that in bird cardiac muscle 80% of the junctional SR is spacially far removed from plasmalemma, the site of electrical activity. This anomalous topography is not in conflict with the notion that translation of the action potential into calcium release may be accomplished by a differential transmitter substance, e.g. calcium. Any hypothesis dealing with this problem must account for the anatomy of the bird heart.

  13. Enrichment and terminal differentiation of striated muscle progenitors in vitro

    SciTech Connect

    Becher, Ulrich M.; Breitbach, Martin; Sasse, Philipp; Garbe, Stephan; Ven, Peter F.M. van der; Fuerst, Dieter O.; Fleischmann, Bernd K.

    2009-10-01

    Enrichment and terminal differentiation of mammalian striated muscle cells is severely hampered by fibroblast overgrowth, de-differentiation and/or lack of functional differentiation. Herein we report a new, reproducible and simple method to enrich and terminally differentiate muscle stem cells and progenitors from mice and humans. We show that a single gamma irradiation of muscle cells induces their massive differentiation into structurally and functionally intact myotubes and cardiomyocytes and that these cells can be kept in culture for many weeks. Similar results are also obtained when treating skeletal muscle-derived stem cells and progenitors with Mitomycin C.

  14. Effect of halothane on isometric twitch and tetanus response and the associated heat production in striated muscle of frogs.

    PubMed

    Price, K A; Matsumoto, Y; Frederickson, E L

    1975-01-01

    The purpose of these investigations was to determine the effect of halothane on isometric contraction of striated muscle and to measure the associated heat production. This basic information is necessary before studies more directly relating to malignant hyperthermia are undertaken. Sartorius muscles were isolate from Rana pipiens during winter and summer months. It appears from these experiments that there is a prolongation of the relaxation phase of the twitch and tetanus responses with low concentrations of halothane, with a more diffuse effect on the contractile process evident at higher administered concentrations. The results of heat measurements, using a sensitive thermopile-galvanometer system, are compatible with the hypotheses that this effect on relaxation could result from either an interference with calcium reuptake by the sarcoplasmic reticulum or an increased affinity of the troponintropomyosin complex for available calcium. PMID:1080024

  15. The Chromatin Remodeling Complex Chd4/NuRD Controls Striated Muscle Identity and Metabolic Homeostasis.

    PubMed

    Gómez-Del Arco, Pablo; Perdiguero, Eusebio; Yunes-Leites, Paula Sofia; Acín-Pérez, Rebeca; Zeini, Miriam; Garcia-Gomez, Antonio; Sreenivasan, Krishnamoorthy; Jiménez-Alcázar, Miguel; Segalés, Jessica; López-Maderuelo, Dolores; Ornés, Beatriz; Jiménez-Borreguero, Luis Jesús; D'Amato, Gaetano; Enshell-Seijffers, David; Morgan, Bruce; Georgopoulos, Katia; Islam, Abul B M M K; Braun, Thomas; de la Pompa, José Luis; Kim, Johnny; Enriquez, José A; Ballestar, Esteban; Muñoz-Cánoves, Pura; Redondo, Juan Miguel

    2016-05-10

    Heart muscle maintains blood circulation, while skeletal muscle powers skeletal movement. Despite having similar myofibrilar sarcomeric structures, these striated muscles differentially express specific sarcomere components to meet their distinct contractile requirements. The mechanism responsible is still unclear. We show here that preservation of the identity of the two striated muscle types depends on epigenetic repression of the alternate lineage gene program by the chromatin remodeling complex Chd4/NuRD. Loss of Chd4 in the heart triggers aberrant expression of the skeletal muscle program, causing severe cardiomyopathy and sudden death. Conversely, genetic depletion of Chd4 in skeletal muscle causes inappropriate expression of cardiac genes and myopathy. In both striated tissues, mitochondrial function was also dependent on the Chd4/NuRD complex. We conclude that an epigenetic mechanism controls cardiac and skeletal muscle structural and metabolic identities and that loss of this regulation leads to hybrid striated muscle tissues incompatible with life.

  16. X-ray Diffraction Studies of Striated Muscles

    SciTech Connect

    Squire, J.M.; Knupp, C.; Roessle, M.; Al-Khayat, H.A.; Irving, T.C.; Eakins, F.; Mok, N.-S.; Harford, J.J.; Reedy, M.K.

    2006-04-24

    In this short review a number of recent X-ray diffraction results on the highly ordered striated muscles in insects and in bony fish have been briefly described. What is clear is that this technique applied to muscles which are amenable to rigorous analysis, taken together with related data from other sources (e.g. protein crystallography, biochemistry, mechanics, computer modelling) can provide not only the best descriptions yet available on the myosin head organisations on different myosin filaments in the relaxed state, but can also show the sequence of molecular events that occurs in the contractile cycle, and may also help to explain such phenomena as stretch-activation. X-ray diffraction is clearly an enormously powerful tool in studies of muscle. It has already provided a wealth of detail on muscle ultrastructure; it is providing ever more fascinating insights into molecular events in the 50-year old sliding filament mechanism, and there remains a great deal more potential that is as yet untapped.

  17. Calcium signaling in insulin action on striated muscle.

    PubMed

    Contreras-Ferrat, A; Lavandero, S; Jaimovich, E; Klip, A

    2014-11-01

    Striated muscles (skeletal and cardiac) are major physiological targets of insulin and this hormone triggers complex signaling pathways regulating cell growth and energy metabolism. Insulin increases glucose uptake into muscle cells by stimulating glucose transporter (GLUT4) translocation from intracellular compartments to the cell surface. The canonical insulin-triggered signaling cascade controlling this process is constituted by well-mapped tyrosine, lipid and serine/threonine phosphorylation reactions. In parallel to these signals, recent findings reveal insulin-dependent Ca(2+) mobilization in skeletal muscle cells and cardiomyocytes. Specifically, insulin activates the sarco-endoplasmic reticulum (SER) channels that release Ca(2+) into the cytosol i.e., the Ryanodine Receptor (RyR) and the inositol 1,4,5-triphosphate receptor (IP3R). In skeletal muscle cells, a rapid, insulin-triggered Ca(2+) release occurs through RyR, that is brought about upon S-glutathionylation of cysteine residues in the channel by reactive oxygen species (ROS) produced by the early activation of the NADPH oxidase (NOX2). In cardiomyocytes insulin induces a fast and transient increase in cytoplasmic [Ca(2+)]i trough L-type Ca(2+) channels activation. In both cell types, a relatively slower Ca(2+) release also occurs through IP3R activation, and is required for GLUT4 translocation and glucose uptake. The insulin-dependent Ca(2+) released from IP3R of skeletal muscle also promotes mitochondrial Ca(2+) uptake. We review here these actions of insulin on intracellular Ca(2+) channel activation and their impact on GLUT4 traffic in muscle cells, as well as other implications of insulin-dependent Ca(2+) release from the SER. PMID:25224502

  18. Muscle Lim Protein isoform negatively regulates striated muscle actin dynamics and differentiation

    PubMed Central

    Vafiadaki, Elizabeth; Arvanitis, Demetrios A.; Papalouka, Vasiliki; Terzis, Gerasimos; Roumeliotis, Theodoros I.; Spengos, Konstantinos; Garbis, Spiros D.; Manta, Panagiota; Kranias, Evangelia G.; Sanoudou, Despina

    2015-01-01

    Muscle Lim Protein (MLP) has emerged as a critical regulator of striated muscle physiology and pathophysiology. Mutations in cysteine and glycine-rich protein 3 (CSRP3), the gene encoding MLP, have been directly associated with human cardiomyopathies, while aberrant expression patterns are reported in human cardiac and skeletal muscle diseases. Increasing evidence suggests that MLP has an important role in both myogenic differentiation and myocyte cytoarchitecture, although the full spectrum of its intracellular roles has not been delineated. We report the discovery of an alternative splice variant of MLP, designated as MLP-b, showing distinct expression in neuromuscular disease and direct roles in actin dynamics and muscle differentiation. This novel isoform originates by alternative splicing of exons 3 and 4. At the protein level, it contains the N-terminus first half LIM domain of MLP and a unique sequence of 22 amino acids. Physiologically it is expressed during early differentiation, whereas its overexpression reduces C2C12 differentiation and myotube formation. This may be mediated through its inhibition of MLP/CFL2-mediated F-actin dynamics. In differentiated striated muscles, MLP-b localizes to the sarcomeres and binds directly to Z-disc components including α-actinin, T-cap and MLP. Our findings unveil a novel player in muscle physiology and pathophysiology that is implicated in myogenesis as a negative regulator of myotube formation, and in differentiated striated muscles as a contributor to sarcomeric integrity. PMID:24860983

  19. Carbonic anhydrase in the sarcoplasmic reticulum of rabbit skeletal muscle.

    PubMed Central

    Bruns, W; Dermietzel, R; Gros, G

    1986-01-01

    Sarcoplasmic reticulum vesicles and mitochondria were prepared from red and white skeletal muscles of the rabbit. The preparations were characterized in terms of their specific activities of citrate synthase, basal (Mg2+-dependent) and Ca2+-dependent ATPase (the latter two in the presence of NaN3 and ouabain), and their specific carbonic anhydrase activities were determined. Skeletal muscle mitochondria had high specific activities of citrate synthase (700-1200 mu. mg protein-1) and low carbonic anhydrase activities (0.1-0.4 u. ml mg protein-1). The latter are likely to be due to a contamination of the preparations with sarcoplasmic reticulum (s.r.) Preparations of s.r. vesicles showed negligible activities of citrate synthase and the expected differing patterns of basal and Ca2+-dependent ATPase in red and white muscles. Specific carbonic anhydrase activities in s.r. from both muscle types were high (2-4 u. ml mg protein-1). The highest carbonic anhydrase activity, 11 u. ml mg protein-1, was found in s.r. from rabbit m. masseter. The inhibition constant of s.r. carbonic anhydrase towards acetazolamide was 4-6 X 10(-8) M and similar but not identical to that of cytosolic carbonic anhydrase II. It appears possible that the carbonic anhydrase II-like enzyme previously found by us in muscle homogenates (Siffert & Gros, 1982) originates from the s.r. Histochemical studies using the dansylsuphonamide method described previously (Dermietzel, Leibstein, Siffert, Zamboglou & Gros, 1985) showed an intracellular pattern of carbonic anhydrase staining compatible with the presence of the enzyme in s.r.: spots homogeneously distributed across the fibre cross-sections in transversely sectioned fibres and thin, longitudinally oriented, bands in longitudinally sectioned fibres. It is estimated that s.r. carbonic anhydrase accelerates CO2 hydration within the s.r. approximately 1000-fold. Thus, CO2 and HCO3- react fast enough to provide a rapid source and sink for protons leaving

  20. Use of flow, electrical, and mechanical stimulation to promote engineering of striated muscles.

    PubMed

    Rangarajan, Swathi; Madden, Lauran; Bursac, Nenad

    2014-07-01

    The field of tissue engineering involves design of high-fidelity tissue substitutes for predictive experimental assays in vitro and cell-based regenerative therapies in vivo. Design of striated muscle tissues, such as cardiac and skeletal muscle, has been particularly challenging due to a high metabolic demand and complex cellular organization and electromechanical function of the native tissues. Successful engineering of highly functional striated muscles may thus require creation of biomimetic culture conditions involving medium perfusion, electrical and mechanical stimulation. When optimized, these external cues are expected to synergistically and dynamically activate important intracellular signaling pathways leading to accelerated muscle growth and development. This review will discuss the use of different types of tissue culture bioreactors aimed at providing conditions for enhanced structural and functional maturation of engineered striated muscles.

  1. Use of flow, electrical, and mechanical stimulation to promote engineering of striated muscles

    PubMed Central

    Rangarajan, Swathi; Madden, Lauran; Bursac, Nenad

    2014-01-01

    The field of tissue engineering involves design of high-fidelity tissue substitutes for predictive experimental assays in vitro and cell-based regenerative therapies in vivo. Design of striated muscle tissues, such as cardiac and skeletal muscle, has been particularly challenging due to a high metabolic demand and complex cellular organization and electromechanical function of the native tissues. Successful engineering of highly functional striated muscles may thus require creation of biomimetic culture conditions involving medium perfusion, electrical and mechanical stimulation. When optimized, these external cues are expected to synergistically and dynamically activate important intracellular signaling pathways leading to accelerated muscle growth and development. This review will discuss the use of different types of tissue culture bioreactors aimed at providing conditions for enhanced structural and functional maturation of engineered striated muscles. PMID:24366526

  2. Segregation of striated and smooth muscle lineages by a Notch-dependent regulatory network

    PubMed Central

    2014-01-01

    Background Lineage segregation from multipotent epithelia is a central theme in development and in adult stem cell plasticity. Previously, we demonstrated that striated and smooth muscle cells share a common progenitor within their epithelium of origin, the lateral domain of the somite-derived dermomyotome. However, what controls the segregation of these muscle subtypes remains unknown. We use this in vivo bifurcation of fates as an experimental model to uncover the underlying mechanisms of lineage diversification from bipotent progenitors. Results Using the strength of spatio-temporally controlled gene missexpression in avian embryos, we report that Notch harbors distinct pro-smooth muscle activities depending on the duration of the signal; short periods prevent striated muscle development and extended periods, through Snail1, promote cell emigration from the dermomyotome towards a smooth muscle fate. Furthermore, we define a Muscle Regulatory Network, consisting of Id2, Id3, FoxC2 and Snail1, which acts in concert to promote smooth muscle by antagonizing the pro-myogenic activities of Myf5 and Pax7, which induce striated muscle fate. Notch and BMP closely regulate the network and reciprocally reinforce each other’s signal. In turn, components of the network strengthen Notch signaling, while Pax7 silences this signaling. These feedbacks augment the robustness and flexibility of the network regulating muscle subtype segregation. Conclusions Our results demarcate the details of the Muscle Regulatory Network, underlying the segregation of muscle sublineages from the lateral dermomyotome, and exhibit how factors within the network promote the smooth muscle at the expense of the striated muscle fate. This network acts as an exemplar demonstrating how lineage segregation occurs within epithelial primordia by integrating inputs from competing factors. PMID:25015411

  3. The significance of striated muscle in the mammary glands of marsupials.

    PubMed Central

    Griffiths, M; Slater, E

    1988-01-01

    The distribution and amounts of striated muscle within the mammary glands of pouched and pouchless marsupials from Australia and South America are described. Invasions into the mammary secretory parenchyma in pouchless marsupials by swathes of striated muscle from the ilio-marsupialis muscle are massive, in some instances concentrated into discrete muscles, which are inserted on to the bases of the teats; the name retractor mammae is proposed for these muscles. In pouched marsupials striated muscle penetrates the parenchyma, but the distribution is diffuse and the muscle strands are not inserted on to teats except in the instance of the glands of the honey possum Tarsipes rostratus. The young of anaesthetised pouchless marsupials hang down from the teats; as anaesthesia wears off they are hauled up tightly into the mammary area. It is concluded that this is a result of contraction of the retractor mammae muscles and that it is a means of protecting the naked young from injury by rough terrain. The mammary gland musculature in pouched marsupials is considered to be vestigial, but its contraction may have the function of initiating a 'tap-response' contraction of myoepithelium acting synergistically with the 'let-down' hormone mesotocin. Mechanisms of imbibition of milk by marsupial neonates, based on observations that they can suck fluid from non-distortable tubes, are discussed. Images Fig. 2 Fig. 3 Fig. 4 Fig. 5 Fig. 6 Fig. 7 Fig. 8 Fig. 9 Fig. 10 PMID:3417541

  4. Z-line formins promote contractile lattice growth and maintenance in striated muscles of C. elegans

    PubMed Central

    Mi-Mi, Lei; Votra, SarahBeth; Kemphues, Kenneth; Bretscher, Anthony

    2012-01-01

    Muscle contraction depends on interactions between actin and myosin filaments organized into sarcomeres, but the mechanism by which actin filaments incorporate into sarcomeres remains unclear. We have found that, during larval development in Caenorhabditis elegans, two members of the actin-assembling formin family, CYK-1 and FHOD-1, are present in striated body wall muscles near or on sarcomere Z lines, where barbed ends of actin filaments are anchored. Depletion of either formin during this period stunted growth of the striated contractile lattice, whereas their simultaneous reduction profoundly diminished lattice size and number of striations per muscle cell. CYK-1 persisted at Z lines in adulthood, and its near complete depletion from adults triggered phenotypes ranging from partial loss of Z line–associated filamentous actin to collapse of the contractile lattice. These results are, to our knowledge, the first genetic evidence implicating sarcomere-associated formins in the in vivo organization of the muscle cytoskeleton. PMID:22753896

  5. Effects of boldine on mouse diaphragm and sarcoplasmic reticulum vesicles isolated from skeletal muscle.

    PubMed

    Kang, J J; Cheng, Y W

    1998-02-01

    The effects of boldine [(S)-2,9-dihydroxy-1,10-dimethoxyaporphine], a major alkaloid in the leaves and bark of boldo (Peumus boldus Mol.), on skeletal muscle were studied using mouse diaphragm and isolated sarcoplasmic reticulum membrane vesicles. Boldine, at 10-200 microM, has little effect on the muscle-evoked twitches; however, the ryanodine-induced contracture was potentiated dose-dependently. At higher concentrations of 300 microM, boldine by itself induced muscle contracture of two phases, which were caused by the influx of extracellular Ca2+ and induction of Ca2+ release from the internal Ca2+ storage site, the sarcoplasmic reticulum, respectively. When tested with isolated sarcoplasmic reticulum membrane vesicles, boldine dose-dependently induced Ca2+ release from actively loaded sarcoplasmic reticulum vesicles isolated from skeletal muscle of rabbit or rat which was inhibited by ruthenium red, suggesting that the release was through the Ca2+ release channel, also known as the ryanodine receptor. Boldine also dose-dependently increased apparent [3H]-ryanodine binding with the EC50 value of 50 microM. In conclusion, we have shown that boldine could sensitize the ryanodine receptor and induce Ca2+ release from the internal Ca2+ storage site of skeletal muscle. PMID:9491763

  6. Porcine malignant hyperthermia susceptibility: increased calcium-sequestering activity of skeletal muscle sarcoplasmic reticulum.

    PubMed Central

    O'Brien, P J

    1986-01-01

    This study tested the hypothesis that calcium-sequestration by isolated sarcoplasmic reticulum was abnormal in skeletal muscle of malignant hyperthermia-susceptible swine. A heavy sarcoplasmic reticulum fraction was isolated from malignant hyperthermia and control muscle using differential and density-gradient centrifugation. Prior to onset of malignant hyperthermia, calcium-sequestering activity (Vmax at 37 degrees C, mumol calcium/mg/min) was twofold increased in malignant hyperthermia sarcoplasmic reticulum compared to control sarcoplasmic reticulum (1.96 +/- 0.50 versus 4.00 +/- 0.87, P less than 0.01), although thermodynamic and kinetic properties of this activity were otherwise indistinguishable between groups. This increased activity of the malignant hyperthermia sarcoplasmic reticulum fraction was associated with twofold increased concentration of Ca-ATPase and calsequestrin protein. When a malignant hyperthermia-reaction developed, calcium-uptake was depressed to less than 5% of control values. These data indicate that malignant hyperthermia is not initiated due to a defect in the calcium-sequestration mechanism, however, loss of calcium-uptake activity occurring after the onset of malignant hyperthermia might result in the propagation and irreversibility of the malignant hyperthermia reaction. Images Fig. 1. PMID:3742368

  7. A striated muscle on the hard palate of rodents and rabbits.

    PubMed

    Pavlíková, H; Witter, K; Mísek, I

    2004-04-01

    Summary A striated muscle of the hard palate has been previously described in some rodents and rabbits. It is not termed in the official veterinary anatomical nomenclature. The aim of this work was to verify the existence of this muscle. Heads of the golden hamster (Mesocricetus auratus), the guinea pig (Cavia aperea f. porcellus), the laboratory rat (Rattus norvegicus var. alba), the field vole (Microtus agrestis) and the domestic rabbit (Oryctolagus cuniculus f. domesticus) have been dissected. Moreover, histological sections have been prepared from heads of the field vole. In all species under study, we could detect a striated muscle of the hard palate composed of an anterior and a posterior muscle. The anterior muscle originated on the os incisivum and diverged in anterior, lateral and posterior directions. The posterior muscle originated on the processus palatinus maxillae and verged into the m. buccinator. Inter-species differences could be detected in shape and position of the muscle. The palatal muscle was innervated by the ramus buccalis of the facial nerve. Whether this muscle should be classified as an individual facial muscle or as a part of the m. buccinator remains to be discussed. PMID:15027950

  8. A Calsequestrin-1 Mutation Associated with a Skeletal Muscle Disease Alters Sarcoplasmic Ca2+ Release.

    PubMed

    D'Adamo, Maria Cristina; Sforna, Luigi; Visentin, Sergio; Grottesi, Alessandro; Servettini, Llenio; Guglielmi, Luca; Macchioni, Lara; Saredi, Simona; Curcio, Maurizio; De Nuccio, Chiara; Hasan, Sonia; Corazzi, Lanfranco; Franciolini, Fabio; Mora, Marina; Catacuzzeno, Luigi; Pessia, Mauro

    2016-01-01

    An autosomal dominant protein aggregate myopathy, characterized by high plasma creatine kinase and calsequestrin-1 (CASQ1) accumulation in skeletal muscle, has been recently associated with a missense mutation in CASQ1 gene. The mutation replaces an evolutionarily-conserved aspartic acid with glycine at position 244 (p.D244G) of CASQ1, the main sarcoplasmic reticulum (SR) Ca2+ binding and storage protein localized at the terminal cisternae of skeletal muscle cells. Here, immunocytochemical analysis of myotubes, differentiated from muscle-derived primary myoblasts, shows that sarcoplasmic vacuolar aggregations positive for CASQ1 are significantly larger in CASQ1-mutated cells than control cells. A strong co-immuno staining of both RyR1 and CASQ1 was also noted in the vacuoles of myotubes and muscle biopsies derived from patients. Electrophysiological recordings and sarcoplasmic Ca2+ measurements provide evidence for less Ca2+ release from the SR of mutated myotubes when compared to that of controls. These findings further clarify the pathogenic nature of the p.D244G variant and point out defects in sarcoplasmic Ca2+ homeostasis as a mechanism underlying this human disease, which could be distinctly classified as "CASQ1-couplonopathy". PMID:27196359

  9. A Calsequestrin-1 Mutation Associated with a Skeletal Muscle Disease Alters Sarcoplasmic Ca2+ Release

    PubMed Central

    D’Adamo, Maria Cristina; Sforna, Luigi; Visentin, Sergio; Grottesi, Alessandro; Servettini, llenio; Guglielmi, Luca; Macchioni, Lara; Saredi, Simona; Curcio, Maurizio; De Nuccio, Chiara; Hasan, Sonia; Corazzi, Lanfranco; Franciolini, Fabio; Mora, Marina; Catacuzzeno, Luigi; Pessia, Mauro

    2016-01-01

    An autosomal dominant protein aggregate myopathy, characterized by high plasma creatine kinase and calsequestrin-1 (CASQ1) accumulation in skeletal muscle, has been recently associated with a missense mutation in CASQ1 gene. The mutation replaces an evolutionarily-conserved aspartic acid with glycine at position 244 (p.D244G) of CASQ1, the main sarcoplasmic reticulum (SR) Ca2+ binding and storage protein localized at the terminal cisternae of skeletal muscle cells. Here, immunocytochemical analysis of myotubes, differentiated from muscle-derived primary myoblasts, shows that sarcoplasmic vacuolar aggregations positive for CASQ1 are significantly larger in CASQ1-mutated cells than control cells. A strong co-immuno staining of both RyR1 and CASQ1 was also noted in the vacuoles of myotubes and muscle biopsies derived from patients. Electrophysiological recordings and sarcoplasmic Ca2+ measurements provide evidence for less Ca2+ release from the SR of mutated myotubes when compared to that of controls. These findings further clarify the pathogenic nature of the p.D244G variant and point out defects in sarcoplasmic Ca2+ homeostasis as a mechanism underlying this human disease, which could be distinctly classified as “CASQ1-couplonopathy”. PMID:27196359

  10. Identification of dantrolene binding sites in porcine skeletal muscle sarcoplasmic reticulum.

    PubMed

    Parness, J; Palnitkar, S S

    1995-08-01

    Dantrolene, an intracellularly acting skeletal muscle relaxant, inhibits Ca2+ release from the sarcoplasmic reticulum during excitation-contraction coupling by an unknown mechanism. The drug is used to treat malignant hyperthermia, a genetic sensitivity to volatile anesthetics which results in the massive release of intracellular Ca2+ from affected skeletal muscle. We hypothesize that determination of the site of action of dantrolene will lead to further understanding of the regulation of sarcoplasmic reticulum calcium release. We report the identification of specific dantrolene binding sites in porcine skeletal muscle sarcoplasmic reticulum using a rapid filtration binding assay for [3H]dantrolene. The binding isotherm in the heavy sarcoplasmic reticulum fraction indicates a single binding site with a Kd of 277 +/- 25 nM and a Bmax of 13.1 +/- 1.5 pmol/mg of protein. Pharmacological specificity is characterized by inhibition of [3H]dantrolene binding with unlabeled dantrolene, or azumolene, a physiologically active congener, but not with aminodantrolene, which is physiologically inactive. Drug binding is maximal at pH 6.5-7.5, requires no Ca2+ or Mg2+, and is inhibited by salt concentrations above 100 mM. [3H]Dantrolene binding is greatest in the sarcoplasmic reticulum, which contains the ryanodine receptor, the primary calcium release channel. No binding is detected in the fractions enriched for sarcolemma or transverse tubules. We suggest that dantrolene inhibits calcium release from the sarcoplasmic reticulum by either direct or indirect interaction with the ryanodine receptor. PMID:7629173

  11. Modulation of the cytosolic androgen receptor in striated muscle by sex steroids

    NASA Technical Reports Server (NTRS)

    Rance, N. E.; Max, S. E.

    1982-01-01

    The influence of orchiectomy (GDX) and steroid administration on the level of the cytosolic androgen receptor in the rat levator ani muscle and in rat skeletal muscles (tibialis anterior and extensor digitorum longus) was studied. Androgen receptor binding to muscle cytosol was measured using H-3 methyltrienolone (R1881) as ligand, 100 fold molar excess unlabeled R1881 to assess nonspecific binding, and 500 fold molar excess of triamcinolone acetonide to prevent binding to glucocorticoid and progestin receptors. Results demonstrate that modification of the levels of sex steroids can alter the content of androgen receptors of rat striated muscle. Data suggest that: (1) cytosolic androgen receptor levels increase after orchiectomy in both levator ani muscle and skeletal muscle; (2) the acute increase in receptor levels is blocked by an inhibitor of protein synthesis; and (3) administration of estradiol-17 beta to castrated animals increases receptor binding in levator ani muscle but not in skeletal muscle.

  12. Myosin filament sliding through the Z-disc relates striated muscle fibre structure to function.

    PubMed

    Rode, Christian; Siebert, Tobias; Tomalka, Andre; Blickhan, Reinhard

    2016-03-16

    Striated muscle contraction requires intricate interactions of microstructures. The classic textbook assumption that myosin filaments are compressed at the meshed Z-disc during striated muscle fibre contraction conflicts with experimental evidence. For example, myosin filaments are too stiff to be compressed sufficiently by the muscular force, and, unlike compressed springs, the muscle fibres do not restore their resting length after contractions to short lengths. Further, the dependence of a fibre's maximum contraction velocity on sarcomere length is unexplained to date. In this paper, we present a structurally consistent model of sarcomere contraction that reconciles these findings with the well-accepted sliding filament and crossbridge theories. The few required model parameters are taken from the literature or obtained from reasoning based on structural arguments. In our model, the transition from hexagonal to tetragonal actin filament arrangement near the Z-disc together with a thoughtful titin arrangement enables myosin filament sliding through the Z-disc. This sliding leads to swivelled crossbridges in the adjacent half-sarcomere that dampen contraction. With no fitting of parameters required, the model predicts straightforwardly the fibre's entire force-length behaviour and the dependence of the maximum contraction velocity on sarcomere length. Our model enables a structurally and functionally consistent view of the contractile machinery of the striated fibre with possible implications for muscle diseases and evolution.

  13. Electron microscopical and histochemical studies on the transverse striated muscles of birds after prolonged hypokinesis

    NASA Technical Reports Server (NTRS)

    Belak, M.; Kocisova, J.; Marcanik, J.; Boda, K.; Skarda, R.

    1981-01-01

    Studies of the gastrocnemius muscle were carried out in 4 month old cockerels of the laying hybrid after hypokinesis lasting 15 and 30 days. It was found that restricted movement resulted in dystrophic changes of myotibrils, enlargement of the sarcoplasmic reticulum and oedem of interfibrillar spaces. Histochemical studies revealed focuses of increased activity of non-specific esterase, decreased activity of dehydrogenase of lactic acid and a positive reaction of acid phosphatase.

  14. Mode of action of prilocaine on sarcoplasmic reticulum in skinned skeletal muscle fibers.

    PubMed

    Saida, K; Suzuki, A

    1981-12-01

    Single fibers were isolated from the semitendinosus muscle of a frog and the skinned fibers were prepared by the removal of the sarcolemma. In the range of several millimolar concentrations, prilocaine caused a contracture of the single fibers in both polarized and depolarized states. It also induced Ca++ release from the sarcoplasmic reticulum and depressed Ca++ uptake by the sarcoplasmic reticulum. Prilocaine increased the release of Ca++ with increasing concentration, but, unlike caffeine, prilocaine could not enhance the Ca++-induced Ca++ release mechanism of the sarcoplasmic reticulum. Like a depolarization-induced Ca++ release, the prilocaine-induced Ca++ release was not inhibited by Mg++, whereas it was inhibited by sucrose. The Ca++ release induced with prilocaine occurred only immediately after the application; thereafter the Ca++ release mechanism seemed to be inactivated by the prolonged presence of the drug. A similar inactivation in the mechanism of prilocaine-induced Ca++ release also occurred when the extent of depolarization of the sarcoplasmic reticulum membrane increased. These results suggest that the mode of action of prilocaine on the sarcoplasmic reticulum could be a depolarization-like action.

  15. Sarcoplasmic masses in the skeletal muscle of a stranded pigmy sperm whale (Kogia breviceps).

    PubMed

    Sierra, Eva; de los Monteros, Antonio Espinosa; Fernández, Antonio; Arbelo, Manuel; Caballero, María José; Rivero, Miguel; Herráez, Pedro

    2013-07-01

    We measured the abundance of sarcoplasmic masses within skeletal muscle myocytes of an adult female stranded pigmy sperm whale (Kogia breviceps). The presence of these masses in other species has been reported in association with myopathies, including myotonic dystrophy, the most frequently related pathology. Other histopathologic muscle changes included a high number of internal nuclei, variations in fiber size and shape, and the predominance of type I fibers.

  16. Amino acid sequence of horseshoe crab, Tachypleus tridentatus, striated muscle troponin C.

    PubMed

    Kobayashi, T; Kagami, O; Takagi, T; Konishi, K

    1989-05-01

    The amino acid sequence of troponin C obtained from horseshoe crab, Tachypleus tridentatus, striated muscle was determined by sequence analysis and alignments of chemically and enzymatically cleaved peptides. Troponin C is composed of 153 amino acid residues with a blocked N-terminus and contains no tryptophan or cysteine residue. The site I, one of the four Ca2+-binding sites, is considered to have lost its ability to bind Ca2+ owing to the replacements of certain amino acid residues.

  17. Age changes in cross striated muscle of the rat

    PubMed Central

    Gutmann, E.; Hanzlíková, V.; Vyskočil, F.

    1971-01-01

    1. Senile muscle atrophy is characterized by a marked reduction in the frequency of spontaneous transmitter release with no electrophysiological evidence of denervation. 2. In spite of the reduced number of muscle fibres, there is no ultrastructural evidence for denervation at the end-plates. There is agglutination of synaptic vesicles, neurotubules and filaments, thickening of the basement membrane, widening of the primary synaptic cleft, and irregular branching of the junctional infoldings, but no axonal degeneration. 3. The contractile process in senile muscles is slowed down as is indicated by a prolongation of contraction time, latency period, maximum rate of twitch tension and relaxation time. 4. The muscle fibres show proliferation of the T system and increased SR but no fragmentation as is observed in denervation atrophy. 5. Senile muscle atrophy thus presents some specific features affecting both pre- and post-synaptic structures, related to a very slow process of deterioration of the neuromuscular contact. ImagesFig. 4Fig. 5Fig. 6Fig. 7Fig. 8Fig. 9Fig. 10Fig. 1Fig. 2Fig. 3 PMID:4326996

  18. Striated muscle involvement in experimental oral infection by herpes simplex virus type 1.

    PubMed

    Gonzalez, María Inés; Sanjuan, Norberto A

    2013-07-01

    Herpes simplex virus type 1 is one of the most frequent causes of oral infection in humans, especially during early childhood. Several experimental models have been developed to study the pathogenesis of this virus but all of them employed adult animals. In this work, we developed an experimental model that uses mice younger than 4 days old, to more closely resemble human infection. Mice were infected subcutaneously with the prototype strain McIntyre of Herpes simplex-1, and the progression of infection was studied by immunoperoxidase. All animals died within 24-72 h post-infection, while viral antigens were found in the oral epithelium, nerves and brain. The most striking result was the finding of viral antigens in the nucleus and cytoplasm of cells belonging to striated muscles. Organotypic cultures of striated muscles were performed, and viral replication was observed in them by immunocytochemistry, electron microscopy and viral isolation. We conclude that the infection of striated muscles is present from the onset of oral infection and, eventually, could explain some clinical observations in humans.

  19. ON THE DIFFERENTIAL RESPONSE OF SARCOPLASM AND MYOPLASM TO DENERVATION IN FROG MUSCLE

    PubMed Central

    Muscatello, Umberto; Margreth, Alfredo; Aloisi, Massimo

    1965-01-01

    Electron microscopic evidence is presented that the early response to denervation ("simple atrophy") of the semitendinosus m. of the frog is characterized by a greater prominence of the sarcoplasmic reticulum and by the presence, in the interfibrillar spaces, of mitochondria which are more numerous and smaller than in normal muscle. In contrast with the dynamic changes of the sarcoplasmic structural components, the myofibrils showed a progressive decrease in diameter after denervation and throughout the period studied. By carrying out tissue fractionation experiments, the yield of microsome-protein was found significantly greater in the denervated muscles, as compared with the contralateral controls, in this initial stage. Under the conditions attending the overdevelopment of the sarcoplasmic reticulum (SR), denervated semitendinosus m. incorporated valine-C14 into proteins more actively than the control pairs. The denervated muscles also showed an increase in the number of freely scattered and membrane-bound ribosomes and of polyribosomes, suggesting a more active synthesis of the SR membranes. Pronounced atrophy of the myofibrils, disorganization of the SR, and an increased number of ribonucleoprotein particles lying in the enlarged interfibrillar spaces were the main ultrastructural features of "degenerative atrophy" in frog muscle in the late periods after denervation. The probably adaptive character of the early changes occurring on denervation of frog muscle is discussed. PMID:5857256

  20. The ultrastructure and contractile properties of a fast-acting, obliquely striated, myosin-regulated muscle: the funnel retractor of squids

    PubMed Central

    Rosenbluth, Jack; Szent-Györgyi, Andrew G.; Thompson, Joseph T.

    2010-01-01

    We investigated the ultrastructure, contractile properties, and in vivo length changes of the fast-acting funnel retractor muscle of the long-finned squid Doryteuthis pealeii. This muscle is composed of obliquely striated, spindle-shaped fibers ~3 μm across that have an abundant sarcoplasmic reticulum, consisting primarily of membranous sacs that form ‘dyads’ along the surface of each cell. The contractile apparatus consists of ‘myofibrils’ ~0.25–0.5 μm wide in cross section arrayed around the periphery of each cell, surrounding a central core that contains the nucleus and large mitochondria. Thick myofilaments are ~25 nm in diameter and ~2.8 μm long. ‘Dense bodies’ are narrow, resembling Z lines, but are discontinuous and are not associated with the cytoskeletal fibrillar elements that are so prominent in slower obliquely striated muscles. The cells approximate each other closely with minimal intervening intercellular connective tissue. Our physiological experiments, conducted at 17°C, showed that the longitudinal muscle fibers of the funnel retractor were activated rapidly (8 ms latent period following stimulation) and generated force rapidly (peak twitch force occurred within 50 ms). The longitudinal fibers had low Vmax (2.15 ±0.26 L0 s−1, where L0 was the length that generated peak isometric force) but generated relatively high isometric stress (270±20 mN mm−2 physiological cross section). The fibers exhibited a moderate maximum power output (49.9 W kg−1), compared with vertebrate and arthropod cross striated fibers, at a V/Vmax of 0.33±0.044. During ventilation of the mantle cavity and locomotion, the funnel retractor muscle operated in vivo over a limited range of strains (+0.075 to −0.15 relative to resting length, LR) and at low strain rates (from 0.16 to 0.91 LR s−1 ), corresponding to a range of V/Vmax from 0.073 to 0.42. During the exhalant phase of the jet the range of strains was even narrower: maximum range less than ±0

  1. Sarcoplasmic-reticulum biogenesis in contraction-inhibited skeletal-muscle cultures.

    PubMed Central

    Charuk, J H; Guerin, C; Holland, P C

    1992-01-01

    We have previously shown that inhibition of the spontaneous contractile activity of cultured embryonic-chick skeletal-muscle fibres with tetrodotoxin (TTX) leads to decreased sarcoplasmic-reticulum Ca(2+)-transport rates and steady-state concentrations of the high-energy Ca(2+)-ATPase phosphoenzyme intermediate [Charuk & Holland (1983) Exp. Cell Res. 144, 143-157]. In the present study we used a monoclonal antibody to the Ca(2+)-ATPase to show that there is a decreased amount of enzyme accumulated by contraction-inhibited myotubes. Indirect immunofluorescence microscopy using the monoclonal antibody to the Ca(2+)-ATPase also revealed a disordered subcellular organization of the sarcotubular system in contraction-inhibited myotubes. The biogenesis of sarcoplasmic-reticulum proteins in TTX-paralysed myofibres was studied by labelling cells with [35S]methionine before isolation of the active Ca(2+)-pump membrane fraction. Protein turnover was selectively increased in that fraction from TTX-treated muscle cultures. Electrophoretic analysis and quantitative fluorography confirmed that decreased accumulation of the Ca(2+)-ATPase enzyme in contraction-inhibited myotubes was associated with increased turnover of this protein. The present results demonstrate that biogenesis of the sarcoplasmic-reticulum Ca(2+)-ATPase is regulated by the contractile activity of skeletal-muscle fibres. Images Fig. 1. Fig. 2. Fig. 3. Fig. 5. Fig. 7. PMID:1312329

  2. Frog striated muscle is permeable to hydroxide and buffer anions.

    PubMed

    Venosa, R A; Kotsias, B A; Horowicz, P

    1994-04-01

    Hydroxide, bicarbonate and buffer anion permeabilities in semitendinosus muscle fibers of Rana pipiens were measured. In all experiments, the fibers were initially equilibrated in isotonic, high K2SO4 solutions at pHo = 7.2 buffered with phosphate. Two different methods were used to estimate permeabilities: (i) membrane potential changes were recorded in response to changes in external ion concentrations, and (ii) intracellular pH changes were recorded in response to changes in external concentrations of ions that alter intracellular pH. Constant field equations were used to calculate relative or absolute permeabilities. In the first method, to increase the size of the membrane potential change produced by a sudden change in anion entry, external K+ was replaced by Cs+ prior to changes of the anion under study. At constant external Cs+ activity, a hyperpolarization results from increasing external pH from 7.2 to 10.0 or higher, using either CAPS (3-[cyclohexylamino]-1-propanesulfonic acid) or CHES (2-[N-cyclohexylamino]-ethanesulfonic acid) as buffer. For each buffer, the protonated form is a zwitterion of zero net charge and the nonprotonated form is an anion. Using reported values of H+ permeability, calculations show that the reduction in [H+]o cannot account for the hyperpolarizations produced by alkaline solutions. Membrane hyperpolarization increases with increasing total external buffer concentration at constant external pH, and with increasing external pH at constant external buffer anion concentration. Taken together, these observations indicate that both OH- and buffer anions permeate the surface membrane. The following relative permeabilities were obtained at pHo = 10.0 +/- 0.3: (POH/PK) = 890 +/- 150, (PCAPS/PK) = 12 +/- 2, (PCHES/PK) = 5.3 +/- 0.9, and (PNO3/PK) = 4.7 +/- 0.5. PNO3/PK was independent of pHo up to 10.75. At pHo = 9.6, (PHCO3/PK) = 0.49 +/- 0.03; at pHo = 8.9, (PCl/PK) = 18 +/- 2 and at pHo = 7.1, (PHEPES/PK) = 20 +/- 2. In the second

  3. Ultrastructural studies of the mitochondriae in the striated muscles of birds with regard to experimental hypokinesis

    NASA Technical Reports Server (NTRS)

    Belak, M.; Kocisova, J.; Boda, K.

    1980-01-01

    Electron microscopic studies were carried out on the mitochrondria of the transversely striated muscles with regard to experimental hypokinesia. As compared to the central group the mitochondria of m. pectoralis thoracicus and the m. iliotibialis posterior in hypokinetic birds reveal marked changes. In filamentous and ovoid mitochondria, vacuoles can be observed which in some cases produced larger light formations with following disappearance of the cristae and destruction of mitochondria. Fat particles located at the poles of the altered mitochondria, sporadically occurring also laterally, presented another finding. The Z-lines of the sarcomere did not form a continuous line, but were somewhat shifted.

  4. Actin capping proteins, CapZ (β-actinin) and tropomodulin in amphioxus striated muscle.

    PubMed

    Bao, Yulong; Kake, Takei; Hanashima, Akira; Nomiya, Yui; Kubokawa, Kaoru; Kimura, Sumiko

    2012-11-15

    CapZ (β-actinin) and tropomodulin (Tmod) are capping proteins involved in the maintenance of thin filaments in vertebrate skeletal muscles. In this study, we focused on amphioxus, the most primitive chordate. We searched for CapZ and Tmod genes in the amphioxus genome and determined their primary structures. Amphioxus possess one CapZα gene (CAPZA) and one CapZβ gene (CAPZB), and the transcripts of these genes were found to be 67%-85% identical to those of human CapZ genes. On the other hand, amphioxus contain one Tmod gene (TMOD), and the product of this gene has an identity of approximately 50% with human Tmod genes 1-4. However, helix 2 of amphioxus Tmod, which is involved in protein-binding to tropomyosin, was highly conserved with approximately 74% identity to human Tmod genes. Western blotting indicated the presence of CapZ and Tmod in the striated muscle of amphioxus. These results suggest that unlike most of vertebrates, such as fish, amphibian, bird, and mammal, CapZ from amphioxus striated muscle is derived from two genes CAPZA and CAPZB, and Tmod is derived from one TMOD gene.

  5. The Popeye Domain Containing Genes and their Function in Striated Muscle

    PubMed Central

    Schindler, Roland FR; Scotton, Chiara; French, Vanessa; Ferlini, Alessandra; Brand, Thomas

    2016-01-01

    The Popeye domain containing (POPDC) genes encode a novel class of cAMP effector proteins, which are abundantly expressed in heart and skeletal muscle. Here we will review their role in striated muscle as deduced from work in cell and animal models and the recent analysis of patients carrying a missense mutation in POPDC1. Evidence suggests that POPDC proteins control membrane trafficking of interacting proteins. Furthermore, we will discuss the current catalogue of established protein-protein interactions. In recent years, the number of POPDC-interacting proteins is rising and currently includes ion channels (TREK-1), sarcolemma-associated proteins serving functions in mechanical stability (Dystrophin), compartmentalization (Caveolin 3), scaffolding (ZO-1), trafficking (NDRG4, VAMP2/3) and repair (Dysferlin), or acting as a guanine nucleotide exchange factor for Rho-family GTPases (GEFT). Recent evidence suggests that POPDC proteins might also control the cellular level of the nuclear proto-oncoprotein c-Myc. These data suggests that this family of cAMP-binding proteins probably serves multiple roles in striated muscle. PMID:27347491

  6. Isoform composition, gene expression and sarcomeric protein phosphorylation in striated muscle of mice after space flight

    NASA Astrophysics Data System (ADS)

    Vikhlyantsev, Ivan; Ulanova, Anna; Salmov, Nikolay; Gritsyna, Yulia; Bobylev, Alexandr; Rogachevsky, Vadim; Shenkman, Boris; Podlubnaya, Zoya

    Using RT-PCR and SDS-PAGE, changes in isoform composition, gene expression, titin and nebulin phosphorylation, as well as changes in isoform composition of myosin heavy chains in striated muscles of mice were studied after 30-day-long space flight onboard the Russian spacecraft “BION-M” No. 1. The muscle fibre-type shift from slow-to-fast was observed in m. gastrocnemius and m. tibialis anterior of animals from “Flight” group. A decrease in the content of the NT and N2A titin isoforms and nebulin in the skeletal muscles of animals from “Flight” group was found. Meanwhile, significant differences in gene expression of these proteins in skeletal muscles of mice from “Flight” and “Control” groups were not observed. Using Pro-Q Diamond stain, an increase in titin phosphorylation in m. gastrocnemius of mice from “Flight” group was detected. The content of the NT, N2BA and N2B titin isoforms in cardiac muscle of mice from “Flight” and “Control” groups did not differ, nevertheless an increase in titin gene expression in the myocardium of the “Flight” group animals was found. The observed changes will be discussed in the context of theirs role in contractile activity of striated muscles of mice in conditions of weightlessness. This work was supported by the Russian Foundation for Basic Research (grants No. 14-04-32240, 14-04-00112). Acknowledgement. We express our gratitude to the teams of Institute of Biomedical Problems RAS and “PROGRESS” Corporation involved in the preparation of the “BION-M” mission.

  7. Comparative studies on troponin, a Ca²⁺-dependent regulator of muscle contraction, in striated and smooth muscles of protochordates.

    PubMed

    Obinata, Takashi; Sato, Naruki

    2012-01-01

    Troponin is well known as a Ca(2+)-dependent regulator of striated muscle contraction and it has been generally accepted that troponin functions as an inhibitor of muscle contraction or actin-myosin interaction at low Ca(2+) concentrations, and Ca(2+) at higher concentrations removes the inhibitory action of troponin. Recently, however, troponin became detectable in non-striated muscles of several invertebrates and in addition, unique troponin that functions as a Ca(2+)-dependent activator of muscle contraction has been detected in protochordate animals, although troponin in vertebrate striated muscle is known as an inhibitor of the contraction in the absence of a Ca(2+). Further studies on troponin in invertebrate muscle, especially in non-striated muscle, would provide new insight into the evolution of regulatory systems for muscle contraction and diverse function of troponin and related proteins. The methodology used for preparation and characterization of functional properties of protochordate striated and smooth muscles will be helpful for further studies of troponin in other invertebrate animals.

  8. [A cause of terminal constipation: hypertonia of the striated pelvic muscle. Value of complementary studies].

    PubMed

    Veyrac, M; Parelon, G; Daures, J P; Bories, P; Michel, H

    1988-12-01

    The aim of this study was to define an other cause of idiopathic constipation with a mathematic automatic classification Fifty patients were studied by colonic transit time, anorectal manometry and defecography. Three classical causes were found: 12 patients had hypertonic upper anal pressure; 8 patients had abnormal rectonanal inhibitory reflex and 12 patients had anatomic conditions reducing stool frequencies Automatic mathematic classification coming from optimised bowls technic has led to the characterization of a new cause. Eight young women (mean age 33 years) with hypertonic striated pelvic muscles on straining were identified. Three items were significantly different: defect of sphincteric relaxation without high sphincteric pressure; closure or non opening anorectal angle, hypertonic levator ani on straining. There was a significant high frequency (p less than 0.05) of urinary disorders in this new group. All abnormalities occurred during defecation. This group can be studied with other methods: balloon expulsion, anal electromyography. Clinical identity with urinary pathology may be explained by common neurologic disorders of pelvic striated muscle innervation.

  9. Overexpression of Striated Muscle Activator of Rho Signaling (STARS) Increases C2C12 Skeletal Muscle Cell Differentiation

    PubMed Central

    Wallace, Marita A.; Della Gatta, Paul A.; Ahmad Mir, Bilal; Kowalski, Greg M.; Kloehn, Joachim; McConville, Malcom J.; Russell, Aaron P.; Lamon, Séverine

    2016-01-01

    Background: Skeletal muscle growth and regeneration depend on the activation of satellite cells, which leads to myocyte proliferation, differentiation and fusion with existing muscle fibers. Skeletal muscle cell proliferation and differentiation are tightly coordinated by a continuum of molecular signaling pathways. The striated muscle activator of Rho signaling (STARS) is an actin binding protein that regulates the transcription of genes involved in muscle cell growth, structure and function via the stimulation of actin polymerization and activation of serum-response factor (SRF) signaling. STARS mediates cell proliferation in smooth and cardiac muscle models; however, whether STARS overexpression enhances cell proliferation and differentiation has not been investigated in skeletal muscle cells. Results: We demonstrate for the first time that STARS overexpression enhances differentiation but not proliferation in C2C12 mouse skeletal muscle cells. Increased differentiation was associated with an increase in the gene levels of the myogenic differentiation markers Ckm, Ckmt2 and Myh4, the differentiation factor Igf2 and the myogenic regulatory factors (MRFs) Myf5 and Myf6. Exposing C2C12 cells to CCG-1423, a pharmacological inhibitor of SRF preventing the nuclear translocation of its co-factor MRTF-A, had no effect on myotube differentiation rate, suggesting that STARS regulates differentiation via a MRTF-A independent mechanism. Conclusion: These findings position STARS as an important regulator of skeletal muscle growth and regeneration. PMID:26903873

  10. Calcium regulation in aortic smooth muscle cells during the initial phase of tunicamycin-induced endo/sarcoplasmic reticulum stress.

    PubMed

    Ziomek, Gabriela; Cheraghi Zanjani, Parisa; Arman, Darian; van Breemen, Cornelis; Esfandiarei, Mitra

    2014-07-15

    Endo/sarcoplasmic reticulum stress and the unfolded protein response have been implicated as underlying mechanisms of cell death in many pathological conditions. We have confirmed that long-term exposure to 10µM tunicamycin induced the endo/sarcoplasmic reticulum stress in cultured vascular smooth muscle cells. Since tunicamycin is reported to induce the stress response by inhibiting protein glycosylation, we attempted to investigate a causal link between accumulation of unfolded proteins and dysregulation of cellular calcium transport. However, we found that tunicamycin caused an immediate release of calcium from the endo/sarcoplasmic reticulum, which was sensitive to thapsigargin, and an influx of calcium through the plasma membrane, resulting in a significant increase in cytoplasmic calcium and depletion of endo/sarcoplasmic reticulum calcium. Furthermore, we observed that tunicamycin also induced contraction in intact vascular smooth muscle. By applying established procedures and antagonists, we established that tunicamycin did not directly activate physiological calcium channels, such as store-operated channels, voltage gated calcium channels, ryanodine receptors or inositol trisphosphate receptors. Instead, we found that its effects on cellular calcium fluxes closely resembled those of the known calcium ionophore, ionomycin. We have concluded that tunicamycin directly permeabilizes the plasma membrane and endo/sarcoplasmic reticulum to calcium, and is, therefore, inappropriate for studying the relationship between accumulation of unfolded proteins and endo/sarcoplasmic reticulum calcium dysregulation during the endo/sarcoplasmic reticulum stress response. In contrast, we also report that two other well-known endo/sarcoplasmic reticulum stress inducers, brefeldin A and dithiothreitol, did not exhibit similar increases in calcium permeability.

  11. Macrophage density in pharyngeal and laryngeal muscles greatly exceeds that in other striated muscles: an immunohistochemical study using elderly human cadavers

    PubMed Central

    Rhee, Sunki; Kitamura, Kei; Masaaki, Kasahara; Katori, Yukio; Murakami, Gen; Abe, Shin-ichi

    2016-01-01

    Macrophages play an important role in aging-related muscle atrophy (i.e., sarcopenia). We examined macrophage density in six striated muscles (cricopharyngeus muscle, posterior cricoarytenoideus muscle, genioglossus muscle, masseter muscle, infraspinatus muscle, and external anal sphincter). We examined 14 donated male cadavers and utilized CD68 immunohistochemistry to clarify macrophage density in muscles. The numbers of macrophages per striated muscle fiber in the larynx and pharynx (0.34 and 0.31) were 5–6 times greater than those in the tongue, shoulder, and anus (0.05–0.07) with high statistical significance. Thick muscle fibers over 80 µm in diameter were seen in the pharynx, larynx, and anal sphincter of two limited specimens. Conversely, in the other sites or specimens, muscle fibers were thinner than 50 µm. We did not find any multinuclear muscle cells suggestive of regeneration. At the beginning of the study, we suspected that mucosal macrophages might have invaded into the muscle layer of the larynx and pharynx, but we found no evidence of inflammation in the mucosa. Likewise, the internal anal sphincter (a smooth muscle layer near the mucosa) usually contained fewer macrophages than the external sphincter. The present result suggest that, in elderly men, thinning and death of striated muscle fibers occur more frequently in the larynx and pharynx than in other parts of the body. PMID:27722010

  12. Revealing T-Tubules in Striated Muscle with New Optical Super-Resolution Microscopy Techniquess

    PubMed Central

    Clowsley, Alexander H.; Munro, Michelle; Hou, Yufeng; Crossman, David J.

    2014-01-01

    The t-tubular system plays a central role in the synchronisation of calcium signalling and excitation-contraction coupling in most striated muscle cells. Light microscopy has been used for imaging t-tubules for well over 100 years and together with electron microscopy (EM), has revealed the three-dimensional complexities of the t-system topology within cardiomyocytes and skeletal muscle fibres from a range of species. The emerging super-resolution single molecule localisation microscopy (SMLM) techniques are offering a near 10-fold improvement over the resolution of conventional fluorescence light microscopy methods, with the ability to spectrally resolve nanometre scale distributions of multiple molecular targets. In conjunction with the next generation of electron microscopy, SMLM has allowed the visualisation and quantification of intricate t-tubule morphologies within large areas of muscle cells at an unprecedented level of detail. In this paper, we review recent advancements in the t-tubule structural biology with the utility of various microscopy techniques. We outline the technical considerations in adapting SMLM to study t-tubules and its potential to further our understanding of the molecular processes that underlie the sub-micron scale structural alterations observed in a range of muscle pathologies. PMID:26913143

  13. Detection of a troponin I-like protein in non-striated muscle of the tardigrades (water bears).

    PubMed

    Obinata, Takashi; Ono, Kanako; Ono, Shoichiro

    2011-03-01

    Tardigrades, also known as water bears, have somatic muscle fibers that are responsible for movement of their body and legs. These muscle fibers contain thin and thick filaments in a non-striated pattern. However, the regulatory mechanism of muscle contraction in tardigrades is unknown. In the absence of extensive molecular and genomic information, we detected a protein of 31 kDa in whole lysates of tardigrades that cross-reacted with the antibody raised against nematode troponin I (TnI). TnI is a component of the troponin complex that regulates actin-myosin interaction in a Ca(2+)-dependent and actin-linked manner. This TnI-like protein was co-extracted with actin in a buffer containing ATP and EGTA, which is known to induce relaxation of a troponin-regulated contractile system. The TnI-like protein was specifically expressed in the somatic muscle fibers in adult animals and partially co-localized with actin filaments in a non-striated manner. Interestingly, the pharyngeal muscle did not express this protein. These observations suggest that the non-striated somatic muscle of tardigrades has an actin-linked and troponin-regulated system for muscle contraction.

  14. Time Course of the Response of Myofibrillar and Sarcoplasmic Protein Metabolism to Unweighting of the Soleus Muscle

    NASA Technical Reports Server (NTRS)

    Munoz, Kathryn A.; Satarug, Soisungwan; Tischler, Marc E.

    1993-01-01

    Contributions of altered in vivo protein synthesis and degradation to unweighting atrophy of the soleus muscle in tail-suspended young female rats were analyzed daily for up to 6 days. Specific changes in myofibrillar and sarcoplasmic proteins were also evaluated to assess their contributions to the loss of total protein. Synthesis of myofibrillar and sarcoplasmic proteins was estimated by intramuscular (IM) injection and total protein by intraperitoneal (IP) injection of flooding doses of H-3-phenylaianine. Total protein loss was greatest during the first 3 days following suspension and was a consequence of the loss of myofibrillar rather than sarcoplasmic proteins. However, synthesis of total myofibrillar and sarcoplasmic proteins diminished in parallel beginning in the first 24 hours. Therefore sarcoplasmic proteins must be spared due to a decrease in their degradation. In contrast, myofibrillar protein degradation increased, thus explaining the elevated degradation of the total pool. Following 72 hours of suspension, protein synthesis remained low, but the rate of myofibrillar protein loss diminished, suggesting a slowing of degradation. These various results show acute loss of protein during unweighting atrophy is a consequence of decreased synthesis and increased degradation of myofibrillar proteins, and sarcoplasmic proteins are spared due to slower degradation, likely explaining the sparing of plasma membrane receptors. Based on other published data, we propose that the slowing of atrophy after the initial response may be attributed to an increased effect of insulin.

  15. Chromosome mapping of five human cardiac and skeletal muscle sarcoplasmic reticulum protein genes

    SciTech Connect

    Otsu, K.; Fujii, J.; MacLennan, D.H. ); Periasamy, M. ); Difilippantonio, M.; Uppender, M.; Ward, D.C. )

    1993-08-01

    Fluorescence in situ hybridization (FISH) experiments were performed using genomic and complementary DNA probes in order to determine the location on human chromosomes for five genes expressed in cardiac and skeletal muscle sarcoplasmic reticulum. The chromosome location of each gene was determined in terms of both cytogenetic bands and fractional chromosome length. The ATP2A2 gene, expressing the SERCA2 isoform of the Ca[sup 2+] pump, maps to bands 12q23-q24.1, the phospholamban gene (PLN) to 6q22.1, the human skeletal muscle calsequestrin gene (CASQ1) to band 1q21, the cardiac calsequestrin gene (CASQ2) to bands 1p11-p13.3, and the cardiac calcium release channel gene (RYR2) to the interval between band 1q42.1 (distal) and band 1q43 (proximal). 13 refs., 1 fig.

  16. Identification of functionally segregated sarcoplasmic reticulum calcium stores in pulmonary arterial smooth muscle.

    PubMed

    Clark, Jill H; Kinnear, Nicholas P; Kalujnaia, Svetlana; Cramb, Gordon; Fleischer, Sidney; Jeyakumar, Loice H; Wuytack, Frank; Evans, A Mark

    2010-04-30

    In pulmonary arterial smooth muscle, Ca(2+) release from the sarcoplasmic reticulum (SR) via ryanodine receptors (RyRs) may induce constriction and dilation in a manner that is not mutually exclusive. We show here that the targeting of different sarcoplasmic/endoplasmic reticulum Ca(2+)-ATPases (SERCA) and RyR subtypes to discrete SR regions explains this paradox. Western blots identified protein bands for SERCA2a and SERCA2b, whereas immunofluorescence labeling of isolated pulmonary arterial smooth muscle cells revealed striking differences in the spatial distribution of SERCA2a and SERCA2b and RyR1, RyR2, and RyR3, respectively. Almost all SERCA2a and RyR3 labeling was restricted to a region within 1.5 microm of the nucleus. In marked contrast, SERCA2b labeling was primarily found within 1.5 microm of the plasma membrane, where labeling for RyR1 was maximal. The majority of labeling for RyR2 lay in between these two regions of the cell. Application of the vasoconstrictor endothelin-1 induced global Ca(2+) waves in pulmonary arterial smooth muscle cells, which were markedly attenuated upon depletion of SR Ca(2+) stores by preincubation of cells with the SERCA inhibitor thapsigargin but remained unaffected after preincubation of cells with a second SERCA antagonist, cyclopiazonic acid. We conclude that functionally segregated SR Ca(2+) stores exist within pulmonary arterial smooth muscle cells. One sits proximal to the plasma membrane, receives Ca(2+) via SERCA2b, and likely releases Ca(2+) via RyR1 to mediate vasodilation. The other is located centrally, receives Ca(2+) via SERCA2a, and likely releases Ca(2+) via RyR3 and RyR2 to initiate vasoconstriction.

  17. Increased cytosolic androgen receptor binding in rat striated muscle following denervation and disuse

    NASA Technical Reports Server (NTRS)

    Bernard, P. A.; Fishman, P. S.; Max, S. R.; Rance, N. E.

    1984-01-01

    The effects of denervation and disuse on cytosolic androgen receptor binding by rat striated muscle are investigated. Denervation of the extensor digitorum longus and tibialis anterior muscles caused by a 40-50-percent increase in cytosolic androgen receptor concentration with no change in apparent binding affinity. This effect was evident at 6 h postdenervation, maximal at 24 h, and declined to 120 percent of the control level 72 h after denervation. A 40-percent increase in cytosolic androgen receptor concentration was also noted 24 hr after denervation of the hormone-sensitive levator ani muscle. The effect of denervation on androgen receptors was blocked by in vivo injection of cycloheximide; therefore, de novo receptor synthesis probably is not involved in the observed increase. Disuse, produced by subperineurial injection of tetrodotoxin into the tibial and common peroneal branches of the sciatic nerve, mimicked the effect of denervation on androgen receptor binding, suggesting that neuromuscular activity is important in regulation of receptor concentration. Possible mechanisms subserving this effect are discussed.

  18. Characterization of beta-connectin (titin 2) from striated muscle by dynamic light scattering.

    PubMed Central

    Higuchi, H; Nakauchi, Y; Maruyama, K; Fujime, S

    1993-01-01

    Connectin (titin) is a large filamentous protein (single peptide) with a molecular mass of approximately 3 MDa, contour length approximately 900 nm, and diameter approximately 4 nm, and resides in striated muscle. Connectin links the thick filaments to the Z-lines in a sarcomere and produces a passive elastic force when muscle fiber is stretched. The aim of this study is to elucidate some aspects of physical properties of isolated beta-connectin (titin 2), a proteolytic fragment of connectin, by means of dynamic light-scattering (DLS) spectroscopy. The analysis of DLS spectra for beta-connectin gave the translational diffusion coefficient of 3.60 x 10(-8) cm2/s at 10 degrees C (or the hydrodynamic radius of 44.1 nm), molecular mass little smaller than 3.0 MDa (for a literature value of sedimentation coefficient), the root-mean-square end-to-end distance of 163 nm (or the radius of gyration of 66.6 nm), and the Kuhn segment number of 30 and segment length of 30 nm (or the persistence length of 15 nm). These results permitted to estimate the flexural rigidity of 6.0 x 10(-20) dyn x cm2 for filament bending, and the elastic constant of 7 dyn/cm for extension of one persistence length. Based on a simple model, implications of the present results in muscle physiology are discussed. Images FIGURE 1 PMID:8298020

  19. Involvement of catecholaminergic neurons in motor innervation of striated muscle in the mouse esophagus.

    PubMed

    van der Keylen, Piet; Garreis, Fabian; Steigleder, Ruth; Sommer, Daniel; Neuhuber, Winfried L; Wörl, Jürgen

    2016-05-01

    Enteric co-innervation is a peculiar innervation pattern of striated esophageal musculature. Both anatomical and functional data on enteric co-innervation related to various transmitters have been collected in different species, although its function remains enigmatic. However, it is unclear whether catecholaminergic components are involved in such a co-innervation. Thus, we examined to identify catecholaminergic neuronal elements and clarify their relationship to other innervation components in the esophagus, using immunohistochemistry with antibodies against tyrosine hydroxylase (TH), vesicular acetylcholine transporter (VAChT), choline acetyltransferase (ChAT) and protein gene product 9.5 (PGP 9.5), α-bungarotoxin (α-BT) and PCR with primers for amplification of cDNA encoding TH and dopamine-β-hydroxylase (DBH). TH-positive nerve fibers were abundant throughout the myenteric plexus and localized on about 14% of α-BT-labelled motor endplates differing from VAChT-positive vagal nerve terminals. TH-positive perikarya represented a subpopulation of only about 2.8% of all PGP 9.5-positive myenteric neurons. Analysis of mRNA showed both TH and DBH transcripts in the mouse esophagus. As ChAT-positive neurons in the compact formation of the nucleus ambiguus were negative for TH, the TH-positive nerve varicosities on motor endplates are presumably of enteric origin, although a sympathetic origin cannot be excluded. In the medulla oblongata, the cholinergic ambiguus neurons were densely supplied with TH-positive varicosities. Thus, catecholamines may modulate vagal motor innervation of esophageal-striated muscles not only at the peripheral level via enteric co-innervation but also at the central level via projections to the nucleus ambiguus. As Parkinson's disease, with a loss of central dopaminergic neurons, also affects the enteric nervous system and dysphagia is prevalent in patients with this disease, investigation of intrinsic catecholamines in the esophagus may

  20. Large-scale Models Reveal the Two-component Mechanics of Striated Muscle

    PubMed Central

    Jarosch, Robert

    2008-01-01

    This paper provides a comprehensive explanation of striated muscle mechanics and contraction on the basis of filament rotations. Helical proteins, particularly the coiled-coils of tropomyosin, myosin and α-actinin, shorten their H-bonds cooperatively and produce torque and filament rotations when the Coulombic net-charge repulsion of their highly charged side-chains is diminished by interaction with ions. The classical “two-component model” of active muscle differentiated a “contractile component” which stretches the “series elastic component” during force production. The contractile components are the helically shaped thin filaments of muscle that shorten the sarcomeres by clockwise drilling into the myosin cross-bridges with torque decrease (= force-deficit). Muscle stretch means drawing out the thin filament helices off the cross-bridges under passive counterclockwise rotation with torque increase (= stretch activation). Since each thin filament is anchored by four elastic α-actinin Z-filaments (provided with force-regulating sites for Ca2+ binding), the thin filament rotations change the torsional twist of the four Z-filaments as the “series elastic components”. Large scale models simulate the changes of structure and force in the Z-band by the different Z-filament twisting stages A, B, C, D, E, F and G. Stage D corresponds to the isometric state. The basic phenomena of muscle physiology, i. e. latency relaxation, Fenn-effect, the force-velocity relation, the length-tension relation, unexplained energy, shortening heat, the Huxley-Simmons phases, etc. are explained and interpreted with the help of the model experiments. PMID:19330099

  1. Inhibition of the Ca sup 2+ -ATPase of vascular smooth muscle sarcoplasmic reticulum by superoxide radicals

    SciTech Connect

    Suzuki, Yuichiro; Ford, G.D. )

    1991-03-15

    The effect of oxygen free radicals generated by hypoxanthine plus xanthine oxidase on the Ca{sup 2+}-ATPase of sarcoplasmic reticulum from bovine aortic smooth muscle were studied. Exogenous hypoxanthine plus xanthine oxidase produced an hypoxanthine concentration dependent inhibition of the Ca{sup 2+}-ATPase. The inhibition could be completely blocked by superoxide dismutase but not by either mannitol or deferoxamine. Direct addition of reagent hydrogen peroxide in the {mu}M range did not cause significant inhibition. These results suggest that superoxide is the primary damaging species. Additionally, 1.16 {plus minus} 0.17 mU/g wet wt of xanthine oxidase activity were detected in the post-nuclear supernatant of bovine aortic smooth muscle, suggesting the existence of a possible intracellular source of superoxide. This value was calculated to be approximately 5 mU/ml by using a usual value of vascular smooth muscle cellular volume. Thus the level of endogenous xanthine oxidase resident in vascular smooth muscle is comparable with the level of exogenous xanthine oxidase used in the present study. These findings suggest a potential role of xanthine oxidase-generated superoxide in free radical injury to vascular smooth muscle.

  2. Drop in endo/sarcoplasmic calcium precedes the unfolded protein response in Brefeldin A-treated vascular smooth muscle cells.

    PubMed

    Ziomek, Gabriela; van Breemen, Cornelis; Esfandiarei, Mitra

    2015-10-01

    The present study addresses the causal relationship between induction of endo/sarcoplasmic reticulum stress and dysregulation of calcium transport, while examining whether the most widely-used experimental endo/sarcoplasmic reticulum stressors can be considered appropriate for elucidating underlying cellular mechanisms involved during the progression of the unfolded protein response in vascular smooth muscle cells. Brefeldin A is most commonly cited as inducing the stress response through an accumulation of unfolded proteins in the lumen as a result of a blockage of protein transport from the endo/sarcoplasmic reticulum to the Golgi apparatus. We investigated the effects of Brefeldin A on cellular calcium regulation during the the unfolded protein response in cultured rat vascular smooth muscle cells. Acute exposure of cells to Brefeldin A caused a small transient increase in cytoplasmic calcium, which did not cause a significant decrease in endo/sarcoplasmic reticulum calcium content. However, over the time course of 0-12 h post-treatment with Brefeldin A, we observed that the endo/sarcoplasmic reticulum of vascular smooth muscle cells exhibited an approximate fifty percent decrease in calcium concentration after the first hour of exposure, which is maintained over the next eleven hours, whereas concentrations of unfolded protein response markers only began to increase significantly around nine to twelve hours post-treatment. We have concluded that the endo/sarcoplasmic reticulum calcium drop, which up to now, has been considered as a characteristic of the late onset of cellular stress response, occurs prior to the initiation of the unfolded protein response, rather than as a result of its many corrective pathways.

  3. Dietary Fish Oil Blocks the Microcirculatory Manifestations of Ischemia- Reperfusion Injury in Striated Muscle in Hamsters

    NASA Astrophysics Data System (ADS)

    Lehr, Hans-Anton; Hubner, Christoph; Nolte, Dirk; Kohlschutter, Alfried; Messmer, Konrad

    1991-08-01

    Epidemiologic observations and experimental studies have demonstrated a protective effect of dietary fish oil on the clinical manifestations of ischemia-reperfusion injury. To investigate the underlying mechanisms, we used the dorsal skinfold chamber model for intravital fluorescence microscopy of the microcirculation in striated muscle of awake hamsters. In control hamsters (n = 7), reperfusion after a 4-hr pressure-induced ischemia to the muscle tissue elicited the adhesion of fluorescently stained leukocytes to the endothelium of postcapillary venules, capillary obstruction, and the breakdown of endothelial integrity. These microvascular manifestations of ischemia-reperfusion injury were significantly attenuated in animals (n = 7) when fed with a fish oil-enriched diet for 4 weeks prior to the experiments. In leukocyte total lipids, the fish oil diet resulted in a substantial displacement of arachidonic acid, the precursor of the potent adhesionpromoting leukotriene (LT) B_4, by fish oil-derived eicosapentaenoic acid, the precursor of biologically less potent LTB_5, emphasizing the mediator role of LTB_4 in ischemia-reperfusion injury. These results suggest that the preservation of microvascular perfusion by dietary fish oil contributes to its protective effects on the clinical manifestations of ischemia-reperfusion injury.

  4. Isoforms of gelsolin from lobster striated muscles differ in calcium-dependence.

    PubMed

    Unger, Andreas; Brunne, Bianka; Hinssen, Horst

    2013-08-01

    Two distinct isoforms of the Ca-dependent actin filament severing protein gelsolin were identified in cross-striated muscles of the American lobster. The variants (termed LG1 and LG2) differ by an extension of 18 AA at the C-terminus of LG1, and by two substitutions at AA735 and AA736, the two C-terminal amino acids of LG2. Functional comparison of the isolated and purified proteins revealed gelsolin-typical properties for both with differences in Ca(2+)-sensitivity, LG2 being activated at significant lower Ca-concentration than LG1: Half maximal activation for both filament severing and G-actin binding was ∼4×10(-7)M Ca(2+) for LG2 vs. ∼2×10(-6)M Ca(2+) for LG1. This indicates a differential activation for the two isoproteins in vivo where they are present in almost equal amounts in the muscle cell. Structure prediction modeling on the basis of the known structure of mammalian gelsolin shows that LG2 lacks the C-terminal alpha-helix which is involved in contact formation between domains G6 and G2. In both mammalian gelsolin and LG1, this "latch bridge" is assumed to play a critical role in Ca(2+)-activation by keeping gelsolin in a closed, inactive conformation at low [Ca(2+)]. In LG2, the reduced contact between G6 and G2 may be responsible for its activation at low Ca(2+)-concentration.

  5. The Intriguing Dual Lattices of the Myosin Filaments in Vertebrate Striated Muscles: Evolution and Advantage

    PubMed Central

    Luther, Pradeep K.; Squire, John M.

    2014-01-01

    Myosin filaments in vertebrate striated muscle have a long roughly cylindrical backbone with cross-bridge projections on the surfaces of both halves except for a short central bare zone. In the middle of this central region the filaments are cross-linked by the M-band which holds them in a well-defined hexagonal lattice in the muscle A-band. During muscular contraction the M-band-defined rotation of the myosin filaments around their long axes influences the interactions that the cross-bridges can make with the neighbouring actin filaments. We can visualise this filament rotation by electron microscopy of thin cross-sections in the bare-region immediately adjacent to the M-band where the filament profiles are distinctly triangular. In the muscles of teleost fishes, the thick filament triangular profiles have a single orientation giving what we call the simple lattice. In other vertebrates, for example all the tetrapods, the thick filaments have one of two orientations where the triangles point in opposite directions (they are rotated by 60° or 180°) according to set rules. Such a distribution cannot be developed in an ordered fashion across a large 2D lattice, but there are small domains of superlattice such that the next-nearest neighbouring thick filaments often have the same orientation. We believe that this difference in the lattice forms can lead to different contractile behaviours. Here we provide a historical review, and when appropriate cite recent work related to the emergence of the simple and superlattice forms by examining the muscles of several species ranging back to primitive vertebrates and we discuss the functional differences that the two lattice forms may have. PMID:25478994

  6. The intriguing dual lattices of the Myosin filaments in vertebrate striated muscles: evolution and advantage.

    PubMed

    Luther, Pradeep K; Squire, John M

    2014-01-01

    Myosin filaments in vertebrate striated muscle have a long roughly cylindrical backbone with cross-bridge projections on the surfaces of both halves except for a short central bare zone. In the middle of this central region the filaments are cross-linked by the M-band which holds them in a well-defined hexagonal lattice in the muscle A-band. During muscular contraction the M-band-defined rotation of the myosin filaments around their long axes influences the interactions that the cross-bridges can make with the neighbouring actin filaments. We can visualise this filament rotation by electron microscopy of thin cross-sections in the bare-region immediately adjacent to the M-band where the filament profiles are distinctly triangular. In the muscles of teleost fishes, the thick filament triangular profiles have a single orientation giving what we call the simple lattice. In other vertebrates, for example all the tetrapods, the thick filaments have one of two orientations where the triangles point in opposite directions (they are rotated by 60° or 180°) according to set rules. Such a distribution cannot be developed in an ordered fashion across a large 2D lattice, but there are small domains of superlattice such that the next-nearest neighbouring thick filaments often have the same orientation. We believe that this difference in the lattice forms can lead to different contractile behaviours. Here we provide a historical review, and when appropriate cite recent work related to the emergence of the simple and superlattice forms by examining the muscles of several species ranging back to primitive vertebrates and we discuss the functional differences that the two lattice forms may have. PMID:25478994

  7. Digital image analysis of striated skeletal muscle tissue injury during reperfusion after induced ischemia

    NASA Astrophysics Data System (ADS)

    Rosero Salazar, Doris Haydee; Salazar Monsalve, Liliana

    2015-01-01

    Conditions such as surgical procedures or vascular diseases produce arterial ischemia and reperfusion injuries, which generate changes in peripheral tissues and organs, for instance, in striated skeletal muscle. To determine such changes, we conducted an experimental method in which 42 male Wistar rat were selected, to be undergone to tourniquet application on the right forelimb and left hind limb, to induce ischemia during one and three hours, followed by reperfusion periods starting at one hour and it was prolonged up to 32 days. Extensor carpi radialis longus and soleus respectively, were obtained to be processed for histochemical and morphometric analysis. By means of image processing and detection of regions of interest, variations of areas occupied by muscle fibers and intramuscular extracellular matrix (IM-ECM) throughout reperfusion were observed. In extensor carpi radialis longus, results shown reduction in the area occupied by muscle fibers; this change is significant between one hour and three hours ischemia followed by 16 hours, 48 hours and 32 days reperfusión (p˂0.005). To compare only periods of reperfusión that continued to three hours ischemia, were found significant differences, as well. For area occupied by IM-ECM, were identified increments in extensor carpi radialis longus by three hours ischemia and eight to 16 days reperfusion; in soleus, was observed difference by one hour ischemia with 42 hours reperfusion, and three hours ischemia followed by four days reperfusion (p˂0.005). Skeletal muscle develops adaptive changes in longer reperfusion, to deal with induced injury. Descriptions beyond 32 days reperfusion, can determine recovering normal pattern.

  8. Characteristics of sarcoplasmic reticulum from normal and denervated rat skeletal muscle.

    PubMed Central

    Palexas, G N; Savage, N; Isaacs, H

    1981-01-01

    Denervation of rat skeletal muscle produces after 14 days a decrease in Ca2+ uptake of a heterogeneous population of sarcoplasmic-reticulum vesicles, when measured in the presence of oxalate. The Mg2+-dependent ATPase (Ca2+-independent) activity increased after the same period and the Ca2+ + Mg2+-dependent ATPase activity decreased. Concomitant with these changes, there was an increase in vesicle size and calcium content. The observations are discussed in terms of changes in altered membrane structure, manifested in the shift of the equilibrium of the ATPase from an enzyme involved in calcium transport to a phosphoenzyme giving rise to an increase in the Mg2+-dependent ATPase activity. PMID:6120692

  9. Sarcoplasmic reticulum and excitation-contraction coupling in mammalian smooth muscles.

    PubMed

    Devine, C E; Somlyo, A V; Somlyo, A P

    1972-03-01

    The sarcoplasmic reticulum (SR) was studied in the smooth muscles of rabbit main pulmonary artery, mesenteric vein, aorta, mesenteric artery, taenia coli, guinea pig mesenteric artery, and human uterus, and correlated with contractions of the smooth muscles in Ca-free media. SR volumes were determined in main pulmonary artery (5.1%), aorta (5%), portal-anterior mesenteric vein (2.2%), taenia coli (2%), and mesenteric artery (1.8%): because of tangentially sectioned membranes these estimates are subject to a correction factor of up to +50% of the values measured. Smooth muscles that contained a relatively large volume of SR maintained significant contractile responses to drugs in the virtual absence of extracellular calcium at room temperatures, while smooth muscles that had less SR did not. The unequal maximal contractions of main pulmonary artery elicited by different drugs were also observed in Ca-free, high potassium-depolarizing solution, indicating that they were secondary to some mechanism independent of changes in membrane potential or calcium influx. Longitudinal tubules of SR run between and are fenestrated about groups of surface vesicles separated from each other by intervening dense bodies. Extracellular markers (ferritin and lanthanum) entered the surface vesicles, but not the SR. The peripheral SR formed couplings with the surface membrane: the two membranes were separated by gaps of approximately 10 nm traversed by electron-opaque connections suggestive of a periodicity of approximately 20-25 nm. These couplings are considered to be the probable sites of electromechanical coupling in twitch smooth muscles. Close contacts between the SR and the surface vesicles may have a similar function, or represent sites of calcium extrusion. The presence of both thick and thin myofilaments and of rough SR in smooth muscles supports the dual, contractile and morphogenetic, function of smooth muscle.

  10. Lessons from mammalian hibernators: molecular insights into striated muscle plasticity and remodeling.

    PubMed

    Tessier, Shannon N; Storey, Kenneth B

    2016-05-01

    Striated muscle shows an amazing ability to adapt its structural apparatus based on contractile activity, loading conditions, fuel supply, or environmental factors. Studies with mammalian hibernators have identified a variety of molecular pathways which are strategically regulated and allow animals to endure multiple stresses associated with the hibernating season. Of particular interest is the observation that hibernators show little skeletal muscle atrophy despite the profound metabolic rate depression and mechanical unloading that they experience during long weeks of torpor. Additionally, the cardiac muscle of hibernators must adjust to low temperature and reduced perfusion, while the strength of contraction increases in order to pump cold, viscous blood. Consequently, hibernators hold a wealth of knowledge as it pertains to understanding the natural capacity of myocytes to alter structural, contractile and metabolic properties in response to environmental stimuli. The present review outlines the molecular and biochemical mechanisms which play a role in muscular atrophy, hypertrophy, and remodeling. In this capacity, four main networks are highlighted: (1) antioxidant defenses, (2) the regulation of structural, contractile and metabolic proteins, (3) ubiquitin proteosomal machinery, and (4) macroautophagy pathways. Subsequently, we discuss the role of transcription factors nuclear factor (erythroid-derived 2)-like 2 (Nrf2), Myocyte enhancer factor 2 (MEF2), and Forkhead box (FOXO) and their associated posttranslational modifications as it pertains to regulating each of these networks. Finally, we propose that comparing and contrasting these concepts to data collected from model organisms able to withstand dramatic changes in muscular function without injury will allow researchers to delineate physiological versus pathological responses. PMID:26982616

  11. Differential expression of beta 1 integrins in nonneoplastic smooth and striated muscle cells and in tumors derived from these cells.

    PubMed Central

    Mechtersheimer, G.; Barth, T.; Quentmeier, A.; Möller, P.

    1994-01-01

    Integrins are a superfamily of transmembrane alpha beta heterodimers that play an important role in cell-matrix and cell-cell interactions by acting as receptors for extracellular matrix proteins and for cell adhesion molecules. Using monoclonal antibodies against beta 1, alpha 1 to alpha 6, and alpha v subunits, the in situ distribution pattern of beta 1 integrins was examined immunohistochemically in nonneoplastic smooth and striated muscle cells and in their tumors. Nonneoplastic smooth muscle cells were beta 1+, alpha 1+, alpha 3+, alpha v+ and, in diverse localizations, also alpha 5+ or even alpha 6+. The expression of the beta 1 chain was conserved in all leiomyomas and leiomyosarcomas. The distribution pattern of the alpha subunits by contrast underwent several changes during malignant transformation of smooth muscle cells. These alterations consisted in a neoexpression of alpha 2, alpha 4, and alpha 6 as well as in an abnormal abrogation of alpha 1 and alpha 3 in some leiomyosarcomas. Except for the absence of alpha 5 in the majority of epithelioid leiomyosarcomas, expression of the alpha 5 and alpha v subunits was mainly conserved. In addition, tumors with epithelioid differentiation differed from typical cases by the absence of alpha 1 and the simultaneous presence of alpha 4. Adult striated muscle cells were beta 1+ but alpha 1- to alpha 6- and alpha v-, whereas fetal striated muscle cells were not only beta 1+ but also alpha 3+/-, alpha 4+/-, alpha 5+ and alpha 6+. In all rhabdomyosarcomas the expression of beta 1 was retained. Furthermore, the majority of cases showed the expression of one or more alpha subunits most of which, ie, alpha 4, alpha 5, and alpha 6, were also found in fetal striated muscle cells. In conclusion, beta 1 integrins exhibited a differential expression pattern along the two lines of myogenic differentiation. This integrin profile underwent characteristic changes during malignant transformation. Nevertheless, the compiled

  12. Testosterone enhances C-14 2-deoxyglucose uptake by striated muscle. [sex hormones and muscle

    NASA Technical Reports Server (NTRS)

    Toop, J.; Max, S. R.

    1982-01-01

    The effect of testosterone propionate (TP) on C-14 2-deoxyglucose (C-14 2DG) uptake was studied in the rat levator ani muscle in vivo using the autoradiographic technique. Following a delay of 1 to 3 h after injecting TP, the rate of C-14 2DG uptake in experimental animals began to increase and continued to increase for at least 20 h. The label, which corresponds to C-14 2-deoxyglucose 6-phosphate, as demonstrated by chromatographic analysis of muscle extracts, was uniformly distributed over the entire muscle and was predominantly in muscle fibers, although nonmuscular elements were also labeled. The 1 to 3 h time lag suggests that the TP effect may be genomic, acting via androgen receptors, rather than directly on muscle membranes. Acceleration of glucose uptake may be an important early event in the anabolic response of the rat levator ani muscle to androgens.

  13. Vanilloid receptor expressed in the sarcoplasmic reticulum of rat skeletal muscle

    SciTech Connect

    Xin Hong; Tanaka, Hideyuki; Yamaguchi, Maki; Takemori, Shigeru; Nakamura, Akio; Kohama, Kazuhiro . E-mail: kohamak@med.gunma-u.ac.jp

    2005-07-08

    Vanilloid receptor subtype 1 (VR1) was cloned as a capsaicin receptor from neuronal cells of dorsal root ganglia. VR1 was subsequently found in a few non-neuronal tissues, including skeletal muscle [Onozawa et al., Tissue distribution of capsaicin receptor in the various organs of rats, Proc. Jpn. Acad. Ser. B 76 (2000) 68-72]. We confirmed the expression of VR1 in muscle cells using the RT-PCR method and Western blot analysis. Immunostaining studies with a confocal microscope and an electron microscope indicated that VR1 was present in the sarcoplasmic reticulum (SR), a store of Ca{sup 2+}. The SR releases Ca{sup 2+} to cause a contraction when a muscle is excited. However, SR still releases a small amount of Ca{sup 2+} under relaxed conditions. We found that this leakage was enhanced by capsaicin and was antagonized by capsazepine, a capsaicin blocker, indicating that leakage of Ca{sup 2+} occurs through a channel composed of VR1.

  14. Static magnetic fields affect capillary flow of red blood cells in striated skin muscle.

    PubMed

    Brix, Gunnar; Strieth, Sebastian; Strelczyk, Donata; Dellian, Marc; Griebel, Jürgen; Eichhorn, Martin E; Andrā, Wilfried; Bellemann, Matthias E

    2008-01-01

    Blood flowing in microvessels is one possible site of action of static magnetic fields (SMFs). We evaluated SMF effects on capillary flow of red blood cells (RBCs) in unanesthetized hamsters, using a skinfold chamber technique for intravital fluorescence microscopy. By this approach, capillary RBC velocities (v(RBC)), capillary diameters (D), arteriolar diameters (D(art)), and functional vessel densities (FVD) were measured in striated skin muscle at different magnetic flux densities. Exposure above a threshold level of about 500 mT resulted in a significant (P < 0.001) reduction of v(RBC) in capillaries as compared to the baseline value. At the maximum field strength of 587 mT, v(RBC) was reduced by more than 40%. Flow reduction was reversible when the field strength was decreased below the threshold level. In contrast, mean values determined at different exposure levels for the parameters D, D(art), and FVD did not vary by more than 5%. Blood flow through capillary networks is affected by strong SMFs directed perpendicular to the vessels. Since the influence of SMFs on blood flow in microvessels directed parallel to the field as well as on collateral blood supply could not be studied, our findings should be carefully interpreted with respect to the setting of safety guidelines.

  15. Role of glycogen availability in sarcoplasmic reticulum Ca2+ kinetics in human skeletal muscle

    PubMed Central

    Ørtenblad, Niels; Nielsen, Joachim; Saltin, Bengt; Holmberg, Hans-Christer

    2011-01-01

    Little is known about the precise mechanism that relates skeletal muscle glycogen to muscle fatigue. The aim of the present study was to examine the effect of glycogen on sarcoplasmic reticulum (SR) function in the arm and leg muscles of elite cross-country skiers (n= 10, 72 ± 2 ml kg−1 min−1) before, immediately after, and 4 h and 22 h after a fatiguing 1 h ski race. During the first 4 h recovery, skiers received either water or carbohydrate (CHO) and thereafter all received CHO-enriched food. Immediately after the race, arm glycogen was reduced to 31 ± 4% and SR Ca2+ release rate decreased to 85 ± 2% of initial levels. Glycogen noticeably recovered after 4 h recovery with CHO (59 ± 5% initial) and the SR Ca2+ release rate returned to pre-exercise levels. However, in the absence of CHO during the first 4 h recovery, glycogen and the SR Ca2+ release rate remained unchanged (29 ± 2% and 77 ± 8%, respectively), with both parameters becoming normal after the remaining 18 h recovery with CHO. Leg muscle glycogen decreased to a lesser extent (71 ± 10% initial), with no effects on the SR Ca2+ release rate. Interestingly, transmission electron microscopy (TEM) analysis revealed that the specific pool of intramyofibrillar glycogen, representing 10–15% of total glycogen, was highly significantly correlated with the SR Ca2+ release rate. These observations strongly indicate that low glycogen and especially intramyofibrillar glycogen, as suggested by TEM, modulate the SR Ca2+ release rate in highly trained subjects. Thus, low glycogen during exercise may contribute to fatigue by causing a decreased SR Ca2+ release rate. PMID:21135051

  16. Kinetic characterization of Channa striatus muscle sarcoplasmic and myofibrillar protein hydrolysates.

    PubMed

    Ghassem, Masomeh; Fern, See Siau; Said, Mamot; Ali, Zainon Mohd; Ibrahim, Saadiah; Babji, Abdul Salam

    2014-03-01

    This study was conducted to evaluate the kinetic characteristics of proteolytic activity of proteases on Channa striatus protein fractions. Degree of hydrolysis (DH), amino acid composition and kinetic parameters of sarcoplasmic and myofibrillar proteins were investigated when incubated with proteinase K and thermolysin, separately. After 30 min incubation with proteases, a decrease in DH of sarcoplasmic protein was observed whereas, hydrolysis of myofibrillar protein with proteases took 2 h with an increase in DH. The major amino acids were glutamic acid (16.6%) in thermolysin- myofibrillar hydrolysate followed by aspartic acid (11.1%) in sarcoplasmic protein fraction with no enzyme treatment and lysine (10%) in thermolysin-myofibrillar hydrolysate. The apparent Michaelis constant of proteinase K was lower than thermolysin for both sarcoplasmic and myofibrillar proteins. However, rate of turnover and enzyme efficiency suggested that sarcoplasmic and myofibrillar proteins are suitable substrates for proteinase K and thermolysin hydrolytic reaction, respectively.

  17. Alteration of Sarcoplasmic Reticulum Ca2+ Release in Skeletal Muscle from Calpain 3-Deficient Mice

    PubMed Central

    Dayanithi, Govindan; Richard, Isabelle; Viero, Cédric; Mazuc, Elsa; Mallie, Sylvie; Valmier, Jean; Bourg, Nathalie; Herasse, Muriel; Marty, Isabelle; Lefranc, Gérard; Mangeat, Paul; Baghdiguian, Stephen

    2009-01-01

    Mutations of Ca2+-activated proteases (calpains) cause muscular dystrophies. Nevertheless, the specific role of calpains in Ca2+ signalling during the onset of dystrophies remains unclear. We investigated Ca2+ handling in skeletal cells from calpain 3-deficient mice. [Ca2+]i responses to caffeine, a ryanodine receptor (RyR) agonist, were decreased in −/− myotubes and absent in −/− myoblasts. The −/− myotubes displayed smaller amplitudes of the Ca2+ transients induced by cyclopiazonic acid in comparison to wild type cells. Inhibition of L-type Ca2+ channels (LCC) suppressed the caffeine-induced [Ca2+]i responses in −/− myotubes. Hence, the absence of calpain 3 modifies the sarcoplasmic reticulum (SR) Ca2+ release, by a decrease of the SR content, an impairment of RyR signalling, and an increase of LCC activity. We propose that calpain 3-dependent proteolysis plays a role in activating support proteins of intracellular Ca2+ signalling at a stage of cellular differentiation which is crucial for skeletal muscle regeneration. PMID:20300593

  18. Porcine malignant hyperthermia susceptibility: hypersensitive calcium-release mechanism of skeletal muscle sarcoplasmic reticulum.

    PubMed Central

    O'Brien, P J

    1986-01-01

    This study tested the hypothesis that calcium-release from sarcoplasmic reticulum isolated from malignant hyperthermia swine had abnormal concentration-dependency on release modulators. Halothane stimulated half-maximal calcium-release at similar concentrations for malignant hyperthermia and control sarcoplasmic reticulum (0.10 +/- 0.04 mM). However, concentrations causing half-maximal calcium-release were lower for malignant hyperthermia sarcoplasmic reticulum (P less than 0.001) by an order of magnitude for Ca2+ (28.1 +/- 8.3 versus 1.23 +/- 0.45 nM), adenosine triphosphate (0.33 +/- 0.09 versus 0.023 +/- 0.014 mM) and caffeine (7.79 +/- 1.56 versus 0.80 +/- 0.44 mM). Half-maximal inhibition by Mg2+ occurred at threefold higher concentrations for malignant hyperthermia sarcoplasmic reticulum (0.23 +/- 0.02 versus 0.78 +/- 0.17 mM). The Ca2+-sensitivity curves for calcium-release by sarcoplasmic reticulum isolated from heterozygotes for the malignant hyperthermia-defect were indistinguishable from the averages of the curves for controls and malignant hyperthermia-homozygotes. Results of this study suggest that malignant hyperthermia is initiated due to a hypersensitive calcium-release mechanism which is inherited in an autosomal, codominant pattern and may be diagnosed using calcium-release sensitivity-tests on isolated sarcoplasmic reticulum. Images Fig. 1. PMID:3742367

  19. The myosin interacting-heads motif is present in the relaxed thick filament of the striated muscle of scorpion.

    PubMed

    Pinto, Antonio; Sánchez, Fredi; Alamo, Lorenzo; Padrón, Raúl

    2012-12-01

    Electron microscopy (EM) studies of 2D crystals of smooth muscle myosin molecules have shown that in the inactive state the two heads of a myosin molecule interact asymmetrically forming a myosin interacting-heads motif. This suggested that inactivation of the two heads occurs by blocking of the actin-binding site of one (free head) and the ATP hydrolysis site of the other (blocked head). This motif has been found by EM of isolated negatively stained myosin molecules of unregulated (vertebrate skeletal and cardiac muscle) and regulated (invertebrate striated and vertebrate smooth muscle) myosins, and nonmuscle myosin. The same motif has also been found in 3D-reconstructions of frozen-hydrated (tarantula, Limulus, scallop) and negatively stained (scallop, vertebrate cardiac) isolated thick filaments. We are carrying out studies of isolated thick filaments from other species to assess how general this myosin interacting-heads motif is. Here, using EM, we have visualized isolated, negatively stained thick filaments from scorpion striated muscle. We modified the iterative helical real space reconstruction (IHRSR) method to include filament tilt, and band-pass filtered the aligned segments before averaging, achieving a 3.3 nm resolution 3D-reconstruction. This reconstruction revealed the presence of the myosin interacting-heads motif (adding to evidence that is widely spread), together with 12 subfilaments in the filament backbone. This demonstrates that conventional negative staining and imaging can be used to detect the presence of the myosin interacting-heads motif in helically ordered thick filaments from different species and muscle types, thus avoiding the use of less accessible cryo-EM and low electron-dose procedures. PMID:22982253

  20. Evolution of the regulatory control of vertebrate striated muscle: the roles of troponin I and myosin binding protein-C.

    PubMed

    Shaffer, Justin F; Gillis, Todd E

    2010-08-01

    Troponin I (TnI) and myosin binding protein-C (MyBP-C) are key regulatory proteins of contractile function in vertebrate muscle. TnI modulates the Ca(2+) activation signal, while MyBP-C regulates cross-bridge cycling kinetics. In vertebrates, each protein is distributed as tissue-specific paralogs in fast skeletal (fs), slow skeletal (ss), and cardiac (c) muscles. The purpose of this study is to characterize how TnI and MyBP-C have changed during the evolution of vertebrate striated muscle and how tissue-specific paralogs have adapted to different physiological conditions. To accomplish this we have completed phylogenetic analyses using the amino acid sequences of all known TnI and MyBP-C isoforms. This includes 99 TnI sequences (fs, ss, and c) from 51 different species and 62 MyBP-C sequences from 26 species, with representatives from each vertebrate group. Results indicate that the role of protein kinase A (PKA) and protein kinase C (PKC) in regulating contractile function has changed during the evolution of vertebrate striated muscle. This is reflected in an increased number of phosphorylatable sites in cTnI and cMyBP-C in endothermic vertebrates and the loss of two PKC sites in fsTnI in a common ancestor of mammals, birds, and reptiles. In addition, we find that His(132), Val(134), and Asn(141) in human ssTnI, previously identified as enabling contractile function during cellular acidosis, are present in all vertebrate cTnI isoforms except those from monotremes, marsupials, and eutherian mammals. This suggests that the replacement of these residues with alternative residues coincides with the evolution of endothermy in the mammalian lineage.

  1. Alterations in the sarcoplasmic protein fraction of beef muscle with postmortem aging and hydrodynamic pressure processing

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Capillary electrophoresis (CE) and reversed-phase high performance liquid chromatography (RP-HPLC) analysis were utilized to detect differences in the sarcoplasmic protein profiles of beef strip loins subjected to aging and hydrodynamic pressure processing (HDP) treatments. At 48 h postmortem, stri...

  2. Calcium release and sarcoplasmic reticulum membrane potential in frog skeletal muscle fibres.

    PubMed Central

    Baylor, S M; Chandler, W K; Marshall, M W

    1984-01-01

    Single twitch fibres were dissected from frog muscle, stretched to a sarcomere spacing greater than or equal to 3.9 micron, then mounted for optical recording. The experiments were carried out at 15-17 degrees C. In some cases D2O Ringer solution was used instead of H2O Ringer solution to reduce movement and any related optical artifacts. Following action potential stimulation, both the amplitude and time course of the change in intrinsic retardation (Baylor & Oetliker's second component, 1975) were found to be approximately independent of wavelength between 480 and 750 nm (D2O Ringer solution). Fibres were injected with the Ca2+-sensitive dye Arsenazo III so that changes in myoplasmic free [Ca2+] could be estimated by measuring changes in dye-related absorbance at 660 nm. The time course of free [Ca2+] was compared with the time course of two other optical signals which have been previously suggested to monitor s.r. (sarcoplasmic reticulum) membrane potential, intrinsic retardation and Nile Blue A fluorescence (Bezanilla & Horowicz, 1975). In D2O Ringer solution the retardation time course was closely similar to that of free [Ca2+] whereas the fluorescence time course was considerably slower. Thus, it is possible that either the retardation signal or Nile Blue A fluorescence (or both) monitors free [Ca2+] rather than s.r. potential. If so, the underlying mechanism which senses Ca2+ must do so very rapidly in the case of retardation and with a delay in the case of Nile Blue A. Changes in Nile Blue A fluorescence were measured in a voltage-clamped fibre (H2O Ringer solution). Only small changes were observed during hyperpolarization or small depolarization whereas relatively large changes were observed near mechanical threshold. These increased e-fold in magnitude every 4-5 mV. This steep voltage dependence, similar to that already shown for intrinsic retardation and [Ca2+], provides additional evidence that Nile Blue A fluorescence monitors a step in excitation

  3. Comparison of the calcium release channel of cardiac and skeletal muscle sarcoplasmic reticulum by target inactivation analysis

    SciTech Connect

    McGrew, S.G.; Inui, Makoto; Chadwick, C.C.; Boucek, R.J. Jr.; Jung, C.Y.; Fleischer, S. )

    1989-02-07

    The calcium release channel of sarcoplasmic reticulum which triggers muscle contraction in excitation-contraction coupling has recently been isolated. The channel has been found to be morphologically identical with the feet structures of the junctional face membrane of terminal cisternae and consists of an oligomer of a unique high molecular weight polypeptide. In this study, the authors compare the target size of the calcium release channel from heart and skeletal muscle using target inactivation analysis. The target molecular weights of the calcium release channel estimated by measuring ryanodine binding after irradiation are similar for heart (139,000) and skeletal muscle (143,000) and are smaller than the monomeric unit (estimated to be about 360,000). The target size, estimated by measuring polypeptide remaining after irradiation, was essentially the same for heart and skeletal muscle, 1,061,000 and 1,070,000, respectively, indicating an oligomeric association of protomers. Thus, the calcium release channel of both cardiac and skeletal muscle reacts uniquely with regard to target inactivation analysis in that (1) the size by ryanodine binding is smaller than the monomeric unit and (2) a single hit leads to destruction of more than one polypeptide, by measuring polypeptide remaining. The target inactivation analysis studies indicate that heart and skeletal muscle receptors are structurally very similar.

  4. Gene expression analyses of essential catch factors in the smooth and striated adductor muscles of larval, juvenile and adult great scallop (Pecten maximus).

    PubMed

    Andersen, Øivind; Torgersen, Jacob S; Pagander, Helene H; Magnesen, Thorolf; Johnston, Ian A

    2009-01-01

    The scallop adductor muscle consists of striated fibres responsible for the fast closure of the shells, and smooth fibres able to maintain tension in a prolonged state of contraction called catch. Formation of the force-bearing catch linkages has been demonstrated to be initiated by dephosphorylation of the key catch-regulating factor twitchin by a calcineurin-like phosphatase, while the involvement of other thick filament proteins is uncertain. Here we report on the development of catchability of the adductor smooth muscle in the great scallop (Pecten maximus) by analysing the spatio-temporal gene expression patterns of the myosin regulatory light chain (MLCr), twitchin, myorod and calcineurin using whole mount in situ hybridization and real-time quantitative PCR. The MLCr signal was identified in the retractor and adductor muscles of the pediveliger larvae, and the juvenile and adult scallop displayed abundant mRNA levels of MLCr in the smooth and striated adductor muscles. Twitchin was mainly expressed in the smooth adductor muscle during metamorphosis, whereas the adult striated adductor muscle contained seven-folds higher twitchin mRNA levels compared to the smooth portion. Calcineurin expression predominated in the gonads and in the smooth adductor, and five-folds higher mRNA levels were measured in the smooth than in the striated fibres at the adult stage. In contrast to the other genes examined, the expression of myorod was confined to the smooth adductor muscle suggesting that myorod plays a permissive role in the molluscan catch muscles, which are first required at the vulnerable settlement stage as a component of the predator defence system.

  5. Correction of Multiple Striated Muscles in Murine Pompe Disease Through Adeno-associated Virus-Mediated Gene Therapy

    PubMed Central

    Sun, Baodong; Young, Sarah P.; Li, Ping; Di, Chunhui; Brown, Talmage; Salva, Maia Z.; Li, Songtao; Bird, Andrew; Yan, Zhen; Auten, Richard; Hauschka, Stephen D.; Koeberl, Dwight D.

    2009-01-01

    Glycogen storage disease type II (GSD-II; Pompe disease; MIM 232300) stems from the deficiency of acid-α-glucosidase (GAA; acid maltase; EC 3.2.1.20), which primarily involves cardiac and skeletal muscles. We hypothesized that systemic administration of an adeno-associated virus (AAV) vector containing a muscle specific regulatory cassette could drive efficacious transgene expression in GAA-knockout (GAA-KO) mice. AAV2/8 vectors containing the muscle creatine kinase (CK1) or hybrid α-myosin heavy chain enhancer-/muscle creatine kinase enhancer-promoter (MHCK7) cassettes were compared. The CK1 reduced glycogen content by approximately 50% in the heart and quadriceps, in comparison to untreated GAA-KO mice, whereas the MHCK7 containing vector reduced glycogen content even further: >95% in heart and >75% in the diaphragm and quadriceps. Administration of the MHCK7-containing vector significantly increased striated muscle function as assessed by increased Rotarod times at 18 weeks post-injection, whereas the CK1-containing vector did not increase Rotarod performance. Transduction efficiency was evaluated with an AAV2/8 vector in which MHCK7 drives alkaline-phosphatase, revealing that many more myofibers were transduced in the quadriceps than in the gastrocnemius. An AAV2/9 vector containing the MHCK7 cassette corrected GAA deficiency in the skeletal muscles of the distal limb, including the gastrocnemius, extensor digitalis longus, and soleus; furthermore, glycogen accumulations were substantially cleared by hGAA expression therein. Importantly, type IIb myofibers in the extensor digitalis longus were transduced, thereby correcting a myofiber type that is unresponsive to enzyme replacement therapy. In summary, AAV8 and AAV9-pseudotyped vectors containing the MHCK7 regulatory cassette achieved enhanced efficacy in Pompe disease mice. PMID:18560415

  6. (−)-EPICATECHIN IMPROVES MITOCHONDRIAL RELATED PROTEIN LEVELS AND AMELIORATES OXIDATIVE STRESS IN DYSTROPHIC DELTA SARCOGLYCAN NULL MOUSE STRIATED MUSCLE

    PubMed Central

    Ramirez-Sanchez, Israel; De los Santos, Sergio; Gonzalez-Basurto, Silvia; Canto, Patricia; Mendoza-Lorenzo, Patricia; Palma-Flores, Carlos; Ceballos-Reyes, Guillermo; Villarreal, Francisco; Zentella-Dehesa, Alejandro; Coral-Vazquez, Ramon

    2014-01-01

    Muscular dystrophies (MD) are a group of heterogeneous genetic disorders characterized by progressive striated muscle wasting and degeneration. Although the genetic basis for many of these disorders has been identified, the exact mechanism for disease pathogenesis remains unclear. The presence of oxidative stress (OS) is known to contribute to the pathophysiology and severity of the MD. Mitochondrial dysfunction is observed in MD and likely represents an important determinant of increased OS. Experimental antioxidant therapies have been implemented with the aim of protecting against disease progression, but results from clinical trials have been disappointing. In this study, we explored the capacity of the cacao flavonoid (−)-epicatechin (Epi) to mitigate OS by acting as a positive regulator of mitochondrial structure/function endpoints and redox balance control systems in skeletal and cardiac muscles of dystrophic, δ-sarcoglycan (δ-SG) null mice. Wild type or δ-SG null 2.5 month old male mice were treated via oral gavage with either water (control animals) or Epi (1 mg/kg, twice/day) for 2 weeks. Results evidence a significant normalization of total protein carbonylation, recovery of reduced/oxidized glutathione (GSH/GSSG ratio) and enhanced superoxide dismutase 2, catalase and citrate synthase activities with Epi treatment. These effects were accompanied by increases in protein levels for thiolredoxin, glutathione peroxidase, superoxide dismutase 2, catalase and mitochondrial endpoints. Furthermore, we evidence decreases in heart and skeletal muscle fibrosis, accompanied with an improvement in skeletal muscle function with treatment. These results warrant the further investigation of Epi as a potential therapeutic agent to mitigate MD associated muscle degeneration. PMID:25284161

  7. (-)-Epicatechin improves mitochondrial-related protein levels and ameliorates oxidative stress in dystrophic δ-sarcoglycan null mouse striated muscle.

    PubMed

    Ramirez-Sanchez, Israel; De los Santos, Sergio; Gonzalez-Basurto, Silvia; Canto, Patricia; Mendoza-Lorenzo, Patricia; Palma-Flores, Carlos; Ceballos-Reyes, Guillermo; Villarreal, Francisco; Zentella-Dehesa, Alejandro; Coral-Vazquez, Ramon

    2014-12-01

    Muscular dystrophies (MDs) are a group of heterogeneous genetic disorders characterized by progressive striated muscle wasting and degeneration. Although the genetic basis for many of these disorders has been identified, the exact mechanism of disease pathogenesis remains unclear. The presence of oxidative stress (OS) is known to contribute to the pathophysiology and severity of the MD. Mitochondrial dysfunction is observed in MD, and probably represents an important determinant of increased OS. Experimental antioxidant therapies have been implemented with the aim of protecting against disease progression, but results from clinical trials have been disappointing. In this study, we explored the capacity of the cacao flavonoid (-)-epicatechin (Epi) to mitigate OS by acting as a positive regulator of mitochondrial structure/function endpoints and redox balance control systems in skeletal and cardiac muscles of dystrophic, δ-sarcoglycan (δ-SG) null mice. Wild-type or δ-SG null 2.5-month-old male mice were treated via oral gavage with either water (controls) or Epi (1 mg·kg(-1) , twice daily) for 2 weeks. The results showed significant normalization of total protein carbonylation, recovery of the glutathione/oxidized glutathione ratio and enhanced superoxide dismutase 2, catalase and citrate synthase activities with Epi treatment. These effects were accompanied by increases in the protein levels of thioredoxin, glutathione peroxidase, superoxide dismutase 2, catalase, and mitochondrial endpoints. Furthermore, we found decreases in heart and skeletal muscle fibrosis, accompanied by an improvement in skeletal muscle function, with treatment. These results warrant further investigation of Epi as a potential therapeutic agent to mitigate MD-associated muscle degeneration. PMID:25284161

  8. Reversible oxidation of vicinal-thiols motif in sarcoplasmic reticulum calcium regulatory proteins is involved in muscle fatigue mechanism.

    PubMed

    Vázquez, Pável; Tirado-Cortés, Aldo; Álvarez, Rocío; Ronjat, Michel; Amaya, Araceli; Ortega, Alicia

    2016-10-01

    The mechanism underlying fatigue in skeletal muscle (SM) related to the redox-potential hypothesis, ranges from a direct effect of oxygen reactive species, to a number of other free radical intermediates targeting specific amino acids in the Ca(2+)-regulatory proteins of the sarcoplasmic reticulum (SR). In the present study, we investigate the selective oxidation/reduction of the protein motif Cys-(Xn=2-6)-Cys, known as a vicinal thiol group (VTG), present in the SR Ca(2+)-ATPase (SERCA) and in the Ca(2+)-channel ryanodine receptor (RyR) which are modified during muscle fatigue in SM. Selective oxidation of VTG with phenyl arsine oxide (PAO) increases fatigue in rat isolated SM and fatigue is prevented when muscle is previously incubated with a VTG selective reducing agent, 2,3-dimercaptopropanol (British anti-Lewisite (BAL)). In isolated SR membranes, PAO [<0.1mM] modifies SERCA conformation and inhibits ATPase activity but does not affect Ca(2+)-release. However, PAO at [>0.1mM] inhibits SERCA and RyR activities in a reversible manner by selectively reducing them. Interestingly, as observed by differential scanning calorimetry, the conformation of SERCA from fatigued muscle changed in a similar manner as when SERCA VTG where oxidized. The addition of BAL to fatigued muscle restored the structural conformation and activity of SERCA with full recovery of muscle force production after fatigue. We conclude that VTG reversible oxidation of SR Ca(2+) regulatory proteins are involved in muscle contraction/relaxation and are a molecular mechanism to be considered for muscle fatigue.

  9. Study of the function of sarcoplasmic reticulum of vascular smooth muscle during activation due to depolarization-induced calcium influx

    SciTech Connect

    Hwang, K.S.

    1987-01-01

    The role of sarcoplasmic reticulum (SR) in vascular smooth muscle was evaluated with respect to regulation of myoplasmic Ca{sup 2+} during the Ca{sup 2+} entry induced by depolarization. Calcium agonist, Bay K8644, stimulated Ca{sup 2+} influx as well as tension in physiological salt solution, (PSS) in contrast to the priming effects due to the depolarization originally reported. Disparity, however, was found between the Ca{sup 2+} entered and tension developed. Correlation between the tension and {sup 45}Ca influx showed a typical threshold phenomenon; the basal Ca{sup 2+} influx can be raised to a certain level (25%) without tension induction, after which a minor increase in Ca{sup 2+} influx produced significant tension. This subthreshold Ca{sup 2+} influx was found accumulated in the caffeine-sensitive Ca stores, the SR. This confirmed the dependency of tension on the rate of Ca{sup 2+} entry demonstrated by a previous report.

  10. Nitric oxide increases gene expression of Ca(2+)-ATPase in myocardial and skeletal muscle sarcoplasmic reticulum: physiological implications.

    PubMed

    Malyshev, I Y; Aymasheva, N P; Malenyuk, E B; Manukhina, E B; Khaspekov, G L; Mikoyan, V D; Kubrina, L N; Vanin, A F

    2000-01-01

    The aim of the study was to verify the hypothesis that NO-dependent regulation of sarcoplasmic reticulum Ca(2+)-ATPase (SERCA) gene expression can play an important role in prevention of calcium overload under the influence of detrimental factors. It was shown that 2 hours after the administration of the NO donor dinitrosyl iron complex (DNIC), the gene expression of myocardial SERCA was increased by 20% as compared to the control. In skeletal muscles, the maximum increase in SERCA expression was observed in 6 hours and amounted to 156% as compared with the initial value. Simultaneously DNIC enhanced the resistance of isolated heart and the organism as a whole to damaging effects of intracellular calcium overload induced by post-ischemic reperfusion or vigorous exercise, respectively. The results obtained confirm the existence of NO-dependent activation of SERCA expression and the important role of this mechanism in restriction of calcium overload. PMID:11208357

  11. The “Goldilocks Zone” from a redox perspective—Adaptive vs. deleterious responses to oxidative stress in striated muscle

    PubMed Central

    Alleman, Rick J.; Katunga, Lalage A.; Nelson, Margaret A. M.; Brown, David A.; Anderson, Ethan J.

    2014-01-01

    Consequences of oxidative stress may be beneficial or detrimental in physiological systems. An organ system's position on the “hormetic curve” is governed by the source and temporality of reactive oxygen species (ROS) production, proximity of ROS to moieties most susceptible to damage, and the capacity of the endogenous cellular ROS scavenging mechanisms. Most importantly, the resilience of the tissue (the capacity to recover from damage) is a decisive factor, and this is reflected in the disparate response to ROS in cardiac and skeletal muscle. In myocytes, a high oxidative capacity invariably results in a significant ROS burden which in homeostasis, is rapidly neutralized by the robust antioxidant network. The up-regulation of key pathways in the antioxidant network is a central component of the hormetic response to ROS. Despite such adaptations, persistent oxidative stress over an extended time-frame (e.g., months to years) inevitably leads to cumulative damages, maladaptation and ultimately the pathogenesis of chronic diseases. Indeed, persistent oxidative stress in heart and skeletal muscle has been repeatedly demonstrated to have causal roles in the etiology of heart disease and insulin resistance, respectively. Deciphering the mechanisms that underlie the divergence between adaptive and maladaptive responses to oxidative stress remains an active area of research for basic scientists and clinicians alike, as this would undoubtedly lead to novel therapeutic approaches. Here, we provide an overview of major types of ROS in striated muscle and the divergent adaptations that occur in response to them. Emphasis is placed on highlighting newly uncovered areas of research on this topic, with particular focus on the mitochondria, and the diverging roles that ROS play in muscle health (e.g., exercise or preconditioning) and disease (e.g., cardiomyopathy, ischemia, metabolic syndrome). PMID:25278906

  12. [Localization of minor proteins and structural changes in the myosin filaments of vertebrate striated muscle].

    PubMed

    Lednev, V V; Srebnitskaia, L K; Kornev, A N; Khromov, A S; Malinchik, S B

    1981-01-01

    The origin of meridional reflections in the X-ray diffraction patterns of vertebrate skeletal muscles in resting and rigor states was studied. The main results may be summarized as follows. 1. Most of the meridional reflections localized in groups at the positions of successive orders of the repeat period of about 430 A are contributed mainly by the C-protein component of thick filaments. 2. The meridional reflections at about 143 and 72 A in the X-ray diffraction pattern of the resting muscle are contributed mainly by the cross-bridge axial repeat period, while in the X-ray diffraction patterns of the rigorized muscle the reflections at approximately the same positions are contributed mainly by C-protein. The change in the positions of these particular reflections accompanying the transition of the muscle from rest to rigor and from rest to contraction cannot be considered as an indication of a change in the axial repeat period of the cross-bridges, as it was earlier suggested by some authors. 3. The transition of the muscle from resting to rigor state is accompanied by substantial changes in the positions of the meridional reflections contributed my minor proteins, which is indicative of the structural transition in the thick filaments. The observed changes may be interpreted as the result of the thick filaments elongation by about 1.5% or, alternatively, as a consequence of the redistribution of electron density of the meridional reflections 215 and 143 A during a single twitch of the muscle (Huxley et al., Nature, 1980 284, 140) may be interpreted as a natural consequence of the structural change in the thick filaments. It is concluded therefore that on stimulation of the vertebrate skeletal muscle the thickness filaments undergo a reversible structural change which may reflect the existence of myosin-linked regulation in that type of muscle.

  13. Triclosan impairs excitation-contraction coupling and Ca2+ dynamics in striated muscle.

    PubMed

    Cherednichenko, Gennady; Zhang, Rui; Bannister, Roger A; Timofeyev, Valeriy; Li, Ning; Fritsch, Erika B; Feng, Wei; Barrientos, Genaro C; Schebb, Nils H; Hammock, Bruce D; Beam, Kurt G; Chiamvimonvat, Nipavan; Pessah, Isaac N

    2012-08-28

    Triclosan (TCS), a high-production-volume chemical used as a bactericide in personal care products, is a priority pollutant of growing concern to human and environmental health. TCS is capable of altering the activity of type 1 ryanodine receptor (RyR1), but its potential to influence physiological excitation-contraction coupling (ECC) and muscle function has not been investigated. Here, we report that TCS impairs ECC of both cardiac and skeletal muscle in vitro and in vivo. TCS acutely depresses hemodynamics and grip strength in mice at doses ≥12.5 mg/kg i.p., and a concentration ≥0.52 μM in water compromises swimming performance in larval fathead minnow. In isolated ventricular cardiomyocytes, skeletal myotubes, and adult flexor digitorum brevis fibers TCS depresses electrically evoked ECC within ∼10-20 min. In myotubes, nanomolar to low micromolar TCS initially potentiates electrically evoked Ca(2+) transients followed by complete failure of ECC, independent of Ca(2+) store depletion or block of RyR1 channels. TCS also completely blocks excitation-coupled Ca(2+) entry. Voltage clamp experiments showed that TCS partially inhibits L-type Ca(2+) currents of cardiac and skeletal muscle, and [(3)H]PN200 binding to skeletal membranes is noncompetitively inhibited by TCS in the same concentration range that enhances [(3)H]ryanodine binding. TCS potently impairs orthograde and retrograde signaling between L-type Ca(2+) and RyR channels in skeletal muscle, and L-type Ca(2+) entry in cardiac muscle, revealing a mechanism by which TCS weakens cardiac and skeletal muscle contractility in a manner that may negatively impact muscle health, especially in susceptible populations.

  14. Alterations in mitochondria and sarcoplasmic reticulum from heart and skeletal muscle of horizontally casted primates

    NASA Technical Reports Server (NTRS)

    Sordahl, L. A.; Stone, H. L.

    1982-01-01

    Horizontally body-casted rhesus monkeys are used as an animal model in order to study the physiological changes known as cardiovascular deconditioning which occur during weightless conditions. No difference was found between the experimental and control animals in heart mitochondrial oxidative phosphorylation which indicates that no apparent changes occurred in the primary energy-producing system of the heart. A marked increase in cytochrome oxidase activity was observed in the casted primate heart mitochondria compared to controls, while a 25% decrease in respiratory substrate-supported calcium uptake was found in casted primate heart mitochondria compared to controls. Sacroplasmic reticulum isolated from the primate hearts revealed marked changes in calcium transport activities. It is concluded that the marked depression in cardiac sarcoplasmic reticulum functions indicates altered calcium homeostasis in the casted-primate heart which could be a factor in cardiovascular deconditioning.

  15. Androgens enhance in vivo 2-deoxyglucose uptake by rat striated muscle

    NASA Technical Reports Server (NTRS)

    Max, S. R.; Toop, J.

    1983-01-01

    It is shown that testosterone propionate (TP) causes a striking increase in the in vivo uptake of 2-deoxyglucose (2-DG) by the levator ani muscle of immature male rats, which was found to be uniformly distributed over the entire muscle. After a single subcutaneous injection of TP, no enhancement of 2-DG was observed before 3.5 hr, at which time uptake was increased 2-fold; maximum enhancement (4-fold) was attained at 12 hr. At 72 hr, 2-DG uptake remained elevated at twice the control value. It was determined that the effect of TP probably is mediated by specific androgen receptors. In addition, it was found that the effect of TP was blocked by the simultaneous administration of an androgen antagonist, cyproterone acetate. TP also was found to enhance the uptake of 2-DG in the bulbocavernosus (253 percent over control) and extensor digitorum longus muscles (150 percent over control), but not in the biceps brachii or soleus. It is suggested that the increased uptake of glucose may be an important early step in the anabolic response of muscle to androgens.

  16. Effects of different activity and inactivity paradigms on myosin heavy chain gene expression in striated muscle

    NASA Technical Reports Server (NTRS)

    Baldwin, K. M.; Haddad, F.

    2001-01-01

    The goal of this mini-review is to summarize findings concerning the role that different models of muscular activity and inactivity play in altering gene expression of the myosin heavy chain (MHC) family of motor proteins in mammalian cardiac and skeletal muscle. This was done in the context of examining parallel findings concerning the role that thyroid hormone (T(3), 3,5,3'-triiodothyronine) plays in MHC expression. Findings show that both cardiac and skeletal muscles of experimental animals are initially undifferentiated at birth and then undergo a marked level of growth and differentiation in attaining the adult MHC phenotype in a T(3)/activity level-dependent fashion. Cardiac MHC expression in small mammals is highly sensitive to thyroid deficiency, diabetes, energy deprivation, and hypertension; each of these interventions induces upregulation of the beta-MHC isoform, which functions to economize circulatory function in the face of altered energy demand. In skeletal muscle, hyperthyroidism, as well as interventions that unload or reduce the weight-bearing activity of the muscle, causes slow to fast MHC conversions. Fast to slow conversions, however, are seen under hypothyroidism or when the muscles either become chronically overloaded or subjected to intermittent loading as occurs during resistance training and endurance exercise. The regulation of MHC gene expression by T(3) or mechanical stimuli appears to be strongly regulated by transcriptional events, based on recent findings on transgenic models and animals transfected with promoter-reporter constructs. However, the mechanisms by which T(3) and mechanical stimuli exert their control on transcriptional processes appear to be different. Additional findings show that individual skeletal muscle fibers have the genetic machinery to express simultaneously all of the adult MHCs, e.g., slow type I and fast IIa, IIx, and IIb, in unique combinations under certain experimental conditions. This degree of

  17. The interactions between mitochondria and sarcoplasmic reticulum and the proteome characterization of mitochondrion-associated membrane from rabbit skeletal muscle.

    PubMed

    Liu, Zhouying; Du, Xiangning; Deng, Jie; Gu, Mingyue; Hu, Hongli; Gui, Miao; Yin, Chang-Cheng; Chang, Zhenzhan

    2015-08-01

    To obtain a comprehensive understanding of proteins involved in mitochondrion-sarcoplasmic reticulum (SR) linking, a catalog of proteins from mitochondrion-associated membrane (MAM) of New Zealand white rabbit skeletal muscle were analyzed by an optimized shotgun proteomic method. The membrane fractions were prepared by differential centrifugation and separated by 1D electrophoresis followed by a highly reproducible, automated LC-MS/MS on the hybrid linear ion trap (LTQ)-Orbitrap mass spectrometer. By integrating as low as 1% false discovery rate as one of the features for quality control method, 459 proteins were identified from both of the two independent MAM preparations. Protein pI value, molecular weight range, and transmembrane region were calculated using bioinformatics softwares. One hundred one proteins were recognized as membrane proteins. This protein database suggested that the MAM preparations composed of proteins from mitochondrion, SR, and transverse-tubule. This result indicated mitochondria physically linked with SR in rabbit skeletal muscle, voltage-dependent anion channel 1 (VDAC1), VDAC2, and VDAC3 might participate in formation of the tethers between SR and mitochondria.

  18. The formation and regression of synapses during the re-innervation of axolotl striated muscles.

    PubMed Central

    Bennett, M R; Raftos, J

    1977-01-01

    1. A study has been made of the formation and regression of synapses formed by spinal nerves 16 and 17 in axolotl hind-limb flexor muscles following the severing of nerve 16, using histological, ultrastructural and electrophysiological techniques. 2. Axolotl hind-limb flexor myofibres possessed 'en plaque' end-plates from either spinal nerve 16 or 17 or both at intervals of about 1000 micronm along their length; the myofibre's length constant was about 700 micronm allowing electrophysiological observations of at least two of these synapses during a single impalement; transmitter release at these synapses could be described by binomial statistics and in a given set of ionic conditions the binomial statistic parameter n was directly proportional to the size of the nerve terminals whilst the binomial statistic parameter p was invariant to changes in nerve terminal size. 3. The distribution of synapses formed by spinal nerves 16 and 17 in different sectors of the axolotl hind-limb flexor muscles was determined from a study of evoked end-plate potentials; the middle and proximal sectors of the flexor muscles contained myofibres which received an innervation from nerve 16 only, whereas the sectors surrounding these contained myofibres innervated either by nerve 16 or nerve 17 or by both nerves. 4. Six days following the severing of spinal nerve 16, evoked transmitter release from the synapses formed by this nerve had failed; transmission was subsequently recorded at a few synapses formed by nerve 17 in the middle and proximal sectors of the flexor muscles which are not normally innervated by this nerve and these synapses had a low n; during the succeeding four weeks the value of n at the synapses increased to a size about 70% that of the terminals normally formed by nerve 16 at these sites. 5. Four weeks after severing nerve 16, myofibres which possessed synapses formed by nerve 17 also possessed synapses from re-innervating nerve 16 and these were sometimes formed at

  19. Modulation of the cytosolic androgen receptor in striated muscle by sex steroids

    NASA Technical Reports Server (NTRS)

    Rance, N. E.; Max, S. R.

    1984-01-01

    The effects of orchiectomy (GDX) and of subsequent administration of testosterone propionate (TP) or 17(beta)-estradiol (E2) on the maximum binding (Bmax) and apparent Kd of the cytosolic androgen receptor in levator ani (LA) and skeletal muscles of adult male Sprague-Dawley rats are investigated experimentally. The results are presented in graphs and discussed. In LA, BMAX is found to rise from a control level of 2.5 fmol/mg protein to 280, 600, 478, and 133 percent of control at 12 h, 14 d, 30 d, and 44 d after GDX, respectively, while Kd increased only insignificantly (from 680 to 960 fM); Bmax is held at control levels for 6 h by cycloheximide given at GDX, is unaffected by TP given at 30 d, and is further increased (by 480 percent at 44 d) by administration of E2 at 30 d. Bmax in skeletal muscles is found to increase to 139, 212, 220, and 158 percent of control at 12 h, 14 d, 30 d, and 44 d, respectively; Bmax is returned to control at 44 d by TP at 30 d but is not affected by E2. The effect of E2 in LA is attributed to either induction of the cytosolic receptor or a decreased rate of receptor degradation.

  20. Dynamic coupling of regulated binding sites and cycling myosin heads in striated muscle.

    PubMed

    Campbell, Kenneth S

    2014-03-01

    In an activated muscle, binding sites on the thin filament and myosin heads switch frequently between different states. Because the status of the binding sites influences the status of the heads, and vice versa, the binding sites and myosin heads are dynamically coupled. The functional consequences of this coupling were investigated using MyoSim, a new computer model of muscle. MyoSim extends existing models based on Huxley-type distribution techniques by incorporating Ca(2+) activation and cooperative effects. It can also simulate arbitrary cross-bridge schemes set by the researcher. Initial calculations investigated the effects of altering the relative speeds of binding-site and cross-bridge kinetics, and of manipulating cooperative processes. Subsequent tests fitted simulated force records to experimental data recorded using permeabilized myocardial preparations. These calculations suggest that the rate of force development at maximum activation is limited by myosin cycling kinetics, whereas the rate at lower levels of activation is limited by how quickly binding sites become available. Additional tests investigated the behavior of transiently activated cells by driving simulations with experimentally recorded Ca(2+) signals. The unloaded shortening profile of a twitching myocyte could be reproduced using a model with two myosin states, cooperative activation, and strain-dependent kinetics. Collectively, these results demonstrate that dynamic coupling of binding sites and myosin heads is important for contractile function.

  1. Twitchin kinase interacts with MAPKAP kinase 2 in Caenorhabditis elegans striated muscle

    PubMed Central

    Matsunaga, Yohei; Qadota, Hiroshi; Furukawa, Miho; Choe, Heejoo (Helen); Benian, Guy M.

    2015-01-01

    In Caenorhabditis elegans, twitchin is a giant polypeptide located in muscle A-bands. The protein kinase of twitchin is autoinhibited by 45 residues upstream (NL) and 60 residues downstream (CRD) of the kinase catalytic core. Molecular dynamics simulation on a twitchin fragment revealed that the NL is released by pulling force. However, it is unclear how the CRD is removed. To identify proteins that may remove the CRD, we performed a yeast two-hybrid screen using twitchin kinase as bait. One interactor is MAK-1, C. elegans orthologue of MAPKAP kinase 2. MAPKAP kinase 2 is phosphorylated and activated by p38 MAP kinase. We demonstrate that the CRD of twitchin is important for binding to MAK-1. mak-1 is expressed in nematode body wall muscle, and antibodies to MAK-1 localize between and around Z-disk analogues and to the edge of A-bands. Whereas unc-22 mutants are completely resistant, mak-1 mutants are partially resistant to nicotine. MAK-1 can phosphorylate twitchin NL-Kin-CRD in vitro. Genetic data suggest the involvement of two other mak-1 paralogues and two orthologues of p38 MAP kinase. These results suggest that MAK-1 is an activator of twitchin kinase and that the p38 MAP kinase pathway may be involved in the regulation of twitchin. PMID:25851606

  2. Acute ethanol inhibits calcium influxes into esophageal smooth but not striated muscle: a possible mechanism for ethanol-induced inhibition of esophageal contractility.

    PubMed

    Keshavarzian, A; Zorub, O; Sayeed, M; Urban, G; Sweeney, C; Winship, D; Fields, J

    1994-09-01

    In both humans and cats, EtOH administered in vivo and acutely decreases contractility of smooth muscle of lower esophageal sphincter (LES) and lower esophagus (LE), but not striated muscle of upper esophagus. To see if these effects are associated with perturbation of Ca++ homeostasis, esophageal muscle slices were incubated in vitro with EtOH and then 45Ca++. At steady-state Ca++ uptake, some slices were exposed to 1 microM carbachol (CCH). Although 100 mM EtOH had no effect on Ca++ uptake into resting or stimulated striated muscle of upper esophagus, it significantly inhibited Ca++ uptake into smooth muscle of LES and LE. For unstimulated LE and resting LES, 100 mM EtOH significantly inhibited both initial uptake and steady-state levels, whereas lower doses had no significant effect. EtOH at 100 mM also affected changes in Ca++ content induced by CCH stimulation. CCH increased total exchangeable tissue Ca++ content in LE, whereas it decreased Ca++ content in LES. EtOH at 100 mM blunted these CCH-induced effects in both LES and LE. In contrast to resting muscle, inhibition of CCH-stimulated LE muscle was not limited to 100 mM EtOH, because substantial and significant inhibition was also seen at EtOH doses of 25 and 50 mM, doses which are relevant even in social drinking. Thus, EtOH inhibition of Ca++ influx into esophageal muscle is selective for smooth muscle, can occur at pharmacologically relevant EtOH doses and could be the underlying mechanism for EtOH's inhibition of contractility of esophageal smooth muscle. PMID:7932153

  3. Metabolic consequences of functional complexes of mitochondria, myofibrils and sarcoplasmic reticulum in muscle cells.

    PubMed

    Andrienko, T; Kuznetsov, A V; Kaambre, T; Usson, Y; Orosco, A; Appaix, F; Tiivel, T; Sikk, P; Vendelin, M; Margreiter, R; Saks, V A

    2003-06-01

    Regulation of mitochondrial respiration both by endogenous and exogenous ADP in the cells in situ was studied in isolated and permeabilized cardiomyocytes, permeabilized cardiac fibers and 'ghost' fibers (all with a diameter of 10-20 micro m) at different (0-3 micro moll(-1)) free Ca(2+) concentrations in the medium. In all these preparations, the apparent K(m) of mitochondrial respiration for exogenous ADP at free Ca(2+) concentrations of 0-0.1 micro moll(-1) was very high, in the range of 250-350 micro moll(-1), in contrast to isolated mitochondria in vitro (apparent K(m) for ADP is approximately 20 micro moll(-1)). An increase in the free Ca(2+) concentration (up to 3 micro moll(-1), which is within physiological range), resulted in a very significant decrease of the apparent K(m) value to 20-30 micro moll(-1), a decrease of V(max) of respiration in permeabilized intact fibers and a strong contraction of sarcomeres. In ghost cardiac fibers, from which myosin was extracted but mitochondria were intact, neither the high apparent K(m) for ADP (300-350 micro moll(-1)) nor V(max) of respiration changed in the range of free Ca(2+) concentration studied, and no sarcomere contraction was observed. The exogenous-ADP-trapping system (pyruvate kinase + phosphoenolpyruvate) inhibited endogenous-ADP-supported respiration in permeabilized cells by no more than 40%, and this inhibition was reversed by creatine due to activation of mitochondrial creatine kinase. These results are taken to show strong structural associations (functional complexes) among mitochondria, sarcomeres and sarcoplasmic reticulum. Inside these complexes, mitochondrial functional state is controlled by channeling of ADP, mostly via energy- and phosphoryl-transfer networks, and apparently depends on the state of sarcomere structures. PMID:12756288

  4. Effect of increasing the level of omega-3 fatty acids on rat skeletal muscle sarcoplasmic reticulum.

    PubMed

    Stubbs, C D; Kisielewski, A E

    1990-09-01

    The effect of dietary supplementation with fish oil as compared to corn oil on the lipid dynamics and calcium ATPase activity of rat skeletal sarcoplasmic reticulum was examined. After four-week supplementation with fish oil, the levels of eicosapentaenoic (20:5 omega 3), docosapentaenoic (22:5 omega 3) and docosahexaenoic (22:6 omega 3) acids in the total lipids were 5.3, 5.5 and 28.1% of the total fatty acids, respectively. In contrast, with corn oil only 22:6 was found (8.9%). The level of these fatty acids in phosphatidylethanolamine from the membranes of animals fed fish oil was 4.2 (20:5), 5.4 (22:5) and 49.1% (22:6); and for phosphatidylcholine it was 5.4 (20:5), 4.6 (22:5) and 17.4% (22:6). Again, in corn oil fed animals, only 22:6 was found in appreciable amounts, namely 28.3% in phosphatidylethanolamine and 1.8% in phosphatidylcholine. The steady state fluorescence anisotropy of 1,6-diphenyl-1,3,5-hexatriene (DPH) was used to assess lipid order and was found to be only slightly less for membranes from animals supplemented with fish oil (0.120) as compared to those supplemented with corn oil (0.124). The calcium ATPase was found to be unaffected by supplementation consistent with the observed modest changes in lipid order as well as with suggestions that the enzyme is relatively insensitive to the level of unsaturation. It could be argued that if large increases in fatty acyl polyunsaturation in mammalian cell membranes would lead to marked alterations in bulk membrane lipid motional properties, this may not be in the interest of preserving physiological function.(ABSTRACT TRUNCATED AT 250 WORDS)

  5. Drug action of benzocaine on the sarcoplasmic reticulum Ca-ATPase from fast-twitch skeletal muscle.

    PubMed

    Di Croce, D; Trinks, P W; Grifo, M B; Takara, D; Sánchez, G A

    2015-11-01

    The effect of the local anesthetic benzocaine on sarcoplasmic reticulum membranes isolated from fast-twitch muscles was tested. The effects on Ca-ATPase activity, calcium binding and uptake, phosphoenzyme accumulation and decomposition were assessed using radioisotopic methods. The calcium binding to the Ca-ATPase was noncompetitively inhibited, and the enzymatic activity decreased in a concentration-dependent manner (IC50 47.1 mM). The inhibition of the activity depended on the presence of the calcium ionophore calcimycin and the membrane protein concentration. The pre-exposure of the membranes to benzocaine enhanced the enzymatic activity in the absence of calcimycin, supporting the benzocaine permeabilizing effect, which was prevented by calcium. Benzocaine also interfered with the calcium transport capability by decreasing the maximal uptake (IC50 40.3 mM) without modification of the calcium affinity for the ATPase. It inhibited the phosphorylation of the enzyme, and at high benzocaine concentration, the dephosphorylation step became rate-limiting as suggested by the biphasic profile of phosphoenzyme accumulation at different benzocaine concentrations. The data reported in this paper revealed a complex pattern of inhibition involving two sites for interaction with low and high benzocaine concentrations. It is concluded that benzocaine not only exerts an indirect action on the membrane permeability to calcium but also affects key steps of the Ca-ATPase enzymatic cycle. PMID:26173386

  6. Effects of age on calcium transport activity of sarcoplasmic reticulum in fast- and slow-twitch rat muscle fibres.

    PubMed Central

    Larsson, L; Salviati, G

    1989-01-01

    1. The calcium transport activity of the sarcoplasmic reticulum (SR) was measured in chemically skinned single fast- and slow-twitch muscle fibres from young (3 months) and old (23-24 months) rats. Contractile properties, the myosin heavy chain (MHC) composition and enzyme histochemical features were studied in relation to the SR characteristics. 2. In fast-twitch single motor units, the contraction time of the isometric twitch increased (P less than 0.001) from 13 +/- 1 ms in young animals to 18 +/- 2 ms in old ones. In the slow-twitch soleus, the contraction (P less than 0.001) and half-relaxation (P less than 0.05) times increased from 30 +/- 5 and 45 +/- 10 ms, respectively, in the young animals to 43 +/- 3 and 55 +/- 4 ms in the old ones. The proportion of slow-twitch (type I) fibres increased (P less than 0.05) with age in the soleus from 92 +/- 6 to 98 +/- 2% and the proportion of fast-twitch fibres (type IIA) decreased (P less than 0.01) from 6 +/- 5 to 0 +/- 0%. 3. The Ca2+ accumulation capacity (an index of SR volume), the rate of Ca2+ uptake and the fractional rate of SR filling (an estimate of the specific activity of the Ca2+ pump) were decreased by 18 (P less than 0.05), 32 (P less than 0.01) and 32% (P less than 0.001), respectively, in the old fast-twitch muscle fibres. In the slow-twitch muscle fibres, on the other hand, no significant age-related changes were observed in the Ca2+ transport activity of the SR. Thus, ageing exerts a differential influence on SR volume and function in fast- and slow-twitch fibres. 4. It is concluded that an age-related impairment of intrinsic SR function and a decrease in SR volume are probable factors underlying the decreased speed of contraction of fast-twitch muscle fibres in old age. In the slow-twitch soleus, on the other hand, one or more other mechanisms are responsible for the age-related decrease in the speed of contraction. The loss of fast-twitch muscle fibres in old soleus is one mechanism, but not the

  7. Mechanical load induces sarcoplasmic wounding and FGF release in differentiated human skeletal muscle cultures

    NASA Technical Reports Server (NTRS)

    Clarke, M. S.; Feeback, D. L.

    1996-01-01

    The transduction mechanism (or mechanisms) responsible for converting a mechanical load into a skeletal muscle growth response are unclear. In this study we have used a mechanically active tissue culture model of differentiated human skeletal muscle cells to investigate the relationship between mechanical load, sarcolemma wounding, fibroblast growth factor release, and skeletal muscle cell growth. Using the Flexcell Strain Unit we demonstrate that as mechanical load increases, so too does the amount of sarcolemma wounding. A similar relationship was also observed between the level of mechanical load inflicted on the cells and the amount of bFGF (FGF2) released into the surrounding medium. In addition, we demonstrate that the muscle cell growth response induced by chronic mechanical loading in culture can be inhibited by the presence of an antibody capable of neutralizing the biological activity of FGF. This study provides direct evidence that mechanically induced, sarcolemma wound-mediated FGF release is an important autocrine mechanism for transducing the stimulus of mechanical load into a skeletal muscle growth response.

  8. Stromal interaction molecule 1 (STIM1) regulates sarcoplasmic/endoplasmic reticulum Ca²⁺-ATPase 1a (SERCA1a) in skeletal muscle.

    PubMed

    Lee, Keon Jin; Hyun, Changdo; Woo, Jin Seok; Park, Chang Sik; Kim, Do Han; Lee, Eun Hui

    2014-05-01

    Stromal interaction molecule 1 (STIM1) mediates Ca2+ movements from the extracellular space to the cytosol through a store-operated Ca2+ entry (SOCE) mechanism in various cells including skeletal muscle cells. In the present study, to reveal the unidentified functional role of the STIM1 C terminus from 449 to 671 amino acids in skeletal muscle, binding assays and quadrupole time-of-flight mass spectrometry were used to identify proteins binding in this region along with proteins that mediate skeletal muscle contraction and relaxation. STIM1 binds to sarcoplasmic/endoplasmic reticulum Ca2+-ATPase 1a (SERCA1a) via this region (called STIM1-SBR). The binding was confirmed in endogenous full-length STIM1 in rabbit skeletal muscle and mouse primary skeletal myotubes via co-immunoprecipitation assay and immunocytochemistry. STIM1 knockdown in mouse primary skeletal myotubes decreased Ca2+ uptake from the cytosol to the sarcoplasmic reticulum (SR) through SERCA1a only at micromolar cytosolic Ca2+ concentrations, suggesting that STIM1 could be required for the full activity of SERCA1a possibly during the relaxation of skeletal muscle. Various Ca2+ imaging experiments using myotubes expressing STIM1-SBR suggest that STIM1 is involved in intracellular Ca2+ distributions between the SR and the cytosol via regulating SERCA1a activity without affecting SOCE. Therefore, in skeletal muscle, STIM1 could play an important role in regulating Ca2+ movements between the SR and the cytosol. PMID:24077737

  9. The Ca2+-release channel/ryanodine receptor is localized in junctional and corbular sarcoplasmic reticulum in cardiac muscle

    PubMed Central

    1993-01-01

    The subcellular distribution of the Ca(2+)-release channel/ryanodine receptor in adult rat papillary myofibers has been determined by immunofluorescence and immunoelectron microscopical studies using affinity purified antibodies against the ryanodine receptor. The receptor is confined to the sarcoplasmic reticulum (SR) where it is localized to interior and peripheral junctional SR and the corbular SR, but it is absent from the network SR where the SR-Ca(2+)-ATPase and phospholamban are densely distributed. Immunofluorescence labeling of sheep Purkinje fibers show that the ryanodine receptor is confined to discrete foci while the SR-Ca(2+)-ATPase is distributed in a continuous network-like structure present at the periphery as well as throughout interior regions of these myofibers. Because Purkinje fibers lack T- tubules, these results indicate that the ryanodine receptor is localized not only to the peripheral junctional SR but also to corbular SR densely distributed in interfibrillar spaces of the I-band regions. We have previously identified both corbular SR and junctional SR in cardiac muscle as potential Ca(2+)-storage/Ca(2+)-release sites by demonstrating that the Ca2+ binding protein calsequestrin and calcium are very densely distributed in these two specialized domains of cardiac SR in situ. The results presented here provide strong evidence in support of the hypothesis that corbular SR is indeed a site of Ca(2+)-induced Ca2+ release via the ryanodine receptor during excitation contraction coupling in cardiac muscle. Furthermore, these results indicate that the function of the cardiac Ca(2+)-release channel/ryanodine receptor is not confined to junctional complexes between SR and the sarcolemma. PMID:8381786

  10. Measurement of Calcium Fluctuations Within the Sarcoplasmic Reticulum of Cultured Smooth Muscle Cells Using FRET-based Confocal Imaging.

    PubMed

    Ziomek, Gabriela; van Breemen, Cornelis; Esfandiarei, Mitra

    2016-01-01

    Maintenance of steady-state calcium (Ca(2+)) levels in the sarcoplasmic reticulum (SR) of vascular smooth muscle cells (VSMCs) is vital to their overall health. A significant portion of intracellular Ca(2+) content is found within the SR stores in VSMCs. As the only intracellular organelle with a close association to the surrounding extracellular space through plasma membrane-SR junctions, the SR can be considered to constitute the first line of response to any irregularity in Ca(2+) transients, or stress experienced by the cell. Among its many functions, one of the most important is its role in the transmission of Ca(2+)-regulated signals throughout the cell to induce further cell-wide reactions downstream. The more common use of cytoplasmic Ca(2+) indicators in this regard is overall insufficient for research into the highly dynamic changes to the intraluminal SR Ca(2+) store that have yet to be fully characterized. Here, we provide a detailed protocol for the direct and clear measurement of luminal SR Ca(2+). This tool is useful for investigation into the nuanced changes in SR Ca(2+) that have significant subsequent effects on the normal function and health of the cell. Fluctuations in SR Ca(2+) content specifically can provide us with a significant amount of information pertaining to cellular responses to disease or stress conditions experienced by the cell. In this method, a modified version of a SR-targeted Ca(2+) indicator, D1SR, is used to detect Ca(2+) fluctuations in response to the introduction of agents to cultured rat aortic smooth muscle cells (SMCs). Following incubation with the D1SR indicator, confocal fluorescence microscopy and fluorescence resonance energy transfer (FRET)-based imaging are used to directly observe changes to intraluminal SR Ca(2+) levels under control conditions and with the addition of agonist agents that function to induce intracellular Ca(2+) movement. PMID:27403723

  11. Simulation of Ca2+ release from the sarcoplasmic reticulum with three-dimensional sarcomere model in cardiac muscle.

    PubMed

    Tameyasu, Tsukasa

    2002-08-01

    A simulation of some basic features of Ca(2+) release from the sarcoplasmic reticulum (SR) in cardiac muscle was made with a model based on the mechanism of Ca(2+)-induced Ca(2+)-release. The half-sarcomere modeled as a circular cylinder was divided into 20 annular elements in the radial, 50 slices in the axial, and 125 slices in the azimuthal direction. The cylindrical surface of the sarcomere was covered by a layer of the SR. The rate of Ca(2+) release from the terminal sac (TS) is proportional to the product of the open probability of the Ca(2+) release channel and the difference of [Ca(2+)] between the TS and an element facing the TS. Ca(2+) moves from element to element by simple diffusion and is taken up by the tubular SR via Ca(2+)-ATPase. Ca(2+) influx (I(ca)) to trigger the TS Ca(2+) release was introduced to either a single element facing the TS (local I(ca)) or to 20 elements aligned at the level of the Z-line (uniform I(ca)). The simulation showed that with both types of I(ca), TS Ca(2+) release is smoothly graded over a wide range of I(ca) with the TS moderately loaded with Ca(2+). The gain determined by dividing the total amount of TS Ca(2+) release by I(ca) was greater with local than with uniform I(ca). Mechanical alternans was simulated with both the local and uniform I(ca) with an appropriate rate of Ca(2+) replenishment to the TS. A Ca(2+) wave was simulated with a model consisting of 8 longitudinally consecutive sarcomeres with TS heavily loaded with Ca(2+). Thus the present model accounted for graded TS Ca(2+) release, mechanical alternans, and Ca(2+) wave in cardiac muscle at the same time. PMID:12519471

  12. Activation and propagation of Ca2+ release from inside the sarcoplasmic reticulum network of mammalian skeletal muscle.

    PubMed

    Cully, Tanya R; Edwards, Joshua N; Launikonis, Bradley S

    2014-09-01

    Skeletal muscle fibres are large and highly elongated cells specialized for producing the force required for posture and movement. The process of controlling the production of force within the muscle, known as excitation-contraction coupling, requires virtually simultaneous release of large amounts of Ca(2+) from the sarcoplasmic reticulum (SR) at the level of every sarcomere within the muscle fibre. Here we imaged Ca(2+) movements within the SR, tubular (t-) system and in the cytoplasm to observe that the SR of skeletal muscle is a connected network capable of allowing diffusion of Ca(2+) within its lumen to promote the propagation of Ca(2+) release throughout the fibre under conditions where inhibition of SR ryanodine receptors (RyRs) was reduced. Reduction of cytoplasmic [Mg(2+)] ([Mg(2+)]cyto) induced a leak of Ca(2+) through RyRs, causing a reduction in SR Ca(2+) buffering power argued to be due to a breakdown of SR calsequestrin polymers, leading to a local elevation of [Ca(2+)]SR. The local rise in [Ca(2+)]SR, an intra-SR Ca(2+) transient, induced a local diffusely rising [Ca(2+)]cyto. A prolonged Ca(2+) wave lasting tens of seconds or more was generated from these events. Ca(2+) waves were dependent on the diffusion of Ca(2+) within the lumen of the SR and ended as [Ca(2+)]SR dropped to low levels to inactivate RyRs. Inactivation of RyRs allowed re-accumulation of [Ca(2+)]SR and the activation of secondary Ca(2+) waves in the persistent presence of low [Mg(2+)]cyto if the threshold [Ca(2+)]SR for RyR opening could be reached. Secondary Ca(2+) waves occurred without an abrupt reduction in SR Ca(2+) buffering power. Ca(2+) release and wave propagation occurred in the absence of Ca(2+)-induced Ca(2+) release. These observations are consistent with the activation of Ca(2+) release through RyRs of lowered cytoplasmic inhibition by [Ca(2+)]SR or store overload-induced Ca(2+) release. Restitution of SR Ca(2+) buffering power to its initially high value required

  13. Detection of the nicotinic acetylcholine receptor alpha-subunit mRNA by in situ hybridization at neuromuscular junctions of 15-day-old chick striated muscles.

    PubMed Central

    Fontaine, B; Sassoon, D; Buckingham, M; Changeux, J P

    1988-01-01

    In adult vertebrate striated muscle, the nicotinic acetylcholine receptor (AChR) is almost exclusively localized in the postsynaptic membrane of the neuromuscular junction. Using in situ hybridization, we show that, in two different chicken muscles [the slow multi-innervated anterior latissimus dorsi (ALD) and the fast singly innervated posterior latissimus dorsi (PLD)], the AChR alpha-subunit mRNA is detected at discrete regions on myofibres and that these regions co-localize (80% correspondence) with neuromuscular junctions identified by histochemical staining for acetylcholinesterase. Moreover, autoradiographic grains densely accumulate on and around subsynaptic nuclei. In contrast, hybridization with an actin probe results in a strong signal distributed over the entire length of the myofibres. Denervation increases the level of AChR alpha-subunit mRNA both in the PLD and to a lesser extent in the ALD. By in situ hybridization we observe that, although a perinuclear pattern is maintained, the labelled nuclei appear randomly distributed among approximately 10% of the nuclei. These results are discussed in a model of AChR gene expression in vertebrate striated muscle fibres. Images PMID:3396533

  14. Merosin, a protein specific for basement membranes of Schwann cells, striated muscle, and trophoblast, is expressed late in nerve and muscle development.

    PubMed Central

    Leivo, I; Engvall, E

    1988-01-01

    We have identified a tissue-specific basement membrane-associated protein by using monoclonal antibodies prepared against a protein fraction of human placenta. In immunofluorescence, the monoclonal antibodies stained basement membranes of Schwann cells, striated muscle, and trophoblast, whereas no reaction was seen with any other basement membrane or tissue structure. In antibody-affinity chromatography of proteolytic digests of human placenta, a 65-kDa polypeptide was bound by these monoclonal antibodies. Rabbit antisera and monoclonal antibodies raised against the isolated 65-kDa polypeptide stained human and monkey tissues identically to the original monoclonal antibodies and reacted with an 80-kDa polypeptide in tissue extracts prepared without proteolysis. The 65-kDa and 80-kDa polypeptides were shown to be immunologically distinct from laminin, type IV collagen, fibronectin, and major serum proteins. They presumably represent a novel basement membrane-associated protein, which we have named merosin. No merosin immunoreactivity could be detected in cultures of any of 28 established cell lines. In developing mouse tissues, merosin staining first appeared at the newborn stage. The restricted tissue distribution and late developmental appearance of merosin suggest that the protein has a tissue-specific function associated with a high level of differentiation. Images PMID:3278318

  15. Calcium release and ionic changes in the sarcoplasmic reticulum of tetanized muscle: an electron-probe study

    PubMed Central

    1981-01-01

    Approximately 60-70% of the total fiber calcium was localized in the terminal cisternae (TC) in resting frog muscle as determined by electron-probe analysis of ultrathin cryosections. During a 1.2 s tetanus, 59% (69 mmol/kg dry TC) of the calcium content of the TC was released, enough to raise total cytoplasmic calcium concentration by approximately 1 mM. This is equivalent to the concentration of binding sites on the calcium-binding proteins (troponin and parvalbumin) in frog muscle. Calcium release was associated with a significant uptake of magnesium and potassium into the TC, but the amount of calcium released exceeded the total measured cation accumulation by 62 mEq/kg dry weight. It is suggested that most of the charge deficit is apparent, and charge compensation is achieved by movement of protons into the sarcoplasmic reticulum (SR) and/or by the movement of organic co- or counterions not measured by energy dispersive electron-probe analysis. There was no significant change in the sodium or chlorine content of the TC during tetanus. The unchanged distribution of a permeant anion, chloride, argues against the existence of a large and sustained transSR potential during tetanus, if the chloride permeability of the in situ SR is as high as suggested by measurements on fractionated SR. The calcium content of the longitudinal SR (LSR) during tetanus did not show the LSR to be a major site of calcium storage and delayed return to the TC. The potassium concentration in the LSR was not significantly different from the adjacent cytoplasmic concentration. Analysis of small areas of I-band and large areas, including several sarcomeres, suggested that chloride is anisotropically distributed, with some of it probably bound to myosin. In contrast, the distribution of potassium in the fiber cytoplasm followed the water distribution. The mitochondrial concentration of calcium was low and did not change significantly during a tetanus. The TC of both tetanized and resting

  16. Evidence of a role for calmodulin in the regulation of calcium release from skeletal muscle sarcoplasmic reticulum

    SciTech Connect

    Meissner, G.

    1986-01-14

    The effect of calmodulin and calmodulin inhibitors on the Ca2+ release channel of heavy skeletal muscle sarcoplasmic reticulum (SR) vesicles was investigated. SR vesicles were passively loaded with 45Ca2+ in the presence of calmodulin and its inhibitors, followed by measurement of 45Ca2+ release rates by means of a rapid-quench-Millipore filtration method. Calmodulin at a concentration of 2-10 microM reduced 45Ca2+ efflux rates from passively loaded vesicles by a factor of 2-3 in media containing 10(-6)-10(-3) M Ca2+. At 10(-9) M Ca2+, calmodulin was without effect. 45Ca2+ release rates were varied 1000-fold (k1 approximately equal to 0.1-100 s-1) by using 10(-5) M Ca2+ with either Mg2+ or the ATP analogue adenosine 5'-(beta,gamma-methylenetriphosphate) in the release medium. In all instances, a similar 2-3-fold reduction in release rates was observed. At 10(-5) M Ca2+, 45Ca2+ release was half-maximally inhibited by about 2 X 10(-7) M calmodulin, and this inhibition was reversible. Heavy SR vesicle fractions contained 0.1-02 micrograms of endogenous calmodulin/mg of vesicle protein. However, the calmodulin inhibitors trifluoperazine, calmidazolium, and compound 48/80 were without significant effect on 45Ca2+ release at concentrations which inhibit calmodulin-mediated reactions in other systems. Studies with actively loaded vesicles also suggested that heavy SR vesicles contain a Ca2+ permeation system that is inhibited by calmodulin.

  17. Ca(2+)-induced folding and aggregation of skeletal muscle sarcoplasmic reticulum calsequestrin. The involvement of the trifluoperazine-binding site.

    PubMed

    He, Z; Dunker, A K; Wesson, C R; Trumble, W R

    1993-11-25

    Calsequestrin is an intermediate affinity, high capacity Ca(2+)-binding protein found in the lumen of the sarcoplasmic reticulum of both skeletal and cardiac muscle cells. Previous sequence analysis suggested that calsequestrin may contain a hydrophobic binding site for the drug trifluoperazine, a site shared by the calmodulin family and shown to play a role in calmodulin/calmodulin receptor interaction. Previous studies showed that, upon Ca2+ binding, calsequestrin undergoes a conformational change, burying the trifluoperazine-binding site, folding into a more compact structure that is trypsin-resistant, and increasing the negative ellipticity of the circular dichroism spectrum. In this study, the structural and functional roles of the trifluoperazine-binding site in the Ca(2+)-induced conformational change of calsequestrin are further studied using the calmodulin antagonists trifluoperazine and melittin. If trifluoperazine or melittin is added to calsequestrin prior to Ca2+ addition, then Ca(2+)-induced folding is inhibited as determined by the changes in circular dichroism spectra and protein sensitivity to trypsin digestion. If, however, Ca2+ is added prior to trifluoperazine or melittin, calsequestrin remains resistant to trypsin digestion, just as if the calmodulin antagonists are not present, suggesting that the conformational change is not affected. Aggregates of calsequestrin that exhibit high Ca2+ binding capacity have previously been shown to occur at high Ca2+ and calsequestrin concentrations. By preventing a prerequisite folding step, trifluoperazine or melittin also prevents the Ca(2+)-induced aggregation of calsequestrin, thus decreasing the maximal Ca2+ binding by calsequestrin. These data suggest that the trifluoperazine-binding site is critically involved in the Ca(2+)-induced intramolecular folding step required for the intermolecular interactions leading to high capacity Ca(2+)-binding by calsequestrin.

  18. The nucleotide-binding site of the sarcoplasmic reticulum Ca-ATPase is conformationally altered in aged skeletal muscle.

    PubMed

    Chen, B; Jones, T E; Bigelow, D J

    1999-11-01

    Cellular conditions in senescent skeletal muscle have been shown to result in the loss of conformational stability of the sarcoplasmic reticulum (SR) Ca-ATPase. To identify underlying structural features of age-modified Ca-ATPase, we have utilized the fluorescence properties of protein-bound probes to assess both local and global structure. We find conformational changes that include an age-related decrease in the apparent binding affinity to high affinity calcium sites detected by fluorescence signals in both tryptophans within nearby membrane-spanning helices and fluorescein isothiocyanate (FITC) bound distally to Lys(515) within the nucleotide-binding site. In addition, a substantial (80%) age-related increase in the accessibility to soluble quenchers of fluorescence of FITC is observed without concomitant changes in bimolecular quenching constants (k(q)) for protein-bound IAEDANS, also within the nucleotide-binding domain, and tryptophans within the membrane. Using fluorescence resonance energy transfer to measure distances between IAEDANS and FITC across the nucleotide-binding domain, we find no significant age-related change in the mean donor-acceptor distance; however, significant increases are observed in the conformational heterogeneity of this domain, as assessed by the width at half-maximum (HW) of the distance distribution, increasing with age from 29.4 +/- 0.8 A to 42.5 +/- 1. 1 A. Circular dichroism indicates that the average secondary structure is unaltered with age. Thus, these data suggest tertiary structural alterations in specific regions around the nucleotide-binding site rather than global conformational changes.

  19. The role of the sarcoplasmic reticulum in neonatal uterine smooth muscle: enhanced role compared to adult rat

    PubMed Central

    Noble, Karen; Wray, Susan

    2002-01-01

    Little is known about contractile activity, response to agonists or excitation-contraction coupling in neonatal smooth muscle. We have therefore investigated 10-day rat uterus to better understand these processes, and compared it to adult uterus to elucidate how control of contractility develops. Spontaneous contractions are present in the 10-day neonatal uterus, although they are not as large or as regular as those present in adult tissues. External Ca2+ entry via L-type Ca2+ channels is the sole source of Ca2+ and is essential for the spontaneous activity. The neonatal uterus was responsive to carbachol or prostaglandin F2α application; it showed a marked stimulation and a clear dissociation between the force and Ca2+ changes. Such sensitization was not apparent in adult rat myometrium. The sarcoplasmic reticulum (SR) had more releasable Ca2+ and contributed more to the response to agonists in neonatal compared to adult tissues. Thus, Ca2+ entry as opposed to SR Ca2+ release contributed much less to the uterine response to agonists in the neonatal, compared to adult tissues. Inhibition of the SR by cyclopiazonic acid also caused a more vigorous increase in Ca2+ and contractile activity, particularly frequency, in the neonatal compared to the adult uterus. Taken together these data suggest that: (1) spontaneous activity is already present by day 10, (2) receptor-coupling and excitation-contraction signalling pathways are functional, (3) the SR and Ca2+ sensitization mechanisms play a more prominent role in the neonate, and (4) there is a shift to a greater reliance on Ca2+ entry and excitability with development of the myometrium. PMID:12456834

  20. A role of the LIN-12/Notch signaling pathway in diversifying the non-striated egg-laying muscles in C. elegans.

    PubMed

    Hale, Jared J; Amin, Nirav M; George, Carolyn; Via, Zachary; Shi, Herong; Liu, Jun

    2014-05-15

    The proper formation and function of an organ is dependent on the specification and integration of multiple cell types and tissues. An example of this is the Caenorhabditis elegans hermaphrodite egg-laying system, which requires coordination between the vulva, uterus, neurons, and musculature. While the genetic constituents of the first three components have been well studied, little is known about the molecular mechanisms underlying the specification of the egg-laying musculature. The egg-laying muscles are non-striated in nature and consist of sixteen cells, four each of type I and type II vulval muscles and uterine muscles. These 16 non-striated muscles exhibit distinct morphology, location, synaptic connectivity and function. Using an RNAi screen targeting the putative transcription factors in the C. elegans genome, we identified a number of novel factors important for the diversification of these different types of egg-laying muscles. In particular, we found that RNAi knockdown of lag-1, which encodes the sole C. elegans ortholog of the transcription factor CSL (CBF1, Suppressor of Hairless, LAG-1), an effector of the LIN-12/Notch pathway, led to the production of extra type I vulval muscles. Similar phenotypes were also observed in animals with down-regulation of the Notch receptor LIN-12 and its DSL (Delta, Serrate, LAG-2) ligand LAG-2. The extra type I vulval muscles in animals with reduced LIN-12/Notch signaling resulted from a cell fate transformation of type II vulval muscles to type I vulval muscles. We showed that LIN-12/Notch was activated in the undifferentiated type II vulval muscle cells by LAG-2/DSL that is likely produced by the anchor cell (AC). Our findings provide additional evidence highlighting the roles of LIN-12/Notch signaling in coordinating the formation of various components of the functional C. elegans egg-laying system. We also identify multiple new factors that play critical roles in the proper specification of the different types

  1. Short-term microvascular response of striated muscle to cp-Ti, Ti-6Al-4V, and Ti-6Al-7Nb.

    PubMed

    Pennekamp, Peter H; Gessmann, Jan; Diedrich, Oliver; Burian, Björn; Wimmer, Markus A; Frauchiger, Vinzenz M; Kraft, Clayton N

    2006-03-01

    Due to excellent mechanical properties and good corrosion resistance, titanium-aluminium-vanadium (Ti-6Al-4V) and titanium-aluminium-niobium (Ti-6Al-7Nb) are extensively used for orthopedic surgery. Concern has been voiced concerning the implications of the constituent vanadium in Ti-6Al-4V on the surrounding environment. Particularly in osteosynthesis where the alloys stand in direct contact to skeletal muscle, undesirable biologic reactions may have severe consequences. In a comparative study, we assessed in vivo nutritive perfusion and leukocytic response of striated muscle to the metals Ti-6Al-4V, Ti-6Al-7Nb, and commercially pure titanium (cpTi), thereby drawing conclusions on their short-term inflammatory potential. In 28 hamsters, utilizing the dorsal skinfold chamber preparation and intravital microscopy, we quantified primary and secondary leukocyte-endothelial cell interaction, leukocyte extravasation, microvascular diameter change, and capillary perfusion in collecting and postcapillary venules of skeletal muscle. A manifest discrepancy between the metals concerning impact on local microvascular parameters was not found. All metals induced an only transient and moderate inflammatory response. Only a slight increase in leukocyte recruitment and a more sluggish recuperation of inflammatory parameters in animals treated with Ti-6Al-4V compared to the other two metals suggested a minor, overall not significant discrepancy in biocompatibility. Gross toxicity of bulk Ti-6Al-4V on surrounding tissue could not be found. Conclusively, the commonly used biomaterials Ti-6Al-4V, Ti-6Al-7Nb, and cpTi induce an only transient inflammatory answer of the skeletal muscle microvascular system. Our results indicate that on the microvascular level the tested bulk Ti-alloys and cpTi do not cause adverse biologic reactions in striated muscle.

  2. Purification of a sarcoplasmic reticulum protein that binds Ca2+ and plasma lipoproteins

    SciTech Connect

    Hofmann, S.L.; Brown, M.S.; Lee, E.; Pathak, R.K.; Anderson, R.G.; Goldstein, J.L. )

    1989-05-15

    A protein in the sarcoplasmic reticulum of rabbit skeletal and cardiac muscle was identified because of its ability to bind 125I-labeled low density lipoprotein (LDL) with high affinity after sodium dodecyl sulfate-polyacrylamide gel electrophoresis. This protein, referred to as the 165-kDa protein, is restricted to striated muscle. It was not detected in 14 other tissues, including several that contain smooth muscle, but it appears in rat L6 myoblasts when they differentiate into myocytes. Immunofluorescence and immunoelectron microscopic studies revealed that the protein is present throughout the sarcoplasmic reticulum and the terminal cisternae. It binds 45Ca2+ on nitrocellulose blots and stains metachromatically with Stains-all, a cationic dye that stains Ca2+-binding proteins. It does not appear to be a glycoprotein, and it appears slightly larger than the 160-kDa glycoprotein previously described in sarcoplasmic reticulum. The 165-kDa protein binds LDL, beta-migrating very low density lipoprotein, and a cholesterol-induced high density lipoprotein particle that contains apoprotein E as its sole apoprotein with much higher affinity than it binds high density lipoprotein. The protein is stable to boiling and to treatment with sodium dodecyl sulfate, but it becomes sensitive to these treatments when its cystine residues are reduced and alkylated. The protein was purified 1300-fold to apparent homogeneity from rabbit skeletal muscle membranes. It differs from the cell surface LDL receptor in that (1) its apparent molecular weight is not changed by reduction and alkylation; (2) it is present in Watanabe-heritable hyperlipidemic rabbits, which lack functional LDL receptors; (3) binding of lipoproteins is not inhibited by EDTA; and (4) it is located within the lumen of the sarcoplasmic reticulum where it has no access to plasma lipoproteins.

  3. Isoform composition and gene expression of thick and thin filament proteins in striated muscles of mice after 30-day space flight.

    PubMed

    Ulanova, Anna; Gritsyna, Yulia; Vikhlyantsev, Ivan; Salmov, Nikolay; Bobylev, Alexander; Abdusalamova, Zarema; Rogachevsky, Vadim; Shenkman, Boris; Podlubnaya, Zoya

    2015-01-01

    Changes in isoform composition, gene expression of titin and nebulin, and isoform composition of myosin heavy chains as well as changes in titin phosphorylation level in skeletal (m. gastrocnemius, m. tibialis anterior, and m. psoas) and cardiac muscles of mice were studied after a 30-day-long space flight onboard the Russian spacecraft "BION-M" number 1. A muscle fibre-type shift from slow-to-fast and a decrease in the content of titin and nebulin in the skeletal muscles of animals from "Flight" group was found. Using Pro-Q Diamond staining, an ~3-fold increase in the phosphorylation level of titin in m. gastrocnemius of mice from the "Flight" group was detected. The content of titin and its phosphorylation level in the cardiac muscle of mice from "Flight" and "Control" groups did not differ; nevertheless an increase (2.2 times) in titin gene expression in the myocardium of flight animals was found. The observed changes are discussed in the context of their role in the contractile activity of striated muscles of mice under conditions of weightlessness.

  4. Isoform Composition and Gene Expression of Thick and Thin Filament Proteins in Striated Muscles of Mice after 30-Day Space Flight

    PubMed Central

    Ulanova, Anna; Gritsyna, Yulia; Vikhlyantsev, Ivan; Salmov, Nikolay; Bobylev, Alexander; Abdusalamova, Zarema; Rogachevsky, Vadim; Shenkman, Boris; Podlubnaya, Zoya

    2015-01-01

    Changes in isoform composition, gene expression of titin and nebulin, and isoform composition of myosin heavy chains as well as changes in titin phosphorylation level in skeletal (m. gastrocnemius, m. tibialis anterior, and m. psoas) and cardiac muscles of mice were studied after a 30-day-long space flight onboard the Russian spacecraft “BION-M” number 1. A muscle fibre-type shift from slow-to-fast and a decrease in the content of titin and nebulin in the skeletal muscles of animals from “Flight” group was found. Using Pro-Q Diamond staining, an ~3-fold increase in the phosphorylation level of titin in m. gastrocnemius of mice from the “Flight” group was detected. The content of titin and its phosphorylation level in the cardiac muscle of mice from “Flight” and “Control” groups did not differ; nevertheless an increase (2.2 times) in titin gene expression in the myocardium of flight animals was found. The observed changes are discussed in the context of their role in the contractile activity of striated muscles of mice under conditions of weightlessness. PMID:25664316

  5. Isoform composition and gene expression of thick and thin filament proteins in striated muscles of mice after 30-day space flight.

    PubMed

    Ulanova, Anna; Gritsyna, Yulia; Vikhlyantsev, Ivan; Salmov, Nikolay; Bobylev, Alexander; Abdusalamova, Zarema; Rogachevsky, Vadim; Shenkman, Boris; Podlubnaya, Zoya

    2015-01-01

    Changes in isoform composition, gene expression of titin and nebulin, and isoform composition of myosin heavy chains as well as changes in titin phosphorylation level in skeletal (m. gastrocnemius, m. tibialis anterior, and m. psoas) and cardiac muscles of mice were studied after a 30-day-long space flight onboard the Russian spacecraft "BION-M" number 1. A muscle fibre-type shift from slow-to-fast and a decrease in the content of titin and nebulin in the skeletal muscles of animals from "Flight" group was found. Using Pro-Q Diamond staining, an ~3-fold increase in the phosphorylation level of titin in m. gastrocnemius of mice from the "Flight" group was detected. The content of titin and its phosphorylation level in the cardiac muscle of mice from "Flight" and "Control" groups did not differ; nevertheless an increase (2.2 times) in titin gene expression in the myocardium of flight animals was found. The observed changes are discussed in the context of their role in the contractile activity of striated muscles of mice under conditions of weightlessness. PMID:25664316

  6. Mechanism of chloride-dependent release of Ca2+ in the sarcoplasmic reticulum of rabbit skeletal muscle.

    PubMed Central

    Sukhareva, M; Morrissette, J; Coronado, R

    1994-01-01

    We investigated the effect of Cl- on the Ca2+ permeability of rabbit skeletal muscle junctional sarcoplasmic reticulum (SR) using 45Ca2+ fluxes and single channel recordings. In 45Ca2+ efflux experiments, the lumen of the SR was passively loaded with solutions of 150 mM univalent salt containing 5 mM 45Ca2+. Release of 45Ca2+ was measured by rapid filtration in the presence of extravesicular 0.4-0.8 microM free Ca2+ and 150 mM of the same univalent salt loaded into the SR lumen. The rate of release was 5-10 times higher when the univalent salt equilibrated across the SR-contained Cl- (Tris-Cl, choline-Cl, KCl) instead of an organic anion or other halides (gluconate-, methanesulfonate-, acetate-, HEPES-, Br-, I-). Cations (K+, Tris+) could be interchanged without a significant effect on the release rate. To determine whether Cl- stimulated ryanodine receptors, we measured the stimulation of release by ATP (5 mM total) and caffeine (20 mM total) and the inhibition by Mg2+ (0.8 mM estimated free) in Cl(-)-free and Cl(-)-containing solutions. The effects of ATP, caffeine, and Mg2+ were the largest in K-gluconate and Tris-gluconate, intermediate in KCl, and notably poor or absent in choline-Cl and Tris-Cl. Procaine (10 mM) inhibited the caffeine-stimulated release measured in K-gluconate, whereas the Cl- channel blocker clofibric acid (10 mM) but not procaine inhibited the caffeine-insensitive release measured in choline-Cl. Ruthenium red (20 microM) inhibited release in all solutions. In SR fused to planar bilayers we identified a nonselective Cl- channel (PCl: PTris: PCa = 1:0.5:0.3) blocked by ruthenium red and clofibric acid but not by procaine. These conductive and pharmacological properties suggested the channel was likely to mediate Cl(-)-dependent SR Ca2+ release. The absence of a contribution of ryanodine receptors to the Cl(-)-dependent release were indicated by the lack of an effect of Cl- on the open probability of this channel, a complete block by procaine

  7. Seasonal changes in isoform composition of giant proteins of thick and thin filaments and titin (connectin) phosphorylation level in striated muscles of bears (Ursidae, Mammalia).

    PubMed

    Salmov, N N; Vikhlyantsev, I M; Ulanova, A D; Gritsyna, Yu V; Bobylev, A G; Saveljev, A P; Makariushchenko, V V; Maksudov, G Yu; Podlubnaya, Z A

    2015-03-01

    Seasonal changes in the isoform composition of thick and thin filament proteins (titin, myosin heavy chains (MyHCs), nebulin), as well as in the phosphorylation level of titin in striated muscles of brown bear (Ursus arctos) and hibernating Himalayan black bear (Ursus thibetanus ussuricus) were studied. We found that the changes that lead to skeletal muscle atrophy in bears during hibernation are not accompanied by a decrease in the content of nebulin and intact titin-1 (T1) isoforms. However, a decrease (2.1-3.4-fold) in the content of T2 fragments of titin was observed in bear skeletal muscles (m. gastrocnemius, m. longissimus dorsi, m. biceps) during hibernation. The content of the stiffer N2B titin isoform was observed to increase relative to the content of its more compliant N2BA isoform in the left ventricles of hibernating bears. At the same time, in spite of the absence of decrease in the total content of T1 in the myocardium of hibernating brown bear, the content of T2 fragments decreased ~1.6-fold. The level of titin phosphorylation only slightly increased in the cardiac muscle of hibernating brown bear. In the skeletal muscles of brown bear, the level of titin phosphorylation did not vary between seasons. However, changes in the composition of MyHCs aimed at increasing the content of slow (I) and decreasing the content of fast (IIa) isoforms of this protein during hibernation of brown bear were detected. Content of MyHCs I and IIa in the skeletal muscles of hibernating Himalayan black bear corresponded to that in the skeletal muscles of hibernating brown bear.

  8. Rapid purification of calsequestrin from cardiac and skeletal muscle sarcoplasmic reticulum vesicles by Ca2+-dependent elution from phenyl-sepharose.

    PubMed

    Cala, S E; Jones, L R

    1983-10-10

    Treatment of cardiac or skeletal muscle sarcoplasmic reticulum vesicles with 0.1 M sodium carbonate selectively extracts both the Ca2+-binding protein calsequestrin and the two "intrinsic glycoproteins," while leaving the Ca2+-dependent ATPase membrane bound. Phenyl-Sepharose chromatography in the presence of ethylene glycol bis(beta-aminoethyl ether)-N,N,N',N'-tetraacetic acid (EGTA) and high salt (0.5 M NaCl) readily fractionates these solubilized proteins into a Ca2+-elutable fraction, which contains purified calsequestrin, and a low ionic strength elutable fraction, which contains one of the two intrinsic glycoproteins. Elution of calsequestrin from phenyl-Sepharose occurs near 1 mM Ca2+. Copurifying with calsequestrin are an homologous set of high molecular weight proteins, which like calsequestrin stain blue with Stains-All. These proteins are present in trace amounts and do not correspond to any sarcoplasmic reticulum proteins previously identified. Elution of calsequestrin from phenyl-Sepharose is consistent with the Ca2+-binding protein losing its hydrophobic character in the presence of millimolar Ca2+. This behavior is converse to that observed for several calmodulin-like proteins, which are eluted from hydrophobic gels in the presence of EGTA. The high yield and purity of calsequestrin prepared by this method makes possible a unique system for studying what may be a distinct class of Ca2+-binding proteins.

  9. Manipulation of mtDNA heteroplasmy in all striated muscles of newborn mice by AAV9-mediated delivery of a mitochondria-targeted restriction endonuclease.

    PubMed

    Bacman, S R; Williams, S L; Duan, D; Moraes, C T

    2012-11-01

    Mitochondrial diseases are frequently caused by heteroplasmic mitochondrial DNA (mtDNA) mutations. As these mutations express themselves only at high relative ratios, any approach able to manipulate mtDNA heteroplasmy can potentially be curative. In this study, we developed a system to manipulate mtDNA heteroplasmy in all skeletal muscles from neonate mice. We selected muscle because it is one of the most clinically affected tissues in mitochondrial disorders. A mitochondria-targeted restriction endonuclease (mito-ApaLI) expressed from AAV9 particles was delivered either by intraperitoneal or intravenous injection in neonate mice harboring two mtDNA haplotypes, only one of which was susceptible to ApaLI digestion. A single injection was able to elicit a predictable and marked change in mtDNA heteroplasmy in all striated muscles analyzed, including heart. No health problems or reduction in mtDNA levels were observed in treated mice, suggesting that this approach could have clinical applications for mitochondrial myopathies.

  10. Identification of striated muscle activator of Rho signaling (STARS) as a novel calmodulin target by a newly developed genome-wide screen.

    PubMed

    Furuya, Yusui; Denda, Miwako; Sakane, Kyohei; Ogusu, Tomoko; Takahashi, Sumio; Magari, Masaki; Kanayama, Naoki; Morishita, Ryo; Tokumitsu, Hiroshi

    2016-07-01

    To search for novel target(s) of the Ca(2+)-signaling transducer, calmodulin (CaM), we performed a newly developed genome-wide CaM interaction screening of 19,676 GST-fused proteins expressed in human. We identified striated muscle activator of Rho signaling (STARS) as a novel CaM target and characterized its CaM binding ability and found that the Ca(2+)/CaM complex interacted stoichiometrically with the N-terminal region (Ala13-Gln35) of STARS in vitro as well as in living cells. Mutagenesis studies identified Ile20 and Trp33 as the essential hydrophobic residues in CaM anchoring. Furthermore, the CaM binding deficient mutant (Ile20Ala, Trp33Ala) of STARS further enhanced its stimulatory effect on SRF-dependent transcriptional activation. These results suggest a connection between Ca(2+)-signaling via excitation-contraction coupling and the regulation of STARS-mediated gene expression in muscles.

  11. Response of the JAK-STAT pathway to mammalian hibernation in 13-lined ground squirrel striated muscle.

    PubMed

    Logan, Samantha M; Tessier, Shannon N; Tye, Joann; Storey, Kenneth B

    2016-03-01

    Over the course of the torpor-arousal cycle, hibernators must make behavioral, physiological, and molecular rearrangements in order to keep a very low metabolic rate and retain organ viability. 13-lined ground squirrels (Ictidomys tridecemlineatus) remain immobile during hibernation, and although the mechanisms of skeletal muscle survival are largely unknown, studies have shown minimal muscle loss in hibernating organisms. Additionally, the ground squirrel heart undergoes cold-stress, reversible cardiac hypertrophy, and ischemia-reperfusion without experiencing fatal impairment. This study examines the role of the Janus kinase-signal transducer and activator of transcription (JAK-STAT) signaling pathway in the regulation of cell stress in cardiac and skeletal muscles, comparing euthermic and hibernating ground squirrels. Immunoblots showed a fivefold decrease in JAK3 expression during torpor in skeletal muscle, along with increases in STAT3 and 5 phosphorylation and suppressors of cytokine signaling-1 (SOCS1) protein levels. Immunoblots also showed coordinated increases in STAT1, 3 and 5 phosphorylation and STAT1 inhibitor protein expression in cardiac muscle during torpor. PCR analysis revealed that the activation of these pro-survival signaling cascades did not result in coordinate changes in downstream genes such as anti-apoptotic B-cell lymphoma-2 (Bcl-2) family gene expression. Overall, these results indicate activation of the JAK-STAT pathway in both cardiac and skeletal muscles, suggesting a response to cellular stress during hibernation. PMID:26885984

  12. Response of the JAK-STAT pathway to mammalian hibernation in 13-lined ground squirrel striated muscle.

    PubMed

    Logan, Samantha M; Tessier, Shannon N; Tye, Joann; Storey, Kenneth B

    2016-03-01

    Over the course of the torpor-arousal cycle, hibernators must make behavioral, physiological, and molecular rearrangements in order to keep a very low metabolic rate and retain organ viability. 13-lined ground squirrels (Ictidomys tridecemlineatus) remain immobile during hibernation, and although the mechanisms of skeletal muscle survival are largely unknown, studies have shown minimal muscle loss in hibernating organisms. Additionally, the ground squirrel heart undergoes cold-stress, reversible cardiac hypertrophy, and ischemia-reperfusion without experiencing fatal impairment. This study examines the role of the Janus kinase-signal transducer and activator of transcription (JAK-STAT) signaling pathway in the regulation of cell stress in cardiac and skeletal muscles, comparing euthermic and hibernating ground squirrels. Immunoblots showed a fivefold decrease in JAK3 expression during torpor in skeletal muscle, along with increases in STAT3 and 5 phosphorylation and suppressors of cytokine signaling-1 (SOCS1) protein levels. Immunoblots also showed coordinated increases in STAT1, 3 and 5 phosphorylation and STAT1 inhibitor protein expression in cardiac muscle during torpor. PCR analysis revealed that the activation of these pro-survival signaling cascades did not result in coordinate changes in downstream genes such as anti-apoptotic B-cell lymphoma-2 (Bcl-2) family gene expression. Overall, these results indicate activation of the JAK-STAT pathway in both cardiac and skeletal muscles, suggesting a response to cellular stress during hibernation.

  13. Multiple effects of ryanodine on intracellular free Ca2+ in smooth muscle cells from bovine and porcine coronary artery: modulation of sarcoplasmic reticulum function.

    PubMed Central

    Wagner-Mann, C.; Hu, Q.; Sturek, M.

    1992-01-01

    1. The effects of ryanodine and caffeine on intracellular free Ca2+ concentration ([Ca2+]i) were studied by use of fura-2 microfluorometry in single smooth muscle cells freshly dispersed from bovine and porcine coronary artery. 2. Bovine and porcine cells demonstrated similar sensitivities to 10 min of exposure to ryanodine in physiological salt solution (PSS), as determined by comparable dose-dependent decreases in the subsequent [Ca2+]i transient induced by 5 mM caffeine. 3. Ryanodine (10 microM) caused a significant increase in [Ca2+]i to a plateau level 27 +/- 3% and 38 +/- 4% above baseline [Ca2+]i (baseline [Ca2+]i = [Ca2+]i at 0 min) in porcine and bovine cells, respectively, when bathed in PSS. In bovine cells the time required to reach 1/2 the plateau level was only 3 min versus 6 min for porcine cells. 4. The ryanodine-induced plateau increase in [Ca2+]i was 35 +/- 5% above baseline for bovine cells bathed in 0 Ca PSS (PSS including 10 microM EGTA with no added Ca2+), but only 7 +/- 3% above baseline in porcine cells during 10 min exposure to 10 microM ryanodine. In bovine cells [Ca2+]i showed proportional increases when extracellular Ca2+ was increased from the normal 2 mM Ca2+ PSS to 5 and 10 mM. 5. Cells pretreated with caffeine in 0 Ca PSS, which depleted the caffeine-sensitive sarcoplasmic reticulum Ca2+ store, showed no increase in [Ca2+]i when challenged with 10 microM ryanodine. The ryanodine-associated increase in [Ca2+]i, which was sustained in 0 Ca PSS during the 10 min ryanodine exposure in cells not pretreated with caffeine, suggests that ryanodine releases Ca2+ from the sarcoplasmic reticulum, but also inhibits Ca2+ efflux.(ABSTRACT TRUNCATED AT 250 WORDS) Images Figure 1 PMID:1504718

  14. Multi-Tasking Role of the Mechanosensing Protein Ankrd2 in the Signaling Network of Striated Muscle

    PubMed Central

    Mittempergher, Lorenza; Campanaro, Stefano; Martinelli, Valentina C.; Mouly, Vincent; Valle, Giorgio; Kojic, Snezana; Faulkner, Georgine

    2011-01-01

    Background Ankrd2 (also known as Arpp) together with Ankrd1/CARP and DARP are members of the MARP mechanosensing proteins that form a complex with titin (N2A)/calpain 3 protease/myopalladin. In muscle, Ankrd2 is located in the I-band of the sarcomere and moves to the nucleus of adjacent myofibers on muscle injury. In myoblasts it is predominantly in the nucleus and on differentiation shifts from the nucleus to the cytoplasm. In agreement with its role as a sensor it interacts both with sarcomeric proteins and transcription factors. Methodology/Principal Findings Expression profiling of endogenous Ankrd2 silenced in human myotubes was undertaken to elucidate its role as an intermediary in cell signaling pathways. Silencing Ankrd2 expression altered the expression of genes involved in both intercellular communication (cytokine-cytokine receptor interaction, endocytosis, focal adhesion, tight junction, gap junction and regulation of the actin cytoskeleton) and intracellular communication (calcium, insulin, MAPK, p53, TGF-β and Wnt signaling). The significance of Ankrd2 in cell signaling was strengthened by the fact that we were able to show for the first time that Nkx2.5 and p53 are upstream effectors of the Ankrd2 gene and that Ankrd1/CARP, another MARP member, can modulate the transcriptional ability of MyoD on the Ankrd2 promoter. Another novel finding was the interaction between Ankrd2 and proteins with PDZ and SH3 domains, further supporting its role in signaling. It is noteworthy that we demonstrated that transcription factors PAX6, LHX2, NFIL3 and MECP2, were able to bind both the Ankrd2 protein and its promoter indicating the presence of a regulatory feedback loop mechanism. Conclusions/Significance In conclusion we demonstrate that Ankrd2 is a potent regulator in muscle cells affecting a multitude of pathways and processes. PMID:22016770

  15. Molluscan visceral muscle fine structure. General structure and sarcolemmal organization in the smooth muscle of the intestinal wall of Buccinum undatum L.

    PubMed

    Hunt, S

    1981-01-01

    Fine structure of intestinal muscle in the gastropod Buccinum undatum is described. Myofibrillar organization is typical of non-pseudostriated molluscan muscles. The dense body system is poorly developed but there are extensive attachments plaques. The sarcolemma is elaborately modified. Deep infoldings of the membrane give the cells an irregular outline. Such infoldings enclose extracellular matrix and are associated with attachment plaques. Arising from these and from the general sarcolemma are numerous tubular membranous invaginations ending blindly at varying depth in the sarcoplasm. These structures have a helical coat of particles on the cytoplasmic face. Associated with both types of invagination are subsarcolemmal vesicles. The possibility that the tubular invaginations are analogues of vertebrate smooth muscle caveolae or striated muscle T-tubules and that the vesicles are the corresponding sarcoplasmic reticulum is discussed. The occurrence of such structures in molluscan muscle and elsewhere is reviewed.

  16. Interaction between endoplasmic/sarcoplasmic reticulum stress (ER/SR stress), mitochondrial signaling and Ca(2+) regulation in airway smooth muscle (ASM).

    PubMed

    Delmotte, Philippe; Sieck, Gary C

    2015-02-01

    Airway inflammation is a key aspect of diseases such as asthma. Several inflammatory cytokines (e.g., TNFα and IL-13) increase cytosolic Ca(2+) ([Ca(2+)]cyt) responses to agonist stimulation and Ca(2+) sensitivity of force generation, thereby enhancing airway smooth muscle (ASM) contractility (hyper-reactive state). Inflammation also induces ASM proliferation and remodeling (synthetic state). In normal ASM, the transient elevation of [Ca(2+)]cyt induced by agonists leads to a transient increase in mitochondrial Ca(2+) ([Ca(2+)]mito) that may be important in matching ATP production with ATP consumption. In human ASM (hASM) exposed to TNFα and IL-13, the transient increase in [Ca(2+)]mito is blunted despite enhanced [Ca(2+)]cyt responses. We also found that TNFα and IL-13 induce reactive oxidant species (ROS) formation and endoplasmic/sarcoplasmic reticulum (ER/SR) stress (unfolded protein response) in hASM. ER/SR stress in hASM is associated with disruption of mitochondrial coupling with the ER/SR membrane, which relates to reduced mitofusin 2 (Mfn2) expression. Thus, in hASM it appears that TNFα and IL-13 result in ROS formation leading to ER/SR stress, reduced Mfn2 expression, disruption of mitochondrion-ER/SR coupling, decreased mitochondrial Ca(2+) buffering, mitochondrial fragmentation, and increased cell proliferation.

  17. Some Relations between Changes in the Linear Electrical Properties of Striated Muscle Fibers and Changes in Ultrastructure

    PubMed Central

    Freygang, W. H.; Rapoport, S. I.; Peachey, L. D.

    1967-01-01

    Some of the linear electrical properties of frog sartorius muscle have been investigated in Ringer's fluid and in a Ringer fluid made hypertonic by the addition of sucrose or NaCl. Electrical constants were determined from measurements of the phase angle of the admittance of a fiber for an applied alternating current, from measurements of the voltage induced by an inward pulse of current, and from measurements of the conduction velocity of the action potential and the time constant of its foot. The dilation of the transverse tubular system induced by the sucrose hypertonic Ringer fluid was correlated with the change in the electrical constants. From this it is concluded that a two time constant equivalent circuit for the membrane, as proposed by Falk and Fatt, is in good agreement with our results. Both the area of the membrane of the transverse tubular system, and the capacity (ce) attributed to it, increased in the sucrose hypertonic Ringer fluid. The resistance re, which is in series with ce, did not fall when the transverse tubular system was dilated and probably is not located in that system. PMID:6063689

  18. Site-directed mutagenesis and deletion of three phosphorylation sites of calsequestrin of skeletal muscle sarcoplasmic reticulum. Effects on intracellular targeting.

    PubMed

    Nori, A; Furlan, S; Patiri, F; Cantini, M; Volpe, P

    2000-10-10

    Calsequestrin (CS) is segregated to the junctional sarcoplasmic reticulum (jSR) of skeletal muscle fibers and is responsible for intraluminal Ca(2+) binding. A chimeric CS-HA1, obtained by adding the nine-amino-acid viral epitope hemagglutinin (HA1) to the carboxy-terminal of CS and shown to be correctly segregated to skeletal muscle jSR in vivo (A. Nori, K. A. Nadalini, A. Martini, R. Rizzuto, A. Villa, and P. Volpe, 1997, Am. J. Physiol. 272, C1420-C1428), is mutagenized in order to identify domains of CS involved in targeting. Since a putative targeting mechanism of CS implies phosphorylation-dependent steps in the endoplasmic reticulum (ER) and/or Golgi complex, five CS-HA1 mutants disrupting the three phosphorylation sites of CS (Thr(189), Thr(229), and Thr(353)) were engineered by either site-directed mutagenesis or deletion: CS-HA1DeltaP1 (Thr(189) --> Ile); CS-HA1DeltaP2 (Thr(229) --> Asn); CS-HA1DeltaP1,2; in which Thr(189) and Thr(229) were changed to Ile and Asn, respectively; and CS-HA1Delta14(COOH) and CS-HA1Delta49 (COOH), in which 14 residues (Glu(354)-Asp(367)) and 49 residues (Asp(319)-Asp(367)), respectively, were deleted at the carboxy-terminal. Mutant cDNAs were transiently transfected in either HeLa cells, cultured myoblasts of rat skeletal muscle, or regenerating soleus muscle fibers of adult rats. Each CS-HA1 mutant was identified by Western blot as a single polypeptide of the predicted molecular weight. The intracellular localization of CS-HA1 mutants was studied by immunofluorescence using specific antibodies against either CS or HA1. CS-HA1 mutants colocalized with ER markers, e.g., calreticulin, and partially overlapped with Golgi complex markers, e.g., alpha-mannosidase II, in HeLa cells and myotubes. CS-HA1 mutants were expressed and retained in ER and ER/SR of HeLa cells and myotubes, respectively, and correctly segregated to jSR of regenerating soleus muscle fibers. Thus, the targeting mechanism of CS in vivo is not affected by

  19. Expression of sarcomeric tropomyosin in striated muscles in axolotl treated with shz-1, a small cardiogenic molecule.

    PubMed

    Nan, Changlong; Dube, Syamalima; Matoq, Amr; Mikesell, Lauren; Abbott, Lynn; Alshiekh-Nasany, Ruham; Chionuma, Henry; Huang, Xupei; Poiesz, Bernard J; Dube, Dipak K

    2015-01-01

    We evaluated the effect of shz-1, a cardiogenic molecule, on the expression of various tropomyosin (TM) isoforms in the Mexican axolotl (Ambystoma mexicanum) hearts. qRT-PCR data show a ~1.5-fold increase in cardiac transcripts of the Nkx2.5 gene, which plays a crucial role in cardiogenesis in vertebrates. Shz-1 augments the expression of transcripts of the total sarcomeric TPM1 (both TPM1α & TPM1κ) and sarcomeric TPM4α. In order to understand the mechanism by which shz-1 augments the expression of sarcomeric TPM transcription in axolotl hearts, we transfected C2C12 cells with pGL3.axolotl. We transfected C2C12 cells with pGL3-axolotl TPM4 promoter constructs containing the firefly luciferase reporter gene. The transfected C2C12 cells were grown in the absence or presence of shz-1 (5 μM). Subsequently, we determined the firefly luciferase activity in the extracts of transfected cells. The results suggest that shz-1 activates the axolotl TPM4 promoter-driven ectopic expression in C2C12 cells. Also, we transfected C2C12 cells with a pGL3.1 vector containing the promoter of the mouse skeletal muscle troponin-I and observed a similar increase in the luciferase activity in shz-1-treated cells. We conclude that shz-1 activates the promoters of a variety of genes including axolotl TPM4. We have quantified the expression of the total sarcomeric TPM1 and observed a 1.5-fold increase in treated cells. Western blot analyses with CH1 monoclonal antibody specific for sarcomeric isoforms show that shz-1 does not increase the expression of TM protein in axolotl hearts, whereas it does in C2C12 cells. These findings support our hypothesis that cardiac TM expression in axolotl undergoes translational control.

  20. Expression of sarcomeric tropomyosin in striated muscles in axolotl treated with shz-1, a small cardiogenic molecule.

    PubMed

    Nan, Changlong; Dube, Syamalima; Matoq, Amr; Mikesell, Lauren; Abbott, Lynn; Alshiekh-Nasany, Ruham; Chionuma, Henry; Huang, Xupei; Poiesz, Bernard J; Dube, Dipak K

    2015-01-01

    We evaluated the effect of shz-1, a cardiogenic molecule, on the expression of various tropomyosin (TM) isoforms in the Mexican axolotl (Ambystoma mexicanum) hearts. qRT-PCR data show a ~1.5-fold increase in cardiac transcripts of the Nkx2.5 gene, which plays a crucial role in cardiogenesis in vertebrates. Shz-1 augments the expression of transcripts of the total sarcomeric TPM1 (both TPM1α & TPM1κ) and sarcomeric TPM4α. In order to understand the mechanism by which shz-1 augments the expression of sarcomeric TPM transcription in axolotl hearts, we transfected C2C12 cells with pGL3.axolotl. We transfected C2C12 cells with pGL3-axolotl TPM4 promoter constructs containing the firefly luciferase reporter gene. The transfected C2C12 cells were grown in the absence or presence of shz-1 (5 μM). Subsequently, we determined the firefly luciferase activity in the extracts of transfected cells. The results suggest that shz-1 activates the axolotl TPM4 promoter-driven ectopic expression in C2C12 cells. Also, we transfected C2C12 cells with a pGL3.1 vector containing the promoter of the mouse skeletal muscle troponin-I and observed a similar increase in the luciferase activity in shz-1-treated cells. We conclude that shz-1 activates the promoters of a variety of genes including axolotl TPM4. We have quantified the expression of the total sarcomeric TPM1 and observed a 1.5-fold increase in treated cells. Western blot analyses with CH1 monoclonal antibody specific for sarcomeric isoforms show that shz-1 does not increase the expression of TM protein in axolotl hearts, whereas it does in C2C12 cells. These findings support our hypothesis that cardiac TM expression in axolotl undergoes translational control. PMID:24958154

  1. Modulation of sarcoplasmic reticulum Ca2+ release in skeletal muscle expressing ryanodine receptor impaired in regulation by calmodulin and S100A1

    PubMed Central

    Yamaguchi, Naohiro; Prosser, Benjamin L.; Ghassemi, Farshid; Xu, Le; Pasek, Daniel A.; Eu, Jerry P.; Hernández-Ochoa, Erick O.; Cannon, Brian R.; Wilder, Paul T.; Lovering, Richard M.; Weber, David; Melzer, Werner; Schneider, Martin F.

    2011-01-01

    In vitro, calmodulin (CaM) and S100A1 activate the skeletal muscle ryanodine receptor ion channel (RyR1) at submicromolar Ca2+ concentrations, whereas at micromolar Ca2+ concentrations, CaM inhibits RyR1. One amino acid substitution (RyR1-L3625D) has previously been demonstrated to impair CaM binding and regulation of RyR1. Here we show that the RyR1-L3625D substitution also abolishes S100A1 binding. To determine the physiological relevance of these findings, mutant mice were generated with the RyR1-L3625D substitution in exon 74, which encodes the CaM and S100A1 binding domain of RyR1. Homozygous mutant mice (Ryr1D/D) were viable and appeared normal. However, single RyR1 channel recordings from Ryr1D/D mice exhibited impaired activation by CaM and S100A1 and impaired CaCaM inhibition. Isolated flexor digitorum brevis muscle fibers from Ryr1D/D mice had depressed Ca2+ transients when stimulated by a single action potential. However, during repetitive stimulation, the mutant fibers demonstrated greater relative summation of the Ca2+ transients. Consistently, in vivo stimulation of tibialis anterior muscles in Ryr1D/D mice demonstrated reduced twitch force in response to a single action potential, but greater summation of force during high-frequency stimulation. During repetitive stimulation, Ryr1D/D fibers exhibited slowed inactivation of sarcoplasmic reticulum Ca2+ release flux, consistent with increased summation of the Ca2+ transient and contractile force. Peak Ca2+ release flux was suppressed at all voltages in voltage-clamped Ryr1D/D fibers. The results suggest that the RyR1-L3625D mutation removes both an early activating effect of S100A1 and CaM and delayed suppressing effect of CaCaM on RyR1 Ca2+ release, providing new insights into CaM and S100A1 regulation of skeletal muscle excitation-contraction coupling. PMID:21289290

  2. Superoxide radicals stimulate IP sub 3 -induced Ca sup 2+ -release from vascular smooth muscle sarcoplasmic reticulum

    SciTech Connect

    Ford, G.D.; Suzuki, Y. )

    1991-03-15

    Oxygen free radicals have been implicated in a variety of pathophysiological conditions and vascular smooth muscle can be a site of damage in such oxygen toxicity. Mechanisms of the effects of these radials on the vascular smooth muscle at the cellular level, however, have not been well studied. In the present study, the authors report that the inositol 1,4,5-trisphosphate (IP{sub 3})-induced Ca{sup 2+}-release from bovine aortic SR was also affected by O{sub 2}{sup {minus}}. Hypoxanthine plus xanthine oxidase in the presence of catalase stimulated the IP{sub 3}-induced Ca{sup 2+}-release from SR monitored using arsenazo III. At 10 {mu}M IP{sub 3}, the release was doubled by O{sub 2}{sup {minus}} treatment. As a consequence of using higher SR protein concentrations required to observe the Ca{sup 2+}-uptake inhibition induced by O{sub 2}{sup {minus}}. Since the effect of O{sub 2}{sup {minus}} was not seen when a non-hydrolyzable analogue of IP{sub 3} is used to induce Ca{sup 2+}-release, O{sub 2}{sup {minus}} may be inhibiting the degradation processes of IP{sub 3} rather than having an influence on the release channel per se.

  3. Skeletal muscle

    Technology Transfer Automated Retrieval System (TEKTRAN)

    There are approximately 650-850 muscles in the human body these include skeletal (striated), smooth and cardiac muscle. The approximation is based on what some anatomists consider separate muscle or muscle systems. Muscles are classified based on their anatomy (striated vs. smooth) and if they are v...

  4. Intermolecular Interactions in the Mechanism of Skeletal Muscle Sarcoplasmic Reticulum Ca2+-ATPase (SERCA1): Evidence for a Tri-protomer*

    PubMed Central

    Mahaney, James E.; Thomas, David D.; Farrance, Iain K.; Froehlich, Jeffrey P.

    2013-01-01

    Native membrane sarcoplasmic reticulum (SR) Ca2+-ATPase isolated from skeletal muscle (SERCA1) exhibits oligomeric kinetic behavior (Mahaney, J.E., Thomas, D.D., and Froehlich, J.P. (2004) Biochemistry 43:4400–4416). In the present study we used quenched-flow mixing, electron paramagnetic resonance (EPR) and chemical cross-linking to probe for intermolecular interactions at physiological (0.1M) and high (0.4 M) KCl. Exposure SR membranes to water- and lipid-soluble cross-linking reagents revealed a mixture of SERCA1 oligomeric species consisting mainly of dimers and trimers. Titration of iodoacetamide spin labeled-SERCA1 with AMPPCP in the presence of 10 µM Ca2+ and 0.1 M KCl revealed high (KD = 45 µM) and low affinity (KD = 315 µM) nucleotide binding sites in a 2:1 ratio, respectively. Raising the [KCl] to 0.4 M increased the fraction of weak binding sites and lowered the KD of the high affinity component (20 µM). Phosphorylation by 10 µM ATP at 21°C and 0.1 M KCl produced an early burst of Pi production without a corresponding decline in the steady state phosphoenzyme (EP) level. The steady state EP level was twice as large as the Pi burst and received equal contributions from E1P and E2P. Chasing the phosphoenzyme at 0.4 M KCl and 2°C with ADP revealed a biphasic time course of E1P formation with a slow phase that matched the kinetics of the transient EPR signal from the spin-labeled Ca2+-ATPase. The absence of a fast component in the EPR signal excludes E1P as its source. Instead, it arises from the slow, KCl-dependent transformation at the start of the cycle which controls the formation of downstream intermediates with an increased mole fraction of rotationally restricted probes. We modeled this behavior with a SERCA1 trimer in which the formation of E1P/E2/E2P from E1ATP/E2P/E1P results from concerted transformations in the subunits coupling phosphorylation (E1ATP → E1P + ADP) to dephosphorylation (E2P → E2 + Pi) and the conversion of E1P to E2P

  5. Striated muscle activator of Rho signalling (STARS) is a PGC-1α/oestrogen-related receptor-α target gene and is upregulated in human skeletal muscle after endurance exercise.

    PubMed

    Wallace, Marita A; Hock, M Benjamin; Hazen, Bethany C; Kralli, Anastasia; Snow, Rod J; Russell, Aaron P

    2011-04-15

    The striated muscle activator of Rho signalling (STARS) is an actin-binding protein specifically expressed in cardiac, skeletal and smooth muscle. STARS has been suggested to provide an important link between the transduction of external stress signals to intracellular signalling pathways controlling genes involved in the maintenance of muscle function. The aims of this study were firstly, to establish if STARS, as well as members of its downstream signalling pathway, are upregulated following acute endurance cycling exercise; and secondly, to determine if STARS is a transcriptional target of peroxisome proliferator-activated receptor gamma co-activator 1-α (PGC-1α) and oestrogen-related receptor-α (ERRα). When measured 3 h post-exercise, STARS mRNA and protein levels as well as MRTF-A and serum response factor (SRF) nuclear protein content, were significantly increased by 140, 40, 40 and 40%, respectively. Known SRF target genes, carnitine palmitoyltransferase-1β (CPT-1β) and jun B proto-oncogene (JUNB), as well as the exercise-responsive genes PGC-1α mRNA and ERRα were increased by 2.3-, 1.8-, 4.5- and 2.7-fold, 3 h post-exercise. Infection of C2C12 myotubes with an adenovirus-expressing human PGC-1α resulted in a 3-fold increase in Stars mRNA, a response that was abolished following the suppression of endogenous ERRα. Over-expression of PGC-1α also increased Cpt-1β, Cox4 and Vegf mRNA by 6.2-, 2.0- and 2.0-fold, respectively. Suppression of endogenous STARS reduced basal Cpt-1β levels by 8.2-fold and inhibited the PGC-1α-induced increase in Cpt-1β mRNA. Our results show for the first time that the STARS signalling pathway is upregulated in response to acute endurance exercise. Additionally, we show in C2C12 myotubes that the STARS gene is a PGC-1α/ERRα transcriptional target. Furthermore, our results suggest a novel role of STARS in the co-ordination of PGC-1α-induced upregulation of the fat oxidative gene, CPT-1β.

  6. Effects of diet on the function of sarcoplasmic reticulum.

    PubMed

    Gould, G W; McWhirter, J M; East, J M; Lee, A G

    1987-08-01

    We have examined the effect of diet on the phospholipid composition of the sarcoplasmic reticulum of rabbit muscle. Enriching the diet with corn or fish oil results in significant changes in the fatty acyl chain composition of the various phospholipid classes, with relatively little change in the relative contents of the phospholipids. These alterations in composition have no significant effect on the ATPase activity of vesicles of sarcoplasmic reticulum or on the pattern of Ca2+ uptake and release.

  7. Striated morphology during electrodeposition

    SciTech Connect

    Jorne, J.; Lee, M.G.

    1996-03-01

    Comparative study of striated morphologies obtained during the electrodeposition of various metals is performed using rotating electrodes. The formation of striation during electrodeposition of zinc, copper, and silver follows the streamlines of the fluid, and is observed to be accompanied by small or negative polarization resistance. The spacings between striae depends strongly on the applied current density. The spacing decreases with current density in the case of zinc and silver, however, a reverse trend is observed in the case of copper. Linear stability analysis shows that spatial multiplicity occurs when ({partial_derivative}i/{partial_derivative}C){sub {eta}} < 0. The experimentally observed spacings between striae (0.1 {approximately} 1 mm) are in agreement with the wavelength obtained from the linear stability analysis. The analysis of polarization and striation data shows fairly good qualitative agreement with respect to the effects of current density, diffusion layer thickness, and electrolyte concentration on striation.

  8. Microvascular response of striated muscle to common arthroplasty-alloys: A comparative in vivo study with CoCrMo, Ti-6Al-4V, and Ti-6Al-7Nb.

    PubMed

    Kraft, Clayton N; Burian, Björn; Diedrich, Oliver; Gessmann, Jan; Wimmer, Markus A; Pennekamp, Peter H

    2005-10-01

    The impairment of skeletal muscle microcirculation by a biomaterial may have profound consequences. Due to excellent physical and corrosion characteristics, CoCrMo-, Ti-6Al-4V-, and Ti-6Al-7Nb-alloys are commonly used in orthopedic surgery. Yet concern has been raised with regard to the implications of inevitable corrosion product of these metals on the surrounding biologic environment, particularly in the case of CoCrMo. We, therefore, studied in vivo nutritive perfusion and leukocytic response of striated muscle to these alloys, thereby drawing conclusions on their inflammatory potential. In 28 hamsters, utilizing the dorsal skinfold chamber preparation and intravital microscopy, we could demonstrate that the implant material CoCrMo has a marked impact on local microvascular parameters. While the Ti-alloys Ti-6Al-4V and Ti-6Al-7Nb induced only a transient and moderate inflammatory response, the implantation of a CoCrMo sample led to a distinct and persistent activation of leukocytes combined with disruption of the microvascular endothelial integrity and marked leukocyte extravasation. Animals with Ti-alloys showed a clear tendency of recuperation, while in all but one CoCrMo-treated animals, a breakdown of microcirculation prior to the scheduled end of the experiment was observed. Overall, the alloy Ti-6Al-7Nb was tolerated slightly better than Ti-6Al-4V under the chosen test conditions, though this discrepancy was not statistically significant. Conclusively, the commonly used biomaterials Ti-6Al-7Nb and Ti-6Al-4V induce a considerably lower inflammatory response in the skeletal muscle microvascular system, compared to a CoCrMo-alloy. With a minimum of adverse host reaction, our results indicate that for this particular model Ti-alloys are better tolerated than CoCrMo implant materials.

  9. Hyperactivation of L-type voltage-gated Ca2+ channels in Caenorhabditis elegans striated muscle can result from point mutations in the IS6 or the IIIS4 segment of the α1 subunit.

    PubMed

    Lainé, Viviane; Ségor, Jean Rony; Zhan, Hong; Bessereau, Jean-Louis; Jospin, Maelle

    2014-11-01

    Several human diseases, including hypokalemic periodic paralysis and Timothy syndrome, are caused by mutations in voltage-gated calcium channels. The effects of these mutations are not always well understood, partially because of difficulties in expressing these channels in heterologous systems. The use of Caenorhabditis elegans could be an alternative approach to determine the effects of mutations on voltage-gated calcium channel function because all the main types of voltage-gated calcium channels are found in C. elegans, a large panel of mutations already exists and efficient genetic tools are available to engineer customized mutations in any gene. In this study, we characterize the effects of two gain-of-function mutations in egl-19, which encodes the L-type calcium channel α1 subunit. One of these mutations, ad695, leads to the replacement of a hydrophobic residue in the IIIS4 segment. The other mutation, n2368, changes a conserved glycine of IS6 segment; this mutation has been identified in patients with Timothy syndrome. We show that both egl-19 (gain-of-function) mutants have defects in locomotion and morphology that are linked to higher muscle tone. Using in situ electrophysiological approaches in striated muscle cells, we provide evidence that this high muscle tone is due to a shift of the voltage dependency towards negative potentials, associated with a decrease of the inactivation rate of the L-type Ca(2+) current. Moreover, we show that the maximal conductance of the Ca(2+) current is decreased in the strongest mutant egl-19(n2368), and that this decrease is correlated with a mislocalization of the channel. PMID:25214488

  10. A sulfated polysaccharide from the sarcoplasmic reticulum of sea cucumber smooth muscle is an endogenous inhibitor of the Ca(2+)-ATPase.

    PubMed

    Landeira-Fernandez, A M; Aiello, K R; Aquino, R S; Silva, L C; Meis, L d; Mourão, P A

    2000-08-01

    Vesicles derived from the endoplasmic reticulum of sea cucumber smooth muscle retain a membrane bound Ca(2+)-ATPase that is able to transport Ca(2+) into the vesicles at the expense of ATP hydrolysis. In contrast with vesicles obtained from rabbit muscles, the activity of the Ca(2+)-dependent ATPase from sea cucumber is dependent on monovalent cations (K(+)>Na(+)>Li(+)). With the addition of highly sulfated polysaccharide to vesicle preparations from rabbit muscle, Ca(2+) uptake decreases sharply and becomes highly sensitive to monovalent cations, as observed with vesicles from sea cucumber muscle. These results led us to investigate the possible occurrence of a highly sulfated polysaccharide on vesicles from the endoplasmic reticulum of sea cucumber smooth muscle, acting as an "endogenous" Ca(2+)-ATPase inhibitor. In fact, vesicles derived from the invertebrate, but not from rabbit muscle, contain a highly sulfated polysaccharide. This compound inhibits Ca(2+) uptake in vesicles obtained from rabbit muscle and the inhibition is antagonized by monovalent cation. In addition, sea cucumber muscles contain high concentrations of another polysaccharide, which surrounds the muscle fibers, and was characterized as a fucosylated chondroitin sulfate. Possibly the occurrence of sulfated polysaccharides in the sea cucumber muscles is related with unique properties of the invertebrate body wall, which can rapidly and reversibly alter its mechanical properties, with change in length by more than 200%.

  11. Immunological localization of ribosomes in striated rat muscle. Evidence for myofibrillar association and ontological changes in the subsarcolemmal:myofibrillar distribution.

    PubMed Central

    Horne, Z; Hesketh, J

    1990-01-01

    Ribosome distribution in skeletal-muscle fibres was investigated immunohistochemically by using polyclonal antibodies raised against large-ribosomal-subunit proteins isolated from rat liver. Immunoblot analysis showed the antibodies to recognize five major proteins of the large subunit; these were identified as L4, L6, L7, L15 and L17 by two-dimensional electrophoresis. Immunohistochemistry of frozen rat skeletal-muscle sections showed staining of both the subsarcolemmal and intermyofibrillar cytoplasm. A distinct banding pattern was observed, and when peroxidase and phase-contrast images of the same field were compared by image analysis the anti-ribosome staining was found to correspond to the A-bands. These results suggest that a proportion of muscle ribosomes are present in the myofibrillar cytoplasm in a regular fashion, possibly associated with myosin. Densitometric analysis of the peroxidase immunostaining showed that the ratio of myofibrillar to sub-sarcolemmal ribosomal material was lower in muscle from 51-day-old rats compared with those from 14-day-old animals. Images Fig. 3. Fig. 4. Fig. 5. Fig. 6. PMID:2188648

  12. [Electric activity of striated muscles of the cervical part of the rabbit esophagus under conditions of hunger, food intake and satiation].

    PubMed

    Kromin, A A

    1990-07-01

    The experiments of free-moving rabbits have shown that the muscles of the proximal zone in the esophageal cervical part functions as the superior esophageal sphincter. In hunger stage the motor unit activity of the sphincter has regular low-amplitude discharge with monomodal distribution of interspike intervals. The process of food satisfaction leads to the appearance of burst-like unit activity with bimodal distribution of interspike intervals. During the food intake reorganization of the motor unit activity of the esophageal cervical part is manifested in characteristic patterns of interspike interval distribution.

  13. Effects of type 1 diabetes, sprint training and sex on skeletal muscle sarcoplasmic reticulum Ca2+ uptake and Ca2+-ATPase activity.

    PubMed

    Harmer, A R; Ruell, P A; Hunter, S K; McKenna, M J; Thom, J M; Chisholm, D J; Flack, J R

    2014-02-01

    Calcium cycling is integral to muscle performance during the rapid muscle contraction and relaxation of high-intensity exercise. Ca(2+) handling is altered by diabetes mellitus, but has not previously been investigated in human skeletal muscle. We investigated effects of high-intensity exercise and sprint training on skeletal muscle Ca(2+) regulation among men and women with type 1 diabetes (T1D, n = 8, 3F, 5M) and matched non-diabetic controls (CON, n = 8, 3F, 5M). Secondarily, we examined sex differences in Ca(2+) regulation. Subjects undertook 7 weeks of three times-weekly cycle sprint training. Before and after training, performance was measured, and blood and muscle were sampled at rest and after high-intensity exercise. In T1D, higher Ca(2+)-ATPase activity (+28%) and Ca(2+) uptake (+21%) than in CON were evident across both times and days (P < 0.05), but performance was similar. In T1D, resting Ca(2+)-ATPase activity correlated with work performed until exhaustion (r = 0.7, P < 0.01). Ca(2+)-ATPase activity, but not Ca(2+) uptake, was lower (-24%, P < 0.05) among the women across both times and days. Intense exercise did not alter Ca(2+)-ATPase activity in T1D or CON. However, sex differences were evident: Ca(2+)-ATPase was reduced with exercise among men but increased among women across both days (time × sex interaction, P < 0.05). Sprint training reduced Ca(2+)-ATPase (-8%, P < 0.05), but not Ca(2+) uptake, in T1D and CON. In summary, skeletal muscle Ca(2+) resequestration capacity was increased in T1D, but performance was not greater than CON. Sprint training reduced Ca(2+)-ATPase in T1D and CON. Sex differences in Ca(2+)-ATPase activity were evident and may be linked with fibre type proportion differences.

  14. Types of muscle tissue (image)

    MedlinePlus

    The 3 types of muscle tissue are cardiac, smooth, and skeletal. Cardiac muscle cells are located in ... heart, appear striated, and are under involuntary control. Smooth muscle fibers are located in walls of hollow ...

  15. Sarcoplasmic reticulum Ca2+ ATPase pump is a major regulator of glucose transport in the healthy and diabetic heart.

    PubMed

    Waller, Amanda P; Kalyanasundaram, Anuradha; Hayes, Summer; Periasamy, Muthu; Lacombe, Véronique A

    2015-05-01

    Despite intensive research, the pathways that mediate calcium (Ca(2+))-stimulated glucose transport in striated muscle remain elusive. Since the sarcoplasmic reticulum calcium ATPase (SERCA) pump tightly regulates cytosolic [Ca(2+)], we investigated whether the SERCA pump is a major regulator of cardiac glucose transport. We used healthy and insulin-deficient diabetic transgenic (TG) mice expressing SERCA1a in the heart. Active cell surface glucose transporter (GLUT)-4 was measured by a biotinylated photolabeled assay in the intact perfused myocardium and isolated myocytes. In healthy TG mice, cardiac-specific SERCA1a expression increased active cell-surface GLUT4 and glucose uptake in the myocardium, as well as whole body glucose tolerance. Diabetes reduced active cell-surface GLUT4 content and glucose uptake in the heart of wild type mice, all of which were preserved in diabetic TG mice. Decreased basal AS160 and increased proportion of calmodulin-bound AS160 paralleled the increase in cell surface GLUT4 content in the heart of TG mice, suggesting that AS160 regulates GLUT trafficking by a Ca(2+)/calmodulin dependent pathway. In addition, cardiac-specific SERCA1a expression partially rescues hyperglycemia during diabetes. Collectively, these data suggested that the SERCA pump is a major regulator of cardiac glucose transport by an AS160 dependent mechanism during healthy and insulin-deficient state. Our data further indicated that cardiac-specific SERCA overexpression rescues diabetes induced-alterations in cardiac glucose transport and improves whole body glucose homeostasis. Therefore, findings from this study provide novel mechanistic insights linking upregulation of the SERCA pump in the heart as a potential therapeutic target to improve glucose metabolism during diabetes.

  16. Differential abundance of sarcoplasmic proteome explains animal effect on beef Longissimus lumborum color stability

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The sarcoplasmic proteome of beef Longissimus lumborum demonstrating animal-to-animal variation in color stability was examined to correlate proteome profile with color. Longissimus lumborum (36 h post-mortem) muscles were obtained from 73 beef carcasses, aged for 13 days, and fabricated to 2.5-cm s...

  17. Strategies to Study Desmin in Cardiac Muscle and Culture Systems.

    PubMed

    Diokmetzidou, Antigoni; Tsikitis, Mary; Nikouli, Sofia; Kloukina, Ismini; Tsoupri, Elsa; Papathanasiou, Stamatis; Psarras, Stelios; Mavroidis, Manolis; Capetanaki, Yassemi

    2016-01-01

    Intermediate filament (IF) cytoskeleton comprises the fine-tuning cellular machinery regulating critical homeostatic mechanisms. In skeletal and cardiac muscle, deficiency or disturbance of the IF network leads to severe pathology, particularly in the latter. The three-dimensional scaffold of the muscle-specific IF protein desmin interconnects key features of the cardiac muscle cells, including the Z-disks, intercalated disks, plasma membrane, nucleus, mitochondria, lysosomes, and potentially sarcoplasmic reticulum. This is crucial for the highly organized striated muscle, in which effective energy production and transmission as well as mechanochemical signaling are tightly coordinated among the organelles and the contractile apparatus. The role of desmin and desmin-associated proteins in the biogenesis, trafficking, and organelle function, as well as the development, differentiation, and survival of the cardiac muscle begins to be enlightened, but the precise mechanisms remain elusive. We propose a set of experimental tools that can be used, in vivo and in vitro, to unravel crucial new pathways by which the IF cytoskeleton facilitates proper organelle function, homeostasis, and cytoprotection and further understand how its disturbance and deficiency lead to disease.

  18. Strategies to Study Desmin in Cardiac Muscle and Culture Systems.

    PubMed

    Diokmetzidou, Antigoni; Tsikitis, Mary; Nikouli, Sofia; Kloukina, Ismini; Tsoupri, Elsa; Papathanasiou, Stamatis; Psarras, Stelios; Mavroidis, Manolis; Capetanaki, Yassemi

    2016-01-01

    Intermediate filament (IF) cytoskeleton comprises the fine-tuning cellular machinery regulating critical homeostatic mechanisms. In skeletal and cardiac muscle, deficiency or disturbance of the IF network leads to severe pathology, particularly in the latter. The three-dimensional scaffold of the muscle-specific IF protein desmin interconnects key features of the cardiac muscle cells, including the Z-disks, intercalated disks, plasma membrane, nucleus, mitochondria, lysosomes, and potentially sarcoplasmic reticulum. This is crucial for the highly organized striated muscle, in which effective energy production and transmission as well as mechanochemical signaling are tightly coordinated among the organelles and the contractile apparatus. The role of desmin and desmin-associated proteins in the biogenesis, trafficking, and organelle function, as well as the development, differentiation, and survival of the cardiac muscle begins to be enlightened, but the precise mechanisms remain elusive. We propose a set of experimental tools that can be used, in vivo and in vitro, to unravel crucial new pathways by which the IF cytoskeleton facilitates proper organelle function, homeostasis, and cytoprotection and further understand how its disturbance and deficiency lead to disease. PMID:26795479

  19. Occurrence and Characteristics of a Rapid Exchange of Phosphate Oxygens Catalyzed by Sarcoplasmic Reticulum Vesicles

    DOE R&D Accomplishments Database

    Kanazawa, T.; Boyer, P. D.

    1972-01-01

    Sarcoplasmic reticulum vesicles isolated from skeletal muscle actively take up Ca{sup ++} from the medium in the presence of Mg{sup ++} and ATP. This transport is coupled to ATP hydrolysis catalyzed by membrane-bound Ca{sup++}, Mg{sup ++}-ATPase which is activated by concurrent presence of Ca{sup ++} and Mg{sup ++}. Considerable informations have accumulated that give insight into the ATPase and its coupling to the calcium transport. The hydrolysis of ATP by this enzyme occurs through a phosphorylated intermediate. Formation and decomposition of the intermediate show vectorial requirements for Ca{sup ++} and Mg{sup ++}, suggesting an intimate involvement of the intermediate in the transport process. ATP synthesis from P{sub i} and ADP coupled to outflow of Ca{sup ++} from sarcoplasmic reticulum vesicles has recently been demonstrated. This indicates the reversibility of the entire process of calcium transport in sarcoplasmic reticulum vesicles.

  20. Obscurin: a multitasking muscle giant.

    PubMed

    Kontrogianni-Konstantopoulos, Aikaterini; Bloch, Robert J

    2005-01-01

    Obscurin (approximately 800 kDa) is the third member of a family of giant proteins expressed in vertebrate striated muscle, along with titin (3-3.7 MDa) and nebulin (approximately 800 kDa). Like its predecessors, it is a multidomain protein composed of tandem adhesion modules and signaling domains. Unlike titin and nebulin, which are integral components of sarcomeres, obscurin is concentrated at the peripheries of Z-disks and M-lines, where it is appropriately positioned to communicate with the surrounding myoplasm. This unique distribution allows obscurin to bind small ankyrin 1, an integral component of the sarcoplasmic reticulum (SR) membrane. Obscurin also associates with the contractile apparatus through its binding to titin, sarcomeric myosin and perhaps other proteins of the contractile apparatus. Overexpression of the COOH-terminus of obscurin in primary myotubes has a dramatic and specific effect on the organization of sarcomeric myosin, indicating a role in the organization and regular assembly of A-bands. Given its ability to associate tightly, selectively and periodically with the periphery of the myofibril, its high affinity for an integral membrane protein of the SR and its close association with thick filaments, we speculate that obscurin is ideally suited to play key roles in modulating the organization and assembly of both the myofibril and the SR.

  1. Striated Acto-Myosin Fibers Can Reorganize and Register in Response to Elastic Interactions with the Matrix

    PubMed Central

    Friedrich, Benjamin M.; Buxboim, Amnon; Discher, Dennis E.; Safran, Samuel A.

    2011-01-01

    The remarkable striation of muscle has fascinated many for centuries. In developing muscle cells, as well as in many adherent, nonmuscle cell types, striated, stress fiberlike structures with sarcomere-periodicity tend to register: Based on several studies, neighboring, parallel fibers at the basal membrane of cultured cells establish registry of their respective periodic sarcomeric architecture, but, to our knowledge, the mechanism has not yet been identified. Here, we propose for cells plated on an elastic substrate or adhered to a neighboring cell, that acto-myosin contractility in striated fibers close to the basal membrane induces substrate strain that gives rise to an elastic interaction between neighboring striated fibers, which in turn favors interfiber registry. Our physical theory predicts a dependence of interfiber registry on externally controllable elastic properties of the substrate. In developing muscle cells, registry of striated fibers (premyofibrils and nascent myofibrils) has been suggested as one major pathway of myofibrillogenesis, where it precedes the fusion of neighboring fibers. This suggests a mechanical basis for the optimal myofibrillogenesis on muscle-mimetic elastic substrates that was recently observed by several groups in cultures of mouse-, human-, and chick-derived muscle cells. PMID:21641316

  2. Muscle Giants: Molecular Scaffolds in Sarcomerogenesis

    PubMed Central

    KONTROGIANNI-KONSTANTOPOULOS, AIKATERINI; ACKERMANN, MAEGEN A.; BOWMAN, AMBER L.; YAP, SOLOMON V.; BLOCH, ROBERT J.

    2011-01-01

    Myofibrillogenesis in striated muscles is a highly complex process that depends on the coordinated assembly and integration of a large number of contractile, cytoskeletal, and signaling proteins into regular arrays, the sarcomeres. It is also associated with the stereotypical assembly of the sarcoplasmic reticulum and the transverse tubules around each sarcomere. Three giant, muscle-specific proteins, titin (3–4 MDa), nebulin (600–800 kDa), and obscurin (~720–900 kDa), have been proposed to play important roles in the assembly and stabilization of sarcomeres. There is a large amount of data showing that each of these molecules interacts with several to many different protein ligands, regulating their activity and localizing them to particular sites within or surrounding sarcomeres. Consistent with this, mutations in each of these proteins have been linked to skeletal and cardiac myopathies or to muscular dystrophies. The evidence that any of them plays a role as a “molecular template,” “molecular blueprint,” or “molecular ruler” is less definitive, however. Here we review the structure and function of titin, nebulin, and obscurin, with the literature supporting a role for them as scaffolding molecules and the contradictory evidence regarding their roles as molecular guides in sarcomerogenesis. PMID:19789381

  3. Objective forensic analysis of striated, quasi-striated and impressed toolmarks

    NASA Astrophysics Data System (ADS)

    Spotts, Ryan E.

    Following the 1993 Daubert v. Merrell Dow Pharmaceuticals, Inc. court case and continuing to the 2010 National Academy of Sciences report, comparative forensic toolmark examination has received many challenges to its admissibility in court cases and its scientific foundations. Many of these challenges deal with the subjective nature in determining whether toolmarks are identifiable. This questioning of current identification methods has created a demand for objective methods of identification - "objective" implying known error rates and statistically reliability. The demand for objective methods has resulted in research that created a statistical algorithm capable of comparing toolmarks to determine their statistical similarity, and thus the ability to separate matching and nonmatching toolmarks. This was expanded to the creation of virtual toolmarking (characterization of a tool to predict the toolmark it will create). The statistical algorithm, originally designed for two-dimensional striated toolmarks, had been successfully applied to striated screwdriver and quasi-striated plier toolmarks. Following this success, a blind study was conducted to validate the virtual toolmarking capability using striated screwdriver marks created at various angles of incidence. Work was also performed to optimize the statistical algorithm by implementing means to ensure the algorithm operations were constrained to logical comparison regions (e.g. the opposite ends of two toolmarks do not need to be compared because they do not coincide with each other). This work was performed on quasi-striated shear cut marks made with pliers - a previously tested, more difficult application of the statistical algorithm that could demonstrate the difference in results due to optimization. The final research conducted was performed with pseudostriated impression toolmarks made with chisels. Impression marks, which are more complex than striated marks, were analyzed using the algorithm to separate

  4. Hyperthyroidism increases the uncoupled ATPase activity and heat production by the sarcoplasmic reticulum Ca2+-ATPase.

    PubMed Central

    Arruda, Ana Paula; Da-Silva, Wagner S; Carvalho, Denise P; De Meis, Leopoldo

    2003-01-01

    The sarcoplasmic reticulum Ca2+-ATPase is able to modulate the distribution of energy released during ATP hydrolysis, so that a portion of energy is used for Ca2+ transport (coupled ATPase activity) and a portion is converted into heat (uncoupled ATPase activity). In this report it is shown that T4 administration to rabbits promotes an increase in the rates of both the uncoupled ATPase activity and heat production in sarcoplasmic reticulum vesicles, and that the degree of activation varies depending on the muscle type used. In white muscles hyperthyroidism promotes a 0.8-fold increase of the uncoupled ATPase activity and in red muscle a 4-fold increase. The yield of vesicles from hyperthyroid muscles is 3-4-fold larger than that obtained from normal muscles; thus the rate of heat production by the Ca2+-ATPase expressed in terms of g of muscle in hyperthyroidism is increased by a factor of 3.6 in white muscles and 12.0 in red muscles. The data presented suggest that the Ca2+-ATPase uncoupled activity may represent one of the heat sources that contributes to the enhanced thermogenesis noted in hyperthyroidism. PMID:12887329

  5. Localisation of AMPK γ subunits in cardiac and skeletal muscles.

    PubMed

    Pinter, Katalin; Grignani, Robert T; Watkins, Hugh; Redwood, Charles

    2013-12-01

    The trimeric protein AMP-activated protein kinase (AMPK) is an important sensor of energetic status and cellular stress, and mutations in genes encoding two of the regulatory γ subunits cause inherited disorders of either cardiac or skeletal muscle. AMPKγ2 mutations cause hypertrophic cardiomyopathy with glycogen deposition and conduction abnormalities; mutations in AMPKγ3 result in increased skeletal muscle glycogen. In order to gain further insight into the roles of the different γ subunits in muscle and into possible disease mechanisms, we localised the γ2 and γ3 subunits, along with the more abundant γ1 subunit, by immunofluorescence in cardiomyocytes and skeletal muscle fibres. The predominant cardiac γ2 variant, γ2-3B, gave a striated pattern in cardiomyocytes, aligning with the Z-disk but with punctate staining similar to T-tubule (L-type Ca(2+) channel) and sarcoplasmic reticulum (SERCA2) markers. In skeletal muscle fibres AMPKγ3 localises to the I band, presenting a uniform staining that flanks the Z-disk, also coinciding with the position of Ca(2+) influx in these muscles. The localisation of γ2-3B- and γ3-containing AMPK suggests that these trimers may have similar functions in the different muscles. AMPK containing γ2-3B was detected in oxidative skeletal muscles which had low expression of γ3, confirming that these two regulatory subunits may be co-ordinately regulated in response to metabolic requirements. Compartmentalisation of AMPK complexes is most likely dependent on the regulatory γ subunit and this differential localisation may direct substrate selection and specify particular functional roles.

  6. Effects of temperature and buffer composition on calcium sequestration by sarcoplasmic reticulum and plasma membrane of rabbit renal artery

    SciTech Connect

    McGuffee, L.J.; Little, S.A.; Mercure, J.V.; Skipper, B.J.; Wheeler-Clark, E.S. )

    1990-11-01

    45Ca electron microscopic autoradiography was used to examine the effects of buffer composition and temperature on the distribution of calcium in rabbit renal artery smooth muscle cells. The results show that the relative distribution of calcium is dependent on both the buffer used (Tris or Krebs) and the temperature of the bathing solution (25 degrees C or 34{degrees}C). Krebs buffer at 34{degrees}C gave the highest relative activity in the plasma membrane, sarcoplasmic reticulum, and mitochondria. Buffer and temperature had little effect on the relative activity of the nucleus or cytoplasm. Next, we identified the cellular sites of calcium accumulation after 5, 15, 30, or 60 min exposure to {sup 45}Ca in Krebs buffer at 34{degrees}C. The results show that sarcoplasmic reticulum and plasma membrane are the primary sites of calcium accumulation during influx into these cells. Although the amount of {sup 45}Ca in the cell continues to increase with longer exposure, the relative distribution of calcium is essentially the same after 5 or 60 min. The data also indicate that the relative activity of plasma membrane + sarcoplasmic reticulum (a combination site that includes sarcoplasmic reticulum within a mean distance of 275 nm of the plasma membrane) is similar to the membrane alone and is lower than the sarcoplasmic reticulum alone.

  7. Distinctive serum miRNA profile in mouse models of striated muscular pathologies.

    PubMed

    Vignier, Nicolas; Amor, Fatima; Fogel, Paul; Duvallet, Angélique; Poupiot, Jérôme; Charrier, Sabine; Arock, Michel; Montus, Marie; Nelson, Isabelle; Richard, Isabelle; Carrier, Lucie; Servais, Laurent; Voit, Thomas; Bonne, Gisèle; Israeli, David

    2013-01-01

    Biomarkers are critically important for disease diagnosis and monitoring. In particular, close monitoring of disease evolution is eminently required for the evaluation of therapeutic treatments. Classical monitoring methods in muscular dystrophies are largely based on histological and molecular analyses of muscle biopsies. Such biopsies are invasive and therefore difficult to obtain. The serum protein creatine kinase is a useful biomarker, which is however not specific for a given pathology and correlates poorly with the severity or course of the muscular pathology. The aim of the present study was the systematic evaluation of serum microRNAs (miRNAs) as biomarkers in striated muscle pathologies. Mouse models for five striated muscle pathologies were investigated: Duchenne muscular dystrophy (DMD), limb-girdle muscular dystrophy type 2D (LGMD2D), limb-girdle muscular dystrophy type 2C (LGMD2C), Emery-Dreifuss muscular dystrophy (EDMD) and hypertrophic cardiomyopathy (HCM). Two-step RT-qPCR methodology was elaborated, using two different RT-qPCR miRNA quantification technologies. We identified miRNA modulation in the serum of all the five mouse models. The most highly dysregulated serum miRNAs were found to be commonly upregulated in DMD, LGMD2D and LGMD2C mouse models, which all exhibit massive destruction of striated muscle tissues. Some of these miRNAs were down rather than upregulated in the EDMD mice, a model without massive myofiber destruction. The dysregulated miRNAs identified in the HCM model were different, with the exception of one dysregulated miRNA common to all pathologies. Importantly, a specific and distinctive circulating miRNA profile was identified for each studied pathological mouse model. The differential expression of a few dysregulated miRNAs in the DMD mice was further evaluated in DMD patients, providing new candidates of circulating miRNA biomarkers for DMD.

  8. Two structurally distinct calcium storage sites in rat cardiac sarcoplasmic reticulum: an electron microprobe analysis study

    SciTech Connect

    Jorgensen, A.O.; Broderick, R.; Somlyo, A.P.; Somlyo, A.V.

    1988-12-01

    The elemental composition of subcellular organelles in resting rat papillary muscle was measured by electron probe x-ray microanalysis of cryosections of flash-frozen tissue. Nonmitochondrial electron-dense structures (50-100 nm in diameter) with a phosphorous concentration larger than 375 mmol/kg dry wt were identified in the interfibrillar spaces of the I band region. They were not visible in the proximity of transverse tubules. The sodium, magnesium, phosphorus, sulfur, chlorine, and potassium content of the electron dense structures showed a normal distribution, consistent with a uniform composition of a specific subcellular organelle. However, the distribution of the calcium concentrations in these electron-dense structures was bimodal, suggesting that they are composed of at least two subpopulations. One subpopulation had relatively high calcium (up to 53 mmol/kg dry wt) content with a mean value of 12.5 +/- 1.1 mmol/kg dry wt, while the other one had a relatively low calcium content with a mean value of 2.8 +/- 0.3 mmol/kg dry wt. The mean calcium concentration in the junctional sarcoplasmic reticulum (j-SR) in rat papillary muscle with calcium concentrations larger than 6 mmol/kg dry wt was 14.6 +/- 2.0 mmol/kg dry wt. We propose that the electron-dense structures described above correspond to nonjunctional sarcoplasmic reticulum and that the population containing relatively high calcium concentrations is calsequestrin-containing corbular sarcoplasmic reticulum (c-SR) confined to the I band region, while the population containing relatively low calcium concentrations corresponds to anastomosing regions of the network sarcoplasmic reticulum that lack calsequestrin.

  9. Interrelated striated elements in vestibular hair cells of the rat

    NASA Technical Reports Server (NTRS)

    Ross, M. D.; Bourne, C.

    1983-01-01

    A series of interrelated striated organelles in types I and II vestibular hair cells of the rat which appear to be less developed in cochlear hair cells have been revealed by unusual fixation procedures, suggesting that contractile elements may play a role in sensory transduction in the inner ear, especially in the vestibular system. Included in the series of interrelated striated elements are the cuticular plate and its basal attachments to the hair cell margins, the connections of the strut array of the kinociliary basal body to the cuticular plate, and striated organelles associated with the plasma membrane and extending below the apical junctional complexes.

  10. Representation of the visual field in the occipital striate cortex.

    PubMed Central

    McFadzean, R; Brosnahan, D; Hadley, D; Mutlukan, E

    1994-01-01

    The representation of the field of vision in the human striate cortex is based on the Holmes map in which about 25% of the surface area of the striate cortex is allocated to the central 15 degrees of vision. Following the introduction of computed tomography of the brain, the accuracy of the Holmes map was apparently confirmed by clinical/radiological correlation, but a revision has been proposed by Horton and Hoyt based on a magnetic resonance imaging study of three patients with visual field defects due to striate lesions. They propose that the central cortical representation of vision occupies a much larger area. This study reviews the perimetric and imaging findings in a larger series of patients with striate cortical disease and provides support for the revised representation. The clinical phenomenon of macular sparing and its relation to representation of the macula at the occipital pole is also discussed. Images PMID:8148333

  11. Insights from diploblasts; the evolution of mesoderm and muscle.

    PubMed

    Burton, Patrick Michael

    2008-01-15

    The origin of both mesoderm and muscle are central questions in metazoan evolution. The majority of metazoan phyla are triploblasts, possessing three discrete germ layers. Attention has therefore been focused on two outgroups to triploblasts, Cnidaria and Ctenophora. Modern texts describe these taxa as diploblasts, lacking a mesodermal germ layer. However, some members of Medusozoa, one of two subphyla within Cnidaria, possess tissue independent of either the ectoderm or endoderm referred to as the entocodon. Furthermore, members of both Cnidaria and Ctenophora have been described as possessing striated muscle, a mesodermal derivative. While it is widely accepted that the ancestor of Eumetazoa was diploblastic, homology of the entocodon and mesoderm as well as striated muscle within Eumetazoa has been suggested. This implies a potential triploblastic ancestor of Eumetazoa possessing striated muscle. In the following review, I examine the evidence for homology of both muscle and mesoderm. Current data support a diploblastic ancestor of cnidarians, ctenophores, and triploblasts lacking striated muscle.

  12. Hand-Held Model of a Sarcomere to Illustrate the Sliding Filament Mechanism in Muscle Contraction

    ERIC Educational Resources Information Center

    Jittivadhna, Karnyupha; Ruenwongsa, Pintip; Panijpan, Bhinyo

    2009-01-01

    From our teaching of the contractile unit of the striated muscle, we have found limitations in using textbook illustrations of sarcomere structure and its related dynamic molecular physiological details. A hand-held model of a striated muscle sarcomere made from common items has thus been made by us to enhance students' understanding of the…

  13. A muscle stem cell for every muscle: variability of satellite cell biology among different muscle groups

    PubMed Central

    Randolph, Matthew E.; Pavlath, Grace K.

    2015-01-01

    The human body contains approximately 640 individual skeletal muscles. Despite the fact that all of these muscles are composed of striated muscle tissue, the biology of these muscles and their associated muscle stem cell populations are quite diverse. Skeletal muscles are affected differentially by various muscular dystrophies (MDs), such that certain genetic mutations specifically alter muscle function in only a subset of muscles. Additionally, defective muscle stem cells have been implicated in the pathology of some MDs. The biology of muscle stem cells varies depending on the muscles with which they are associated. Here we review the biology of skeletal muscle stem cell populations of eight different muscle groups. Understanding the biological variation of skeletal muscles and their resident stem cells could provide valuable insight into mechanisms underlying the susceptibility of certain muscles to myopathic disease. PMID:26500547

  14. Effects of tetrandrine on calcium transport, protein fluorescences and membrane fluidity of sarcoplasmic reticulum

    PubMed Central

    Chen, Lan-Ying; Chen, Xi; Tian, Xiao-Li; Yu, Xiao-Hong

    2000-01-01

    To understand whether the molecular mechanism of Tetrandrine (Tet)'s pharmacological effects is concerned with sarcoplasmic reticulum calcium transport so as to be involved in myocardial contractility, we observed the effects of Tet on calcium transport and membrane structure of rabbit skeletal muscle sarcoplasmic reticulum vesicles (SR) and rat cardiac sarcoplasmic reticulum vesicles (CSR).Calcium uptake was monitored with a dual-wavelength spectrophotometer. Protein conformation and fluorescence polarization were measured by fluospectrophotometric method and membrane lipids labelled with fluorescence probes for SR, respectively.128 μmol l−1 Tet reduced the initial rate of calcium uptake to 59% of control 6 min after reaction. Tet un-competitively inhibited SR Ca2+,Mg2+-ATPase activity, causing the stoichiometric ratio of SR Ca2+/ATP to decrease to 1.43 from 2.0 of control.Inhibitory rates on SR Ca2+,Mg2+-ATPase by Tet were reduced from 60% in the absence of phosphate to 50% in the presence of phosphate and reduced from 92% in 1 mmol l−1 ATP to 60% in 5 mmol l−1 ATP.Tet markedly reduced SR intrinsic protein fluorescence, while it slightly decreased the thiol(SH)-modified protein fluorescence of SR labelled with N-(3-pyrene)-maleimide.Tet slightly increased fluorescence polarization in the middle and deep layers of SR membrane lipids labelled with 7- or 12-(9-anthroyloxy) stearic acid (AS) probes, whereas it did not change that of SR labelled with 1,6-diphenyl-1,3,5-hexatrine (DPH).These results revealed that prevention of SR calcium uptake by Tet was due to inhibition of the SR calcium pump Ca2+,Mg2+-ATPase, changes in spatial conformation of the pumps protein molecules and a decrease in the extent of motion of membrane lipid molecules, thus altering the regulation of [Ca2+]i and myocardial contractility. PMID:11015304

  15. Colour and sarcoplasmic protein evaluation of pork following water bath and ohmic cooking.

    PubMed

    Dai, Yan; Miao, Jing; Yuan, Shan-Zhen; Liu, Yi; Li, Xing-Min; Dai, Rui-Tong

    2013-04-01

    The objective of this study was to investigate the effects of ohmic (OH) and waterbath (WB) cooking on colour attributes and sarcoplasmic changes of porcine longissimus dorsi muscle at the same endpoint temperatures (EPTs; range 10°C-80°C). The OH treatment was carried out at 10 Vcm(-1), and the WB temperature at 85°C. The colour parameters, deoxymyoglobin% (DeoMb) and metmyoglobin% (MetMb) of the OH-cooked meat were significantly lower (P<0.05) than those obtained by WB-cooking at the same EPTs (range 60°C-80°C). SDS-PAGE analysis showed that the meat treated with WB-cooking had a lower sarcoplasmic protein solubility (5.97 mg/g vs.14.89 mg/g, P<0.05) and fainter protein bands than that of OH-cooking thus, indicating paler colour, and lower water-holding capacity especially in WB-cooked meat at EPTs above 40°C. Strong correlations among lightness, browness, metmyoglobin% and soluble proteins were observed in meat following OH-cooking.

  16. Differential abundance of sarcoplasmic proteome explains animal effect on beef Longissimus lumborum color stability.

    PubMed

    Canto, Anna C V C S; Suman, Surendranath P; Nair, Mahesh N; Li, Shuting; Rentfrow, Gregg; Beach, Carol M; Silva, Teofilo J P; Wheeler, Tommy L; Shackelford, Steven D; Grayson, Adria; McKeith, Russell O; King, D Andy

    2015-04-01

    The sarcoplasmic proteome of beef Longissimus lumborum demonstrating animal-to-animal variation in color stability was examined to correlate proteome profile with color. Longissimus lumborum (36 h post-mortem) muscles were obtained from 73 beef carcasses, aged for 13 days, and fabricated to 2.5-cm steaks. One steak was allotted to retail display, and another was immediately vacuum packaged and frozen at -80°C. Aerobically packaged steaks were stored under display, and color was evaluated on days 0 and 11. The steaks were ranked based on redness and color stability on day 11, and ten color-stable and ten color-labile carcasses were identified. Sarcoplasmic proteome of frozen steaks from the selected carcasses was analyzed. Nine proteins were differentially abundant in color-stable and color-labile steaks. Three glycolytic enzymes (phosphoglucomutase-1, glyceraldehyde-3-phosphate dehydrogenase, and pyruvate kinase M2) were over-abundant in color-stable steaks and positively correlated (P<0.05) to redness and color stability. These results indicated that animal variations in proteome contribute to differences in beef color.

  17. Mechanical Properties of Respiratory Muscles

    PubMed Central

    Sieck, Gary C.; Ferreira, Leonardo F.; Reid, Michael B.; Mantilla, Carlos B.

    2014-01-01

    Striated respiratory muscles are necessary for lung ventilation and to maintain the patency of the upper airway. The basic structural and functional properties of respiratory muscles are similar to those of other striated muscles (both skeletal and cardiac). The sarcomere is the fundamental organizational unit of striated muscles and sarcomeric proteins underlie the passive and active mechanical properties of muscle fibers. In this respect, the functional categorization of different fiber types provides a conceptual framework to understand the physiological properties of respiratory muscles. Within the sarcomere, the interaction between the thick and thin filaments at the level of cross-bridges provides the elementary unit of force generation and contraction. Key to an understanding of the unique functional differences across muscle fiber types are differences in cross-bridge recruitment and cycling that relate to the expression of different myosin heavy chain isoforms in the thick filament. The active mechanical properties of muscle fibers are characterized by the relationship between myoplasmic Ca2+ and cross-bridge recruitment, force generation and sarcomere length (also cross-bridge recruitment), external load and shortening velocity (cross-bridge cycling rate), and cross-bridge cycling rate and ATP consumption. Passive mechanical properties are also important reflecting viscoelastic elements within sarcomeres as well as the extracellular matrix. Conditions that affect respiratory muscle performance may have a range of underlying pathophysiological causes, but their manifestations will depend on their impact on these basic elemental structures. PMID:24265238

  18. Nandrolone decanoate treatment affects sarcoplasmic reticulum Ca(2+) ATPase function in skinned rat slow- and fast-twitch fibres.

    PubMed

    Bouhlel, Aicha; Joumaa, Wissam H; Léoty, Claude

    2003-09-01

    The effects of anabolic-androgenic steroid administration on the function of the sarcoplasmic reticulum (SR) pump were investigated in chemically skinned fibres from the extensor digitorum longus (EDL) and soleus muscles of sedentary rats. Twenty male rats were divided into two groups, one group received an intramuscular injection of nandrolone decanoate (15 mg x kg(-1)) weekly for 8 weeks, the second received similar weekly doses of vehicle (sterile peanut oil). Compared with control muscles, nandrolone decanoate treatment reduced SR Ca(2+) loading in EDL and soleus fibres by 49% and 29%, respectively. In control and treated muscles, the rate of Ca(2+) leakage depended on the quantity of Ca(2+) loaded. Furthermore, for similar SR Ca(2+) contents, the Ca(2+) leakage rate was not significantly modified by nandrolone decanoate treatment. Nandrolone decanoate treatment thus affects Ca (2+) uptake by the SR in a fibre-type dependent manner.

  19. Abnormal expression of the calmodulin gene in muscle from the dystrophic chicken

    SciTech Connect

    Hudecki, M.S.; Kibler, P.K.; Pollina, C.M.; Thacore, H.R.; Davis, P.J.; Davis, F.B.

    1986-05-29

    Compared to that of genetically-related normal chickens, pectoralis muscle from the dystrophic chicken contained increased calmodulin measured by radioimmunoassay. Determined by the dot blot procedure, expression of the calmodulin gene was enhanced in muscle from affected animals. The bioactivity of the gene product was normal. Together with previous studies reporting of increased sarcoplasmic calmodulin suggest the latter is a cellular response to defective Ca/sup 2 +/ transport at the level of cell efflux or intracellular organelle (sarcoplasmic reticulum) uptake.

  20. The sarcoplasmic reticulum and the evolution of the vertebrate heart.

    PubMed

    Shiels, Holly A; Galli, Gina L J

    2014-11-01

    The sarcoplasmic reticulum (SR) is crucial for contraction and relaxation of the mammalian cardiomyocyte, but its role in other vertebrate classes is equivocal. Recent evidence suggests differences in SR function across species may have an underlying structural basis. Here, we discuss how SR recruitment relates to the structural organization of the cardiomyocyte to provide new insight into the evolution of cardiac design and function in vertebrates.

  1. Histochemical characteristics of the striated inclusions of adrenoleukodystrophy.

    PubMed

    Johnson, A B; Schaumburg, H H; Powers, J M

    1976-06-01

    The straited accumulations in adrenal cortical cells and brain macrophages that are characteristic of adrenoleukodystrophy have been studied histochemically in cryostat sections to seek leads for the biochemical identification of the striated material. It stained pale pink with oil red O and did not stain with the Schultz cholesterol procedure or periodic acid-Schiff technique. By utilizing the birefringence of the accumulations as a marker, it was determined that, unlike natural cholesterol and cholesterol esters, the striated material was resistant to acetone and ethanol extraction. It was readily soluble, however, in nonpolar solvents such as n-hexane and chloroform. These findings indicated that the material was most probably a lipid, and they suggested that sequential extraction of adrenoleukodystrophy adrenal and brain with acetone and then n-hexane could be used to isolate this material in relatively pure form. Based on this lead, biochemical studies have just revealed a fatty acid abnormality in adrenoleukodystrophy which appears to be unique to this genetic disease.

  2. Virtual and simulated striated toolmarks for forensic applications.

    PubMed

    Baiker, Martin; Petraco, Nicholas D K; Gambino, Carol; Pieterman, René; Shenkin, Peter; Zoon, Peter

    2016-04-01

    Large numbers of experimental toolmarks of screwdrivers are often required in casework of toolmark examiners and in research environments alike, to be able to recover the angle of attack of a crime scene mark and to determine statistically meaningful properties of toolmarks respectively. However, in practice the number of marks is limited by the time needed to create them. In this article, we present an approach to predict how a striated mark of a particular tool would look like, using 3D surface datasets of screwdrivers. We compare these virtual toolmarks qualitatively and quantitatively with real experimental marks in wax and show that they are very similar. In addition we study toolmark similarity, dependent on the angle of attack, with a very high angular resolution of 1°. The results show that for the tested type of screwdriver, our toolmark comparison framework yields known match similarity scores that are above the mean known non-match similarity scores, even for known match differences in angle of attack of up to 40°. In addition we demonstrate an approach to automatically recover the angle of attack of an experimental toolmark and experiments yield high accuracy and precision of 0.618 ± 4.179°. Furthermore, we present a strategy to study the structural elements of striated toolmarks using wavelet analysis, and show how to use the results to simulate realistic toolmarks.

  3. Allergic Interstitial Nephritis Manifesting as a Striated Nephrogram

    PubMed Central

    Moinuddin, Irfan; Bracamonte, Erika; Thajudeen, Bijin; Sussman, Amy; Madhrira, Machaiah; Costello, James

    2015-01-01

    Allergic interstitial nephritis (AIN) is an underdiagnosed cause of acute kidney injury (AKI). Guidelines suggest that AIN should be suspected in a patient who presents with an elevated serum creatinine and a urinalysis that shows white cells, white cell casts, or eosinophiluria. Drug-induced AIN is suspected if AKI is temporally related to the initiation of a new drug. However, patients with bland sediment and normal urinalysis can also have AIN. Currently, a definitive diagnosis of AIN is made by renal biopsy which is invasive and fraught with risks such as bleeding, infection, and hematoma. Additionally, it is frequently unclear when a kidney biopsy should be undertaken. We describe a biopsy proven case of allergic interstitial nephritis which manifested on contrast enhanced Magnetic Resonance Imaging (MRI) as a striated nephrogram. Newer and more stable macrocyclic gadolinium contrast agents have a well-demonstrated safety profile. Additionally, in the presentation of AKI, gadolinium contrast agents are safe to administer in patients who demonstrate good urine output and a downtrending creatinine. We propose that the differential for a striated nephrogram may include AIN. In cases in which the suspicion for AIN is high, this diagnostic consideration may be further characterized by contrast enhanced MRI. PMID:26664405

  4. Topography of Striate-Extrastriate Connections in Neonatally Enucleated Rats

    PubMed Central

    Laing, Robyn J.; Lasiene, Jurate; Olavarria, Jaime F.

    2013-01-01

    It is known that retinal input is necessary for the normal development of striate cortex and its corticocortical connections, but there is little information on the role that retinal input plays in the development of retinotopically organized connections between V1 and surrounding visual areas. In nearly all lateral extrastriate areas, the anatomical and physiological representation of the nasotemporal axis of the visual field mirrors the representation of this axis in V1. To determine whether the mediolateral topography of striate-extrastriate projections is preserved in neonatally enucleated rats, we analyzed the patterns of projections resulting from tracer injections placed at different sites along the mediolateral axis of V1. We found that the correlation between the distance from injection sites to the lateral border of V1 and the distance of the labeling patterns in area 18a was strong in controls and much weaker in enucleates. Data from pairs of injections in the same animal revealed that the separation of area 18a projection fields for a given separation of injection sites was more variable in enucleated than in control rats. Our analysis of single and double tracer injections suggests that neonatal bilateral enucleation weakens, but not completely abolishes, the mediolateral topography in area 18a. PMID:24199194

  5. Theoretical study of interactions between striated cylindrical particles and membrane

    NASA Astrophysics Data System (ADS)

    Wang, Jing-Jing; Jia-Wei, Feng; Ren, Chun-Lai

    2015-08-01

    The interaction of nanoparticles with cell membranes is of great importance because of their potential biomedical applications. In this paper, we investigate the adhesion of stripe-patterned cylinders to a fluid membrane with a full consideration of the Helfrich free energy. Three situations are considered: one striated cylindrical particle, two pure cylindrical particles, and two Janus cylindrical particles. It is found that, with the adhesion of a single sparse striated cylinder, there are a variety of steady-states with energy barriers and the stable state is determined by the pattern of the cylinder. However, when the particle is densely striped, it has no effect on the stable state. By comparing the wrapping degree of two cylindrical particles with that of a single cylindrical particle, we find that two pure cylindrical particles can promote or suppress their interaction with the membrane under different situations. However, two Janus cylindrical particles can only inhibit their interaction with the membrane. Besides, this interaction is related to a first-order transition which is a shallow-to-deep wrapping transition for two pure cylinders while it is a shallow-to-half wrapping transition for two Janus cylinders. Furthermore, the position where the transition happens as a function of adhesion energy is given for fixed membrane tension and the precondition of the transition is presented. Project supported by the National Natural Science Foundation of China (Grant Nos. 91027040 and 21274062).

  6. CARDIAC MUSCLE

    PubMed Central

    Sommer, Joachim R.; Johnson, Edward A.

    1968-01-01

    With light and electron microscopy a comparison has been made of the morphology of ventricular (V) and Purkinje (P) fibers of the hearts of guinea pig, rabbit, cat, dog, goat, and sheep. The criteria, previously established for the rabbit heart, that V fibers are distinguished from P fibers by the respective presence and absence of transverse tubules is shown to be true for all animals studied. No evidence was found of a permanent connection between the sarcoplasmic reticulum and the extracellular space. The sarcoplasmic reticulum (SR) of V fibers formed couplings with the sarcolemma of a transverse tubule (interior coupling) and with the peripheral sarcolemma (peripheral coupling), whereas in P fibers the SR formed only peripheral couplings. The forms of the couplings were identical. The significance, with respect to excitation-contraction coupling, of the difference in the form of the couplings in cardiac versus skeletal muscle is discussed together with the electrophysiological implications of the differing geometries of bundles of P fibers from different animals. PMID:5645545

  7. Localization of phospholamban in smooth muscle using immunogold electron microscopy

    PubMed Central

    1988-01-01

    Phospholamban, the putative regulator of the Ca2+-ATPase in cardiac sarcoplasmic reticulum, was immunolocalized in canine visceral and vascular smooth muscle. Gently disrupted tissues were labeled with an affinity-purified phospholamban polyclonal antibody and indirect immunogold, using preembedding techniques. The sarcoplasmic reticulum of smooth muscle cells was specifically labeled with patches of immunogold distributed in a nonuniform fashion, while the sarcolemma did not appear to contain any phospholamban. The outer nuclear envelopes were also observed to be heavily labeled with the affinity- purified phospholamban polyclonal antibody. These findings suggest that phospholamban may play a role in the regulation of cytoplasmic and intranuclear calcium levels in smooth muscle cells. PMID:3417762

  8. Calcium-Binding Properties of Sarcoplasmic Reticulum As Influenced by ATP, Caffeine, Quinine, and Local Anesthetics

    PubMed Central

    Carvalho, Arselio P.

    1968-01-01

    Calcium retained at binding sites of the sarcoplasmic reticulum membranes isolated from rabbit skeletal muscle requires 10-5 - 10-4 M ATP to exchange with 45Ca added to the medium. The ATP requirement for Ca exchangeability was observed with respect to the "intrinsic" Ca of the reticulum membranes and the fraction of Ca that is "actively" bound in the presence of ATP. Furthermore, a concentration of free Ca in the medium higher than 10-8 M is required for ATP to promote Ca exchangeability. This exchangeability is not influenced by caffeine, quinine, procaine, and tetracaine, and Ca that is either nonexchangeable (in the absence of ATP) or exchangeable (in the presence of ATP) is released by 1–5 mM quinine or tetracaine, but neither caffeine (6 mM) nor procaine (2–5 mM) has this effect. Quinine or tetracaine also releases Ca and Mg bound passively to the reticulum membranes. A possible role of ATP in maintaining the integrity of cellular membranes is discussed, and the effects of caffeine, quinine, and of local anesthetics on the binding of Ca by the isolated reticulum are related to the effects of these agents on 45Ca fluxes and on the twitch output observed in whole muscles. PMID:5682486

  9. Weakly nonlinear ion waves in striated electron temperatures.

    PubMed

    Guio, P; Pécseli, H L

    2016-04-01

    The existence of low-frequency waveguide modes of electrostatic ion acoustic waves is demonstrated in magnetized plasmas for cases where the electron temperature is striated along magnetic field lines. For low frequencies, the temperature striation acts as waveguide that supports a trapped mode. For conditions where the ion cyclotron frequency is below the ion plasma frequency we find a dispersion relation having also a radiative frequency band, where waves can escape from the striation. Arguments for the formation and propagation of an equivalent of electrostatic shocks are presented and demonstrated numerically for these conditions. The shock represents here a balance between an external energy input maintained by ion injection and a dissipation mechanism in the form of energy leakage of the harmonics generated by nonlinear wave steepening. This is a reversible form for energy loss that can replace the time-irreversible losses in a standard Burgers equation. PMID:27176415

  10. On the water-holding of myofibrils: Effect of sarcoplasmic protein denaturation.

    PubMed

    Liu, Jiao; Arner, Anders; Puolanne, Eero; Ertbjerg, Per

    2016-09-01

    The role of heat-denatured sarcoplasmic proteins in water-holding is not well understood. Here we propose a new hypothesis that in PSE-like conditions denatured sarcoplasmic proteins aggregate within and outside myofilaments, improving the water-holding of denatured myofibrils. The process is compartmentalized: 1) within the filaments the denatured sarcoplasmic proteins shrink the lattice space and water is expelled; and 2) between the myofibrils and in the extracellular space, the coagulated sarcoplasmic proteins trap the expelled water from interfilamental space. The effect of sarcoplasmic proteins on the water-holding of myofibrils following incubation for 1h at 21 to 44°C was investigated. Our results were consistent with the new hypothesis. Myofibrils without sarcoplasm had the poorest water-holding. With increasing amount of denatured sarcoplasmic proteins, the water-holding of heat-denatured myofibrils improved proportionally. X-ray diffraction was used to measure the lattice space between the filaments. Precipitated sarcoplasmic proteins shrank (P<0.001) the lattice spacing by 6.3% at 44°C. PMID:27129081

  11. Cooked sausage batter cohesiveness as affected by sarcoplasmic proteins.

    PubMed

    Farouk, M M; Wieliczko, K; Lim, R; Turnwald, S; Macdonald, G A

    2002-05-01

    In the first trial, m. semitendinosus and m. biceps femoris were held at 0, 10 and 35 °C until they entered rigor, and in the second trial, minced m. semitendinosus was washed in water for 15, 30, 45 or 60 min. The samples from both the trials were then used to make a finely comminuted sausage batter. Soluble sarcoplasmic protein (SSP) levels decreased with increasing rigor temperature (P < 0.05) or washing (P < 0.01). Cooked batter shear stress was not affected by SSP level, but batter shear strain decreased with the decreasing SSP level associated with an increasing rigor temperature (P < 0.05) or washing (P < 0.01). Reducing the SSP content lowered the cook yield (P < 0.05) and emulsion stability (P < 0.01) of the batter from the washed samples compared to that of controls. The results suggest that sarcoplasmic proteins are important in determining the strain values (cohesiveness) of cooked sausage batter.

  12. Laser raman characterization of conformational charges in sarcoplasmic reticulum induced by termperature, Ca/sup 2 +/, and Mg/sup 2 +/

    SciTech Connect

    Lippert, J.L.; Lindsay, R.M.; Schultz, R.

    1981-12-10

    Laser Raman spectroscopy has been used to examine the conformations of the protein and phospholipid components of sarcoplasmic reticulum from rabbit white skeletal muscle. The phospholipid component is shown to have the conformation of fluid, liquid-crystalline lipids, even at 10/sup o/C, and no breaks in the lipid conformation are observed in the range of 10-37/sup o/C. Protein (predominantly the Ca/sup 2 +/-dependent ATPase) conformation is shown to contain very little ..beta..-sheet structure under all conditions. Absolute content of ..cap alpha..-helix and random coil or ..beta..-turn could not be determined because of interference in the amide I and III regions. However, the Ca/sup 2 +/-ATPase in sarcoplasmic reticulum appears to undergo a conformational change at 15-18/sup o/C which involves removal of a portion of the tryptophan residues from an aqueous environment and an increase in ..cap alpha..-helical content. This conformation change coincides with a change in slope of Arrhenius plots of ATP hydrolysis activity. Increasing concentrations of Ca/sup 2 +/ and Mg/sup 2 +/ appear to slightly decrease the ..cap alpha..-helical content of sarcoplasmic reticulum protein.

  13. Trimeric intracellular cation channels and sarcoplasmic/endoplasmic reticulum calcium homeostasis.

    PubMed

    Zhou, Xinyu; Lin, Peihui; Yamazaki, Daiju; Park, Ki Ho; Komazaki, Shinji; Chen, S R Wayne; Takeshima, Hiroshi; Ma, Jianjie

    2014-02-14

    Trimeric intracellular cation channels (TRIC) represents a novel class of trimeric intracellular cation channels. Two TRIC isoforms have been identified in both the human and the mouse genomes: TRIC-A, a subtype predominantly expressed in the sarcoplasmic reticulum (SR) of muscle cells, and TRIC-B, a ubiquitous subtype expressed in the endoplasmic reticulum (ER) of all tissues. Genetic ablation of either TRIC-A or TRIC-B leads to compromised K(+) permeation and Ca(2+) release across the SR/ER membrane, supporting the hypothesis that TRIC channels provide a counter balancing K(+) flux that reduces SR/ER membrane depolarization for maintenance of the electrochemical gradient that drives SR/ER Ca(2+) release. TRIC-A and TRIC-B seem to have differential functions in Ca(2+) signaling in excitable and nonexcitable cells. Tric-a(-/-) mice display defective Ca(2+) sparks and spontaneous transient outward currents in arterial smooth muscle and develop hypertension, in addition to skeletal muscle dysfunction. Knockout of TRIC-B results in abnormal IP3 receptor-mediated Ca(2+) release in airway epithelial cells, respiratory defects, and neonatal lethality. Double knockout mice lacking both TRIC-A and TRIC-B show embryonic lethality as a result of cardiac arrest. Such an aggravated lethality indicates that TRIC-A and TRIC-B share complementary physiological functions in Ca(2+) signaling in embryonic cardiomyocytes. Tric-a(-/-) and Tric-b(+/-) mice are viable and susceptible to stress-induced heart failure. Recent evidence suggests that TRIC-A directly modulates the function of the cardiac ryanodine receptor 2 Ca(2+) release channel, which in turn controls store-overload-induced Ca(2+) release from the SR. Thus, the TRIC channels, in addition to providing a countercurrent for SR/ER Ca(2+) release, may also function as accessory proteins that directly modulate the ryanodine receptor/IP3 receptor channel functions.

  14. NUCLEOSIDE PHOSPHATASE ACTIVITIES IN RAT CARDIAC MUSCLE.

    PubMed

    ESSNER, E; NOVIKOFF, A B; QUINTANA, N

    1965-05-01

    Localizations of aldehyde-resistant nucleoside phosphatase activities in frozen sections of rat cardiac muscle have been studied by electron microscopy. Activities are higher after fixation with formaldehyde than with glutaraldehyde. After incubation with adenosine triphosphate or inosine diphosphate at pH 7.2, reaction product is found in the "terminal cisternae" or "transverse sacs" of the sarcoplasmic reticulum, which, together with the "intermediary vesicles" (T system), constitute the "dyads" or "triads". Reaction product is also present at the membranes of micropinocytotic vacuoles which apparently form from the plasma membrane of capillary endothelial cells and from the sarcolemma. In certain regions of the intercalated discs, reaction product is found within the narrow spaces between sarcolemmas of adjacent cells and within micropinocytotic vacuoles that seem to form from the sarcolemma. With inosine diphosphate, reaction product is also found in other parts of the sarcoplasmic reticulum. After incubation with cytidine monophosphate at pH 5, reaction product is present in the transverse sacs of sarcoplasmic reticulum, in micropinocytotic vacuoles in capillary endothelium, and in lysosomes of muscle fibers and capillaries. The possible significance of the sarcoplasmic reticulum phosphatases is discussed in relation to the role the reticulum probably plays in moving calcium ions and thereby controlling contraction and relaxation of the muscle fiber.

  15. Microdomains of endoplasmic reticulum within the sarcoplasmic reticulum of skeletal myofibers

    SciTech Connect

    Kaakinen, Mika; Papponen, Hinni; Metsikkoe, Kalervo

    2008-01-15

    The relationship between the endoplasmic reticulum (ER) and the sarcoplasmic reticulum (SR) of skeletal muscle cells has remained obscure. In this study, we found that ER- and SR-specific membrane proteins exhibited diverse solubility properties when extracted with mild detergents. Accordingly, the major SR-specific protein Ca{sup 2+}-ATPase (SERCA) remained insoluble in Brij 58 and floated in sucrose gradients while typical ER proteins were partially or fully soluble. Sphingomyelinase treatment rendered SERCA soluble in Brij 58. Immunofluorescence staining for resident ER proteins revealed dispersed dots over I bands contrasting the continuous staining pattern of SERCA. Infection of isolated myofibers with enveloped viruses indicated that interfibrillar protein synthesis occurred. Furthermore, we found that GFP-tagged Dad1, able to incorporate into the oligosaccharyltransferase complex, showed the dot-like structures but the fusion protein was also present in membranes over the Z lines. This behaviour mimics that of cargo proteins that accumulated over the Z lines when blocked in the ER. Taken together, the results suggest that resident ER proteins comprised Brij 58-soluble microdomains within the insoluble SR membrane. After synthesis and folding in the ER-microdomains, cargo proteins and non-incorporated GFP-Dad1 diffused into the Z line-flanking compartment which likely represents the ER exit sites.

  16. A model for the uptake and release of Ca2+ by sarcoplasmic reticulum.

    PubMed

    Gould, G W; McWhirter, J M; East, J M; Lee, A G

    1987-08-01

    On addition of ATP to vesicles derived from the sarcoplasmic reticulum (SR) of skeletal muscle, Ca2+ is accumulated from the external medium. Following uptake, spontaneous release of Ca2+ occurs in the presence or in the absence of ATP. These processes of Ca2+ uptake and release were simulated by using the models derived for ATPase activity [Gould, East, Froud, McWhirter, Stefanova & Lee (1986) Biochem. J. 237, 217-227; Stefanova, Napier, East & Lee (1987) Biochem. J. 245, 723-730] and for Ca2+ release from passively loaded vesicles [McWhirter, Gould, East & Lee (1987) Biochem. J. 245, 713-722]. The simulations are consistent with measurements of the effects of pH, K+, Ca2+ and Mg2+ on uptake and release of Ca2+. The increase in maximal Ca2+ accumulation observed in the presence of maleate is explained in terms of complexing of Ca2+ and maleate within the SR. The calculated concentration of ADP generated by hydrolysis of ATP has a large effect on the simulations. The effects of an ATP-regenerating system on the measured Ca2+ uptake is explained in terms of both removal of ADP and precipitation of Ca3(PO4)2 within the vesicles. It is concluded that both the process of Ca2+ uptake and the process of Ca2+ release seen with SR vesicles can be interpreted quantitatively in terms solely of the properties of the Ca2+ + Mg2+-activated ATPase.

  17. Effect of halothane on the oligomerization of the sarcoplasmic reticulum Ca(2+)-ATPase.

    PubMed

    Brennan, L K; Froemming, G R; Ohlendieck, K

    2000-05-19

    The exact molecular mechanism of inhalational anesthetics remains obscure. Since the enzyme activity of the sarcoplasmic reticulum Ca(2+)-ATPase from skeletal muscle fibres is modified by halothane and because protein-protein interactions play an important role in the regulation of Ca(2+)-regulatory proteins, we investigated the effect of this volatile drug on the oligomerization of the fast-twitch Ca(2+)-ATPase. Using electrophoretic separation following incubation with halothane, increases in relative molecular mass were determined by immunoblotting with a monoclonal antibody to the SERCA1 isoform of the Ca(2+)-ATPase. Distinct drug-induced decreases in electrophoretic mobility indicated oligomerization of the native Ca(2+)-pump by halothane, comparable to crosslinking-mediated formation of homo-tetramers. Determination of the effect of halothane on enzyme activity suggested that halothane-mediated protein aggregation triggers a partial inhibition of Ca(2+)-pump units. Thus, halothane appears to exert its action via specific peptide binding sites and not indirectly by lipid perturbation. These findings support the protein theory of anesthetic action.

  18. Striated spectral activity in Jovian and Saturnian radio emission

    NASA Astrophysics Data System (ADS)

    Thieman, James R.; Alexander, Joseph K.; Arias, Tomas A.; Staelin, David H.

    1988-09-01

    Examination of high time resolution frequency-time spectrograms of radio emission measured near the Voyager 1 and 2 encounters with Jupiter reveals occasional striation patterns within the normally diffuse hectometric radiation. The patterns are characterized by distinctive banded structures of enhanced intensity meandering in frequency over time scales of minutes to tens of minutes. This banded form of striated spectral activity (SSA) has an occurrence probability of the order of 5 percent during the three weeks before and after Jupiter encounters. Plots of single 6-s frequency sweeps often exhibit a slow rise in intensity followed by a sharp drop-off in each band as frequency decreases. Banded SSA is often preceded or followed by chaotic SSA in which banding of the emission becomes discontinuous or unrecognizable, although the intensity modulation is still evident. Although SSA normally occurs in the frequency range of roughly 0.2-1.0 MHz, similar but longer-lasting patterns have been found occasionally in decametric emission above 10 MHz. Analogous modulation has also been observed in the Saturnian radio emission, suggesting that SSA may be a common feature intrinsic to the radio emission at both planets.

  19. Spatially congruent model for the striate visual cortex

    NASA Astrophysics Data System (ADS)

    da Fontoura Costa, Luciano

    1994-05-01

    A spatially congruent new model for the striate visual cortex (SVC) is proposed which accounts for some of the known functional and organizational properties of the superior mammalian SVC. Even though there is a broad consensus that the topographical representation of the visual field is one of the principal structuring principles underlying the SVC organization, the orientation maps in the SVC have often been described as non-topographical maps. In the present model, the adopted foot-of-normal representation of straight lines has allowed full congruency between the visual field topographic map and the orientation maps in the SVC. The proposed computational model includes three neural layers and assumes that the ocular dominance columns are already established at birth; three possibilities of neural mechanisms leading to orientation encoding are outlined and discussed. The model provides reasonable explanation to some of the most intriguing recently verified properties of the SVC such as the increased neural activity at the cytochrome oxidase blobs, the reduced orientation selectivity at these same places, and the pinwheel-like organization of the orientation selectivity in the SVC.

  20. Neuronal basis of perceptual learning in striate cortex

    PubMed Central

    Ren, Zhen; Zhou, Jiawei; Yao, Zhimo; Wang, Zhengchun; Yuan, Nini; Xu, Guangwei; Wang, Xuan; Zhang, Bing; Hess, Robert F.; Zhou, Yifeng

    2016-01-01

    It is well known that, in humans, contrast sensitivity training at high spatial frequency (SF) not only leads to contrast sensitivity improvement, but also results in an improvement in visual acuity as assessed with gratings (direct effect) or letters (transfer effect). However, the underlying neural mechanisms of this high spatial frequency training improvement remain to be elucidated. In the present study, we examined four properties of neurons in primary visual cortex (area 17) of adult cats that exhibited significantly improved acuity after contrast sensitivity training with a high spatial frequency grating and those of untrained control cats. We found no difference in neuronal contrast sensitivity or tuning width (Width) between the trained and untrained cats. However, the trained cats showed a displacement of the cells’ optimal spatial frequency (OSF) to higher spatial frequencies as well as a larger neuronal signal-to-noise ratio (SNR). Furthermore, both the neuronal differences in OSF and SNR were significantly correlated with the improvement of acuity measured behaviorally. These results suggest that striate neurons might mediate the perceptual learning-induced improvement for high spatial frequency stimuli by an alteration in their spatial frequency representation and by an increased SNR. PMID:27094565

  1. Voltage-Independent Calcium Release in Heart Muscle

    NASA Astrophysics Data System (ADS)

    Niggli, Ernst; Lederer, W. Jonathan

    1990-10-01

    The Ca2+ that activates contraction in heart muscle is regulated as in skeletal muscle by processes that depend on voltage and intracellular Ca2+ and involve a positive feedback system. How the initial electrical signal is amplified in heart muscle has remained controversial, however. Analogous protein structures from skeletal muscle and heart muscle have been identified physiologically and sequenced; these include the Ca2+ channel of the sarcolemma and the Ca2+ release channel of the sarcoplasmic reticulum. Although the parallels found in cardiac and skeletal muscles have provoked valuable experiments in both tissues, separation of the effects of voltage and intracellular Ca2+ on sarcoplasmic reticulum Ca2+ release in heart muscle has been imperfect. With the use of caged Ca2+ and flash photolysis in voltage-clamped heart myocytes, effects of membrane potential in heart muscle cells on Ca2+ release from intracellular stores have been studied. Unlike the response in skeletal muscle, voltage across the sarcolemma of heart muscle does not affect the release of Ca2+ from the sarcoplasmic reticulum, suggesting that other regulatory processes are needed to control Ca2+-induced Ca2+ release.

  2. High skeletal muscle adenylate cyclase in malignant hyperthermia.

    PubMed Central

    Willner, J H; Cerri, C G; Wood, D S

    1981-01-01

    Malignant hyperthermia occurs in humans with several congenital myopathies, usually in response to general anesthesia. Commonly, individuals who develop this syndrome lack symptoms of muscle disease, and their muscle lacks specific pathological changes. A biochemical marker for this myopathy has not previously been available; we found activity of adenylate cyclase and content of cyclic AMP to be abnormally high in skeletal muscle. Secondary modification of protein phosphorylation could explain observed abnormalities of phosphorylase activation and sarcoplasmic reticulum function. PMID:6271806

  3. Role of glucocorticoids in the response of rat leg muscles to reduced activity

    NASA Technical Reports Server (NTRS)

    Jaspers, Stephen R.; Tischler, Marc E.

    1986-01-01

    Adrenalectomy did not prevent atrophy of rat soleus muscle during 6 days of tail cast suspension. Cortisol treatment enhanced the atrophy and caused atrophy of the weight-bearing soleus and both extensor digitorum longus (EDL) muscles. Unloading led to increased sarcoplasmic protein concentration in the soleus but cortisol administration increased the myhofibrillar (+stromal) protein concentration in both muscles. Suspension of hindlimbs of adrenalectomized animals led to faster protein degradation, slower sarcoplasmic protein degradation, and faster myofibrillar protein synthesis in the isolated soleus, whereas with cortisol-treated animals, the difference in synthesis of myofibrillar proteins was enhanced and that of sarcoplasmic proteins was abolished. Both soleus and EDL of suspended, cortisol-treated animals showed faster protein degradation. It is unlikely that any elevation in circulating glucocorticoids was solely responsible for atrophy of the soleus in this model, but catabolic amounts of glucocorticoids could alter the response of muscle to unloading.

  4. Receptive fields and functional architecture of monkey striate cortex.

    PubMed

    Hubel, D H; Wiesel, T N

    1968-03-01

    1. The striate cortex was studied in lightly anaesthetized macaque and spider monkeys by recording extracellularly from single units and stimulating the retinas with spots or patterns of light. Most cells can be categorized as simple, complex, or hypercomplex, with response properties very similar to those previously described in the cat. On the average, however, receptive fields are smaller, and there is a greater sensitivity to changes in stimulus orientation. A small proportion of the cells are colour coded.2. Evidence is presented for at least two independent systems of columns extending vertically from surface to white matter. Columns of the first type contain cells with common receptive-field orientations. They are similar to the orientation columns described in the cat, but are probably smaller in cross-sectional area. In the second system cells are aggregated into columns according to eye preference. The ocular dominance columns are larger than the orientation columns, and the two sets of boundaries seem to be independent.3. There is a tendency for cells to be grouped according to symmetry of responses to movement; in some regions the cells respond equally well to the two opposite directions of movement of a line, but other regions contain a mixture of cells favouring one direction and cells favouring the other.4. A horizontal organization corresponding to the cortical layering can also be discerned. The upper layers (II and the upper two-thirds of III) contain complex and hypercomplex cells, but simple cells are virtually absent. The cells are mostly binocularly driven. Simple cells are found deep in layer III, and in IV A and IV B. In layer IV B they form a large proportion of the population, whereas complex cells are rare. In layers IV A and IV B one finds units lacking orientation specificity; it is not clear whether these are cell bodies or axons of geniculate cells. In layer IV most cells are driven by one eye only; this layer consists of a mosaic

  5. Signaling in muscle contraction.

    PubMed

    Kuo, Ivana Y; Ehrlich, Barbara E

    2015-02-02

    Signaling pathways regulate contraction of striated (skeletal and cardiac) and smooth muscle. Although these are similar, there are striking differences in the pathways that can be attributed to the distinct functional roles of the different muscle types. Muscles contract in response to depolarization, activation of G-protein-coupled receptors and other stimuli. The actomyosin fibers responsible for contraction require an increase in the cytosolic levels of calcium, which signaling pathways induce by promoting influx from extracellular sources or release from intracellular stores. Rises in cytosolic calcium stimulate numerous downstream calcium-dependent signaling pathways, which can also regulate contraction. Alterations to the signaling pathways that initiate and sustain contraction and relaxation occur as a consequence of exercise and pathophysiological conditions.

  6. Proteins related to green algal striated fiber assemblin are present in stramenopiles and alveolates.

    PubMed

    Harper, John D I; Thuet, Jacques; Lechtreck, Karl F; Hardham, Adrienne R

    2009-07-01

    In green algae, striated fiber assemblin (SFA) is the major protein of the striated microtubule-associated fibers that are structural elements in the flagellar basal apparatus. Using Basic Local Alignment Search Tool (BLAST) searches of recently established databases, SFA-like sequences were detected in the genomes not only of green algal species but also of a range of other protists. These included species in two alveolate subgroups, the ciliates (Tetrahymena thermophila, Paramecium tetraurelia) and the dinoflagellates (Perkinsus marinus), and two stramenopile subgroups, the oomycetes (Phytophthora sojae, Phytophthora ramorum, Phytophthora infestans) and the diatoms (Thalassiosira pseudonana, Phaeodactylum tricornutum). Together with earlier identification of SFA-like sequences in the apicomplexans, these results indicate that homologs of SFA are present across the alveolates and stramenopiles. Antibodies raised against SFA from the green alga, Spermatozopsis similis, react in immunofluorescence assays with the two basal bodies and an anteriorly directed striated fiber in the flagellar apparatus of biflagellate Phytophthora zoospores.

  7. Muscle assembly: a titanic achievement?

    PubMed

    Gregorio, C C; Granzier, H; Sorimachi, H; Labeit, S

    1999-02-01

    The formation of perfectly aligned myofibrils in striated muscle represents a dramatic example of supramolecular assembly in eukaryotic cells. Recently, considerable progress has been made in deciphering the roles that titin, the third most abundant protein in muscle, has in this process. An increasing number of sarcomeric proteins (ligands) are being identified that bind to specific titin domains. Titin may serve as a molecular blueprint for sarcomere assembly and turnover by specifying the precise position of its ligands within each half-sarcomere in addition to functioning as a molecular spring that maintains the structural integrity of the contracting myofibrils.

  8. Ryanodine receptor fragmentation and sarcoplasmic reticulum Ca2+ leak after one session of high-intensity interval exercise.

    PubMed

    Place, Nicolas; Ivarsson, Niklas; Venckunas, Tomas; Neyroud, Daria; Brazaitis, Marius; Cheng, Arthur J; Ochala, Julien; Kamandulis, Sigitas; Girard, Sebastien; Volungevičius, Gintautas; Paužas, Henrikas; Mekideche, Abdelhafid; Kayser, Bengt; Martinez-Redondo, Vicente; Ruas, Jorge L; Bruton, Joseph; Truffert, Andre; Lanner, Johanna T; Skurvydas, Albertas; Westerblad, Håkan

    2015-12-15

    High-intensity interval training (HIIT) is a time-efficient way of improving physical performance in healthy subjects and in patients with common chronic diseases, but less so in elite endurance athletes. The mechanisms underlying the effectiveness of HIIT are uncertain. Here, recreationally active human subjects performed highly demanding HIIT consisting of 30-s bouts of all-out cycling with 4-min rest in between bouts (≤3 min total exercise time). Skeletal muscle biopsies taken 24 h after the HIIT exercise showed an extensive fragmentation of the sarcoplasmic reticulum (SR) Ca(2+) release channel, the ryanodine receptor type 1 (RyR1). The HIIT exercise also caused a prolonged force depression and triggered major changes in the expression of genes related to endurance exercise. Subsequent experiments on elite endurance athletes performing the same HIIT exercise showed no RyR1 fragmentation or prolonged changes in the expression of endurance-related genes. Finally, mechanistic experiments performed on isolated mouse muscles exposed to HIIT-mimicking stimulation showed reactive oxygen/nitrogen species (ROS)-dependent RyR1 fragmentation, calpain activation, increased SR Ca(2+) leak at rest, and depressed force production due to impaired SR Ca(2+) release upon stimulation. In conclusion, HIIT exercise induces a ROS-dependent RyR1 fragmentation in muscles of recreationally active subjects, and the resulting changes in muscle fiber Ca(2+)-handling trigger muscular adaptations. However, the same HIIT exercise does not cause RyR1 fragmentation in muscles of elite endurance athletes, which may explain why HIIT is less effective in this group.

  9. Ryanodine receptor fragmentation and sarcoplasmic reticulum Ca2+ leak after one session of high-intensity interval exercise.

    PubMed

    Place, Nicolas; Ivarsson, Niklas; Venckunas, Tomas; Neyroud, Daria; Brazaitis, Marius; Cheng, Arthur J; Ochala, Julien; Kamandulis, Sigitas; Girard, Sebastien; Volungevičius, Gintautas; Paužas, Henrikas; Mekideche, Abdelhafid; Kayser, Bengt; Martinez-Redondo, Vicente; Ruas, Jorge L; Bruton, Joseph; Truffert, Andre; Lanner, Johanna T; Skurvydas, Albertas; Westerblad, Håkan

    2015-12-15

    High-intensity interval training (HIIT) is a time-efficient way of improving physical performance in healthy subjects and in patients with common chronic diseases, but less so in elite endurance athletes. The mechanisms underlying the effectiveness of HIIT are uncertain. Here, recreationally active human subjects performed highly demanding HIIT consisting of 30-s bouts of all-out cycling with 4-min rest in between bouts (≤3 min total exercise time). Skeletal muscle biopsies taken 24 h after the HIIT exercise showed an extensive fragmentation of the sarcoplasmic reticulum (SR) Ca(2+) release channel, the ryanodine receptor type 1 (RyR1). The HIIT exercise also caused a prolonged force depression and triggered major changes in the expression of genes related to endurance exercise. Subsequent experiments on elite endurance athletes performing the same HIIT exercise showed no RyR1 fragmentation or prolonged changes in the expression of endurance-related genes. Finally, mechanistic experiments performed on isolated mouse muscles exposed to HIIT-mimicking stimulation showed reactive oxygen/nitrogen species (ROS)-dependent RyR1 fragmentation, calpain activation, increased SR Ca(2+) leak at rest, and depressed force production due to impaired SR Ca(2+) release upon stimulation. In conclusion, HIIT exercise induces a ROS-dependent RyR1 fragmentation in muscles of recreationally active subjects, and the resulting changes in muscle fiber Ca(2+)-handling trigger muscular adaptations. However, the same HIIT exercise does not cause RyR1 fragmentation in muscles of elite endurance athletes, which may explain why HIIT is less effective in this group. PMID:26575622

  10. Polymorphism in high-affinity calcium-binding proteins from crustacean sarcoplasm.

    PubMed

    Wnuk, W; Jauregui-Adell, J

    1983-03-01

    The sarcoplasmic calcium-binding proteins (SCP) from crayfish, lobster and shrimp myogen have been purified to homogeneity. These proteins exist as dimers and dissociate in the presence of sodium dodecyl sulfate or urea in subunits of 22000 molecular weight. During the last step of purification (DEAE-cellulose chromatography), SCP emerges in three peaks in the ratio of 14:1.5:1 for crayfish, of 7:2:1 for lobster and of 3:2:1 for shrimp. Gel electrophoresis and isoelectrofocusing experiments, under native and denaturing conditions, indicate that among the three SCP isotypes there are only two different polypeptide chains, alpha and beta, which appear in the form of three dimers: alpha 2, alpha beta and beta 2. The alpha and beta subunits differ slightly in polypeptide chain composition as found by amino acid analyses of the crayfish and lobster SCPs, and also by comparison of tryptic peptides for crayfish SCPs. The polymorphism observed in crustacean SCPs, which is increased by their ability to form dimers, contrasts with the situation prevailing among other invertebrate SCPs and vertebrate parvalbumins where only monomeric isotypes are found. Equilibrium binding studies show that all three SCP isotypes from both crayfish and lobster display the same metal-binding properties. They have in their dimeric form six Ca2+-binding sites: two calcium-specific sites, two Ca/Mg sites that interact with positive cooperativity and two Ca/Mg sites that interact with negative cooperativity. Interactions between the two subunits of SCP seem to result in cooperative binding of Ca2+, which in turn may control more efficiently Ca2+ fluxes in crustacean muscle. PMID:6832140

  11. Sarcoplasmic reticulum is an intermediary of mitochondrial and myofibrillar growth at the intercalated disc.

    PubMed

    Bennett, Pauline M; Ehler, Elisabeth; Wilson, Amanda J

    2016-06-01

    In cardiomyocytes columns of intermyofibrillar mitochondria run up to the intercalated disc (ID); half are collinear with those in the neighbouring cell, suggesting coordinated addition of sarcomeres and mitochondria both within and between cells during cardiomyocyte growth. Recent evidence for an association between sarcoplasmic reticulum (SR) and mitochondria indicates that the SR may be an intermediary in this coordinated behaviour. For this reason we have investigated the arrangement of SR and t tubules with respect to mitochondria and myofibrils, particularly at the ID. In the body of the cardiomyocyte the mitochondrial columns are frequently intersected by transverse tubules. In addition, we find that a majority of axial tubules are sandwiched between mitochondria and myofibril. No tubules are found at the ID. SR coats mitochondrial columns and fibrils throughout their length and reaches towards the peaks of the ID membrane where it attaches in the form of junctional (j)SR. These peripheral ID couplings are often situated between mitochondria and ID membrane, suggesting an SR connection between the two. In dilated cardiomyopathy (DCM) the mitochondria are somewhat disordered and clumped. In a mouse model for DCM, the muscle LIM protein KO, we find that there is a lack of mitochondria near the ID, suggesting the uncoupling of the myofibril/mitochondria organisation during growth. SR still coats the fibrils and reaches the ID folds in a jSR coupling. Unlike in control tissue, however, loops and long fingers of ID membrane penetrate into the proximal sarcomere suggesting a possible intermediary state in cardiomyocyte growth. PMID:27329158

  12. Potentiation of fractional sarcoplasmic reticulum calcium release by total and free intra-sarcoplasmic reticulum calcium concentration.

    PubMed Central

    Shannon, T R; Ginsburg, K S; Bers, D M

    2000-01-01

    Our aim was to measure the influence of sarcoplasmic reticulum (SR) calcium content ([Ca](SRT)) and free SR [Ca] ([Ca](SR)) on the fraction of SR calcium released during voltage clamp steps in isolated rabbit ventricular myocytes. [Ca](SRT), as measured by caffeine application, was progressively increased by conditioning pulses. Sodium was absent in both the intracellular and in the extracellular solutions to block sodium/calcium exchange. Total cytosolic calcium flux during the transient was inferred from I(Ca), [Ca](SRT), [Ca](i), and cellular buffering characteristics. Fluxes via the calcium current (I(Ca)), the SR calcium pump, and passive leak from the SR were evaluated to determine SR calcium release flux (J(rel)). Excitation-contraction (EC) coupling was characterized with respect to both gain (integral J(rel)/integral I(Ca)) and fractional SR calcium release. Both parameters were virtually zero for a small, but measurable [Ca](SRT). Gain and fractional SR calcium release increased steeply and nonlinearly with both [Ca](SRT) and [Ca](SR). We conclude that potentiation of EC coupling can be correlated with both [Ca](SRT) and [Ca](SR). While fractional SR calcium release was not linearly dependent upon [Ca](SR), intra-SR calcium may play a crucial role in regulating the SR calcium release process. PMID:10620297

  13. Lipogenesis mitigates dysregulated sarcoplasmic reticulum calcium uptake in muscular dystrophy.

    PubMed

    Paran, Christopher W; Zou, Kai; Ferrara, Patrick J; Song, Haowei; Turk, John; Funai, Katsuhiko

    2015-12-01

    Muscular dystrophy is accompanied by a reduction in activity of sarco/endoplasmic reticulum Ca(2+)-ATPase (SERCA) that contributes to abnormal Ca(2+) homeostasis in sarco/endoplasmic reticulum (SR/ER). Recent findings suggest that skeletal muscle fatty acid synthase (FAS) modulates SERCA activity and muscle function via its effects on SR membrane phospholipids. In this study, we examined muscle's lipid metabolism in mdx mice, a mouse model for Duchenne muscular dystrophy (DMD). De novo lipogenesis was ~50% reduced in mdx muscles compared to wildtype (WT) muscles. Gene expressions of lipogenic and other ER lipid-modifying enzymes were found to be differentially expressed between wildtype (WT) and mdx muscles. A comprehensive examination of muscles' SR phospholipidome revealed elevated phosphatidylcholine (PC) and PC/phosphatidylethanolamine (PE) ratio in mdx compared to WT mice. Studies in primary myocytes suggested that defects in key lipogenic enzymes including FAS, stearoyl-CoA desaturase-1 (SCD1), and Lipin1 are likely contributing to reduced SERCA activity in mdx mice. Triple transgenic expression of FAS, SCD1, and Lipin1 (3TG) in mdx myocytes partly rescued SERCA activity, which coincided with an increase in SR PE that normalized PC/PE ratio. These findings implicate a defect in lipogenesis to be a contributing factor for SERCA dysfunction in muscular dystrophy. Restoration of muscle's lipogenic pathway appears to mitigate SERCA function through its effects on SR membrane composition.

  14. Calcium and proton dependence of sarcoplasmic reticulum ATPase.

    PubMed Central

    Inesi, G; Hill, T L

    1983-01-01

    The influence of Ca2+ and H+ concentrations on the sequential reactions of the ATPase cycle was studied by a series of pre-steady state and steady state experiments with sarcoplasmic reticulum vesicles. It is shown that H+ competition with calcium binding results in a reduced population of activated enzyme, which is manifested by a lower level of phosphorylated enzyme intermediate following addition of ATP. Further effects of Ca2+ and H+ are demonstrated on the progression of the phosphoenzyme through the reaction cycle and on the final hydrolytic cleavage of Pi. The overall dependence of steady state ATP flux on Ca2+ and H+ concentrations in leaky vesicles is expressed by a series of curves showing that as the H+ concentration is raised higher Ca2+ concentrations are required to obtain half-maximal ATP fluxes. At saturating Ca2+, maximal ATP fluxes are observed at an intermediate H+ concentration (pH 7.2), while lower levels are obtained as the H+ concentration is reduced (to pH 8) or increased (to pH 6). A preliminary model is then proposed based on the presence of two interacting domains permitting competitive binding of Ca2+ or H+, per each catalytic site undergoing phosphorylation by ATP. The model considers three main states and thirteen substates (depending on the occupancy of the binding sites in each state by Ca2+, H+, or neither) in the progression of the ATP cycle, coupled to transport of Ca2+ and counter transport of H+ in leaky vesicles. Considering the preliminary nature of the model and the experimental scatter, a rather satisfactory agreement is noted between a family of curves generated by theoretical analysis and the ATP flux curves obtained experimentally. PMID:6317076

  15. Regulation of RYR2 by sarcoplasmic reticulum Ca(2+).

    PubMed

    Zhang, Joe Z; Waddell, Helen M M; Jones, Peter P

    2015-06-01

    Ca(2+) is arguably the most important ion involved in the contraction of the heart. The cardiac ryanodine receptor (RyR2), the major Ca(2+) release channel located in the sarcoplasmic reticulum (SR) membrane, is responsible for releasing the bulk of Ca(2+) required for contraction. Moreover, RyR2 is also crucial for maintaining SR Ca(2+) homeostasis by releasing Ca(2+) from the SR when it becomes overloaded with Ca(2+) . During normal contraction, RyR2 is activated by cytosolic Ca(2+) , whereas during store overload conditions, the opening of RyR2 is governed by SR Ca(2+) . Although the process of the cytosolic control of RyR2 is well established, the molecular mechanism by which SR luminal Ca(2+) regulates RyR2 has only recently been elucidated and remains controversial. In addition to the activation of RyR2, SR luminal Ca(2+) also determines when the RyR2 channel closes. RyR2-mediated Ca(2+) release from the SR does not continue until the SR is completely depleted. Rather, it ceases when SR luminal Ca(2+) falls below a certain level. Given the importance of SR Ca(2+) , it is not surprising that the SR luminal Ca(2+) level is tightly controlled by SR Ca(2+) -buffering proteins. Consequently, the opening and closing of RyR2 is heavily influenced by the presence of such proteins, particularly those associated with RyR2, such as calsequestrin and the histidine-rich Ca(2+) -binding protein. These proteins appear to indirectly alter RyR2 activity by modifying the microdomain SR Ca(2+) level surrounding RyR2. PMID:25603835

  16. Striated boulder pavements within glaciomarine diamicts of the Yakataga Formation, Middleton Island, Alaska

    SciTech Connect

    Eyles, C.H.

    1985-01-01

    The presence of striated boulder pavements in glacial sequences is often cited as evidence of transport and deposition by grounded glacier ice. However, recent reports show that striated pavements also form in non-glacial environments by the abrasion of boulder lag surfaces by floating glacier and seasonal ice. Several striated boulder pavements are identified within Early Pleistocene upper Yakataga Formation sediments exposed on Middleton Island close to the southern edge of the Gulf of Alaska continental shelf. The sequence is dominated by thick stratiform units of massive and stratified diamict formed by the settling of fine-grained sands and muds from suspension together with ice-rafted debris. Boulder pavements outcrop as extensive planar horizons within the diamicts, can be traced for several kilometers along strike and consist of single lines of clasts with faceted upper surfaces showing consistently oriented striation directions. Clasts are not preferentially aligned, however, and do not have the characteristic bullet shape of boulders transported at a glacier base and deposited by lodgement processes. Striated boulder pavements on Middleton Island appear to have formed as boulder lag surfaces generated by wave and tidal current reworking of diamict on relatively shallow banks. Lags were then overridden and abraded by a grounding ice shelf. The glacially-abraded boulder pavements on Middleton Island record the repeated expansion of a continuous ice shelf to the edge of the Gulf of Alaska continental shelf during the Early Pleistocene.

  17. Ultrastructure of geniculocortical synaptic connections in the tree shrew striate cortex.

    PubMed

    Familtsev, Dmitry; Quiggins, Ranida; Masterson, Sean P; Dang, Wenhao; Slusarczyk, Arkadiusz S; Petry, Heywood M; Bickford, Martha E

    2016-04-15

    To determine whether thalamocortical synaptic circuits differ across cortical areas, we examined the ultrastructure of geniculocortical terminals in the tree shrew striate cortex to compare directly the characteristics of these terminals with those of pulvinocortical terminals (examined previously in the temporal cortex of the same species; Chomsung et al. [] Cereb Cortex 20:997-1011). Tree shrews are considered to represent a prototype of early prosimian primates but are unique in that sublaminae of striate cortex layer IV respond preferentially to light onset (IVa) or offset (IVb). We examined geniculocortical inputs to these two sublayers labeled by tracer or virus injections or an antibody against the type 2 vesicular glutamate antibody (vGLUT2). We found that layer IV geniculocortical terminals, as well as their postsynaptic targets, were significantly larger than pulvinocortical terminals and their postsynaptic targets. In addition, we found that 9-10% of geniculocortical terminals in each sublamina contacted GABAergic interneurons, whereas pulvinocortical terminals were not found to contact any interneurons. Moreover, we found that the majority of geniculocortical terminals in both IVa and IVb contained dendritic protrusions, whereas pulvinocortical terminals do not contain these structures. Finally, we found that synaptopodin, a protein uniquely associated with the spine apparatus, and telencephalin (TLCN, or intercellular adhesion molecule type 5), a protein associated with maturation of dendritic spines, are largely excluded from geniculocortical recipient layers of the striate cortex. Together our results suggest major differences in the synaptic organization of thalamocortical pathways in striate and extrastriate areas.

  18. Magnetization transfer imaging reveals geniculocalcarine and striate area degeneration in primary glaucoma: a preliminary study

    PubMed Central

    Zhang, Yan; Liang, Wenwen; Wu, Guijun; Zhang, Xuelin

    2016-01-01

    Background Glaucoma is a neurodegenerative disease that affects both the retina and central visual pathway. Magnetization transfer imaging (MTI) is a sensitive magnetic resonance imaging (MRI) technique that can detect degenerative changes in the brain. Purpose To investigate the geniculocalcarine (GCT) and striate areas in primary glaucoma patients using region of interest (ROI) analysis of magnetization transfer ratio (MTR). Material and Methods Twenty patients with primary glaucoma in both eyes were compared with 31 healthy control patients. All of the participants were examined on a 3.0 T scanner using a three-dimensional T1-weighted spoiled gradient recalled acquisition (SPGR) with and without a MT saturation pulse. A two-sample t-test was used to evaluate the MTR difference between the groups. P < 0.05 was used to determine statistical significance. Results The MTR of the glaucoma group was lower than the healthy controls in both the bilateral GCT (t = 3.781, P = 0.001) and striate areas (t = 4.177, P = 0.000). Conclusion The MTR reductions in the bilateral GCT and striate areas suggest that there is GCT demyelination and striate area degeneration in primary glaucoma. These neurodegenerative effects may be induced as a direct effect of retrograde axonal degeneration along with the indirect effect of anterograde trans-synaptic degeneration.

  19. The retinotopic organization of striate cortex is well predicted by surface topology

    PubMed Central

    Benson, Noah C.; Butt, Omar H.; Datta, Ritobrato; Radoeva, Petya D.; Brainard, David H.; Aguirre, Geoffrey Karl

    2012-01-01

    Summary In 1918, Gordon Holmes combined observations of visual field scotomas across brain lesioned soldiers to produce a schematic map of the projection of the visual field upon the striate cortex [1]. One limit to the precision of his result, and the mapping of anatomy to retinotopy generally, is the substantial individual variation in the size [2,3], volumetric position [4], and cortical magnification [5] of area V1. When viewed within the context of the curvature of the cortical surface, however, the boundaries of striate cortex fall at a consistent location across individuals [6]. We asked if the surface topology of the human brain can be used to accurately predict the internal, retinotopic function of striate cortex as well. We used fMRI to measure polar angle and eccentricity in 25 participants and combined their maps within a left-right, transform-symmetric representation of the cortical surface [7]. These data were then fit using a deterministic, algebraic model of visual field representation [8]. We found that an anatomical image alone can be used to predict the retinotopic organization of striate cortex for an individual as accurately as 10–25 minutes of functional mapping. This indicates tight developmental linkage of structure and function within a primary, sensory cortical area. PMID:23041195

  20. Structure and protein composition of the striated flagellar rootlets of some protists.

    PubMed

    Dingle, A D; Larson, D E

    1981-01-01

    The striated rootlets of different protists are extremely diverse and, on the basis of structural organization, can be assigned to no fewer than four major types. In light of this extreme variation in fine-structure is it reasonable to expect that all striated rootlets may share common protein species? Using the flagellar rootlet of Naegleria gruberi strain NB-1 as reference, we compared rootlet size, structure, and protein composition to another Naegleria strain, another amoebo-flagellate, Tetramitus rostratus, and to the ciliate Tetrahymena pyriformis. Although differing in size and distribution in the cells, the rootlets of all three amoebo-flagellates appeared very similar in structure, periodicity, and in the presence of a common 170 000 Dalton subunit. Kinetodesmal fibres of Tetrahymena differed markedly in detailed fine-structure, in periodicity, and in the apparent absence of the 170 000 Dalton subunit as tested either by SDS gel electrophoresis or by indirect immunofluorescence staining using a specific antiserum directed against the NB-1 major rootlet protein. Consideration of literature describing striated rootlet structure in a wide variety of ciliated and flagellated cells led to the speculation that striated rootlets arose subsequent to primitive flagella and likely evolved along two major pathways: a narrow-period rootlet similar to those discussed above, which developed from the microtubule rootlets of algal flagellates, and a wide-band, contractile rootlet which originated from the primitive interbasal body connector prominent in both algal and protozoan species.

  1. Magnetization transfer imaging reveals geniculocalcarine and striate area degeneration in primary glaucoma: a preliminary study

    PubMed Central

    Zhang, Yan; Liang, Wenwen; Wu, Guijun; Zhang, Xuelin

    2016-01-01

    Background Glaucoma is a neurodegenerative disease that affects both the retina and central visual pathway. Magnetization transfer imaging (MTI) is a sensitive magnetic resonance imaging (MRI) technique that can detect degenerative changes in the brain. Purpose To investigate the geniculocalcarine (GCT) and striate areas in primary glaucoma patients using region of interest (ROI) analysis of magnetization transfer ratio (MTR). Material and Methods Twenty patients with primary glaucoma in both eyes were compared with 31 healthy control patients. All of the participants were examined on a 3.0 T scanner using a three-dimensional T1-weighted spoiled gradient recalled acquisition (SPGR) with and without a MT saturation pulse. A two-sample t-test was used to evaluate the MTR difference between the groups. P < 0.05 was used to determine statistical significance. Results The MTR of the glaucoma group was lower than the healthy controls in both the bilateral GCT (t = 3.781, P = 0.001) and striate areas (t = 4.177, P = 0.000). Conclusion The MTR reductions in the bilateral GCT and striate areas suggest that there is GCT demyelination and striate area degeneration in primary glaucoma. These neurodegenerative effects may be induced as a direct effect of retrograde axonal degeneration along with the indirect effect of anterograde trans-synaptic degeneration. PMID:27651931

  2. Changes in meat quality traits and sarcoplasmic proteins during aging in three different cattle breeds.

    PubMed

    Marino, R; Albenzio, M; della Malva, A; Caroprese, M; Santillo, A; Sevi, A

    2014-10-01

    The effects of breed and aging time (1, 7, 14, 21d) were evaluated on physical meat properties and on sarcoplasmic protein changes in 24 young bulls from Romagnola×Podolian, Podolian and Friesian breeds. Aging affects lightness showing an increase in all breeds while changes in redness varied according to the breed. Podolian breed showed meat with the darkest and the reddest color and the lowest drip loss compared to the other breeds. Extending aging to 21d reduced drip loss from meat. SDS-PAGE and 2DE showed that many changes in the sarcoplasmic proteins occurred among breeds and during aging. During post-mortem some sarcoplasmic proteins decline in intensity after 21d highlighting that they were susceptible to aging. Protein identification and western blotting showed the presence of myosin light chains, Troponin T and tropomyosin proteins during aging, suggesting a degradation of myofibers and a more intense proteolysis especially in the Podolian breed.

  3. Preservative solution for skeletal muscle biopsy samples

    PubMed Central

    Kurt, Yasemin Gulcan; Kurt, Bulent; Ozcan, Omer; Topal, Turgut; Kilic, Abdullah; Muftuoglu, Tuba; Acikel, Cengizhan; Sener, Kenan; Sahiner, Fatih; Yigit, Nuri; Aydin, Ibrahim; Alay, Semih; Ekinci, Safak

    2015-01-01

    Context: Muscle biopsy samples must be frozen with liquid nitrogen immediately after excision and maintained at -80°C until analysis. Because of this requirement for tissue processing, patients with neuromuscular diseases often have to travel to centers with on-site muscle pathology laboratories for muscle biopsy sample excision to ensure that samples are properly preserved. Aim: Here, we developed a preservative solution and examined its protectiveness on striated muscle tissues for a minimum of the length of time that would be required to reach a specific muscle pathology laboratory. Materials and Methods: A preservative solution called Kurt-Ozcan (KO) solution was prepared. Eight healthy Sprague-Dawley rats were sacrificed; striated muscle tissue samples were collected and divided into six different groups. Muscle tissue samples were separated into groups for morphological, enzyme histochemical, molecular, and biochemical analysis. Statistical method used: Chi-square and Kruskal Wallis tests. Results: Samples kept in the KO and University of Wisconsin (UW) solutions exhibited very good morphological scores at 3, 6, and 18 hours, but artificial changes were observed at 24 hours. Similar findings were observed for the evaluated enzyme activities. There were no differences between the control group and the samples kept in the KO or UW solution at 3, 6, and 18 hours for morphological, enzyme histochemical, and biochemical features. The messenger ribonucleic acid (mRNA) of β-actin gene was protected up to 6 hours in the KO and UW solutions. Conclusion: The KO solution protects the morphological, enzyme histochemical, and biochemical features of striated muscle tissue of healthy rats for 18 hours and preserves the mRNA for 6 hours. PMID:26019417

  4. Specific binding of (/sup 3/H)LY186126, an analogue of indolidan (LY195115), to cardiac membranes enriched in sarcoplasmic reticulum vesicles

    SciTech Connect

    Kauffman, R.F.; Utterback, B.G.; Robertson, D.W.

    1989-05-01

    LY186126 was found to be a potent inhibitor of type IV cyclic AMP phosphodiesterase located in the sarcoplasmic reticulum of canine cardiac muscle. This compound, a close structural analogue of indolidan (LY195115), was prepared in high specific activity, tritiated form to study the positive inotropic receptor(s) for cardiotonic phosphodiesterase inhibitors such as indolidan and milrinone. A high-affinity binding site for (/sup 3/H)LY186126 was observed (Kd = 4 nM) in purified preparations of canine cardiac sarcoplasmic reticulum vesicles. Binding was proportional to vesicle protein, was inactivated by subjecting membranes to proteolysis or boiling, and was dependent on added Mg2+. Scatchard analysis suggested the presence of a single class of binding sites in the membrane preparation. Indolidan, milrinone, and LY186126 (all at 1 microM) produced essentially complete displacement of bound (/sup 3/H)LY186126, while nifedipine, propranolol, and prazosin had little or no effect at this concentration. This represents the first reported use of a radioactive analogue to label the inotropic receptor for cardiotonic phosphodiesterase inhibitors. The results suggest that (/sup 3/H)LY186126 is a useful radioligand for examining the subcellular site(s) responsible for positive inotropic effects of these drugs.

  5. Short-term effects of β2-AR blocker ICI 118,551 on sarcoplasmic reticulum SERCA2a and cardiac function of rats with heart failure

    PubMed Central

    Gong, Haibin; Li, Yanfei; Wang, Lei; Lv, Qian; Wang, Xiuli

    2016-01-01

    The study was conducted to examine the effects of ICI 118,551 on the systolic function of cardiac muscle cells of rats in heart failure and determine the molecular mechanism of selective β2-adrenergic receptor (β2-AR) antagonist on these cells. The chronic heart failure model for rats was prepared through abdominal aortic constriction and separate cardiac muscle cells using the collagenase digestion method. The rats were then divided into Sham, HF and HF+ICI 50 nM goups and cultivated for 48 h. β2-AR, Gi/Gs and sarcoplasmic reticulum Ca2+-ATPase (SERCA2a) protein expression levels in the cardiac muscle cells were evaluated by western blotting and changes in the systolic function of cardiac muscle cells based on the boundary detection system of contraction dynamics for individual cells was measured. The results showed that compared with the Sham group, the survival rate, percentage of basic contraction and maximum contraction amplitude percentage of cardiac muscle cells with heart failure decreased, Gi protein expression increased while Gs and SERCA2a protein expression decreased. Compared with the HF group, the maximum contraction amplitude percentage of cardiac muscle cells in group HF+ICI 50 nM decreased, the Gi protein expression level increased while the SERCA2a protein expression level decreased. Following the stimulation of Ca2+ and ISO, the maximum contraction amplitude percentage of cardiac muscle cells in the HF+ICI 50 nM group was lower than that in group HF. This indicated that ICI 118,551 has negative inotropic effects on cardiac muscle cells with heart failure, which may be related to Gi protein. Systolic function of cardiac muscle cells with heart failure can therefore be reduced by increasing Gi protein expression and lowering SERCA2a protein expression.

  6. Short-term effects of β2-AR blocker ICI 118,551 on sarcoplasmic reticulum SERCA2a and cardiac function of rats with heart failure

    PubMed Central

    Gong, Haibin; Li, Yanfei; Wang, Lei; Lv, Qian; Wang, Xiuli

    2016-01-01

    The study was conducted to examine the effects of ICI 118,551 on the systolic function of cardiac muscle cells of rats in heart failure and determine the molecular mechanism of selective β2-adrenergic receptor (β2-AR) antagonist on these cells. The chronic heart failure model for rats was prepared through abdominal aortic constriction and separate cardiac muscle cells using the collagenase digestion method. The rats were then divided into Sham, HF and HF+ICI 50 nM goups and cultivated for 48 h. β2-AR, Gi/Gs and sarcoplasmic reticulum Ca2+-ATPase (SERCA2a) protein expression levels in the cardiac muscle cells were evaluated by western blotting and changes in the systolic function of cardiac muscle cells based on the boundary detection system of contraction dynamics for individual cells was measured. The results showed that compared with the Sham group, the survival rate, percentage of basic contraction and maximum contraction amplitude percentage of cardiac muscle cells with heart failure decreased, Gi protein expression increased while Gs and SERCA2a protein expression decreased. Compared with the HF group, the maximum contraction amplitude percentage of cardiac muscle cells in group HF+ICI 50 nM decreased, the Gi protein expression level increased while the SERCA2a protein expression level decreased. Following the stimulation of Ca2+ and ISO, the maximum contraction amplitude percentage of cardiac muscle cells in the HF+ICI 50 nM group was lower than that in group HF. This indicated that ICI 118,551 has negative inotropic effects on cardiac muscle cells with heart failure, which may be related to Gi protein. Systolic function of cardiac muscle cells with heart failure can therefore be reduced by increasing Gi protein expression and lowering SERCA2a protein expression. PMID:27602067

  7. Muscle as a “Mediator“ of Systemic Metabolism

    PubMed Central

    Baskin, Kedryn K.; Winders, Benjamin R.; Olson, Eric N.

    2015-01-01

    Skeletal and cardiac muscles play key roles in the regulation of systemic energy homeostasis and display remarkable plasticity in their metabolic responses to caloric availability and physical activity. In this Perspective we discuss recent studies highlighting transcriptional mechanisms that govern systemic metabolism by striated muscles. We focus on the participation of the Mediator complex in this process, and suggest that tissue-specific regulation of Mediator subunits impacts metabolic homeostasis. PMID:25651178

  8. Assessment of the angiotensin-I-converting enzyme (ACE-I) inhibitory and antioxidant activities of hydrolysates of bovine brisket sarcoplasmic proteins produced by papain and characterisation of associated bioactive peptidic fractions.

    PubMed

    Di Bernardini, Roberta; Mullen, Anne Maria; Bolton, Declan; Kerry, Joseph; O'Neill, Eileen; Hayes, Maria

    2012-01-01

    The main objective was to investigate the angiotensin-I-converting enzyme (ACE-I) inhibitory and antioxidant activities of sarcoplasmic proteins isolated from the brisket muscle (Pectoralis profundus) of 3 (Bos taurus) cattle and hydrolysed with papain for 24 h at 37°C. Sarcoplasmic protein hydrolysates were ultra-filtered using molecular weight cut off (MWCO) membranes and 10-kDa and 3-kDa filtrates were obtained. The total sarcoplasmic protein extracts and the 3-kDa filtrates were tested for angiotensin I-converting enzyme inhibitory (ACE-I) activities. The total hydrolysates, 10-kDa and 3-kDa filtrates were also tested for their associated antioxidant activities using the 2,2-diphenyl-1-picrylhydrazyl (DPPH) radical scavenging activity assay, the ferric ion reducing antioxidant power (FRAP) assay and the Fe(2+) metal chelating ability assay. The peptidic content of the total hydrolysates, the 10-kDa and the 3-kDa filtrates were analysed using an ORBITRAP mass spectrometer, and mass spectral data obtained were analysed using TurboSEQUEST. Eleven peptides were characterised from the total hydrolysates, fifteen from the 10-kDa filtrate fractions, whilst nine peptides were characterised from the 3-kDa filtrate fractions. Similarities between the amino acid sequences of the peptides identified in this study and previously identified antioxidant and ACE-I inhibitory peptides detailed in the BIOPEP database were outlined. PMID:21880436

  9. Sarcoplasmic Reticulum Calcium Release Channels in Ventricles of Older Adult Hamsters

    ERIC Educational Resources Information Center

    Nicholl, Peter A.; Howlett, Susan E.

    2006-01-01

    Whether the density of sarcoplasmic reticulum (SR) calcium release channels/ryanodine receptors in the heart declines with age is not clear. We investigated age-related changes in the density of [3H]-ryanodine receptors in crude ventricular homogenates, which contained all ligand binding sites in heart and in isolated junctional SR membranes.…

  10. Effect of hydrodynamic pressure processing and aging on sarcoplasmic proteins of beef strip loins

    Technology Transfer Automated Retrieval System (TEKTRAN)

    This study evaluated the effects of hydrodynamic pressure processing (HDP) and aging on the sarcoplasmic proteins of beef strip loins. Loins (n=12) were halved at 48 h postmortem and assigned to HDP or control treatments. Following treatment, each half was divided into three portions for aging (0, 5...

  11. Effects of postmortem aging and hydrodynamic pressure processing on sarcoplasmic proteins and beef tenderness

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The objective of this study was to determine the effects of hydrodynamic pressure processing (HDP) and postmortem aging on the sarcoplasmic protein fraction of beef strip loins. Loins (n=12) were halved at 48 h postmortem and assigned to HDP or control treatments. Following treatment, each half was ...

  12. Cytoplasmic nanojunctions between lysosomes and sarcoplasmic reticulum are required for specific calcium signaling

    PubMed Central

    Fameli, Nicola; Ogunbayo, Oluseye A.

    2014-01-01

    Herein we demonstrate how nanojunctions between lysosomes and sarcoplasmic reticulum (L-SR junctions) serve to couple lysosomal activation to regenerative, ryanodine receptor-mediated cellular Ca 2+ waves. In pulmonary artery smooth muscle cells (PASMCs) it has been proposed that nicotinic acid adenine dinucleotide phosphate (NAADP) triggers increases in cytoplasmic Ca 2+ via L-SR junctions, in a manner that requires initial Ca 2+ release from lysosomes and subsequent Ca 2+-induced Ca 2+ release (CICR) via ryanodine receptor (RyR) subtype 3 on the SR membrane proximal to lysosomes. L-SR junction membrane separation has been estimated to be < 400 nm and thus beyond the resolution of light microscopy, which has restricted detailed investigations of the junctional coupling process. The present study utilizes standard and tomographic transmission electron microscopy to provide a thorough ultrastructural characterization of the L-SR junctions in PASMCs. We show that L-SR nanojunctions are prominent features within these cells and estimate that the junctional membrane separation and extension are about 15 nm and 300 nm, respectively. Furthermore, we develop a quantitative model of the L-SR junction using these measurements, prior kinetic and specific Ca 2+ signal information as input data. Simulations of NAADP-dependent junctional Ca 2+ transients demonstrate that the magnitude of these signals can breach the threshold for CICR via RyR3. By correlation analysis of live cell Ca 2+ signals and simulated Ca 2+ transients within L-SR junctions, we estimate that “trigger zones” comprising 60–100 junctions are required to confer a signal of similar magnitude. This is compatible with the 110 lysosomes/cell estimated from our ultrastructural observations. Most importantly, our model shows that increasing the L-SR junctional width above 50 nm lowers the magnitude of junctional [Ca 2+] such that there is a failure to breach the threshold for CICR via RyR3. L-SR junctions are

  13. Non-conscious recognition of affect in the absence of striate cortex.

    PubMed

    de Gelder, B; Vroomen, J; Pourtois, G; Weiskrantz, L

    1999-12-16

    Functional neuroimaging experiments have shown that recognition of emotional expressions does not depend on awareness of visual stimuli and that unseen fear stimuli can activate the amygdala via a colliculopulvinar pathway. Perception of emotional expressions in the absence of awareness in normal subjects has some similarities with the unconscious recognition of visual stimuli which is well documented in patients with striate cortex lesions (blindsight). Presumably in these patients residual vision engages alternative extra-striate routes such as the superior colliculus and pulvinar. Against this background, we conjectured that a blindsight subject (GY) might recognize facial expressions presented in his blind field. The present study now provides direct evidence for this claim. PMID:10716205

  14. Effects of calmodulin and calmodulin inhibitors on Ca uptake by sarcoplasmic reticulum of saponin skinned caudal artery

    SciTech Connect

    Stout, M.A.; Silver, P.J.

    1986-03-05

    Calmodulin (CaM) stimulates plasma membrane transport in many cell types, however, its role in Ca regulation by the sarcoplasmic reticulum (SR) in smooth muscle has not been established. /sup 45/Ca uptake was studied in saponin skinned strips of rat caudal artery as a function of CaM and the CaM inhibitors, W-7, calmidazolium (CaMZ), and trifluoperazine (TFP). Although caudal artery strips lose approximately 30% of total tissue CaM during skinning, 0.3 - 2 ..mu..M CaM did not increase /sup 45/Ca uptake over a wide range of free Ca concentrations (10/sup -8/ - 10/sup -6/M). Neither W-7 nor CaMZ at concentration of 10/sup -4/ - 2 x 10/sup -4/M inhibited the MgATP-dependent Ca uptake. Ca uptake was not affected by 50 ..mu..M TFP but a significant inhibition was produced by 500 ..mu..M. Studies of the effects of TFP on /sup 45/Ca efflux indicated that TFP concentrations which inhibited Ca uptake also significantly increased the rate of Ca release. The results suggest that total Ca uptake in caudal artery depends mainly upon MgATP and is not modulated by exogenous CaM or affected by these CaM inhibitors. They cannot preclude that CaM may affect initial velocities or that the CaM inhibitors failed to reach active sites.

  15. Modulation of sarcoplasmic/endoplasmic reticulum Ca(2+)-ATPase activity and oxidative modification during the development of adjuvant arthritis.

    PubMed

    Strosova, Miriam K; Karlovska, Janka; Zizkova, Petronela; Kwolek-Mirek, Magdalena; Ponist, Silvester; Spickett, Corinne M; Horakova, Lubica

    2011-07-01

    Adjuvant arthritis (AA) was induced by intradermal administration of Mycobacterium butyricum to the tail of Lewis rats. In sarcoplasmic reticulum (SR) of skeletal muscles, we investigated the development of AA. SR Ca(2+)-ATPase (SERCA) activity decreased on day 21, suggesting possible conformational changes in the transmembrane part of the enzyme, especially at the site of the calcium binding transmembrane part. These events were associated with an increased level of protein carbonyls, a decrease in cysteine SH groups, and alterations in SR membrane fluidity. There was no alteration in the nucleotide binding site at any time point of AA, as detected by a FITC fluorescence marker. Some changes observed on day 21 appeared to be reversible, as indicated by SERCA activity, cysteine SH groups, SR membrane fluidity, protein carbonyl content and fluorescence of an NCD-4 marker specific for the calcium binding site. The reversibility may represent adaptive mechanisms of AA, induced by higher relative expression of SERCA, oxidation of cysteine, nitration of tyrosine and presence of acidic phospholipids such as phosphatidic acid. Nitric oxide may regulate cytoplasmic Ca(2+) level through conformational alterations of SERCA, and decreasing levels of calsequestrin in SR may also play regulatory role in SERCA activity and expression. PMID:21531199

  16. Distinct regions of triadin are required for targeting and retention at the junctional domain of the sarcoplasmic reticulum.

    PubMed

    Rossi, Daniela; Bencini, Cristina; Maritati, Marina; Benini, Francesca; Lorenzini, Stefania; Pierantozzi, Enrico; Scarcella, Angela Maria; Paolini, Cecilia; Protasi, Feliciano; Sorrentino, Vincenzo

    2014-03-01

    Ca2+ release, which is necessary for muscle contraction, occurs at the j-SR (junctional domain of the sarcoplasmic reticulum). It requires the assembly of a large multiprotein complex containing the RyR (ryanodine receptor) and additional proteins, including triadin and calsequestrin. The signals which drive these proteins to the j-SR and how they assemble to form this multiprotein complex are poorly understood. To address aspects of these questions we studied the localization, dynamic properties and molecular interactions of triadin. We identified three regions, named TR1 (targeting region 1), TR2 and TR3, that contribute to the localization of triadin at the j-SR. FRAP experiments showed that triadin is stably associated with the j-SR and that this association is mediated by TR3. Protein pull-down experiments indicated that TR3 contains binding sites for calsequestrin-1 and that triadin clustering can be enhanced by binding to calsequestrin-1. These findings were confirmed by FRET experiments. Interestingly, the stable association of triadin to the j-SR was significantly decreased in myotubes from calsequestrin-1 knockout mice. Taken together, these results identify three regions in triadin that mediate targeting to the j-SR and reveal a role for calsequestrin-1 in promoting the stable association of triadin to the multiprotein complex associated with RyR. PMID:24325401

  17. Localization of the hinge region of the Ca(2+)-ATPase of sarcoplasmic reticulum using resonance energy transfer.

    PubMed

    Baker, K J; East, J M; Lee, A G

    1994-06-01

    The Ca(2+)-ATPase of skeletal muscle sarcoplasmic reticulum can be labelled at Cys-670 and Cys-674 with 5-[[2-[(iodoacetyl) amino]ethyl]amino]naphthalene-1-sulphonic acid (IAEDANS). Resonance energy transfer has been used to measure the distance between Cys-670/Cys-674 and Glu-439 labelled with 5-(bromomethyl)fluorescein as 40 A. The height of Cys-670/Cys-674 above the phospholipid/water interface has been measured by resonance energy transfer between IAEDANS-labelled ATPase and fluorescein-labelled phosphatidylethanolamine as 54 A. This locates the hinge region of the ATPase close to the mouth of the pore observed in the cytoplasmic region of the ATPase in electron micrographs. No significant changes in these distances can be detected by resonance energy transfer on binding Ca2+ or vanadate. The height of the IAEDANS label above the phospholipid/water interface is the same for bilayers of dimyristoleoylphosphatidylcholine and dioleoylphosphatidylcholine. Conformation changes on the Ca(2+)-ATPase appear to be localised to small regions of the ATPase.

  18. Thermodynamic and kinetic studies of the gating behavior of a K+- selective channel from the sarcoplasmic reticulum membrane

    PubMed Central

    1980-01-01

    A voltage-dependent, K+-selective ionic channel from sarcoplasmic reticulum of rabbit skeletal muscle has been studied in a planar phospholipid bilayer membrane. The puppose of this work is to study the mechanism by which the channel undergoes transitions between its conducting and nonconducting states. Thermodynamic studies show that the "open" and "closed" states of the channel exist in a voltage- dependent equilibrium, and that the channel displays only a single open state; the channel conductance is 120 pmho in 0.1 M K+. The channel's gating process follows single exponential kinetics at all voltages tested, and the individual opening and closing rate constants are exponentially dependent on voltage. The individual rate constants may also be determined from a stochastic analysis of channel fluctuations among multiple conductance levels. Neither the thermodynamic nor the kinetic parameters of gating depend on the absolute concentration of channels in the bilayer. The results are taken as evidence that the channel gates by an unusually simple two-state conformational mechanism in which the equivalent of 1.1 net charges are moved across the membrane during the formation of the open channel. PMID:6255061

  19. Modulation of sarcoplasmic/endoplasmic reticulum Ca(2+)-ATPase activity and oxidative modification during the development of adjuvant arthritis.

    PubMed

    Strosova, Miriam K; Karlovska, Janka; Zizkova, Petronela; Kwolek-Mirek, Magdalena; Ponist, Silvester; Spickett, Corinne M; Horakova, Lubica

    2011-07-01

    Adjuvant arthritis (AA) was induced by intradermal administration of Mycobacterium butyricum to the tail of Lewis rats. In sarcoplasmic reticulum (SR) of skeletal muscles, we investigated the development of AA. SR Ca(2+)-ATPase (SERCA) activity decreased on day 21, suggesting possible conformational changes in the transmembrane part of the enzyme, especially at the site of the calcium binding transmembrane part. These events were associated with an increased level of protein carbonyls, a decrease in cysteine SH groups, and alterations in SR membrane fluidity. There was no alteration in the nucleotide binding site at any time point of AA, as detected by a FITC fluorescence marker. Some changes observed on day 21 appeared to be reversible, as indicated by SERCA activity, cysteine SH groups, SR membrane fluidity, protein carbonyl content and fluorescence of an NCD-4 marker specific for the calcium binding site. The reversibility may represent adaptive mechanisms of AA, induced by higher relative expression of SERCA, oxidation of cysteine, nitration of tyrosine and presence of acidic phospholipids such as phosphatidic acid. Nitric oxide may regulate cytoplasmic Ca(2+) level through conformational alterations of SERCA, and decreasing levels of calsequestrin in SR may also play regulatory role in SERCA activity and expression.

  20. THE STRUCTURE OF INSECT FIBRILLAR FLIGHT MUSCLE

    PubMed Central

    Smith, David S.

    1961-01-01

    The fine structure of fibrillar flight muscle of the mature adult beetle Tenebrio molitor is described. Although the very high frequency of contraction of fibrillar muscle has previously been in part accounted for as the result of mechanical specialization of the wing-bearing segment rather than of a correspondingly high rate of motor impulse supply, the problem of the nature of the pathway by which excitation is conducted into these large fibers remained. Therefore, particular attention has been given to the disposition and relationships of the plasma membrane and sarcoplasmic reticulum in this tissue. The invading tracheoles draw with them a sheath of plasma membrane from the surface to all depths in the fiber, and it is suggested that these sheaths, together with the extensive tubular arborisations arising from them, reduce the maximum plasma membrane-to-fibril distance from the radius of the fiber to a value of less than 2 µ. The evidence presented here confirms Veratti's contention that in fibrillar muscle the "reticulum" is associated with, though entirely distinct from the fibrils. Unlike other muscles so far examined, these flight muscle fibers contain a plasma membrane reticulum only, but it is possible that elsewhere the general "sarcoplasmic reticulum" includes a component derived from the plasma membrane, likewise acting as the pathway for inward conduction of excitation. Profiles of the internalised plasma membrane in Tenebrio showing the usual triple-layered 25-25-25 A organization are frequently seen, in sections, in close association with isolated vesicles (defined by "simple" 50 A membranes) which are here considered to represent, in vestigial form, the portion of the sarcoplasmic reticulum which in other types of muscle is complex and highly developed. Such associations, in Tenebrio, between these two dissimilar elements are here termed "dyads" and the possible morphological and functional homology between these and the "triads" of other types of

  1. Neural correlates of visual motion processing without awareness in patients with striate cortex and pulvinar lesions.

    PubMed

    Barleben, Maria; Stoppel, Christian M; Kaufmann, Jörn; Merkel, Christian; Wecke, Thoralf; Goertler, Michael; Heinze, Hans-Jochen; Hopf, Jens-Max; Schoenfeld, Mircea A

    2015-04-01

    Patients with striate cortex lesions experience visual perception loss in the contralateral visual field. In few patients, however, stimuli within the blind field can lead to unconscious (blindsight) or even conscious perception when the stimuli are moving (Riddoch syndrome). Using functional magnetic resonance imaging (fMRI), we investigated the neural responses elicited by motion stimulation in the sighted and blind visual fields of eight patients with lesions of the striate cortex. Importantly, repeated testing ensured that none of the patients exhibited blindsight or a Riddoch syndrome. Three patients had additional lesions in the ipsilesional pulvinar. For blind visual field stimulation, great care was given that the moving stimulus was precisely presented within the borders of the scotoma. In six of eight patients, the stimulation within the scotoma elicited hemodynamic activity in area human middle temporal (hMT) while no activity was observed within the ipsilateral lesioned area of the striate cortex. One of the two patients in whom no ipsilesional activity was observed had an extensive lesion including massive subcortical damage. The other patient had an additional focal lesion within the lateral inferior pulvinar. Fiber-tracking based on anatomical and functional markers (hMT and Pulvinar) on individual diffusion tensor imaging (DTI) data from each patient revealed the structural integrity of subcortical pathways in all but the patient with the extensive subcortical lesion. These results provide clear evidence for the robustness of direct subcortical pathways from the pulvinar to area hMT in patients with striate cortex lesions and demonstrate that ipsilesional activity in area hMT is completely independent of conscious perception. PMID:25529748

  2. Visual receptive fields in the striate-recipient zone of the lateral posterior-pulvinar complex.

    PubMed

    Chalupa, L M; Abramson, B P

    1989-01-01

    The lateral posterior (LP)-pulvinar complex of the cat is known to contain multiple visual areas. In the present study, we examined the receptive field properties of single neurons isolated in the lateral division of this complex (the LPI). The LPI is designated the striate-recipient zone because it is the only region of the LP-pulvinar receiving cortical projections from areas 17 and 18. The recordings revealed that the striate-recipient zone of LP comprises 2 subareas, which we have termed LPI-1 and LPI-2. In the main segment (LPI-1), virtually all cells responded securely to visual stimuli. The vast majority of these neurons were binocular, with relatively small and well-defined receptive fields. More than half of the cells were found to be directionally selective, and almost this many were orientation specific. The orientation tuning of these cells was found to be quite precise, comparable to complex cells in area 17. In contrast, in the small dorsolateral segment of the striate-recipient zone (the LPI-2), a substantial proportion of cells could not be visually activated. Here, the visual cells had very large receptive fields, and relatively few were direction or orientation selective. The LPI-2 receives subcortical inputs from the superficial layers of the superior colliculus, the hypothalamus, and cerebellum, while the LPI-1 is innervated only by cortical axons. It is suggested that the subcortical connections of the LPI-2 account for the differences in the response properties of the 2 striate-recipient areas. The present results, in conjunction with our previous findings on the principal tectorecipient zone (Chalupa et al., 1983), permit 2 generalizations regarding the functional organization of the cat's LP-pulvinar complex. First, there are clear differences among the visual areas of the LP-pulvinar in the cellular processing of visual information. Second, these functional differences can be related to the principal sources of visual input to the various

  3. Two-dimensional separation of the membrane protein sarcoplasmic reticulum Ca-ATPase for high-performance liquid chromatography-tandem mass spectrometry analysis of posttranslational protein modifications.

    PubMed

    Sharov, Victor S; Galeva, Nadezhda A; Knyushko, Tatyana V; Bigelow, Diana J; Williams, Todd D; Schöneich, Christian

    2002-09-15

    For the characterization of posttranslational modifications of the sarcoplasmic/endoplasmic reticulum Ca-ATPase (SERCA), we developed a two-dimensional separation protocol based on reversed-phase HPLC followed by SDS-PAGE and LC-MS/MS analysis of in-gel tryptic digests. Representative experiments are shown for the rabbit fast-twitch skeletal muscle isoform SERCA1. Matrix-assisted laser desorption-ionization and electrospray ionization-mass spectrometry analyses of SERCA1 tryptic digests revealed ca. 66% coverage of the protein sequence. This approach was used for the detection and quantitation of nitrotyrosine formation after exposure of SERCA1 to peroxynitrite in vitro. At molar ratios of nitrotyrosine to protein of 0.23 we confirmed by LC-MS/MS the nitration of predominantly Tyr(122) in the SERCA1 sequence. PMID:12419347

  4. Food control by applied biochemistry of marine organisms: Comparison of proteins and metabolites from fish and invertebrate muscle

    NASA Astrophysics Data System (ADS)

    Rehbein, H.

    1995-03-01

    Most fishery products consist of muscle tissue from fish and invertebrates. Differences in the molecular structure and in metabolism of muscles can be utilized to characterize and identify various seafood. Creatine and arginine were found to be useful for the differentiation between imitation crab/shrimp meat and real crustacean meat. Octopine served as an indicator for the meat of cephalopods and mussels. In order to identify the animal species of a fishery product, several electrophoretic methods were used. It depended on the type of product, whether sarcoplasmic or myofibrillar proteins were better suited. Raw products were best analysed by isoelectric focusing of sarcoplasmic proteins. Two types of sarcoplasmic calcium-binding proteins, parvalbumins of fish and soluble calcium-binding proteins of invertebrates, were especially useful for species identification. Due to their thermal stability, these proteins gave species-specific patterns for cooked products, too. Two other techniques were also investigated: urea gel isoelectric focusing, and sodium dodecyl sulphate — polyacrylamide gel electrophoresis. These methods were applied in the analysis of products where the sarcoplasmic proteins had been removed by washing steps, i.e. imitation crab meat made from surimi, and of other raw and cooked products. The myosin light chains gave protein patterns that were characteristic for many species. Paramyosin, which is absent from vertebrate muscle, indicated the presence of mollusc muscle. It was shown that the determination, of arginine kinase activity enabled differentiation between raw fish muscle and invertebrate muscles.

  5. Identification and localization of caldesmon in cardiac muscle.

    PubMed Central

    Scott-Woo, G C; Walsh, M P; Ikebe, M; Kargacin, G J

    1998-01-01

    Caldesmon has been detected in smooth muscle and in a number of non-muscle cells. It binds both actin and myosin and may act as a regulator of contraction or a structural element in smooth muscle. The presence of caldesmon in striated muscle has not been well established. To address this issue, polyclonal antibodies and a panel of monoclonal antibodies were raised against chicken gizzard smooth muscle caldesmon and used to demonstrate that caldesmon is present in adult cardiac muscle of a variety of mammalian species. Western-blot analysis revealed the presence of caldesmon in ventricular myocytes isolated from rat heart. The epitopes for the individual monoclonal antibodies were mapped to the caldesmon primary structure using chymotryptic and 2-nitro-5-thiocyanatobenzoic acid fragments. Bovine and rat cardiac caldesmons were recognized only by a subset of these monoclonal antibodies, indicating primary sequence differences from the chicken smooth muscle protein. Immunofluorescence labelling of isolated myocytes from rat, rabbit and guinea pig cardiac muscle revealed a striated pattern of fluorescence labelling. Dual labelling of caldesmon and myosin or caldesmon and alpha-actinin demonstrated that caldesmon was present at the centre of the I-band rather than in the A-band, as might have been expected from the myosin binding properties of the smooth muscle protein. These results suggest a structural role for caldesmon in cardiac muscle cells. PMID:9693116

  6. A new hypothesis for Ca2+ flows in skeletal muscle and its implications for other cell types.

    PubMed

    Islam, Mohammad Naimul; Ochs, Raymond S

    2006-01-01

    We offer a new hypothesis to explain calcium flows in skeletal muscle cells. Our model accounts for the uptake of Ca2+ from the extracellular fluid, and the release of Ca2+ from the sarcoplasmic reticulum (SR/ER) (the endoplasmic reticulum in muscle is named sarcoplasmic reticulum); this has engendered difficulty in reviews encompassing both muscle and nonmuscle cells. Here we will typically refer to the organelle as ER, except when specifically discussing muscle cells. The broad consideration of two major, still unexplained properties of skeletal muscle function, namely excitation contraction coupling and capacitative calcium entry are accounted for in a unitary hypothesis. This model allows a reinterpretation of existing data, and points to areas where new investigation may be fruitful. While primarily aimed at explaining Ca2+ flows in skeletal muscle, we consider findings of other systems to explore the implications of this hypothesis for other cell types.

  7. Methods for Creating and Animating a Computer Model Depicting the Structure and Function of the Sarcoplasmic Reticulum Calcium ATPase Enzyme.

    ERIC Educational Resources Information Center

    Chen, Alice Y.; McKee, Nancy

    1999-01-01

    Describes the developmental process used to visualize the calcium ATPase enzyme of the sarcoplasmic reticulum which involves evaluating scientific information, consulting scientists, model making, storyboarding, and creating and editing in a computer medium. (Author/CCM)

  8. Altered calcium pump and secondary deficiency of γ-sarcoglycan and microspan in sarcoplasmic reticulum membranes isolated from δ-sarcoglycan knockout mice

    PubMed Central

    Solares-Pérez, Alhondra; Álvarez, Rocío; Crosbie, Rachelle H.; Vega-Moreno, Jesús; Medina-Monares, Joel; Estrada, Francisco J.; Ortega, Alicia; Coral-Vazquez, Ramón

    2016-01-01

    Sarcoglycans (SGs) and sarcospan (SSPN) are transmembrane proteins of the dystrophin-glycoprotein complex. Mutations in the genes encoding SGs cause many inherited forms of muscular dystrophy. In this study, using purified membranes of wild-type (WT) and δ-SG knockout (KO) mice, we found the specific localization of the SG-SSPN isoforms in transverse tubules (TT) and sarcoplasmic reticulum (SR) membranes. Immunoblotting revealed that the absence of δ-SG isoforms in TT and SR results in a secondary deficiency of γ-SG and µSPN. Our results showed augmented ATP hydrolytic activity, ATP-dependent calcium uptake and passive calcium efflux, probably through SERCA1 in KO compared to WT mice. Furthermore, we found a conformational change in SERCA1 isolated from KO muscle as demonstrated by calorimetric analysis. Following these alterations with mechanical properties, we found an increase in force in KO muscle with the same rate of fatigue but with a decreased fatigue recovery compared to WT. Together our observations suggest, for the first time, that the δ-SG isoforms may stabilize the expression of γ-SG and µSPN in the TT and SR membranes and that this possible complex may play a role in the maintenance of a stable level of resting cytosolic calcium concentration in skeletal muscle. PMID:20638123

  9. Biochemical heterogeneity of skeletal-muscle microsomal membranes. Membrane origin, membrane specificity and fibre types

    PubMed Central

    Salviati, Giovanni; Volpe, Pompeo; Salvatori, Sergio; Betto, Romeo; Damiani, Ernesto; Margreth, Alfredo; Pasquali-Ronchetti, Ivonne

    1982-01-01

    1. Microsomes were isolated from rabbit fast-twitch and slow-twitch muscle and were separated into heavy and light fractions by centrifugation in a linear (0.3–2m) sucrose density gradient. The membrane origin of microsomal vesicles was investigated by studying biochemical markers of the sarcoplasmic-reticulum membranes and of surface and T-tubular membranes, as well as their freeze-fracture properties. 2. Polyacrylamide-gel electrophoresis showed differences in the Ca2+-dependent ATPase/calsequestrin ratio between heavy and light fractions, which were apparently consistent with their respective origin from cisternal and longitudinal sarcoplasmic reticulum, as well as unrelated differences, such as peptides specific to slow-muscle microsomes (mol.wts. 76000, 60000, 56000 and 45000). 3. Freeze-fracture electron microscopy of muscle microsomes demonstrated that vesicles truly derived from the sarcoplasmic reticulum, with an average density of 9nm particles on the concave face of about 3000/μm2 for both fast and slow muscle, were admixed with vesicles with particle densities below 1000/μm2. 4. As determined in the light fractions, the sarcoplasmic-reticulum vesicles accounted for 84% and 57% of the total number of microsomal vesicles, for fast and slow muscle respectively. These values agreed closely with the percentage values of Ca2+-dependent ATPase protein obtained by gel densitometry. 5. The T-tubular origin of vesicles with a smooth concave fracture face in slow-muscle microsomes is supported by their relative high content in total phospholipid and cholesterol, compared with the microsomes of fast muscle, and by other correlative data, such as the presence of (Na++K+)-dependent ATPase activity and of low amounts of Na+-dependent membrane phosphorylation. 6. Among intrinsic sarcoplasmic-reticulum membrane proteins, a proteolipid of mol.wt. 12000 is shown to be identical in the microsomes of both fast and slow muscle and the Ca2+-dependent ATPase to be

  10. Subcellular distribution of phospholipids in different types of skeletal muscle.

    PubMed

    Domonkos, J; Heiner, L; Vargha, M

    1975-01-01

    Subcellular distribution of choline and non-choline phosphatides has been studied in tetanic (fast-twitch) and tonic (slow-twitch) muscles of rabbits. The choline phosphatide content of the subcellular fraction including the sarcolemma was greater in the tetanic than in the tonic muscle. On the other hand, the choline phosphatide content of the mitochondria-free sarcoplasmic fraction was greater in the tonic than in the tetanic muscle. A greater amount of non-choline phosphatide was found in each subcellular fraction of the tonic muscle as compared with those of the tetanic one. There was more fatty aldehyde in the non-choline phosphatides of each subcellular fraction of the tetanic muscle, than in those of the tonic one, of this type being much smaller in the tetanic muscle. There is not such an expressed difference in the fatty aldehyde contents of choline phosphatides of the subcellular fractions between the two kinds of muscle.

  11. Chemical modification and fluorescence labeling study of Ca2+,Mg2+-adenosine triphosphatase of sarcoplasmic reticulum using iodoacetamide and its N-substituted derivatives.

    PubMed

    Baba, A; Nakamura, T; Kawakita, M

    1986-11-01

    Sarcoplasmic reticulum membrane vesicles from rabbit skeletal muscle were treated with iodoacetamide (IAA) at pH 7.0 and 30 degrees C. At 1.0 mM IAA, 1 mol of IAA per mol of ATPase peptide was bound in 1 h. Under these conditions, IAA was attached specifically to the B-tryptic fragment portion of the peptide. The binding of IAA did not affect the Ca2+-transporting activity of ATPase. Three fluorescent derivatives of iodoacetamide, 5-(2-acetamidoethyl)aminonaphthalene-1-sulfonate (IAEDANS), 5-iodoacetamido fluorescein (IAF), and 5-iodoacetamido eosin (IAE), were also tested for reactivity toward sarcoplasmic reticulum ATPase at 30 degrees C and pH 7.0. In 1 h at 50 microM concentration, each of these fluorescent labels modified ATPase to a labeling density of 1 mol per mol of ATPase. Neither IAEDANS nor IAF at this labeling density affected Ca2+-transporting activity, but IAE reduced it to 20% of the untreated control. The target site of IAEDANS at this labeling density was located exclusively on the B-fragment portion, as was the case with IAA, but IAF label was found on both A1 and B fragments after limited tryptic digestion. IAEDANS was used as a B-fragment portion-directed conformational probe of Ca2+-transport ATPase, and an increase in fluorescence intensity accompanying E1Ca-P formation was detected. The fluorescence enhancement was abolished when E1Ca-P X ADP beta S was formed by adding ADP beta S to preformed E1Ca-P. This suggests that the conformation of ATPase in the neighborhood of the IAEDANS binding site may be altered in response to the dissociation of ADP from the phosphorylated intermediate.

  12. MRI in occipital lobe infarcts: classification by involvement of the striate cortex.

    PubMed

    Kitajima, M; Korogi, Y; Kido, T; Ikeda, O; Morishita, S; Takahashi, M

    1998-11-01

    We reviewed the MRI studies of 25 patients with occipital lobe infarcts to clarify the distribution of infarcts in the posterior cerebral arterial territory, focussing on their relationship to the striate cortex. Visual field defects and MRI findings were also correlated in 16 patients. On coronal and/or sagittal images, the distribution of the infarct and its relationship to the striate cortex were classified. Involvement of the cortex of both upper and lower lips of the calcarine fissure was observed in 10 patients, and involvement of the lower lip alone in 15. The upper cortical lesions were always accompanied by lower cortical lesions. The visual field defects were complete hemianopia in nine patients, superior quadrantanopia in six and hemianopia with a preserved temporal crescent in one. All patients with superior quadrantanopia had involvement of the lower cortex alone; there were no cases of inferior quadrantanopia. The characteristic vascular anatomy, and poor development of the collateral circulation in the lower cortical area, may explain the vulnerability of this area to infarcts.

  13. YAP-Mediated Mechanotransduction in Skeletal Muscle

    PubMed Central

    Fischer, Martina; Rikeit, Paul; Knaus, Petra; Coirault, Catherine

    2016-01-01

    Skeletal muscle is not only translating chemical energy into mechanical work, it is also a highly adaptive and regenerative tissue whose architecture and functionality is determined by its mechanical and physical environment. Processing intra- and extracellular mechanical signaling cues contributes to the regulation of cell growth, survival, migration and differentiation. Yes-associated Protein (YAP), a transcriptional coactivator downstream of the Hippo pathway and its paralog, the transcriptional co-activator with PDZ-binding motif (TAZ), were recently found to play a key role in mechanotransduction in various tissues including skeletal muscle. Furthermore, YAP/TAZ modulate myogenesis and muscle regeneration and abnormal YAP activity has been reported in muscular dystrophy and rhabdomyosarcoma. Here, we summarize the current knowledge of mechanosensing and -signaling in striated muscle. We highlight the role of YAP signaling and discuss the different routes and hypotheses of its regulation in the context of mechanotransduction. PMID:26909043

  14. Artificial muscle: facts and fiction.

    PubMed

    Schaub, Marcus C

    2011-01-01

    Mechanical devices are sought to support insufficient or paralysed striated muscles including the failing heart. Nickel-titanium alloys (nitinol) present the following two properties: (i) super-elasticity, and (ii) the potential to assume different crystal structures depending on temperature and/or stress. Starting from the martensite state nitinol is able to resume the austenite form (state of low potential energy and high entropy) even against an external resistance. This one-way shape change is deployed in self-expanding vascular stents. Heating induces the force generating transformation from martensite to the austenite state while cooling induces relaxation back to the martensite state. This two-way shape change oscillating between the two states may be used in cyclically contracting support devices of silicon-coated nitinol wires. Such a contractile device sutured to the right atrium has been tested in vitro in a bench model and in vivo in sheep. The contraction properties of natural muscles, specifically of the myocardium, and the tight correlation with ATP production by oxidative phosphorylation in the mitochondria is briefly outlined. Force development by the nitinol device cannot be smoothly regulated as in natural muscle. Its mechanical impact is forced onto the natural muscle regardless of the actual condition with regard to metabolism and Ca2+-homeostasis. The development of artificial muscle on the basis of nitinol wires is still in its infancy. The nitinol artificial muscle will have to prove its viability in the various clinical settings. PMID:22183715

  15. Artificial muscle: facts and fiction.

    PubMed

    Schaub, Marcus C

    2011-12-19

    Mechanical devices are sought to support insufficient or paralysed striated muscles including the failing heart. Nickel-titanium alloys (nitinol) present the following two properties: (i) super-elasticity, and (ii) the potential to assume different crystal structures depending on temperature and/or stress. Starting from the martensite state nitinol is able to resume the austenite form (state of low potential energy and high entropy) even against an external resistance. This one-way shape change is deployed in self-expanding vascular stents. Heating induces the force generating transformation from martensite to the austenite state while cooling induces relaxation back to the martensite state. This two-way shape change oscillating between the two states may be used in cyclically contracting support devices of silicon-coated nitinol wires. Such a contractile device sutured to the right atrium has been tested in vitro in a bench model and in vivo in sheep. The contraction properties of natural muscles, specifically of the myocardium, and the tight correlation with ATP production by oxidative phosphorylation in the mitochondria is briefly outlined. Force development by the nitinol device cannot be smoothly regulated as in natural muscle. Its mechanical impact is forced onto the natural muscle regardless of the actual condition with regard to metabolism and Ca2+-homeostasis. The development of artificial muscle on the basis of nitinol wires is still in its infancy. The nitinol artificial muscle will have to prove its viability in the various clinical settings.

  16. Your Muscles

    MedlinePlus

    ... Homework? Here's Help White House Lunch Recipes Your Muscles KidsHealth > For Kids > Your Muscles Print A A ... and skeletal (say: SKEL-uh-tul) muscle. Smooth Muscles Smooth muscles — sometimes also called involuntary muscles — are ...

  17. An anomalous muscle linking superior and inferior rectus muscles in the orbit.

    PubMed

    Kakizaki, Hirohiko; Zako, Masahiro; Nakano, Takashi; Asamoto, Ken; Miyaishi, Osamu; Iwaki, Masayoshi

    2006-09-01

    Dissections of the bilateral orbits in a 45-year-old female cadaver, who had no ocular movement disorders in her lifetime, revealed anomalous muscles linking the superior and inferior rectus muscles. The muscles, situated between the optic nerve and the lateral rectus muscle, originated from the annulus of Zinn and branched off two heads; one inserted into the medial inferior side of the superior rectus muscle and the other inserted into the central superior side of the inferior rectus muscle. Each insertion was located on a distal site of the myoneural junction of each rectus muscle. Histological investigations showed that the muscles had a striated muscle structure. No definite nerve insertion was observed in the muscles. Although this type of anomalous muscle has been reported in a few Caucasian cases, the present study is the first report in an Asian person. Anomalous orbital structures, which are a rare cause of strabismus, are important in the differential diagnosis of intra-orbital space-occupying lesions, rather than the differential diagnosis of strabismus.

  18. Muscle glycogen stores and fatigue.

    PubMed

    Ørtenblad, Niels; Westerblad, Håkan; Nielsen, Joachim

    2013-09-15

      Studies performed at the beginning of the last century revealed the importance of carbohydrate as a fuel during exercise, and the importance of muscle glycogen on performance has subsequently been confirmed in numerous studies. However, the link between glycogen depletion and impaired muscle function during fatigue is not well understood and a direct cause-and-effect relationship between glycogen and muscle function remains to be established. The use of electron microscopy has revealed that glycogen is not homogeneously distributed in skeletal muscle fibres, but rather localized in distinct pools. Furthermore, each glycogen granule has its own metabolic machinery with glycolytic enzymes and regulating proteins. One pool of such glycogenolytic complexes is localized within the myofibrils in close contact with key proteins involved in the excitation-contraction coupling and Ca2+ release from the sarcoplasmic reticulum (SR). We and others have provided experimental evidence in favour of a direct role of decreased glycogen, localized within the myofibrils, for the reduction in SR Ca2+ release during fatigue. This is consistent with compartmentalized energy turnover and distinctly localized glycogen pools being of key importance for SR Ca2+ release and thereby affecting muscle contractility and fatigability.

  19. Muscle glycogen stores and fatigue

    PubMed Central

    Ørtenblad, Niels; Westerblad, Håkan; Nielsen, Joachim

    2013-01-01

    Studies performed at the beginning of the last century revealed the importance of carbohydrate as a fuel during exercise, and the importance of muscle glycogen on performance has subsequently been confirmed in numerous studies. However, the link between glycogen depletion and impaired muscle function during fatigue is not well understood and a direct cause-and-effect relationship between glycogen and muscle function remains to be established. The use of electron microscopy has revealed that glycogen is not homogeneously distributed in skeletal muscle fibres, but rather localized in distinct pools. Furthermore, each glycogen granule has its own metabolic machinery with glycolytic enzymes and regulating proteins. One pool of such glycogenolytic complexes is localized within the myofibrils in close contact with key proteins involved in the excitation–contraction coupling and Ca2+ release from the sarcoplasmic reticulum (SR). We and others have provided experimental evidence in favour of a direct role of decreased glycogen, localized within the myofibrils, for the reduction in SR Ca2+ release during fatigue. This is consistent with compartmentalized energy turnover and distinctly localized glycogen pools being of key importance for SR Ca2+ release and thereby affecting muscle contractility and fatigability. PMID:23652590

  20. Morphometric variability of minicolumns in the striate cortex of Homo sapiens, Macaca mulatta, and Pan troglodytes.

    PubMed

    Casanova, Manuel F; Trippe, Juan; Tillquist, Christopher; Switala, Andrew E

    2009-02-01

    Radially oriented ensembles of neurons and their projections, termed minicolumns, are hypothesized to be the basic microcircuit of mammalian cerebral cortex. Minicolumns can be divided into a core and a peripheral neuropil space compartment. The core of minicolumns is constrained by the migratory path of pyramidal cells and their attendant radially oriented projections. Variation in minicolumnar morphometry and density is observed both within and across species. Using a scale-independent measure of variability in minicolumnar width (V(CW)), we demonstrated a significant increase in V(CW) in layers III-V of striate cortex in humans relative to macaques and chimpanzees. Despite changes in minicolumnar width (CW) across species, their core space (w) remained the same. Given that cellular elements and processes within the peripheral neuropil space of minicolumns are derived from assorted sources, cross-species differences in VCW may result from genetic and epigenetic influences acting primarily on this compartment of the minicolumn.

  1. Analytical study of striated nozzle flow with small radius of curvature ratio throats

    NASA Technical Reports Server (NTRS)

    Norton, D. J.; White, R. E.

    1972-01-01

    An analytical method was developed which is capable of estimating the chamber and throat conditions in a nozzle with a low radius of curvature throat. The method was programmed using standard FORTRAN 4 language and includes chemical equilibrium calculation subprograms (modified NASA Lewis program CEC71) as an integral part. The method determines detailed and gross rocket characteristics in the presence of striated flows and gives detailed results for the motor chamber and throat plane with as many as 20 discrete zones. The method employs a simultaneous solution of the mass, momentum, and energy equations and allows propellant types, 0/F ratios, propellant distribution, nozzle geometry, and injection schemes to be varied so to predict spatial velocity, density, pressure, and other thermodynamic variable distributions in the chamber as well as the throat. Results for small radius of curvature have shown good comparison to experimental results. Both gaseous and liquid injection may be considered with frozen or equilibrium flow calculations.

  2. Aging and regenerative capacity of skeletal muscle in rats.

    PubMed

    Kaasik, Priit; Aru, Maire; Alev, Karin; Seene, Teet

    2012-07-01

    The objective of the study was to examine skeletal muscle regeneration capacity of young and very old rats during autotransplantation. In 3.5 and 30 month-old Wistar rats, gastrocnemius muscle was removed and grafted back to its original bed. Incorporation of 3H leucine into myofibrillar and sarcoplasmic protein fractions, their relative contents in autografts and synthesis rate of MyHC and actin were recorded. The relative muscle mass of old rats was about 67% of that of young rats; the absolute mass of autografted muscle was 61% intact in the young rat group and 51% in the old rat group. Content of myofibrillar protein in the autografts of young rats was 46% of the intact muscle content, and 39% in the old rat group. In conclusion, the difference in skeletal muscle regeneration capacity of young and very old rats is about ten percent. In the autografts of both young and old rats, the regeneration of the contractile apparatus is less effective in comparison with the sarcoplasmic compartment.

  3. Neurons in Striate Cortex Signal Disparity in Half-Matched Random-Dot Stereograms

    PubMed Central

    Read, Jenny C. A.; Cumming, Bruce G.

    2016-01-01

    Human stereopsis can operate in dense “cyclopean” images containing no monocular objects. This is believed to depend on the computation of binocular correlation by neurons in primary visual cortex (V1). The observation that humans perceive depth in half-matched random-dot stereograms, although these stimuli have no net correlation, has led to the proposition that human depth perception in these stimuli depends on a distinct “matching” computation possibly performed in extrastriate cortex. However, recording from disparity-selective neurons in V1 of fixating monkeys, we found that they are in fact able to signal disparity in half-matched stimuli. We present a simple model that explains these results. This reinstates the view that disparity-selective neurons in V1 provide the initial substrate for perception in dense cyclopean stimuli, and strongly suggests that separate correlation and matching computations are not necessary to explain existing data on mixed correlation stereograms. SIGNIFICANCE STATEMENT The initial step in stereoscopic 3D vision is generally thought to be a correlation-based computation that takes place in striate cortex. Recent research has argued that there must be an additional matching computation involved in extracting stereoscopic depth in random-dot stereograms. This is based on the observation that humans can perceive depth in stimuli with a mean binocular correlation of zero (where a correlation-based mechanism should not signal depth). We show that correlation-based cells in striate cortex do in fact signal depth here because they convert fluctuations in the correlation level into a mean change in the firing rate. Our results reinstate the view that these cells provide a sufficient substrate for the perception of stereoscopic depth. PMID:27559177

  4. Deletion of muscle GRP94 impairs both muscle and body growth by inhibiting local IGF production

    PubMed Central

    Barton, Elisabeth R.; Park, SooHyun; James, Jose K.; Makarewich, Catherine A.; Philippou, Anastassios; Eletto, Davide; Lei, Hanqin; Brisson, Becky; Ostrovsky, Olga; Li, Zihai; Argon, Yair

    2012-01-01

    Insulin-like growth factors (IGFs) are critical for development and growth of skeletal muscles, but because several tissues produce IGFs, it is not clear which source is necessary or sufficient for muscle growth. Because it is critical for production of both IGF-I and IGF-II, we ablated glucose-regulated protein 94 (GRP94) in murine striated muscle to test the necessity of local IGFs for normal muscle growth. These mice exhibited smaller skeletal muscles with diminished IGF contents but with normal contractile function and no apparent endoplasmic reticulum stress response. This result shows that muscles rely on GRP94 primarily to support local production of IGFs, a pool that is necessary for normal muscle growth. In addition, body weights were ∼30% smaller than those of littermate controls, and circulating IGF-I also decreased significantly, yet glucose homeostasis was maintained with little disruption to the growth hormone pathway. The growth defect was complemented on administration of recombinant IGF-I. Thus, unlike liver production of IGF-I, muscle IGF-I is necessary not only locally but also globally for whole-body growth.—Barton, E. R., Park, S., James, J. K., Makarewich, C. A., Philippou, A., Eletto, D., Lei, H., Brisson, B., Ostrovsky, O., Li, Z., Argon, Y. Deletion of muscle GRP94 impairs both muscle and body growth by inhibiting local IGF production. PMID:22649033

  5. Design Principles of Reptilian Muscles: Calcium Cycling Strategies.

    PubMed

    Perni, Stefano; Close, Matthew; Franzini-Armstrong, Clara

    2016-03-01

    The ultrastructure of the sarcoplasmic reticulum (SR) in skeletal muscles was compared among different reptile species (watersnake, boa constrictor, lizard, and turtle) and a mammal (mouse). Morphometric analysis demonstrates a pattern of increasing calsequestrin (CASQ) content in the lumen of SR from turtle to lizard, watersnake, and boa constrictor, and this content is in all cases higher than in mouse. In all reptiles sampled except turtle, CASQ is not confined to the junctional sarcoplasmic reticulum (jSR) cisternae as it is in other species. It instead fills the entire longitudinal (free) SR (fSR) regions, and in the extreme case of snakes, the shape of the SR is modified around the extra CASQ. We suggest that high CASQ content may represent an ATP-saving adaptation that permits relatively low metabolic rates during prolonged periods of fasting and inactivity, particularly in watersnake and boa constrictor.

  6. Genetically enhancing mitochondrial antioxidant activity improves muscle function in aging.

    PubMed

    Umanskaya, Alisa; Santulli, Gaetano; Xie, Wenjun; Andersson, Daniel C; Reiken, Steven R; Marks, Andrew R

    2014-10-21

    Age-related skeletal muscle dysfunction is a leading cause of morbidity that affects up to half the population aged 80 or greater. Here we tested the effects of increased mitochondrial antioxidant activity on age-dependent skeletal muscle dysfunction using transgenic mice with targeted overexpression of the human catalase gene to mitochondria (MCat mice). Aged MCat mice exhibited improved voluntary exercise, increased skeletal muscle specific force and tetanic Ca(2+) transients, decreased intracellular Ca(2+) leak and increased sarcoplasmic reticulum (SR) Ca(2+) load compared with age-matched wild type (WT) littermates. Furthermore, ryanodine receptor 1 (the sarcoplasmic reticulum Ca(2+) release channel required for skeletal muscle contraction; RyR1) from aged MCat mice was less oxidized, depleted of the channel stabilizing subunit, calstabin1, and displayed increased single channel open probability (Po). Overall, these data indicate a direct role for mitochondrial free radicals in promoting the pathological intracellular Ca(2+) leak that underlies age-dependent loss of skeletal muscle function. This study harbors implications for the development of novel therapeutic strategies, including mitochondria-targeted antioxidants for treatment of mitochondrial myopathies and other healthspan-limiting disorders. PMID:25288763

  7. The STARS signaling pathway: a key regulator of skeletal muscle function.

    PubMed

    Lamon, Séverine; Wallace, Marita A; Russell, Aaron P

    2014-09-01

    During the last decade, the striated muscle activator of Rho signaling (STARS), a muscle-specific protein, has been proposed to play an increasingly important role in skeletal muscle growth, metabolism, regeneration and stress adaptation. STARS influences actin dynamics and, as a consequence, regulates the myocardin-related transcription factor A/serum response factor (MRTF-A/SRF) transcriptional program, a well-known pathway controlling skeletal muscle development and function. Muscle-specific stress conditions, such as exercise, positively regulates, while disuse and degenerative muscle diseases are associated with a downregulation of STARS and its downstream partners, suggesting a pivotal role for STARS in skeletal muscle health. This review provides a comprehensive overview of the known role and regulation of STARS and the members of its signaling pathway, RhoA, MRTF-A and SRF, in skeletal muscle.

  8. Interaction of phosphatidic acid and phosphatidylserine with the Ca2+-ATPase of sarcoplasmic reticulum and the mechanism of inhibition.

    PubMed

    Dalton, K A; East, J M; Mall, S; Oliver, S; Starling, A P; Lee, A G

    1998-02-01

    The sarcoplasmic reticulum of skeletal muscle contains anionic phospholipids as well as the zwitterionic phosphatidylcholine and phosphatidylethanolamine. Here we study the effects of anionic phospholipids on the activity of the Ca2+-ATPase purified from the membrane. Reconstitution of the Ca2+-ATPase into dioleoylphosphatidylserine [di(C18:1)PS] or dioleoylphosphatidic acid [di(C18:1)PA] leads to a decrease in ATPase activity. Measurements of the quenching of the tryptophan fluorescence of the ATPase by brominated phospholipids give a relative binding constant for the anionic lipids compared with dioleoylphosphatidylcholine close to 1 and suggest that phosphatidic acid only binds to the ATPase at the bulk lipid sites around the ATPase. Addition of di(C18:1)PS or di(C18:1)PA to the ATPase in the short-chain dimyristoleoylphosphatidylcholine [di(C14:1)PC] reverse the effects of the short-chain lipid on ATPase activity and on Ca2+ binding, as revealed by the response of tryptophan fluorescence intensity to Ca2+ binding. It is concluded that the lipid headgroup and lipid fatty acyl chains have separate effects on the function of the ATPase. The anionic phospholipids have no significant effect on Ca2+ binding to the ATPase; the level of Ca2+ binding to the ATPase, the affinity of binding and the rate of dissociation of Ca2+ are unchanged by reconstitution into di(C18:1)PA. The major effect of the anionic lipids is a reduction in the maximal level of binding of MgATP. This is attributed to the formation of oligomers of the Ca2+-ATPase, in which only one molecule of the ATPase can bind MgATP dimers in di(C18:1)PS and trimers or tetramers in di(C18:1)PA. The rates of phosphorylation and dephosphorylation for the proportion of the ATPase still able to bind ATP are unaffected by reconstitution. Larger changes were observed in the level of phosphorylation of the ATPase by Pi, which became very low in the anionic phospholipids. The fluorescence response to Mg2+ for the ATPase

  9. Effects of pH-treated Fish Sarcoplasmic Proteins on the Functional Properties of Chicken Myofibrillar Protein Gel Mediated by Microbial Transglutaminase

    PubMed Central

    Hemung, Bung-Orn

    2014-01-01

    pH adjustment would be of advantage in improving the water holding capacity of muscle proteins. The objective of this study was to evaluate the addition of fish sarcoplasmic protein (SP) solution, which was adjusted to pH 3.0 or 12.0, neutralized to pH 7.0, and lyophilized to obtain the acid- and alkaline-treated SP samples, on the functional properties of the chicken myofibrillar protein induced by microbial transglutaminase (MTG). The solubility of alkaline-treated SP was higher than that of the acid counterpart; however, those values of the two pH-treated samples were lower than that of normal SP (p<0.05). All SP solutions were mixed with myofibrillar proteins (MP) extracted from chicken breast, and incubated with MTG. The shear stresses of MP with acid- and alkaline-treated SP were higher than that of normal SP. The thermal stability of MP mixture reduced upon adding SP, regardless of the pH treatment. The breaking force of MP gels with acid-treated SP increased more than those of alkaline-treated SP, while normal SP showed the highest value. The MP gel lightness increased, but cooking loss reduced, with the addition of SP. Smooth microstructure of the gel surface was observed. These results indicated that adjusting the pH of SP improved the water holding capacity of chicken myofibrillar proteins induced by MTG. PMID:26761171

  10. Interactive effects among several stimulus parameters on the responses of striate cortical complex cells.

    PubMed

    Gawne, T J; Richmond, B J; Optican, L M

    1991-08-01

    1. Although neurons within the visual system are often described in terms of their responses to particular patterns such as bars and edges, they are actually sensitive to many different stimulus features, such as the luminances making up the patterns and the duration of presentation. Many different combinations of stimulus parameters can result in the same neuronal response, raising the problem of how the nervous system can extract information about visual stimuli from such inherently ambiguous responses. It has been shown that complex cells transmit significant amounts of information in the temporal modulation of their responses, raising the possibility that different stimulus parameters are encoded in different aspects of the response. To find out how much information is actually available about individual stimulus parameters, we examined the interactions among three stimulus parameters in the temporally modulated responses of striate cortical complex cells. 2. Sixteen black and white patterns were presented to two awake monkeys at each of four luminance-combinations and five durations, giving a total of 320 unique stimuli. Complex cells were recorded in layers 2 and 3 of striate cortex, with the stimuli centered on the receptive fields as determined by mapping with black and white bars. 3. An analysis of variance (ANOVA) was applied to these data with the three stimulus parameters of pattern, the luminance-combinations, and duration as the independent variables. The ANOVA was repeated with the magnitude and three different aspects of the temporal modulation of the response as the dependent variables. For the 19 neurons studied, many of the interactions between the different stimulus parameters were statistically significant. For some response measures the interactions accounted for more than one-half of the total response variance. 4. We also analyzed the stimulus-response relationships with the use of information theoretical techniques. We defined input codes

  11. Mesodermal iPSC–derived progenitor cells functionally regenerate cardiac and skeletal muscle

    PubMed Central

    Quattrocelli, Mattia; Swinnen, Melissa; Giacomazzi, Giorgia; Camps, Jordi; Barthélemy, Ines; Ceccarelli, Gabriele; Caluwé, Ellen; Grosemans, Hanne; Thorrez, Lieven; Pelizzo, Gloria; Muijtjens, Manja; Verfaillie, Catherine M.; Blot, Stephane; Janssens, Stefan; Sampaolesi, Maurilio

    2015-01-01

    Conditions such as muscular dystrophies (MDs) that affect both cardiac and skeletal muscles would benefit from therapeutic strategies that enable regeneration of both of these striated muscle types. Protocols have been developed to promote induced pluripotent stem cells (iPSCs) to differentiate toward cardiac or skeletal muscle; however, there are currently no strategies to simultaneously target both muscle types. Tissues exhibit specific epigenetic alterations; therefore, source-related lineage biases have the potential to improve iPSC-driven multilineage differentiation. Here, we determined that differential myogenic propensity influences the commitment of isogenic iPSCs and a specifically isolated pool of mesodermal iPSC-derived progenitors (MiPs) toward the striated muscle lineages. Differential myogenic propensity did not influence pluripotency, but did selectively enhance chimerism of MiP-derived tissue in both fetal and adult skeletal muscle. When injected into dystrophic mice, MiPs engrafted and repaired both skeletal and cardiac muscle, reducing functional defects. Similarly, engraftment into dystrophic mice of canine MiPs from dystrophic dogs that had undergone TALEN-mediated correction of the MD-associated mutation also resulted in functional striatal muscle regeneration. Moreover, human MiPs exhibited the same capacity for the dual differentiation observed in murine and canine MiPs. The findings of this study suggest that MiPs should be further explored for combined therapy of cardiac and skeletal muscles. PMID:26571398

  12. Independent specialisation of myosin II paralogues in muscle vs. non-muscle functions during early animal evolution: a ctenophore perspective

    PubMed Central

    2012-01-01

    Background Myosin II (or Myosin Heavy Chain II, MHCII) is a family of molecular motors involved in the contractile activity of animal muscle cells but also in various other cellular processes in non-muscle cells. Previous phylogenetic analyses of bilaterian MHCII genes identified two main clades associated respectively with smooth/non-muscle cells (MHCIIa) and striated muscle cells (MHCIIb). Muscle cells are generally thought to have originated only once in ancient animal history, and decisive insights about their early evolution are expected to come from expression studies of Myosin II genes in the two non-bilaterian phyla that possess muscles, the Cnidaria and Ctenophora. Results We have uncovered three MHCII paralogues in the ctenophore species Pleurobrachia pileus. Phylogenetic analyses indicate that the MHCIIa / MHCIIb duplication is more ancient than the divergence between extant metazoan lineages. The ctenophore MHCIIa gene (PpiMHCIIa) has an expression pattern akin to that of "stem cell markers" (Piwi, Vasa…) and is expressed in proliferating cells. We identified two MHCIIb genes that originated from a ctenophore-specific duplication. PpiMHCIIb1 represents the exclusively muscular form of myosin II in ctenophore, while PpiMHCIIb2 is expressed in non-muscle cells of various types. In parallel, our phalloidin staining and TEM observations highlight the structural complexity of ctenophore musculature and emphasize the experimental interest of the ctenophore tentacle root, in which myogenesis is spatially ordered and strikingly similar to striated muscle formation in vertebrates. Conclusion MHCIIa expression in putative stem cells/proliferating cells probably represents an ancestral trait, while specific involvement of some MHCIIa genes in smooth muscle fibres is a uniquely derived feature of the vertebrates. That one ctenophore MHCIIb paralogue (PpiMHCIIb2) has retained MHCIIa-like expression features furthermore suggests that muscular expression of the

  13. Isolation, purification and characterization of antioxidant peptidic fractions from a bovine liver sarcoplasmic protein thermolysin hydrolyzate.

    PubMed

    Di Bernardini, Roberta; Rai, Dilip K; Bolton, Declan; Kerry, Joseph; O'Neill, Eileen; Mullen, Anne Maria; Harnedy, Pádraigín; Hayes, Maria

    2011-02-01

    Sarcoplasmic proteins isolated from bovine livers were hydrolyzed using the enzyme thermolysin at 37°C for 2h. The hydrolyzates were filtered through molecular weight cut off membranes (MWCO) and filtrates were obtained. The water activity (a(w)) of unhydrolysed sarcoplasmic protein, full hydrolyzates, 10-kDa and 3-kDa filtrates were below the limit necessary for microbial growth. The antioxidant activities of both filtrates and fractions were assessed using the 2,2-diphenyl-1-picrylhydrazyl (DPPH) radical scavenging activity assay, the ferric ion reducing antioxidant power (FRAP) assay and the Fe(2+) chelating ability assay. RP-HPLC was used for purification of the full hydrolyzates, the 10-kDa and the 3-kDa filtrates. The peptidic content of the full hydrolyzates, the 10-kDa and the 3-kDa filtrates were assessed using the Dumas method and peptide contents of each fraction were characterized using electrospray quadrupole time-of-flight (ESI-Q-TOF) mass spectrometry with the resultant spectrum analysed using the software programmes Protein Lynx Global Server 2.4. and TurboSEQUEST. Similarities between the amino acid composition of characterized peptides from each fraction and previously reported antioxidant peptides were found. This study demonstrates that meat by-product such as liver can be utilised as raw material for the generation of bioactive peptides with demonstrated antioxidant activities in vitro using the enzyme thermolysin. It is significant as it presents a potential opportunity for meat processors to use their waste streams for the generation of bioactive peptides for potential functional food use. PMID:21129427

  14. Two dust populations of particle fragments in the striated tail of Comet MRKOS 1957 V

    NASA Technical Reports Server (NTRS)

    Sekanina, Z.; Farrell, J. A.

    1982-01-01

    The striated tail of Comet Mrkos 1957 V is investigated. The formation and motions of the striae are described by the same particle-fragmentation model that has been successfully applied to Comet West. Two new kinds of striae are discovered, one made of absorbing particles and the other of essentially nonabsorbing ones. The grains appear to range in size from 0.1 and 0.3 micron. Tentative evidence is found for an excess of dielectric particles with radii near 0.1 micron, whereas among absorbing grains such sizes are relatively scarce. These results illustrate a high degree of sensitivity of the brightness variation along a stria to the particle size distribution. The regular spacing of the bursts of parent particles suggests that all the striae originated in three emission areas on the comet's nucleus rotating with a period of 19.7 hr. The apparent pairing of striae in the tail is a by-product of the existence of more than one discrete source on the nucleus surface and of the different lifetimes of the parent particles.

  15. Two dust populations of particle fragments in the striated tail of Comet Mrkos 1957 V

    SciTech Connect

    Sekanina, Z.; Farrell, J.A.

    1982-12-01

    A total of 26 striae in the dust tail of Comet Mrkos 1957 V is investigated on five small-scale photographs taken 14--17 August 1957. The dynamical parameters of the striae are determined on the premise that these formations are products of the ejection of dust particles that subsequently break up in the tail. We establish the existence of two kinds of striae, consisting of submicron-sized particle fragments made of strongly absorbing and essentially dielectric materials. Although particles in the striae of both kinds are apparently restricted to sizes between 0.1 and 0.3 ..mu..m, inferred size distribution functions of the two dust populations are quite different. A calculated sequence of ejection times indicates that the striae originated in three isolated emission areas on the nucleus rotating with a 19.7-hr period. The discrete dust sources also account for an observed ''doublet'' structure of the striated pattern. We point out differences between the dust tails of Comets West 1976 VI and Mrkos and compare the particle-fragmentation model with a hypothesis that advocates a strong coupling between charged dust and comet plasma.

  16. Magnetization ac loss reduction in HTS CORC® cables made of striated coated conductors

    NASA Astrophysics Data System (ADS)

    Vojenčiak, M.; Kario, A.; Ringsdorf, B.; Nast, R.; van der Laan, D. C.; Scheiter, J.; Jung, A.; Runtsch, B.; Gömöry, F.; Goldacker, W.

    2015-10-01

    High temperature superconductors (HTSs), like for instance REBCO (RE = rare earth) coated conductors, are of high potential for building large superconducting magnets. Some magnets, such as accelerator magnets, require the use of superconducting cables to allow fast ramping, and low magnetization loss to mitigate field quality issues. One of the methods to lower ac loss is to divide the superconducting layer in the tape into filaments. In this paper, conductors with copper stabilization for practical applications are laser scribed into narrow filaments. Striated tapes are then wound into conductor on round core (CORC®) cables. The critical current and magnetization ac loss of single tapes were measured. We found that the stabilizing copper layer causes difficulties for laser scribing. The degradation of the critical current is more pronounced than in the case of non-stabilized tapes. The selection of the number of filaments is therefore a compromise between critical current degradation and reduction of ac loss. Based on the results obtained from single tape experiments, the optimum number of filaments in 4 mm wide tapes was chosen, and CORC® cables with 2, 3 and 4 layers of tapes with and without filaments were manufactured. Magnetization ac loss measurements at 77 K showed a reduction of ac loss in the cables with filaments. This reduction corresponds almost to the number of filaments. Measurement at different frequencies also showed that the coupling loss in CORC® cables with a short twist-pitch is relatively small in comparison to hysteretic loss.

  17. The extracellular compartments of frog skeletal muscle.

    PubMed Central

    Neville, M C; Mathias, R T

    1979-01-01

    1. Detailed studies of solute efflux from frog sartorius muscle and single muscle fibres were carried out in order to characterize a 'special region' (Harris, 1963) in the extracellular space of muscle and determine whether this 'special region' is the sarcoplasmic reticulum. 2. The efflux of radioactive Na, Cl, glusose, 3-O-methylglucose, xylose, glycine, leucine, cycloleucine, Rb, K, inulin (mol. wt. 5000) and dextran (mol. wt. 17,000) from previously loaded muscles was studied. In all cases except dextran the curve had three components, a rapid (A) component which could be equated with efflux from the extracellular space proper, a slow (C) component representing cellular solute and an intermediate (B) component. The distribution space for the B component was 8% of muscle volume in summer frogs and 12% in winter frogs and appeared to be equal for all compounds studied. We tested the hypothesis that the B component originated from the sarcoplasmic reticulum. 3. The C component was missing from the dextran curves. Both dextran and inulin entered the compartment of origin of the B component (compartment B) to the same extent as small molecules. 4. For all compounds studies, the efflux rate constant for the A component could be predicted from the diffusion coefficient. For the B component the efflux rate constant was 6--10 times slower than that for the A component but was still proportional to the diffusion coefficient for the solute in question. 5. When Na and sucrose efflux from single fibres was followed, a B component was usually observed. The average distribution space for this component was small, averaging 1.5% of fibre volume. There was no difference between the average efflux rate constants for Na and sucrose. 6. In an appendix, the constraints placed on the properties of a hypothetical channel between the sarcoplasmic reticulum and the T-system by the linear electrical parameters of frog skeletal muscle are derived. It is shown that the conductance of such

  18. The length-tension curve in muscle depends on lattice spacing

    SciTech Connect

    Williams, C. D.; Salcedo, M. K.; Irving, T. C.; Regnier, M.; Daniel, T. L.

    2013-07-10

    Classic interpretations of the striated muscle length–tension curve focus on how force varies with overlap of thin (actin) and thick (myosin) filaments. New models of sarcomere geometry and experiments with skinned synchronous insect flight muscle suggest that changes in the radial distance between the actin and myosin filaments, the filament lattice spacing, are responsible for between 20% and 50% of the change in force seen between sarcomere lengths of 1.4 and 3.4 µm. Thus, lattice spacing is a significant force regulator, increasing the slope of muscle's force–length dependence.

  19. The length–tension curve in muscle depends on lattice spacing

    PubMed Central

    Williams, C. David; Salcedo, Mary K.; Irving, Thomas C.; Regnier, Michael; Daniel, Thomas L.

    2013-01-01

    Classic interpretations of the striated muscle length–tension curve focus on how force varies with overlap of thin (actin) and thick (myosin) filaments. New models of sarcomere geometry and experiments with skinned synchronous insect flight muscle suggest that changes in the radial distance between the actin and myosin filaments, the filament lattice spacing, are responsible for between 20% and 50% of the change in force seen between sarcomere lengths of 1.4 and 3.4 µm. Thus, lattice spacing is a significant force regulator, increasing the slope of muscle's force–length dependence. PMID:23843386

  20. Virus-like particles in paraspinal muscle in scoliosis.

    PubMed Central

    Webb, J N; Gillespie, W J

    1976-01-01

    Biopsy material from the skeletal muscle (paraxials) of 21 patients with scoliosis was examined by light and electron microscopy. Virus-like particles, 17 nm in diameter with a crystalline structure, were identified in the skeletal muscle fibres of four patients. Associated changes in the sarcoplasm included swelling of mitochondria, presence of lipid droplets, and vesicular structures. Serological studies and culture for virus isolation gave negative results. An excess of lipid (predominantly in type 1 fibres) was noted in the skeletal muscle of several other cases. The significance of these findings is obscure, but the morphology of the paraxial muscles of patients with scoliosis and controls is currently being investigated in greater detail. Images FIG 1 FIG 2 FIG 3 FIG 4 PMID:974659

  1. Muscle Cramps

    MedlinePlus

    Muscle cramps are sudden, involuntary contractions or spasms in one or more of your muscles. They often occur after exercise or at night, ... to several minutes. It is a very common muscle problem. Muscle cramps can be caused by nerves ...

  2. Muscle Disorders

    MedlinePlus

    Your muscles help you move and help your body work. Different types of muscles have different jobs. There are many problems that can affect muscles. Muscle disorders can cause weakness, pain or even ...

  3. Muscle atrophy

    MedlinePlus

    Muscle wasting; Wasting; Atrophy of the muscles ... There are two types of muscle atrophy: disuse and neurogenic. Disuse atrophy is caused by not using the muscles enough . This type of atrophy can often be ...

  4. Muscle biopsy

    MedlinePlus

    ... the removal of a small piece of muscle tissue for examination. ... dystrophy Myopathic changes (destruction of the muscle) Necrosis (tissue death) of muscle Necrotizing vasculitis Traumatic muscle damage Polymyositis Additional conditions ...

  5. Early changes of type 2B fibers after denervation of rat EDL skeletal muscle.

    PubMed

    Germinario, Elena; Esposito, Alessandra; Megighian, Aram; Midrio, Menotti; Biral, Donatella; Betto, Romeo; Danieli-Betto, Daniela

    2002-05-01

    Skeletal muscle type 2B fibers normally receive a moderate level of motoneuron discharge. As a consequence, we hypothesize that type 2B fiber properties should be less sensitive to the absence of the nerve. Therefore, we have investigated the response of sarcoplasmic reticulum and myofibrillar proteins of type 2B fibers isolated from rat extensor digitorum longus muscle after denervation (2 and 7 days). Single fibers were identified by SDS-PAGE of myosin heavy chain isoforms. Electrophysiological and isometric contractile properties of the whole muscle were also analyzed. The pCa-tension relationship of type 2B single fibers was shifted to the left at 2 days and to right at 7 days after denervation, with significant differences in the Hill coefficients and pCa threshold values in 2- vs. 7-day-denervated fibers. The sarcoplasmic reticulum Ca2+ uptake capacity and rate significantly decreased after 2 days of denervation, whereas both increased at 7 days. Caffeine sensitivity of sarcoplasmic reticulum Ca2+ release was transitory and markedly increased in 2-day-denervated fibers. Our results indicate that type 2B fiber functional properties are highly sensitive to the interruption of nerve supply. Moreover, most of 2-day-denervated changes were reverted at 7 days. PMID:11960956

  6. Oxidation of sarcoplasmic proteins during processing of Cantonese sausage in relation to their aggregation behaviour and in vitro digestibility.

    PubMed

    Sun, Weizheng; Zhao, Mouming; Yang, Bao; Zhao, Haifeng; Cui, Chun

    2011-07-01

    The physicochemical changes of sarcoplasmic proteins, especially oxidation behaviour, were measured to determine their mechanism of action on in vitro protein digestibility during Cantonese sausage processing. The results indicated that carbonyl level increased (p<0.05) during the process. The fluorescence loss of tryptophan residues was a direct consequence of the oxidative degradation. All the parameters of protein aggregation were highly (p<0.05) correlated with carbonyl level and protein surface hydrophobicity (H(0)), indicating that protein oxidation and thermal denaturation could induce protein aggregation, leading to secondary structural changes. The analysis of in vitro digestibility showed no correlation between pepsin activity and protein oxidation, due to the biphasic response of sarcoplasmic proteins toward proteolysis. However, a highly significant (p<0.05) correlation was observed with trypsin and α-chymotrypsin activity, indicating that protein oxidation induced the changes in H(0), protein aggregation and secondary structure, which further influenced in vitro digestibility. PMID:21353394

  7. Skeletal muscle atrophy and the E3 ubiquitin ligases MuRF1 and MAFbx/atrogin-1

    PubMed Central

    Baehr, Leslie M.

    2014-01-01

    Muscle RING finger 1 (MuRF1) and muscle atrophy F-box (MAFbx)/atrogin-1 were identified more than 10 years ago as two muscle-specific E3 ubiquitin ligases that are increased transcriptionally in skeletal muscle under atrophy-inducing conditions, making them excellent markers of muscle atrophy. In the past 10 years much has been published about MuRF1 and MAFbx with respect to their mRNA expression patterns under atrophy-inducing conditions, their transcriptional regulation, and their putative substrates. However, much remains to be learned about the physiological role of both genes in the regulation of mass and other cellular functions in striated muscle. Although both MuRF1 and MAFbx are enriched in skeletal, cardiac, and smooth muscle, this review will focus on the current understanding of MuRF1 and MAFbx in skeletal muscle, highlighting the critical questions that remain to be answered. PMID:25096180

  8. Biphasic decay of the Ca transient results from increased sarcoplasmic reticulum Ca leak

    PubMed Central

    Sankaranarayanan, Rajiv; Li, Yatong; Greensmith, David J.; Eisner, David A.

    2016-01-01

    Key points Ca leak from the sarcoplasmic reticulum through the ryanodine receptor (RyR) reduces the amplitude of the Ca transient and slows its rate of decay.In the presence of β‐adrenergic stimulation, RyR‐mediated Ca leak produces a biphasic decay of the Ca transient with a fast early phase and a slow late phase.Two forms of Ca leak have been studied, Ca‐sensitising (induced by caffeine) and non‐sensitising (induced by ryanodine) and both induce biphasic decay of the Ca transient.Only Ca‐sensitising leak can be reversed by traditional RyR inhibitors such as tetracaine.Ca leak can also induce Ca waves. At low levels of leak, waves occur. As leak is increased, first biphasic decay and then slowed monophasic decay is seen. The level of leak has major effects on the shape of the Ca transient. Abstract In heart failure, a reduction in Ca transient amplitude and contractile dysfunction can by caused by Ca leak through the sarcoplasmic reticulum (SR) Ca channel (ryanodine receptor, RyR) and/or decreased activity of the SR Ca ATPase (SERCA). We have characterised the effects of two forms of Ca leak (Ca‐sensitising and non‐sensitising) on calcium cycling and compared with those of SERCA inhibition. We measured [Ca2+]i with fluo‐3 in voltage‐clamped rat ventricular myocytes. Increasing SR leak with either caffeine (to sensitise the RyR to Ca activation) or ryanodine (non‐sensitising) had similar effects to SERCA inhibition: decreased systolic [Ca2+]i, increased diastolic [Ca2+]i and slowed decay. However, in the presence of isoproterenol, leak produced a biphasic decay of the Ca transient in the majority of cells while SERCA inhibition produced monophasic decay. Tetracaine reversed the effects of caffeine but not of ryanodine. When caffeine (1 mmol l−1) was added to a cell which displayed Ca waves, the wave frequency initially increased before waves disappeared and biphasic decay developed. Eventually (at higher caffeine concentrations), the

  9. Atrogin-1, a muscle-specific F-box protein highly expressed during muscle atrophy

    NASA Technical Reports Server (NTRS)

    Gomes, M. D.; Lecker, S. H.; Jagoe, R. T.; Navon, A.; Goldberg, A. L.

    2001-01-01

    Muscle wasting is a debilitating consequence of fasting, inactivity, cancer, and other systemic diseases that results primarily from accelerated protein degradation by the ubiquitin-proteasome pathway. To identify key factors in this process, we have used cDNA microarrays to compare normal and atrophying muscles and found a unique gene fragment that is induced more than ninefold in muscles of fasted mice. We cloned this gene, which is expressed specifically in striated muscles. Because this mRNA also markedly increases in muscles atrophying because of diabetes, cancer, and renal failure, we named it atrogin-1. It contains a functional F-box domain that binds to Skp1 and thereby to Roc1 and Cul1, the other components of SCF-type Ub-protein ligases (E3s), as well as a nuclear localization sequence and PDZ-binding domain. On fasting, atrogin-1 mRNA levels increase specifically in skeletal muscle and before atrophy occurs. Atrogin-1 is one of the few examples of an F-box protein or Ub-protein ligase (E3) expressed in a tissue-specific manner and appears to be a critical component in the enhanced proteolysis leading to muscle atrophy in diverse diseases.

  10. The role of bacterial fermentation in the hydrolysis and oxidation of sarcoplasmic and myofibrillar proteins in Harbin dry sausages.

    PubMed

    Chen, Qian; Kong, Baohua; Han, Qi; Liu, Qian; Xu, Li

    2016-11-01

    Pediococcus pentosaceus, Lactobacillus curvatus, Lactobacillus sake and Staphylococcus xylosus were evaluated to determine their role in the hydrolysis and oxidation of sarcoplasmic and myofibrillar proteins in Harbin dry sausages. Electrophoresis analysis showed that the hydrolysis of sarcoplasmic and myofibrillar proteins in dry sausages inoculated with bacterial strains was more severe than that in the non-inoculated control. The predominant free amino acids at the end of the fermentation were glutamic acid and alanine, both of which are involved in creating a desirable taste. The inoculation of dry sausages with bacterial strains, especially mixed strains, significantly decreased carbonyl formation and sulfhydryl loss in sausages (P<0.05). Both hydrolysis and oxidation led to increased surface hydrophobicity in sarcoplasmic and myofibrillar proteins. Fermentation of dry sausage with multiple bacterial strains could contribute to flavour formation via flavour precursors. The results demonstrate that Harbin dry sausage can be inoculated with a starter culture mixture of P. pentosaceus, L. curvatus and S. xylosus to improve flavour formation. PMID:27341621

  11. Molecular mechanisms responsible for alcohol-induced myopathy in skeletal muscle and heart.

    PubMed

    Lang, Charles H; Frost, Robert A; Summer, Andrew D; Vary, Thomas C

    2005-10-01

    Chronic alcohol abuse has the potential to modulate striated muscle physiology and function. The skeletal muscle alcoholic myopathy is characterized by muscle weakness and difficulties in gait and locomotion, while chronic alcohol consumption ultimately leads to a decrease in cardiac contractility and output. In both tissues a loss of protein mass results in part from a decreased protein synthesis that initially manifests as a defect in translational efficiency. This review focuses on recent developments in understanding the cellular and molecular mechanisms by which alcohol impairs mRNA translation in skeletal and cardiac muscle, including identification of the signaling pathways and biochemical sites negatively impacted. Defective signaling potentially results from resistance to the normal stimulating effects of anabolic hormones (insulin and insulin-like growth factor-I) and nutrients (leucine) as well as increased production of several negative regulators of muscle mass. Overall, the biochemical mechanisms contributing to the pathogenesis of loss of skeletal and cardiac muscle are reviewed.

  12. Sarcoplasmic reticulum K(+) (TRIC) channel does not carry essential countercurrent during Ca(2+) release.

    PubMed

    Guo, Tao; Nani, Alma; Shonts, Stephen; Perryman, Matthew; Chen, Haiyan; Shannon, Thomas; Gillespie, Dirk; Fill, Michael

    2013-09-01

    The charge translocation associated with sarcoplasmic reticulum (SR) Ca(2+) efflux is compensated for by a simultaneous SR K(+) influx. This influx is essential because, with no countercurrent, the SR membrane potential (Vm) would quickly (<1 ms) reach the Ca(2+) equilibrium potential and SR Ca(2+) release would cease. The SR K(+) trimeric intracellular cation (TRIC) channel has been proposed to carry the essential countercurrent. However, the ryanodine receptor (RyR) itself also carries a substantial K(+) countercurrent during release. To better define the physiological role of the SR K(+) channel, we compared SR Ca(2+) transport in saponin-permeabilized cardiomyocytes before and after limiting SR K(+) channel function. Specifically, we reduced SR K(+) channel conduction 35 and 88% by replacing cytosolic K(+) for Na(+) or Cs(+) (respectively), changes that have little effect on RyR function. Calcium sparks, SR Ca(2+) reloading, and caffeine-evoked Ca(2+) release amplitude (and rate) were unaffected by these ionic changes. Our results show that countercurrent carried by SR K(+) (TRIC) channels is not required to support SR Ca(2+) release (or uptake). Because K(+) enters the SR through RyRs during release, the SR K(+) (TRIC) channel most likely is needed to restore trans-SR K(+) balance after RyRs close, assuring SR Vm stays near 0 mV. PMID:24010658

  13. Hofmeister effect of anions on calcium translocation by sarcoplasmic reticulum Ca2+-ATPase

    NASA Astrophysics Data System (ADS)

    Tadini-Buoninsegni, Francesco; Moncelli, Maria Rosa; Peruzzi, Niccolò; Ninham, Barry W.; Dei, Luigi; Nostro, Pierandrea Lo

    2015-10-01

    The occurrence of Hofmeister (specific ion) effects in various membrane-related physiological processes is well documented. For example the effect of anions on the transport activity of the ion pump Na+, K+-ATPase has been investigated. Here we report on specific anion effects on the ATP-dependent Ca2+ translocation by the sarcoplasmic reticulum Ca2+-ATPase (SERCA). Current measurements following ATP concentration jumps on SERCA-containing vesicles adsorbed on solid supported membranes were carried out in the presence of different potassium salts. We found that monovalent anions strongly interfere with ATP-induced Ca2+ translocation by SERCA, according to their increasing chaotropicity in the Hofmeister series. On the contrary, a significant increase in Ca2+ translocation was observed in the presence of sulphate. We suggest that the anions can affect the conformational transition between the phosphorylated intermediates E1P and E2P of the SERCA cycle. In particular, the stabilization of the E1P conformation by chaotropic anions seems to be related to their adsorption at the enzyme/water and/or at the membrane/water interface, while the more kosmotropic species affect SERCA conformation and functionality by modifying the hydration layers of the enzyme.

  14. Impaired Sarcoplasmic Reticulum Calcium Uptake and Release Promote Electromechanically and Spatially Discordant Alternans: A Computational Study

    PubMed Central

    Weinberg, Seth H.

    2016-01-01

    Cardiac electrical dynamics are governed by cellular-level properties, such as action potential duration (APD) restitution and intracellular calcium (Ca) handling, and tissue-level properties, including conduction velocity restitution and cell–cell coupling. Irregular dynamics at the cellular level can lead to instabilities in cardiac tissue, including alternans, a beat-to-beat alternation in the action potential and/or the intracellular Ca transient. In this study, we incorporate a detailed single cell coupled map model of Ca cycling and bidirectional APD-Ca coupling into a spatially extended tissue model to investigate the influence of sarcoplasmic reticulum (SR) Ca uptake and release properties on alternans and conduction block. We find that an intermediate SR Ca uptake rate and larger SR Ca release resulted in the widest range of stimulus periods that promoted alternans. However, both reduced SR Ca uptake and release promote arrhythmogenic spatially and electromechanically discordant alternans, suggesting a complex interaction between SR Ca handling and alternans characteristics at the cellular and tissue level. PMID:27385917

  15. Design and characterization of self-assembled fish sarcoplasmic protein-alginate nanocomplexes.

    PubMed

    Stephansen, Karen; Mattebjerg, Maria; Wattjes, Jasper; Milisavljevic, Ana; Jessen, Flemming; Qvortrup, Klaus; Goycoolea, Francisco M; Chronakis, Ioannis S

    2015-05-01

    Macrostructures based on natural polymers are subject to large attention, as the application range is wide within the food and pharmaceutical industries. In this study we present nanocomplexes (NCXs) made from electrostatic self-assembly between negatively charged alginate and positively charged fish sarcoplasmic proteins (FSP), prepared by bulk mixing. A concentration screening revealed that there was a range of alginate and FSP concentrations where stable NCXs with similar properties were formed, rather than two exact concentrations. The size of the NCXs was 293 ± 3 nm, and the zeta potential was -42 ± 0.3 mV. The NCXs were stable in water, gastric buffer, intestinal buffer and HEPES buffered glycose, and at all pH values from 2 to 9 except pH 3, where they aggregated. When proteolytic enzymes were present in the buffer, the NCXs were degraded. Only at high concentrations the NCXs caused a decreased viability in HeLa and U2OS cell lines. The simple processing procedure and the high stability of the NCXs, makes them excellent candidates for use in the food and pharmaceutical industry.

  16. Free radical-induced protein modification and inhibition of Ca2+-ATPase of cardiac sarcoplasmic reticulum.

    PubMed

    Kaplan, Peter; Babusikova, Eva; Lehotsky, Jan; Dobrota, Dusan

    2003-06-01

    The effect of oxidative stress on the Ca2+-ATPase activity, lipid peroxidation and protein modification of cardiac sarcoplasmic reticulum (SR) membranes was investigated. Isolated SR vesicles were exposed to FeSO4/EDTA (0.2 micromol Fe2+ per mg of protein) at 37 degrees C for 1 h in the presence or absence of antioxidants. FeSO4/EDTA decreased the maximum velocity of Ca2+-ATPase reaction without a change of affinity for Ca2+ or Hill coefficient. Treatment with radical-generating system led also to conjugated diene formation, loss of sulfhydryl groups, changes in tryptophan and bityrosine fluorescences and to production of lysine conjugates with lipid peroxidation end-products. Lipid antioxidants butylated hydroxytoluene (BHT) and stobadine partially prevented inhibition of Ca2+-ATPase and decrease in tryptophan fluorescence, while the loss of -SH groups and formation of bityrosines or lysine conjugates were completely prevented. Glutathione also partially protected Ca2+-ATPase activity and decreased formation of bityrosine, but it was not able to prevent oxidative modification of tryptophan and lysine. These findings suggest that combination of amino acid modifications, rather than oxidation of amino acids of one kind, is responsible for inhibition of SR Ca2+-ATPase activity. PMID:12870653

  17. A continuous spectrophotometric assay for simultaneous measurement of calcium uptake and ATP hydrolysis in sarcoplasmic reticulum.

    PubMed

    Karon, B S; Nissen, E R; Voss, J; Thomas, D D

    1995-05-20

    A continuous, spectrophotometric assay to simultaneously measure Ca uptake and ATP hydrolysis has been developed, in order to assess the function of the Ca-ATPase in skeletal and cardiac sarcoplasmic reticulum (SR) vesicles. The absorbance of Fura Red was measured continuously at 490 nm, in EGTA-buffered solutions containing initial free ionized calcium concentrations of 300 nM, 500 nM, 790 nM, and 2 microM, during assays of oxalate-facilitated or phosphate-facilitated active calcium uptake in skeletal SR. Simultaneous measurement of ATP hydrolysis during the measurement of phosphate-facilitated Ca uptake was accomplished by measuring the disappearance of NADH at 340 nm, coupled to the hydrolysis of ATP by an enzyme-linked, continuous ATPase assay. This new method, unlike the standard 45Ca-filtration assay, measures calcium uptake in real time and eliminates the need for radioactivity. Moreover, the rates of calcium uptake and ATP hydrolysis are measured simultaneously, allowing the direct quantitative comparison of the two parameters. This assay will facilitate the characterization of Ca-ATPase function and malfunction in skeletal and cardiac SR and advances the methodology for comparison of normal and physically, chemically, or biologically altered Ca-ATPase.

  18. The calcium uptake of the rat heart sarcoplasmic reticulum is altered by dietary lipid.

    PubMed

    Taffet, G E; Pham, T T; Bick, D L; Entman, M L; Pownall, H J; Bick, R J

    1993-01-01

    Small amounts of dietary n-3 fatty acids can have dramatic physiological effects, including the reduction of plasma triglycerides and an elevation of cellular eicosapentanoic (EPA) and docosahexanoic acids (DHA) at the expense of arachidonic acid (AA). We investigated the effects of alterations in the fatty acid compositions of cardiac sarcoplasmic reticulum (CSR) produced by dietary manipulation on the calcium pump protein that is required for energy dependent calcium transport. CSR was isolated from rats fed menhaden oil, which is rich in n-3 fatty acids, and from control animals that were given corn oil. Relative to control membranes, those isolated from rats fed menhaden oil, had a lower content of saturated phospholipids, an increased DHA/AA ratio, and an increased ratio of n-3 to n-6 fatty acids. These changes were associated with a 30% decrease in oxalate-facilitated, ATP-dependent calcium uptake and concomitant decreased Ca-ATPase activity in the membranes from the animals fed menhaden oil. In contrast, there was no alteration in active pump sites as measured by phosphoenzyme formation. Thus, the CSR Ca-ATPase function can be altered by dietary interventions that change the composition, and possibly structure, of the phospholipid membranes thereby affecting enzyme turnover.

  19. Vectorially oriented monolayers of detergent-solubilized Ca(2+) -ATPase from sarcoplasmic reticulum.

    PubMed Central

    Prokop, L A; Stongin, R M; Smith, A B; Blasie, J K; Peticolas, L J; Bean, J C

    1996-01-01

    A method for tethering proteins to solid surfaces has been utilized to form vectorially oriented monolayers of the detergent-solubilized integral membrane protein Ca(2+) -ATPase from the sarcoplasmic reticulum (SR). Bifunctional, organic self-assembled monolayers (SAMs) possessing "headgroup" binding specificity for the substrate and "endgroup" binding specificity for the enzyme were utilized to tether the enzyme to the substrate. Specifically, an amine-terminated 11-siloxyundecaneamine SAM was found to bind the Ca(2+)-ATPase primarily electrostatically. The Ca(2+)-ATPase was labeled with the fluorescent probe 5-(2-[(iodoacetyl)amino]ethyl)aminonaphthalene-1-sulfonic acid before monolayer formation. Consequently, fluorescence measurements performed on amine-terminated SAM/enzyme monolayers formed on quartz substrates served to establish the nature of protein binding. Formation of the monolayers on inorganic multilayer substrates fabricated by molecular beam epitaxy made it possible to use x-ray interferometry to determine the profile structure for the system, which was proved correct by x-ray holography. The profile structures established the vectorial orientation of the Ca(2+)-ATPase within these monolayers, to a spatial resolution of approximately 12 A. Such vectorially oriented monolayers of detergent-solubilized Ca(2+)-ATPase from SR make possible a wide variety of correlative structure/function studies, which would serve to elucidate the mechanism of Ca(2+) transport by this enzyme. Images FIGURE 8 PMID:9172737

  20. Effect of short-chain primary alcohols on fluidity and activity of sarcoplasmic reticulum membranes.

    PubMed

    Almeida, L M; Vaz, W L; Stümpel, J; Madeira, V M

    1986-08-26

    Intramolecular excimer formation with the fluorescent probe 1,3-di(1-pyrenyl)propane, differential scanning calorimetry, and X-ray diffraction were used to assess the effect of ethanol, 1-butanol, and 1-hexanol on the bilayer organization in model membranes, sarcoplasmic reticulum (SR) lipids and native SR membranes. These alcohols have fluidizing effects on membranes and lower the main transition temperature of dimyristoylphosphatidylcholine (DMPC), but only 1-hexanol alters the cooperativity of the phase transition and significantly increases the thickness of DMPC bilayers. The interaction of the three alcohols with the SR Ca2+ pump was also investigated. Hydrolysis of ATP and coupled Ca2+ uptake are differently sensitive to the three alcohols. Whereas ethanol and 1-butanol inhibited the Ca2+ uptake, 1-hexanol stimulated it. Nevertheless, the energetic efficiency of the pump (Ca2+/ATP) is not significantly affected by ethanol or 1-hexanol, but uncoupling was observed with 1-butanol at high concentrations. The different effects of alcohols on the activity of SR membranes rule out an unitary mechanism of action on the basis of fluidity changes induced in the lipid bilayer. Depending on the chain length, the alcohols interact with the SR membranes in different domains, perturbing differently the Ca2+-pump activity.

  1. Calsequestrins in skeletal and cardiac muscle from adult Danio rerio.

    PubMed

    Furlan, Sandra; Mosole, Simone; Murgia, Marta; Nagaraj, Nagarjuna; Argenton, Francesco; Volpe, Pompeo; Nori, Alessandra

    2016-04-01

    Calsequestrin (Casq) is a high capacity, low affinity Ca(2+)-binding protein, critical for Ca(2+)-buffering in cardiac and skeletal muscle sarcoplasmic reticulum. All vertebrates have multiple genes encoding for different Casq isoforms. Increasing interest has been focused on mammalian and human Casq genes since mutations of both cardiac (Casq2) and skeletal muscle (Casq1) isoforms cause different, and sometime severe, human pathologies. Danio rerio (zebrafish) is a powerful model for studying function and mutations of human proteins. In this work, expression, biochemical properties cellular and sub-cellular localization of D. rerio native Casq isoforms are investigated. By quantitative PCR, three mRNAs were detected in skeletal muscle and heart with different abundances. Three zebrafish Casqs: Casq1a, Casq1b and Casq2 were identified by mass spectrometry (Data are available via ProteomeXchange with identifier PXD002455). Skeletal and cardiac zebrafish calsequestrins share properties with mammalian Casq1 and Casq2. Skeletal Casqs were found primarily, but not exclusively, at the sarcomere Z-line level where terminal cisternae of sarcoplasmic reticulum are located. PMID:26585961

  2. Beta-amyloid protein-containing inclusions in skeletal muscle of apolipoprotein-E-deficient mice.

    PubMed Central

    Robertson, T. A.; Dutton, N. S.; Martins, R. N.; Roses, A. D.; Kakulas, B. A.; Papadimitriou, J. M.

    1997-01-01

    The tibialis anterior muscle and soleus muscle of apolipoprotein-E-deficient mice were examined by light and electron microscopy. By light microscopy, sarcoplasmic inclusions were seen in tibialis anterior muscle and 40% of type 2 myofibers were affected in all animals over 8 months of age. These inclusions reacted for nonspecific esterase, cytochrome oxidase, and myoadenylate deaminase and were also periodic acid Schiff positive and stained basophilic with hematoxylin. Moreover, they reacted immunocytochemically with an antibody specific to fragment 17 to 24 of the published sequence of Alzheimer's cerebrovascular amyloid peptide. Immunoreactivity was lost when the antibody was adsorbed with the appropriate synthetic peptide. Ultrastructurally, the inclusions consisted of tubular arrays and were similar to those observed in human muscle in several pathological conditions. In type 1 myofibers of both tibialis anterior and soleus muscle, however, mitochondrial abnormalities including an increase in their number and size were detected, but tubular aggregates were not seen. These large mitochondria possessed an electron-dense inner chamber with an increased number of tightly packed cristae. The results obtained suggest that in these mice there is a disturbed lipid metabolism in skeletal muscle fibers that manifests itself with an accumulation of phospholipid in the form of sarcoplasmic reticulum tubules in the type 2 fibers and enlarged mitochondria with tightly packed cristae in the type 1 fibers. In addition, beta-amyloid protein was closely associated with the accumulated tubules and vesicles of sarcoplasmic reticulum and may represent dysregulation of amyloid precursor protein metabolism. Images Figure 1 Figure 2 Figure 3 PMID:9033257

  3. Protein carbonylation and heat shock proteins in human skeletal muscle: relationships to age and sarcopenia.

    PubMed

    Beltran Valls, Maria R; Wilkinson, Daniel J; Narici, Marco V; Smith, Kenneth; Phillips, Bethan E; Caporossi, Daniela; Atherton, Philip J

    2015-02-01

    Aging is associated with a gradual loss of muscle mass termed sarcopenia, which has significant impact on quality-of-life. Because oxidative stress is proposed to negatively impact upon musculoskeletal aging, we investigated links between human aging and markers of oxidative stress, and relationships to muscle mass and strength in young and old nonsarcopenic and sarcopenic adults. Sixteen young and 16 old males (further subdivided into "old" and "old sarcopenic") were studied. The abundance of protein carbonyl adducts within skeletal muscle sarcoplasmic, myofibrillar, and mitochondrial protein subfractions from musculus vastus lateralis biopsies were determined using Oxyblot immunoblotting techniques. In addition, concentrations of recognized cytoprotective proteins (eg, heat shock proteins [HSP], αβ-crystallin) were also assayed. Aging was associated with increased mitochondrial (but not myofibrillar or sarcoplasmic) protein carbonyl adducts, independently of (stage-I) sarcopenia. Correlation analyses of all subjects revealed that mitochondrial protein carbonyl abundance negatively correlated with muscle strength ([1-repetition maximum], p = .02, r (2) = -.16), but not muscle mass (p = .13, r (2) = -.08). Abundance of cytoprotective proteins, including various HSPs (HSP 27 and 70), were unaffected by aging/sarcopenia. To conclude, these data reveal that mitochondrial protein carbonylation increases moderately with age, and that this increase may impact upon skeletal muscle function, but is not a hallmark of (stage-I) sarcopenia, per se.

  4. Compartmentalization of NO signaling cascade in skeletal muscles

    SciTech Connect

    Buchwalow, Igor B. . E-mail: buchwalo@uni-muenster.de; Minin, Evgeny A.; Samoilova, Vera E.; Boecker, Werner; Wellner, Maren; Schmitz, Wilhelm; Neumann, Joachim

    2005-05-06

    Skeletal muscle functions regulated by NO are now firmly established. However, the literature on the compartmentalization of NO signaling in myocytes is highly controversial. To address this issue, we examined localization of enzymes engaged in L-arginine-NO-cGMP signaling in the rat quadriceps muscle. Employing immunocytochemical labeling complemented with tyramide signal amplification and electron microscopy, we found NO synthase expressed not only in the sarcolemma, but also along contractile fibers, in the sarcoplasmic reticulum and mitochondria. The expression pattern of NO synthase in myocytes showed striking parallels with the enzymes engaged in L-arginine-NO-cGMP signaling (arginase, phosphodiesterase, and soluble guanylyl cyclase). Our findings are indicative of an autocrine fashion of NO signaling in skeletal muscles at both cellular and subcellular levels, and challenge the notion that the NO generation is restricted to the sarcolemma.

  5. [Application of ultrasonic investigation in estimation of morphofunctional state of muscles in patients with ischemic contracture of foot].

    PubMed

    Liabakh, A P; Vovchenko, H Ia

    2002-03-01

    Results of ultrasonic investigation of the shin and foot muscles were analyzed in 14 patients with an ischemic contracture of foot (ICF). All the patients were operated on, clinical significance of ICF ultrasonic semiotics was estimated. Structural changes were demonstrated by diffuse strengthening of echogeneicity, striateness loss, ischemized muscles thickness reduction, origination of hypoechogenic (of hydrophilic nature) portions in places of necrosis of muscular tissues. Hyperechogeneicity is caused by fibrosis of an ischemized muscles, which during isometric contraction somewhat thickened, their echogeneicity did not change.

  6. Muscle Deoxygenation Causes Muscle Fatigue

    NASA Technical Reports Server (NTRS)

    Murthy, G.; Hargens, A. R.; Lehman, S.; Rempel, D.

    1999-01-01

    Muscle fatigue is a common musculoskeletal disorder in the work place, and may be a harbinger for more disabling cumulative trauma disorders. Although the cause of fatigue is multifactorial, reduced blood flow and muscle oxygenation may be the primary factor in causing muscle fatigue during low intensity muscle exertion. Muscle fatigue is defined as a reduction in muscle force production, and also occurs among astronauts who are subjected to postural constraints while performing lengthy, repetitive tasks. The objectives of this research are to: 1) develop an objective tool to study the role of decreased muscle oxygenation on muscle force production, and 2) to evaluate muscle fatigue during prolonged glovebox work.

  7. Muscle disorder

    MedlinePlus

    Myopathic changes; Myopathy; Muscle problem ... Blood tests sometimes show abnormally high muscle enzymes. If a muscle disorder might also affect other family members, genetic testing may be done. When someone has symptoms and signs ...

  8. Regulation of muscle contraction by Drebrin-like protein 1 probed by atomic force microscopy

    NASA Astrophysics Data System (ADS)

    Garces, Renata; Butkevich, Eugenia; Platen, Mitja; Schmidt, Christoph F.; Biophysics Team

    Sarcomeres are the fundamental contractile units of striated muscle cells. They are composed of a variety of structural and regulatory proteins functioning in a precisely orchestrated fashion to enable coordinated force generation in striated muscles. Recently, we have identified a C. elegans drebrin-like protein 1 (DBN-1) as a novel sarcomere component, which stabilizes actin filaments during muscle contraction. To further characterize the function of DBN-1 in muscle cells, we generated a new dbn-1 loss-of-function allele. Absence of DBN-1 resulted in a unique worm movement phenotype, characterized by hyper-bending. It is not clear yet if DBN-1 acts to enhance or reduce the capacity for contraction. We present here an experimental mechanical study on C. elegans muscle mechanics. We measured the stiffness of the worm by indenting living C. eleganswith a micron-sized sphere adhered to the cantilever of an atomic force microscope (AFM). Modeling the worm as a pressurized elastic shell allows us to monitor the axial tension in the muscle through the measured stiffness. We compared responses of wild-type and mutant C. elegans in which DBN-1 is not expressed..

  9. Protein machines and self assembly in muscle organization

    NASA Technical Reports Server (NTRS)

    Barral, J. M.; Epstein, H. F.

    1999-01-01

    The remarkable order of striated muscle is the result of a complex series of protein interactions at different levels of organization. Within muscle, the thick filament and its major protein myosin are classical examples of functioning protein machines. Our understanding of the structure and assembly of thick filaments and their organization into the regular arrays of the A-band has recently been enhanced by the application of biochemical, genetic, and structural approaches. Detailed studies of the thick filament backbone have shown that the myosins are organized into a tubular structure. Additional protein machines and specific myosin rod sequences have been identified that play significant roles in thick filament structure, assembly, and organization. These include intrinsic filament components, cross-linking molecules of the M-band and constituents of the membrane-cytoskeleton system. Muscle organization is directed by the multistep actions of protein machines that take advantage of well-established self-assembly relationships. Copyright 1999 John Wiley & Sons, Inc.

  10. Electron microscope observations on Ca2+-ATPase microcrystals in detergent-solubilized sarcoplasmic reticulum.

    PubMed

    Taylor, K A; Mullner, N; Pikula, S; Dux, L; Peracchia, C; Varga, S; Martonosi, A

    1988-04-15

    Crystalline arrays of Ca2+-ATPase molecules develop in detergent-solubilized sarcoplasmic reticulum during incubation for several weeks at 2 degrees C under nitrogen in a medium of 0.1 M KCl, 10 mM K-3-(N-morpholino)propanesulfonate, pH 6.0, 3 mM MgCl2, 20 mM CaCl2, 20% glycerol, 3 mM NaN3, 5 mM dithiothreitol, 25 IU/ml Trasylol, 2 micrograms/ml 1,6-di-tert-butyl-p-cresol, 2 mg/ml protein, and 2-4 mg of detergent/mg of protein. Electron microscopy of sectioned, negatively stained, freeze-fractured, and frozen-hydrated Ca2+-ATPase crystals indicates that they consist of stacked lamellar arrays of Ca2+-ATPase molecules. Prominent periodicities of ATPase molecules within the lamellae arise from a centered rectangular lattice of dimensions 164 x 55.5 A. The association of lamellae into three-dimensional stacks is assumed to involve interactions between the exposed hydrophilic headgroups of ATPase molecules, that is promoted by glycerol and 20 mM Ca2+. Similar Ca2+-induced crystals were observed with purified or purified and delipidated Ca2+-ATPase preparations at lower detergent/protein ratios. Cross-linking of Ca2+-ATPase crystals with glutaraldehyde protects the structure against conditions such as low Ca2+, high pH, elevated temperature, SH group reagents, high concentration of detergents, and removal of phospholipids by extraction with organic solvents that disrupt unfixed preparations. PMID:2965700

  11. The calcium stored in the sarcoplasmic reticulum acts as a safety mechanism in rainbow trout heart

    PubMed Central

    Cros, Caroline; Sallé, Laurent; Warren, Daniel E.; Shiels, Holly A.

    2014-01-01

    Cardiomyocyte contraction depends on rapid changes in intracellular Ca2+. In mammals, Ca2+ influx as L-type Ca2+ current (ICa) triggers the release of Ca2+ from sarcoplasmic reticulum (SR) and Ca2+-induced Ca2+ release (CICR) is critical for excitation-contraction coupling. In fish, the relative contribution of external and internal Ca2+ is unclear. Here, we characterized the role of ICa to trigger SR Ca2+ release in rainbow trout ventricular myocytes using ICa regulation by Ca2+ as an index of CICR. ICa was recorded with a slow (EGTA) or fast (BAPTA) Ca2+ chelator in control and isoproterenol conditions. In the absence of β-adrenergic stimulation, the rate of ICa inactivation was not significantly different in EGTA and BAPTA (27.1 ± 1.8 vs. 30.3 ± 2.4 ms), whereas with isoproterenol (1 μM), inactivation was significantly faster with EGTA (11.6 ± 1.7 vs. 27.3 ± 1.6 ms). When barium was the charge carrier, inactivation was significantly slower in both conditions (61.9 ± 6.1 vs. 68.0 ± 8.7 ms, control, isoproterenol). Quantification revealed that without isoproterenol, only 39% of ICa inactivation was due to Ca2+, while with isoproterenol, inactivation was Ca2+-dependent (∼65%) and highly reliant on SR Ca2+ (∼46%). Thus, SR Ca2+ is not released in basal conditions, and ICa is the main trigger of contraction, whereas during a stress response, SR Ca2+ is an important source of cytosolic Ca2+. This was not attributed to differences in SR Ca2+ load because caffeine-induced transients were not different in both conditions. Therefore, Ca2+ stored in SR of trout cardiomyocytes may act as a safety mechanism, allowing greater contraction when higher contractility is required, such as stress or exercise. PMID:25377479

  12. Critical roles of interdomain interactions for modulatory ATP binding to sarcoplasmic reticulum Ca2+-ATPase.

    PubMed

    Clausen, Johannes D; Holdensen, Anne Nyholm; Andersen, Jens Peter

    2014-10-17

    ATP has dual roles in the reaction cycle of sarcoplasmic reticulum Ca(2+)-ATPase. Upon binding to the Ca2E1 state, ATP phosphorylates the enzyme, and by binding to other conformational states in a non-phosphorylating modulatory mode ATP stimulates the dephosphorylation and other partial reaction steps of the cycle, thereby ensuring a high rate of Ca(2+) transport under physiological conditions. The present study elucidates the mechanism underlying the modulatory effect on dephosphorylation. In the intermediate states of dephosphorylation the A-domain residues Ser(186) and Asp(203) interact with Glu(439) (N-domain) and Arg(678) (P-domain), respectively. Single mutations to these residues abolish the stimulation of dephosphorylation by ATP. The double mutation swapping Asp(203) and Arg(678) rescues ATP stimulation, whereas this is not the case for the double mutation swapping Ser(186) and Glu(439). By taking advantage of the ability of wild type and mutant Ca(2+)-ATPases to form stable complexes with aluminum fluoride (E2·AlF) and beryllium fluoride (E2·BeF) as analogs of the E2·P phosphoryl transition state and E2P ground state, respectively, of the dephosphorylation reaction, the mutational effects on ATP binding to these intermediates are demonstrated. In the wild type Ca(2+)-ATPase, the ATP affinity of the E2·P phosphoryl transition state is higher than that of the E2P ground state, thus explaining the stimulation of dephosphorylation by nucleotide-induced transition state stabilization. We find that the Asp(203)-Arg(678) and Ser(186)-Glu(439) interdomain bonds are critical, because they tighten the interaction with ATP in the E2·P phosphoryl transition state. Moreover, ATP binding and the Ser(186)-Glu(439) bond are mutually exclusive in the E2P ground state. PMID:25193668

  13. The calcium stored in the sarcoplasmic reticulum acts as a safety mechanism in rainbow trout heart.

    PubMed

    Cros, Caroline; Sallé, Laurent; Warren, Daniel E; Shiels, Holly A; Brette, Fabien

    2014-12-15

    Cardiomyocyte contraction depends on rapid changes in intracellular Ca(2+). In mammals, Ca(2+) influx as L-type Ca(2+) current (ICa) triggers the release of Ca(2+) from sarcoplasmic reticulum (SR) and Ca(2+)-induced Ca(2+) release (CICR) is critical for excitation-contraction coupling. In fish, the relative contribution of external and internal Ca(2+) is unclear. Here, we characterized the role of ICa to trigger SR Ca(2+) release in rainbow trout ventricular myocytes using ICa regulation by Ca(2+) as an index of CICR. ICa was recorded with a slow (EGTA) or fast (BAPTA) Ca(2+) chelator in control and isoproterenol conditions. In the absence of β-adrenergic stimulation, the rate of ICa inactivation was not significantly different in EGTA and BAPTA (27.1 ± 1.8 vs. 30.3 ± 2.4 ms), whereas with isoproterenol (1 μM), inactivation was significantly faster with EGTA (11.6 ± 1.7 vs. 27.3 ± 1.6 ms). When barium was the charge carrier, inactivation was significantly slower in both conditions (61.9 ± 6.1 vs. 68.0 ± 8.7 ms, control, isoproterenol). Quantification revealed that without isoproterenol, only 39% of ICa inactivation was due to Ca(2+), while with isoproterenol, inactivation was Ca(2+)-dependent (∼65%) and highly reliant on SR Ca(2+) (∼46%). Thus, SR Ca(2+) is not released in basal conditions, and ICa is the main trigger of contraction, whereas during a stress response, SR Ca(2+) is an important source of cytosolic Ca(2+). This was not attributed to differences in SR Ca(2+) load because caffeine-induced transients were not different in both conditions. Therefore, Ca(2+) stored in SR of trout cardiomyocytes may act as a safety mechanism, allowing greater contraction when higher contractility is required, such as stress or exercise. PMID:25377479

  14. Defective sarcoplasmic reticulum–mitochondria calcium exchange in aged mouse myocardium

    PubMed Central

    Fernandez-Sanz, C; Ruiz-Meana, M; Miro-Casas, E; Nuñez, E; Castellano, J; Loureiro, M; Barba, I; Poncelas, M; Rodriguez-Sinovas, A; Vázquez, J; Garcia-Dorado, D

    2014-01-01

    Mitochondrial alterations are critically involved in increased vulnerability to disease during aging. We investigated the contribution of mitochondria–sarcoplasmic reticulum (SR) communication in cardiomyocyte functional alterations during aging. Heart function (echocardiography) and ATP/phosphocreatine (NMR spectroscopy) were preserved in hearts from old mice (>20 months) with respect to young mice (5–6 months). Mitochondrial membrane potential and resting O2 consumption were similar in mitochondria from young and old hearts. However, maximal ADP-stimulated O2 consumption was specifically reduced in interfibrillar mitochondria from aged hearts. Second generation proteomics disclosed an increased mitochondrial protein oxidation in advanced age. Because energy production and oxidative status are regulated by mitochondrial Ca2+, we investigated the effect of age on mitochondrial Ca2+ uptake. Although no age-dependent differences were found in Ca2+ uptake kinetics in isolated mitochondria, mitochondrial Ca2+ uptake secondary to SR Ca2+ release was significantly reduced in cardiomyocytes from old hearts, and this effect was associated with decreased NAD(P)H regeneration and increased mitochondrial ROS upon increased contractile activity. Immunofluorescence and proximity ligation assay identified the defective communication between mitochondrial voltage-dependent anion channel and SR ryanodine receptor (RyR) in cardiomyocytes from aged hearts associated with altered Ca2+ handling. Age-dependent alterations in SR Ca2+ transfer to mitochondria and in Ca2+ handling could be reproduced in cardiomyoctes from young hearts after interorganelle disruption with colchicine, at concentrations that had no effect in aged cardiomyocytes or isolated mitochondria. Thus, defective SR–mitochondria communication underlies inefficient interorganelle Ca2+ exchange that contributes to energy demand/supply mistmach and oxidative stress in the aged heart. PMID:25522267

  15. Detection, Properties, and Frequency of Local Calcium Release from the Sarcoplasmic Reticulum in Teleost Cardiomyocytes

    PubMed Central

    Alvarez-Lacalle, Enrique; Tort, Lluis; Benítez, Raul; Hove-Madsen, Leif

    2011-01-01

    Calcium release from the sarcoplasmic reticulum (SR) plays a central role in the regulation of cardiac contraction and rhythm in mammals and humans but its role is controversial in teleosts. Since the zebrafish is an emerging model for studies of cardiovascular function and regeneration we here sought to determine if basic features of SR calcium release are phylogenetically conserved. Confocal calcium imaging was used to detect spontaneous calcium release (calcium sparks and waves) from the SR. Calcium sparks were detected in 16 of 38 trout atrial myocytes and 6 of 15 ventricular cells. The spark amplitude was 1.45±0.03 times the baseline fluorescence and the time to half maximal decay of sparks was 27±3 ms. Spark frequency was 0.88 sparks µm−1 min−1 while calcium waves were 8.5 times less frequent. Inhibition of SR calcium uptake reduced the calcium transient (F/F0) from 1.77±0.17 to 1.12±0.18 (p = 0.002) and abolished calcium sparks and waves. Moreover, elevation of extracellular calcium from 2 to 10 mM promoted early and delayed afterdepolarizations (from 0.6±0.3 min−1 to 8.1±2.0 min−1, p = 0.001), demonstrating the ability of SR calcium release to induce afterdepolarizations in the trout heart. Calcium sparks of similar width and duration were also observed in zebrafish ventricular myocytes. In conclusion, this is the first study to consistently report calcium sparks in teleosts and demonstrate that the basic features of calcium release through the ryanodine receptor are conserved, suggesting that teleost cardiac myocytes is a relevant model to study the functional impact of abnormal SR function. PMID:21897853

  16. Structural effects of substrate utilization on the adenosinetriphosphatase chains of sarcoplasmic reticulum.

    PubMed

    Watanabe, T; Inesi, G

    1982-07-01

    Addition of ATP to suspensions of fragmented sarcoplasmic reticulum (SR) containing low concentrations of a detergent that does not by itself produce major vesicular disruption is followed by a transient reduction in turbidity accompanied by solubilization of the vesicles. The effect of ATP is Ca2+ dependent and proceeds in parallel with utilization of the nucleotide as a substrate for the SR ATPase. Analogous effects are observed with other substrates producing enzyme phosphorylation at the catalytic site. The effect of ATP can also be detected in studies of fluorescence energy transfer between enzyme chains, by using the technique of Vanderkooi et al. [Vanderkooi, J., Ierokomas, A., Nakamura, H., & Martonosi, A. (1977) Biochemistry 16, 1262]. For this purpose, ATPase chains are labeled separately with N-(iodoacetyl)-N'-(5-sulfo-1-naphthyl)ethylenediamine (IAEDANS) and 6-(iodoacetamido) fluorescein (IAF). Samples of vesicles uniformly labeled with either IAEDANS or IAF, mixtures of two populations of vesicles uniformly labeled with either fluorophore, and vesicles containing randomized chains labeled with either fluorophore are used as experimental systems. In the last system, significant energy transfer from IAEDANS (donor) to IAF (acceptor) is revealed by fluorescence spectra and measurements of donor fluorescence intensity and lifetime. This is attributed to close interactions between ATPase chains within the membrane bilayer. It is then found that in the presence of low detergent concentrations, ATP changes the extent of energy transfer between labeled ATPase chains, consistent with destabilization of the interaction of chains. The observed effects are attributed to a reversible structural transition concomitant with enzyme phosphorylation and related to catalytic and transport function.

  17. Topology of sarcoplasmic reticulum Ca2+-ATPase: an infrared study of thermal denaturation and limited proteolysis.

    PubMed

    Echabe, I; Dornberger, U; Prado, A; Goñi, F M; Arrondo, J L

    1998-05-01

    Sarcoplasmic reticulum Ca2+-ATPase structure and organization in the membrane has been studied by infrared spectroscopy by decomposition of the amide I band. Besides the component bands assignable to secondary structure elements such as alpha-helix, beta-sheet, etc...., two unusual bands, one at 1,645 cm(-1) in H2O buffer and the other at 1,625 cm(-1) in D2O buffer are present. By perturbing the protein using temperature and limited proteolysis, the band at 1,645 cm(-1) is tentatively assigned to alpha-helical segments located in the cytoplasmic domain and coupled to beta-sheet structure, whereas the band at 1,625 cm(-1) arises probably from monomer-monomer contacts in the native oligomeric protein. The secondary structure obtained is 33% alpha-helical segments in the transmembrane plus stalk domain; 20% alpha-helix and 22% beta-sheet in the cytoplasmic domain plus 19% turns and 6% unordered structure. Thermal unfolding of Ca2+-ATPase is a complex process that cannot be described as a two-state denaturation. The results obtained are compatible with the idea that the protein is an oligomer at room temperature. The loss of the 1,625 cm(-1) band upon heating would be consistent with a disruption of the oligomers in a process that later gives rise to aggregates (appearance of the 1,618 cm(-1) band). This picture would also be compatible with early results suggesting that processes governing Ca2+ accumulation and ATPase activity are uncoupled at temperatures above 37 degrees C, so that while ATPase activity proceeds at high rates, Ca2+ accumulation is inhibited.

  18. Topology of sarcoplasmic reticulum Ca2+-ATPase: an infrared study of thermal denaturation and limited proteolysis.

    PubMed Central

    Echabe, I.; Dornberger, U.; Prado, A.; Goñi, F. M.; Arrondo, J. L.

    1998-01-01

    Sarcoplasmic reticulum Ca2+-ATPase structure and organization in the membrane has been studied by infrared spectroscopy by decomposition of the amide I band. Besides the component bands assignable to secondary structure elements such as alpha-helix, beta-sheet, etc...., two unusual bands, one at 1,645 cm(-1) in H2O buffer and the other at 1,625 cm(-1) in D2O buffer are present. By perturbing the protein using temperature and limited proteolysis, the band at 1,645 cm(-1) is tentatively assigned to alpha-helical segments located in the cytoplasmic domain and coupled to beta-sheet structure, whereas the band at 1,625 cm(-1) arises probably from monomer-monomer contacts in the native oligomeric protein. The secondary structure obtained is 33% alpha-helical segments in the transmembrane plus stalk domain; 20% alpha-helix and 22% beta-sheet in the cytoplasmic domain plus 19% turns and 6% unordered structure. Thermal unfolding of Ca2+-ATPase is a complex process that cannot be described as a two-state denaturation. The results obtained are compatible with the idea that the protein is an oligomer at room temperature. The loss of the 1,625 cm(-1) band upon heating would be consistent with a disruption of the oligomers in a process that later gives rise to aggregates (appearance of the 1,618 cm(-1) band). This picture would also be compatible with early results suggesting that processes governing Ca2+ accumulation and ATPase activity are uncoupled at temperatures above 37 degrees C, so that while ATPase activity proceeds at high rates, Ca2+ accumulation is inhibited. PMID:9605321

  19. Estrogen upregulates the sarcoplasmic reticulum Ca2+-ATPase pump in coronary arteries

    PubMed Central

    Hill, Brent J.F.; Muldrew, Edwin

    2014-01-01

    The presence of circulating plasma 17β-estradiol (E2) is beneficial in women against abnormal vascular tone development, such as coronary arterial vasospasms. Several vascular diseases have demonstrated that an increased expression of the sarcoplasmic reticulum Ca2+-ATPase pump (SERCA2b) serves to limit the excessive accumulation of intracellular Ca2+. Therefore, the hypothesis of this study was that E2 would increase SERCA2b expression in the coronary vasculature. Coronary arteries were dissected from hearts obtained from mature female pigs. Artery segments were cultured for 24 hrs in E2 (1 pM or 1 nM) and homogenized for Western blot analysis. E2 (1 nM) induced a ~ 50% increase in the immunoreactivity for SERCA2b. E2 also increased the protein expression of the known SERCA regulatory proteins, protein kinase A (PKA) and protein kinase G (PKG). The E2-induced increase in SERCA2b was attenuated when the culture media was supplemented with the α/β estrogen receptor antagonist, ICI 182,780, and the PKG antagonist, KT5823. The PKA antagonist (KT5720) had no effect on SERCA2b expression. Removal of the endothelium (using a wooden toothpick) decreased the E2-mediated increase in SERCA2b and PKG expression by 45% and 47%, respectively. Overall, these findings suggest that one of the potential cardiovascular benefits of E2 in women is the upregulation of SERCA2b, via the activation of the classical α and β estrogen receptor pathway. PMID:24684418

  20. Bis-quaternary ammonium blockers as structural probes of the sarcoplasmic reticulum K+ channel

    PubMed Central

    1982-01-01

    A series of n-alkyl-bis-alpha,omega-trimethylammonium (bisQn) compounds was synthesized, and their ability to block K+ currents through a K+ channel from sarcoplasmic reticulum was studied. K+ channels were inserted into planar phospholipid membranes, and single-channel K+ currents were measured in the presence of the blocking cations. These bisQn compounds block K+ currents only from the side of the membrane opposite to the addition of SR vesicles (the trans side). The block is dependent on transmembrane voltage, and the effective valence of the block (a measure of this voltage dependence) varies with the methylene chain length. For short chains (bisQ2-bisQ5), the effective valence decreases with chain length from 1.1 to 0.65; it then remains constant at approximately 0.65 for bisQ5 to bisQ8; the effective valence abruptly increases to 1.2-1.3 for chains of nine carbons and longer. For the compounds of nine carbons and longer, the discrete nature of the block can be observed directly as 'flickering noise" on the open channel. The kinetics of the block were studied for these long-chain blockers. Both blocking and unblocking rates of the blockers vary with chain length, with the blocking rate showing the strongest variation-- an increase of 2.8-fold per added methylene group. All of the voltage dependence of the binding equilibrium resides in the blocking rate, and none in the unblocking rate. The results imply that 65% of the voltage drop within the channel occurs over a distance of 6-7A, and that the short-chain blockers bind in a bent-over conformation with both charges deeply inside the channel. PMID:6284862

  1. The effect of high hydrostatic pressure on the muscle proteins of rainbow trout (Oncorhynchus mykiss Walbaum) fillets wrapped with chitosan-based edible film during cold storage (4±1°C)

    NASA Astrophysics Data System (ADS)

    Günlü, Ali; Sipahioǧlu, Sinem; Alpas, Hami

    2014-01-01

    This study was to determine the effects of changes that occurred in the muscle proteins of fresh rainbow trout (Oncorhynchus mykiss) fillets during storage at 4±1°C as a result of packaging in vacuum (C), subject to high pressure after packaging in vacuum high hydrostatic pressue (HHP), packaged in vacuum after wrapping with chitosan film (CFW) and subject to high pressure after wrapping with chitosan-based film and packaged in vacuum (HHP+CFW ). Samples were subjected to SDS-PAGE in four-day intervals and the densitometric analyses of the gels were carried out. According to the results, minor changes were determined in the major bands of the sarcoplasmic and myofibrillar muscle fractions of trouts as a result of HHP application and CFW. The most important change occurred in the myofibrillar muscle fraction as a decrease in the densities of the bands at 200 and 31.4 kDa after HHP application. Similarly, the sarcoplasmic muscle fraction of trout fillet decreased in the densities of the bands at 39.3, 26.6 and 23.3 kDa after HHP application. In addition, it is thought that the bands that occur at 30 kDa in the myofibrillar muscle fraction and at 20.7 kDa at the sarcoplasmic muscle fraction may be related with the degradation of trouts during cold storage.

  2. Protein diffusion in living skeletal muscle fibers: dependence on protein size, fiber type, and contraction.

    PubMed Central

    Papadopoulos, S; Jürgens, K D; Gros, G

    2000-01-01

    Sarcoplasmic protein diffusion was studied under different conditions, using microinjection in combination with microspectrophotometry. Six globular proteins with molecular masses between 12 and 3700 kDa, with diameters from 3 to 30 nm, were used for the experiments. Proteins were injected into single, intact skeletal muscle fibers taken from either soleus or extensor digitorum longus (edl) muscle of adult rats. No correlation was found between sarcomere spacing and the sarcoplasmic diffusion coefficient (D) for all proteins studied. D of the smaller proteins cytochrome c (diameter 3.1 nm), myoglobin (diameter 3.5 nm), and hemoglobin (diameter 5.5 nm) amounted to only approximately 1/10 of their value in water and was not increased by auxotonic fiber contractions. D for cytochrome c and myoglobin was significantly higher in fibers from edl (mainly type II fibers) compared to fibers from soleus (mainly type I fibers). Measurements of D for myoglobin at 37 degrees C in addition to 22 degrees C led to a Q(10) of 1.46 for this temperature range. For the larger proteins catalase (diameter 10.5 nm) and ferritin (diameter 12.2 nm), a decrease in D to approximately 1/20 and approximately 1/50 of that in water was observed, whereas no diffusive flux at all of earthworm hemoglobin (diameter 30 nm) along the fiber axis could be detected. We conclude that 1) sarcoplasmic protein diffusion is strongly impaired by the presence of the myofilamental lattice, which also gives rise to differences in diffusivity between different fiber types; 2) contractions do not cause significant convection in sarcoplasm and do not lead to increased diffusional transport; and 3) in addition to the steric hindrance that slows down the diffusion of smaller proteins, diffusion of large proteins is further hindered when their dimensions approach the interfilament distances. This molecular sieve property progressively reduces intracellular diffusion of proteins when the molecular diameter increases to

  3. Muscle changes in the neuroleptic malignant syndrome

    PubMed Central

    Behan, W; Madigan, M; Clark, B; Goldberg, J; McLellan, D

    2000-01-01

    Aims—To characterise the skeletal muscle changes in the neuroleptic malignant syndrome (NMS). Methods—Detailed light and ultrastructural examination was carried out on skeletal muscle from three cases of NMS, two associated with recreational drugs (3,4-methlenedioxymethylamphetamine (MDMA, Ecstasy) and lysergic acid diethylamide (LSD)) and one with antipsychotic drugs (fluoxetine (Prozac) and remoxipride hydrochloride monohydrate (Roxiam)). Results—The muscles were grossly swollen and oedematous in all cases, in one with such severe local involvement that the diagnosis of sarcoma was considered. On microscopy, there was conspicuous oedema. In some fascicles less than 10% of fibres were affected whereas in others more than 50% were pale and enlarged. There was a spectrum of changes: tiny to large vacuoles replaced most of the sarcoplasm and were associated with necrosis. A striking feature in some fibres was the presence of contraction bands separating segments of oedematous myofibrils. Severe endomysial oedema was also detectable. There was a scanty mononuclear infiltrate but no evidence of regeneration. Conclusions—The muscle changes associated with NMS are characteristic and may be helpful in differential diagnosis. Key Words: myopathy • neuroleptic malignant syndrome • fluoxetine • remoxipride hydrochloride monohydrate • Ecstasy • LSD PMID:10823143

  4. Atypical 'fibrillar' flight muscle in strepsiptera.

    PubMed

    Smith, D S; Kathirithamby, J

    1984-01-01

    The fine structure of the principal and ancillary metathoracic flight muscle fibres in the adult male of a strepsipteran, Elenchus tenuicornis, is described. Power-producing dorsal longitudinal and dorso-ventral flight muscles show features consistent with myoneural asynchrony: myofibrils are large and discrete and are separated by large closely packed mitochondria; the sarcoplasmic reticulum is very reduced but engages with T-system membranes in dyads at the mid-sarcomere H-band level. With respect to other asynchronous insect flight muscles, the fibres of Elenchus are anomalous (i) in the small fibre diameter, (ii) in the variable contour of the myofibrils and (iii) in the absence of tracheolar invagination. The functional significance of these structural features is discussed. Ancillary metathoracic muscles are structurally comparable with other synchronous fibres in possessing an extensive SR compartment. Structural evidence for asynchrony in the flight mechanism of Strepsiptera is considered in the context of the evolution of this mechanism throughout the insect Orders. PMID:6531780

  5. Drosophila UNC-45 accumulates in embryonic blastoderm and in muscles, and is essential for muscle myosin stability.

    PubMed

    Lee, Chi F; Melkani, Girish C; Yu, Qin; Suggs, Jennifer A; Kronert, William A; Suzuki, Yoko; Hipolito, Lori; Price, Maureen G; Epstein, Henry F; Bernstein, Sanford I

    2011-03-01

    UNC-45 is a chaperone that facilitates folding of myosin motor domains. We have used Drosophila melanogaster to investigate the role of UNC-45 in muscle development and function. Drosophila UNC-45 (dUNC-45) is expressed at all developmental stages. It colocalizes with non-muscle myosin in embryonic blastoderm of 2-hour-old embryos. At 14 hours, it accumulates most strongly in embryonic striated muscles, similarly to muscle myosin. dUNC-45 localizes to the Z-discs of sarcomeres in third instar larval body-wall muscles. We produced a dunc-45 mutant in which zygotic expression is disrupted. This results in nearly undetectable dUNC-45 levels in maturing embryos as well as late embryonic lethality. Muscle myosin accumulation is robust in dunc-45 mutant embryos at 14 hours. However, myosin is dramatically decreased in the body-wall muscles of 22-hour-old mutant embryos. Furthermore, electron microscopy showed only a few thick filaments and irregular thick-thin filament lattice spacing. The lethality, defective protein accumulation, and ultrastructural abnormalities are rescued with a wild-type dunc-45 transgene, indicating that the mutant phenotypes arise from the dUNC-45 deficiency. Overall, our data indicate that dUNC-45 is important for myosin accumulation and muscle function. Furthermore, our results suggest that dUNC-45 acts post-translationally for proper myosin folding and maturation.

  6. Comparative Sensitivity Analysis of Muscle Activation Dynamics.

    PubMed

    Rockenfeller, Robert; Günther, Michael; Schmitt, Syn; Götz, Thomas

    2015-01-01

    We mathematically compared two models of mammalian striated muscle activation dynamics proposed by Hatze and Zajac. Both models are representative for a broad variety of biomechanical models formulated as ordinary differential equations (ODEs). These models incorporate parameters that directly represent known physiological properties. Other parameters have been introduced to reproduce empirical observations. We used sensitivity analysis to investigate the influence of model parameters on the ODE solutions. In addition, we expanded an existing approach to treating initial conditions as parameters and to calculating second-order sensitivities. Furthermore, we used a global sensitivity analysis approach to include finite ranges of parameter values. Hence, a theoretician striving for model reduction could use the method for identifying particularly low sensitivities to detect superfluous parameters. An experimenter could use it for identifying particularly high sensitivities to improve parameter estimation. Hatze's nonlinear model incorporates some parameters to which activation dynamics is clearly more sensitive than to any parameter in Zajac's linear model. Other than Zajac's model, Hatze's model can, however, reproduce measured shifts in optimal muscle length with varied muscle activity. Accordingly we extracted a specific parameter set for Hatze's model that combines best with a particular muscle force-length relation. PMID:26417379

  7. Calcium oscillations in human mesenteric vascular smooth muscle.

    PubMed

    Navarro-Dorado, Jorge; Garcia-Alonso, Mauricio; van Breemen, Cornelis; Tejerina, Teresa; Fameli, Nicola

    2014-02-28

    Phenylephrine (PE)-induced oscillatory fluctuations in intracellular Ca(2+) concentration ([Ca(2+)]i) of vascular smooth muscle have been observed in many blood vessels isolated from a wide variety of mammals. Paradoxically, until recently similar observations in humans have proven elusive. In this study, we report for the first time observations of adrenergically-stimulated [Ca(2+)]i oscillations in human mesenteric artery smooth muscle. In arterial segments preloaded with Fluo-4 AM and mounted on a myograph on the stage of a confocal microscope, we observed PE-induced oscillations in [Ca(2+)]i, which initiated and maintained vasoconstriction. These oscillations present some variability, possibly due to compromised health of the tissue. This view is corroborated by our ultrastructural analysis of the cells, in which we found only (5 ± 2)% plasma membrane-sarcoplasmic reticulum apposition, markedly less than measured in healthy tissue from laboratory animals. We also partially characterized the oscillations by using the inhibitory drugs 2-aminoethoxydiphenyl borate (2-APB), cyclopiazonic acid (CPA) and nifedipine. After PE contraction, all drugs provoked relaxation of the vessel segments, sometimes only partial, and reduced or inhibited oscillations, except CPA, which rarely caused relaxation. These preliminary results point to a potential involvement of the sarcoplasmic reticulum Ca(2+) and inositol 1,4,5-trisphosphate receptor (IP3R) in the maintenance of the Ca(2+) oscillations observed in human blood vessels. PMID:24508261

  8. Compliance Accelerates Relaxation in Muscle by Allowing Myosin Heads to Move Relative to Actin.

    PubMed

    Campbell, Kenneth S

    2016-02-01

    The mechanisms that limit the speed at which striated muscles relax are poorly understood. This work presents, to our knowledge, novel simulations that show that the time course of relaxation is accelerated by interfilamentary movement resulting from series compliance; force drops faster when myosin heads move relative to actin during relaxation. This insight was obtained by using cross-bridge distribution techniques to simulate the mechanical behavior of half-sarcomeres that were connected in series with springs of varying stiffness. (The springs mimic the combined effects of half-sarcomere heterogeneity and muscle's series elastic component.) Half-sarcomeres that shortened by >∼10 nm when they were activated subsequently relaxed with a biphasic profile; force initially declined slowly and approximately linearly before collapsing with a fast exponential time course. Stretches imposed during the linear phase quickened relaxation, while shortening movements prolonged the time course. These predictions are consistent with data from experiments performed by many other groups using single muscle fibers and isolated myofibrils. When half-sarcomeres were linked to stiff springs (so that they did not shorten appreciably during the simulations), force relaxed with a slow exponential time course and did not show biphasic behavior. Together, these results suggest that fast relaxation of striated muscle is an emergent property that reflects multiscale interactions within the muscle architecture. The nonlinear behavior during relaxation reflects perturbations to the dynamic coupling of regulated binding sites and cycling myosin heads that are induced by interfilamentary movement.

  9. Changes in skeletal muscle properties following hindlimb suspension

    NASA Technical Reports Server (NTRS)

    Patterson, G. T.; Dettbarn, W.-D.

    1985-01-01

    Changes in AChE regulation, the functions of the sarcoplasmic reticulum (SR), and muscle fiber-type population in the extensor digitorium longus (EDL) and soleus of male rats suspended in a nonweight-bearing position are studied. Muscle weight, AChE activity, protein content, SR calcium loading rates, and muscle types in the suspended and control rats are evaluated and compared. It is observed that following 1-4 weeks of suspension the EDL is unaffected; however, the soleus muscle weight, and protein content decrease and the AChE activity increases. The SR calcium loading rates of EDL decrease 10 percent and the soleus increase 50 percent in the suspended rats. The data reveal that the population of muscle type 1 (slow fibers) decreases and type 2 (fast fibers) increases for the soleus, and the distribution is unchanged for the EDL of the suspended rats. It is detected that a nonweight-bearing position affects the soleus (a fast-twitch muscle), but not the EDL (a slow-twitch muscle).

  10. Potential dynamics of the human striate cortex cerebrum realistic neural network under the influence of an external signal

    NASA Astrophysics Data System (ADS)

    Melnikov, Leonid A.; Novosselova, Anna V.; Blinova, Nadejda V.; Vinitsky, Sergey I.; Serov, Vladislav V.; Bakutkin, Valery V.; Camenskich, T. G.; Guileva, E. V.

    2000-03-01

    In this work the numerical investigations of a potential dynamics of a neural network as the non-linear system and dynamics of the visual nerve which connect the eye retina receptors with the striate cortex cerebrum as the answer to the through-skin excitement of the eye retina by the electrical signal were realized. The visual evoked potential is the answer and characterizes the human brain state over the structures retina state and the conduction of the visual nerve fibers. The results of these investigations were presented. Specific features of the neural network, such as the excitation and depression, we took into account too. The discussion about the model parameters, used at the time of the numerical investigation, was made. The comparative analysis of the retina potential data and the data of the external signal filing by the brain hemicerebrum visual centers was made too.

  11. Computed tomography of cerebral infarction along the distribution of the basal perforating arteries. Part 1. Striate arterial group

    SciTech Connect

    Takahashi, S.; Goto, K.; Fukasawa, H.; Kawata, Y.; Uemura, K.; Suzuki, K.

    1985-04-01

    Computed tomographic (CT) manifestations of cerebral infarction along the distribution of the basal perforating arteries were reviewed in correlation with cerebral angiography. Infarcts in the territories of perforators were demonstrated individually based on knowledge of their three dimensional distribution as demonstrated by microangiography of cadavers. In Part I of the study, the areas supplied by the medial (MSA) and lateral striate arteries (LSA) were examined. Infarction along the branches of the MSA usually involved the antero-inferior portion of the corpus striatum, immediately posterolateral to the most inferior part of the frontal horn of the lateral ventricle. Infarcts along the branches of the LSA abutted the territory of the MSA posteriorly and superiorly and involved the posterolateral region of the corpus striatum. Clinical and neuroradiological correlations are discussed.

  12. Co-localization of glutamic acid decarboxylase and vesicular GABA transporter in cytochrome oxidase patches of macaque striate cortex.

    PubMed

    Adams, Daniel L; Economides, John R; Horton, Jonathan C

    2015-01-01

    The patches in primary visual cortex constitute hot spots of metabolic activity, manifested by enhanced levels of cytochrome oxidase (CO) activity. They are also labeled preferentially by immunostaining for glutamic acid decarboxylase (GAD), γ-aminobutyric acid (GABA), and parvalbumin. However, calbindin shows stronger immunoreactivity outside patches. In light of this discrepancy, the distribution of the vesicular GABA transporter (VGAT) was examined in striate cortex of two normal macaques. VGAT immunoreactivity was strongest in layers 4B, 4Cα, and 5. In tangential sections, the distribution of CO, GAD, and VGAT was compared in layer 2/3. There was a close match between all three labels. This finding indicates that GABA synthesis is enriched in patches, and that inhibitory synapses are more active in patches than interpatches. PMID:26579566

  13. Relationship between large-aperture optical components of striated surface shape and focal spot characteristics in the far-field

    NASA Astrophysics Data System (ADS)

    Lei, Zemin; Sun, Xiaoyan; Yin, Xianhua; Lv, Fengnian; Zhang, Zhen; Lu, Xingqiang; Fan, Dianyuan

    2015-07-01

    Surface shape of optical components is an essential factor of the laser beam quality. Different types of surface correspond to different characteristics of the laser focal spot. Striated surface shape is one of common and typical cases of optical component surfaces in laser facilities, which have attracted great attention. For learning the impact of the component on focal spot in the far-field, a model component with the similar features was introduced in the study. Intensity distributions of focal spot in the far-field was simulated after laser beam went through the model component. Effects of the modulation depth and the modulation period on spot morphology were presented. Furthermore, the relations between these optical specifications and focal spots with some requirements had been analyzed. The results can enhance our understanding about striae degrees of optical elements and have reference values to guide the processing and the use of large-aperture components correctly.

  14. A Study of the Reinnervation of Fast and Slow Mammalian Muscles

    PubMed Central

    McArdle, Joseph J.; Albuquerque, Edson X.

    1973-01-01

    Miniature end plate potential (mepp) frequency in innervated extensor muscle is significantly higher than in soleus muscle. 9 days after nerve crush mepps of low amplitude and prolonged duration reappeared at a frequency of 2% of control and were similar to normal muscles after 35 days. Membrane potential began to increase 9–10 days after nerve crush and at 30 days was similar to controls. The region most sensitive to ACh in denervated and reinnervated muscles was the end plate. Caffeine (20 mM, 23°C) induced contracture in innervated soleus but not in extensor muscles. After denervation the extensor became sensitive to caffeine while the soleus muscles decreased in sensitivity to the drug; 4–5 days after reinnervation the effect of caffeine on these muscles was similar to control. The events during reinnervation are: (a) reappearance of mepps at the same time as end plate potential and muscle twitch; (b) partial restoration of the membrane potential; (c) return of caffeine-induced contracture to normal levels in the soleus and its absence in the extensor muscles; (d) return of membrane resistance to normal values in both muscles at about 25 days; and (e) return of ACh-sensitivity to control levels at about 30 days in both muscles. Although these results suggest that the membrane potential and sarcoplasmic reticulum are under neural influence, it remains to be established whether or not separate neurotrophic factors are involved. PMID:4683095

  15. Morphofunctional responses to anaemia in rat skeletal muscle

    PubMed Central

    Esteva, Santiago; Panisello, Pere; Casas, Mireia; Torrella, Joan Ramon; Pagés, Teresa; Viscor, Ginés

    2008-01-01

    Adult male Sprague-Dawley rats were randomly assigned to two groups: control and anaemic. Anaemia was induced by periodical blood withdrawal. Extensor digitorum longus and soleus muscles were excised under pentobarbital sodium total anaesthesia and processed for transmission electron microscopy, histochemical and biochemical analyses. Mitochondrial volume was determined by transmission electron microscopy in three different regions of each muscle fibre: pericapillary, sarcolemmal and sarcoplasmatic. Muscle samples sections were also stained with histochemical methods (SDH and m-ATPase) to reveal the oxidative capacity and shortening velocity of each muscle fibre. Determinations of fibre and capillary densities and fibre type composition were made from micrographs of different fixed fields selected in the equatorial region of each rat muscle. Determination of metabolites (ATP, inorganic phosphate, creatine, creatine phosphate and lactate) was done using established enzymatic methods and spectrophotometric detection. Significant differences in mitochondrial volumes were found between pericapillary, sarcolemmal and sarcoplasmic regions when data from animal groups were tested independently. Moreover, it was verified that anaemic rats had significantly lower values than control animals in all the sampled regions of both muscles. These changes were associated with a significantly higher proportion of fast fibres in anaemic rat soleus muscles (slow oxidative group = 63.8%; fast glycolytic group = 8.2%; fast oxidative glycolytic group = 27.4%) than in the controls (slow oxidative group = 79.0%; fast glycolytic group = 3.9%; fast oxidative glycolytic group = 17.1%). No significant changes were detected in the extensor digitorum longus muscle. A significant increase was found in metabolite concentration in both the extensor digitorum longus and soleus muscles of the anaemic animals as compared to the control group. In conclusion, hypoxaemic hypoxia causes a reduction in

  16. A peptide encoded by a transcript annotated as long noncoding RNA enhances SERCA activity in muscle.

    PubMed

    Nelson, Benjamin R; Makarewich, Catherine A; Anderson, Douglas M; Winders, Benjamin R; Troupes, Constantine D; Wu, Fenfen; Reese, Austin L; McAnally, John R; Chen, Xiongwen; Kavalali, Ege T; Cannon, Stephen C; Houser, Steven R; Bassel-Duby, Rhonda; Olson, Eric N

    2016-01-15

    Muscle contraction depends on release of Ca(2+) from the sarcoplasmic reticulum (SR) and reuptake by the Ca(2+)adenosine triphosphatase SERCA. We discovered a putative muscle-specific long noncoding RNA that encodes a peptide of 34 amino acids and that we named dwarf open reading frame (DWORF). DWORF localizes to the SR membrane, where it enhances SERCA activity by displacing the SERCA inhibitors, phospholamban, sarcolipin, and myoregulin. In mice, overexpression of DWORF in cardiomyocytes increases peak Ca(2+) transient amplitude and SR Ca(2+) load while reducing the time constant of cytosolic Ca(2+) decay during each cycle of contraction-relaxation. Conversely, slow skeletal muscle lacking DWORF exhibits delayed Ca(2+) clearance and relaxation and reduced SERCA activity. DWORF is the only endogenous peptide known to activate the SERCA pump by physical interaction and provides a means for enhancing muscle contractility. PMID:26816378

  17. The role of nitric oxide in muscle fibers with oxidative phosphorylation defects

    SciTech Connect

    Tengan, Celia H. . E-mail: chtengan@neuro.epm.br; Kiyomoto, Beatriz H.; Godinho, Rosely O.; Gamba, Juliana; Neves, Afonso C.; Schmidt, Beny; Oliveira, Acary S.B.; Gabbai, Alberto A.

    2007-08-03

    NO has been pointed as an important player in the control of mitochondrial respiration, especially because of its inhibitory effect on cytochrome c oxidase (COX). However, all the events involved in this control are still not completely elucidated. We demonstrate compartmentalized abnormalities on nitric oxide synthase (NOS) activity on muscle biopsies of patients with mitochondrial diseases. NOS activity was reduced in the sarcoplasmic compartment in COX deficient fibers, whereas increased activity was found in the sarcolemma of fibers with mitochondrial proliferation. We observed increased expression of neuronal NOS (nNOS) in patients and a correlation between nNOS expression and mitochondrial content. Treatment of skeletal muscle culture with an NO donor induced an increase in mitochondrial content. Our results indicate specific roles of NO in compensatory mechanisms of muscle fibers with mitochondrial deficiency and suggest the participation of nNOS in the signaling process of mitochondrial proliferation in human skeletal muscle.

  18. Ligands for FKBP12 Increase Ca2+ Influx and Protein Synthesis to Improve Skeletal Muscle Function*

    PubMed Central

    Lee, Chang Seok; Georgiou, Dimitra K.; Dagnino-Acosta, Adan; Xu, Jianjun; Ismailov, Iskander I.; Knoblauch, Mark; Monroe, Tanner O.; Ji, RuiRui; Hanna, Amy D.; Joshi, Aditya D.; Long, Cheng; Oakes, Joshua; Tran, Ted; Corona, Benjamin T.; Lorca, Sabina; Ingalls, Christopher P.; Narkar, Vihang A.; Lanner, Johanna T.; Bayle, J. Henri; Durham, William J.; Hamilton, Susan L.

    2014-01-01

    Rapamycin at high doses (2–10 mg/kg body weight) inhibits mammalian target of rapamycin complex 1 (mTORC1) and protein synthesis in mice. In contrast, low doses of rapamycin (10 μg/kg) increase mTORC1 activity and protein synthesis in skeletal muscle. Similar changes are found with SLF (synthetic ligand for FKBP12, which does not inhibit mTORC1) and in mice with a skeletal muscle-specific FKBP12 deficiency. These interventions also increase Ca2+ influx to enhance refilling of sarcoplasmic reticulum Ca2+ stores, slow muscle fatigue, and increase running endurance without negatively impacting cardiac function. FKBP12 deficiency or longer treatments with low dose rapamycin or SLF increase the percentage of type I fibers, further adding to fatigue resistance. We demonstrate that FKBP12 and its ligands impact multiple aspects of muscle function. PMID:25053409

  19. Mononuclear muscle cells in Drosophila ovaries revealed by GFP protein traps

    PubMed Central

    Hudson, Andrew M.; Petrella, Lisa N.; Tanaka, Akemi J.; Cooley, Lynn

    2008-01-01

    Genetic analysis of muscle specification, formation and function in model systems has provided valuable insight into human muscle physiology and disease. Studies in Drosophila have been particularly useful for discovering key genes involved in muscle specification, myoblast fusion, and sarcomere organization. The muscles of the Drosophila female reproductive system have received little attention despite extensive work on oogenesis. We have used newly available GFP protein trap lines to characterize of ovarian muscle morphology and sarcomere organization. The muscle cells surrounding the oviducts are multinuclear with highly organized sarcomeres typical of somatic muscles. In contrast, the two muscle layers of the ovary, which are derived from gonadal mesoderm, have a mesh-like morphology similar to gut visceral muscle. Protein traps in the Fasciclin 3 gene produced Fas3::GFP that localized in dots around the periphery of epithelial sheath cells, the muscle surrounding ovarioles. Surprisingly, the epithelial sheath cells each contain a single nucleus, indicating these cells do not undergo myoblast fusion during development. Consistent with this observation, we were able to use the Flp/FRT system to efficiently generate genetic mosaics in the epithelial sheath, suggesting these cells provide a new opportunity for clonal analysis of adult striated muscle. PMID:18199432

  20. The Structure and Regulation of Human Muscle α-Actinin

    PubMed Central

    Ribeiro, Euripedes de Almeida; Pinotsis, Nikos; Ghisleni, Andrea; Salmazo, Anita; Konarev, Petr V.; Kostan, Julius; Sjöblom, Björn; Schreiner, Claudia; Polyansky, Anton A.; Gkougkoulia, Eirini A.; Holt, Mark R.; Aachmann, Finn L.; Žagrović, Bojan; Bordignon, Enrica; Pirker, Katharina F.; Svergun, Dmitri I.; Gautel, Mathias; Djinović-Carugo, Kristina

    2014-01-01

    Summary The spectrin superfamily of proteins plays key roles in assembling the actin cytoskeleton in various cell types, crosslinks actin filaments, and acts as scaffolds for the assembly of large protein complexes involved in structural integrity and mechanosensation, as well as cell signaling. α-actinins in particular are the major actin crosslinkers in muscle Z-disks, focal adhesions, and actin stress fibers. We report a complete high-resolution structure of the 200 kDa α-actinin-2 dimer from striated muscle and explore its functional implications on the biochemical and cellular level. The structure provides insight into the phosphoinositide-based mechanism controlling its interaction with sarcomeric proteins such as titin, lays a foundation for studying the impact of pathogenic mutations at molecular resolution, and is likely to be broadly relevant for the regulation of spectrin-like proteins. PMID:25433700

  1. Modeling Muscles

    ERIC Educational Resources Information Center

    Goodwyn, Lauren; Salm, Sarah

    2007-01-01

    Teaching the anatomy of the muscle system to high school students can be challenging. Students often learn about muscle anatomy by memorizing information from textbooks or by observing plastic, inflexible models. Although these mediums help students learn about muscle placement, the mediums do not facilitate understanding regarding integration of…

  2. The use of label-free mass spectrometry for relative quantification of sarcoplasmic proteins during the processing of dry-cured ham.

    PubMed

    Gallego, Marta; Mora, Leticia; Concepción Aristoy, M; Toldrá, Fidel

    2016-04-01

    The aim of this work was to quantify changes in the abundance of the major sarcoplasmic proteins throughout the ham dry-curing process by using a label-free mass spectrometry methodology based on the measurement of mass spectral peak intensities obtained from the extracted ion chromatogram. For this purpose, extraction of sarcoplasmic proteins was followed by trypsin digestion and analysis by nanoliquid chromatography coupled to tandem mass spectrometry (Q/TOF) for the identification and relative quantification of sarcoplasmic proteins through individual quantification of trypsinised peptides. In total, 20 proteins, including 12 glycolytic enzymes, were identified and quantified. The accuracy of the protocol was based on MS/MS replicates, and beta-lactoglobulin protein was used to normalise data and correct possible variations during sample preparation or LC-MS/MS analysis. Mass spectrometry-based proteomics provides precise identification and quantification of proteins in comparison with traditional methodologies based on gel electrophoresis, especially in the case of overlapping proteins. Moreover, the label-free approach used in this study proved to be a simple, fast, reliable method for evaluating proteolytic degradation of sarcoplasmic proteins during the processing of dry-cured ham.

  3. Alterations in Skeletal Muscle Microcirculation of Head-Down Tilted Rats

    NASA Technical Reports Server (NTRS)

    Musacchia, X. J.; Stepke, Bernhard; Fleming, John T.; Joshua, Irving G.

    1992-01-01

    In this study we assessed the function of microscopic blood vessels in skeletal muscle (cremaster muscle) for alterations which may contribute to the observed elevation of blood pressure associated with head-down tilted whole body suspension (HDT/WBS), a model of weightlessness. Arteriolar baseline diameters, vasoconstrictor responses to norepinephrine (NE) and vasodilation to nitroprusside (NP) were assessed in control rats, rats suspended for 7 or 14 day HDT/WBS rats, and rats allowed to recover for 1 day after 7 days HDT/WBS. Neither baseline diameters nor ability to dilate were influenced by HDT/WBS. Maximum vasoconstriction to norepinephrine was significantly greater in arterioles of hypertensive 14 day HDT/WBS rats. This first study of the intact microvasculature in skeletal muscle indicates that an elevated contractility of arterioles to norepinephrine in suspended rats, and suggests an elevated peripheral resistance in striated muscle may contribute to the increase in blood pressures among animals subjected to HDT/WBS.

  4. Titin, a Central Mediator for Hypertrophic Signaling, Exercise-Induced Mechanosignaling and Skeletal Muscle Remodeling

    PubMed Central

    Krüger, Martina; Kötter, Sebastian

    2016-01-01

    Titin is a giant scaffold protein with multiple functions in striated muscle physiology. Due to the elastic I-band domains and the filament-like integration in the half-sarcomere titin is an important factor for sarcomere assembly and serves as an adaptable molecular spring that determines myofilament distensibility. Protein-interactions e.g., with muscle ankyrin repeat proteins or muscle LIM-protein link titin to hypertrophic signaling and via p62 and Muscle Ring Finger proteins to mechanisms that control protein quality control. This review summarizes our current knowledge on titin as a central node for exercise-induced mechanosignaling and remodeling and further highlights the pathophysiological implications. PMID:26973541

  5. Independent flexible motion of submolecular domains of the Ca2+,Mg2+-ATPase of sarcoplasmic reticulum measured by time-resolved fluorescence depolarization of site-specifically attached probes.

    PubMed

    Suzuki, S; Kawato, S; Kouyama, T; Kinosita, K; Ikegami, A; Kawakita, M

    1989-09-19

    The Ca2+-transporting ATPase of rabbit skeletal muscle sarcoplasmic reticulum was site-specifically labeled with either N-(1-anilinonaphth-4-yl)maleimide (ANM) or 5-[[(iodoacetamido)-ethyl]amino]naphthalene-1-sulfonate (IAEDANS), and the segmental motion of submolecular domains of the ATPase molecule was examined by means of time-resolved and steady-state fluorescence anisotropy measurements. The ANM-binding domain showed wobbling with a rotational relaxation time phi = 69 ns in the absence of free Ca2+ without any independent wobbling of the ANM moiety. The IAEDANS-binding domain showed a significantly slower wobbling with phi = 190 ns in the absence of Ca2+. The present results demonstrated for the first time that the ATPase molecule is composed of distinct domains whose mobilities are considerably different from each other. The binding of Ca2+ to the transport site increased the segmental motion of ANM-labeled domain, leading to a phi value of 65 ns. Solubilization of the ANM-labeled SR membranes by deoxycholate led to a further increase in the segmental flexibility (phi = 48 ns in the absence of free Ca2+), indicating that the mobility of the ANM-binding domain was considerably restricted through interaction with the membrane. The mobility of the ANM-binding domain of solubilized ATPase was also increased to some extent upon binding of Ca2+.

  6. Myopathic changes in murine skeletal muscle lacking synemin

    PubMed Central

    García-Pelagio, Karla P.; Muriel, Joaquin; O'Neill, Andrea; Desmond, Patrick F.; Lovering, Richard M.; Lund, Linda; Bond, Meredith

    2015-01-01

    Diseases of striated muscle linked to intermediate filament (IF) proteins are associated with defects in the organization of the contractile apparatus and its links to costameres, which connect the sarcomeres to the cell membrane. Here we study the role in skeletal muscle of synemin, a type IV IF protein, by examining mice null for synemin (synm-null). Synm-null mice have a mild skeletal muscle phenotype. Tibialis anterior (TA) muscles show a significant decrease in mean fiber diameter, a decrease in twitch and tetanic force, and an increase in susceptibility to injury caused by lengthening contractions. Organization of proteins associated with the contractile apparatus and costameres is not significantly altered in the synm-null. Elastimetry of the sarcolemma and associated contractile apparatus in extensor digitorum longus myofibers reveals a reduction in tension consistent with an increase in sarcolemmal deformability. Although fatigue after repeated isometric contractions is more marked in TA muscles of synm-null mice, the ability of the mice to run uphill on a treadmill is similar to controls. Our results suggest that synemin contributes to linkage between costameres and the contractile apparatus and that the absence of synemin results in decreased fiber size and increased sarcolemmal deformability and susceptibility to injury. Thus synemin plays a moderate but distinct role in fast twitch skeletal muscle. PMID:25567810

  7. Electrostatic forces in muscle and cylindrical gel systems

    SciTech Connect

    Millman, B.M.; Nickel, B.G.

    1980-10-01

    Repulsive pressure has been measured as a function of lattice spacing in gels of tobacco mosaic virus (TMV) and in the filament lattice of vertebrate striated muscle. External pressures up to ten atm have been applied to these lattices by an osmotic stress method. Numerical solutions to the Poisson-Boltzmann equation in hexagonal lattices have been obtained and compared to the TMV and muscle data. The theoretical curves using values for kappa calculated from the ionic strength give a good fit to experimental data from TMV gels, and an approximate fit to that from the muscle lattice, provided that a charge radius for the muscle thick filaments of approx. 16 nm is assumed. Variations in ionic strength, sarcomere length and state of the muscle give results which agree qualitatively with the theory, though a good fit between experiment and theory in the muscle case will clearly require consideration of other types of forces. We conclude that Poisson-Boltzmann theory can provide a good first approximation to the long-range electrostatic forces operating in such biological gel systems.

  8. Muscle RING‐finger 2 and 3 maintain striated‐muscle structure and function

    PubMed Central

    Lodka, Dörte; Pahuja, Aanchal; Geers‐Knörr, Cornelia; Scheibe, Renate J.; Nowak, Marcel; Hamati, Jida; Köhncke, Clemens; Purfürst, Bettina; Kanashova, Tamara; Schmidt, Sibylle; Glass, David J.; Morano, Ingo; Heuser, Arnd; Kraft, Theresia; Bassel‐Duby, Rhonda; Olson, Eric N.; Dittmar, Gunnar; Sommer, Thomas

    2015-01-01

    Abstract Background The Muscle‐specific RING‐finger (MuRF) protein family of E3 ubiquitin ligases is important for maintenance of muscular structure and function. MuRF proteins mediate adaptation of striated muscles to stress. MuRF2 and MuRF3 bind to microtubules and are implicated in sarcomere formation with noticeable functional redundancy. However, if this redundancy is important for muscle function in vivo is unknown. Our objective was to investigate cooperative function of MuRF2 and MuRF3 in the skeletal muscle and the heart in vivo. Methods MuRF2 and MuRF3 double knockout mice (DKO) were generated and phenotypically characterized. Skeletal muscle and the heart were investigated by morphological measurements, histological analyses, electron microscopy, immunoblotting, and real‐time PCR. Isolated muscles were subjected to in vitro force measurements. Cardiac function was determined by echocardiography and working heart preparations. Function of cardiomyocytes was measured in vitro. Cell culture experiments and mass‐spectrometry were used for mechanistic analyses. Results DKO mice showed a protein aggregate myopathy in skeletal muscle. Maximal force development was reduced in DKO soleus and extensor digitorum longus. Additionally, a fibre type shift towards slow/type I fibres occurred in DKO soleus and extensor digitorum longus. MuRF2 and MuRF3‐deficient hearts showed decreased systolic and diastolic function. Further analyses revealed an increased expression of the myosin heavy chain isoform beta/slow and disturbed calcium handling as potential causes for the phenotype in DKO hearts. Conclusions The redundant function of MuRF2 and MuRF3 is important for maintenance of skeletal muscle and cardiac structure and function in vivo. PMID:27493870

  9. The effect of quinine on tension development, membrane potentials and excitation-contraction coupling of crab skeletal muscle fibres

    PubMed Central

    Huddart, H.

    1971-01-01

    1. The effect of quinine on tension development and membrane potentials of crab skeletal muscle was examined using strain gauges and intracellular electrodes. 2. In low concentrations (0·1-0·5 mM), quinine caused transient potentiation of twitch tension which then rapidly declined along with progressive depression of the tetanus. These actions are correlated with the decline of both action and resting potentials during quinine treatment. 3. In moderate concentrations (1-5 mM), quinine induced phasic contractures, but the attendant depolarization made the muscles refractory to stimulation and potassium activation, but not to caffeine activation. 4. Quinine did not induce contractures in depolarized muscle, which suggests that the action of quinine in inducing calcium release from the sarcoplasmic reticulum may be blocked by potassium depolarization, unlike the calcium-releasing action of caffeine. Quinine appeared to have no effect on the mechanical threshold of crab skeletal muscle fibres. 5. To explain its depression of contractility in crab muscle, it is suggested that quinine may deplete the calcium store of the sarcoplasmic reticulum, leading to extinction of the terminal stages of the excitation—contraction coupling process and loss of contractility. PMID:5565642

  10. Disruption of muscle architecture and myocardial degeneration in mice lacking desmin

    PubMed Central

    1996-01-01

    Desmin, the muscle specific intermediate filament (IF) protein encoded by a single gene, is expressed in all muscle tissues. In mature striated muscle, desmin IFs surround the Z-discs, interlink them together and integrate the contractile apparatus with the sarcolemma and the nucleus. To investigate the function of desmin in all three muscle types in vivo, we generated desmin null mice through homologous recombination. Surprisingly, desmin null mice are viable and fertile. However, these mice demonstrated a multisystem disorder involving cardiac, skeletal, and smooth muscle. Histological and electron microscopic analysis in both heart and skeletal muscle tissues revealed severe disruption of muscle architecture and degeneration. Structural abnormalities included loss of lateral alignment of myofibrils and abnormal mitochondrial organization. The consequences of these abnormalities were most severe in the heart, which exhibited progressive degeneration and necrosis of the myocardium accompanied by extensive calcification. Abnormalities of smooth muscle included hypoplasia and degeneration. The present data demonstrate the essential role of desmin in the maintenance of myofibril, myofiber, and whole muscle tissue structural and functional integrity, and show that the absence of desmin leads to muscle degeneration. PMID:8794866

  11. Impaired mitochondrial respiration and decreased fatigue resistance followed by severe muscle weakness in skeletal muscle of mitochondrial DNA mutator mice.

    PubMed

    Yamada, Takashi; Ivarsson, Niklas; Hernández, Andrés; Fahlström, Andreas; Cheng, Arthur J; Zhang, Shi-Jin; Bruton, Joseph D; Ulfhake, Brun; Westerblad, Håkan

    2012-12-01

    Mitochondrial dysfunction can drastically impair muscle function, with weakness and exercise intolerance as key symptoms. Here we examine the time course of development of muscle dysfunction in a mouse model of premature ageing induced by defective proofreading function of mitochondrial DNA (mtDNA) polymerase (mtDNA mutator mouse). Isolated fast-twitch muscles and single muscle fibres from young (3-5 months) and end-stage (11 months) mtDNA mutator mice were compared to age-matched control mice. Force and free myoplasmic [Ca(2+)] ([Ca(2+)](i)) were measured under resting conditions and during fatigue induced by repeated tetani. Muscles of young mtDNA mutator mice displayed no weakness in the rested state, but had lower force and [Ca(2+)](i) than control mice during induction of fatigue. Muscles of young mtDNA mutator mice showed decreased activities of citrate synthase and β-hydroxyacyl-coenzyme A dehydrogenase, reduced expression of cytochrome c oxidase, and decreased expression of triggers of mitochondrial biogenesis (PGC-1α, PPARα, AMPK). Muscles from end-stage mtDNA mutator mice showed weakness under resting conditions with markedly decreased tetanic [Ca(2+)](i), force per cross-sectional area and protein expression of the sarcoplasmic reticulum Ca(2+) pump (SERCA1). In conclusion, fast-twitch muscles of prematurely ageing mtDNA mutator mice display a sequence of deleterious mitochondrial-to-nucleus signalling with an initial decrease in oxidative capacity, which was not counteracted by activation of signalling to increase mitochondrial biogenesis. This was followed by severe muscle weakness in the end stage. These results have implication for normal ageing and suggest that decreased mitochondrial oxidative capacity due to a sedentary lifestyle may predispose towards muscle weakness developing later in life.

  12. Ca2+ scraps: local depletions of free [Ca2+] in cardiac sarcoplasmic reticulum during contractions leave substantial Ca2+ reserve.

    PubMed

    Shannon, Thomas R; Guo, Tao; Bers, Donald M

    2003-07-11

    Free [Ca2+] inside the sarcoplasmic reticulum ([Ca2+]SR) is difficult to measure yet critically important in controlling many cellular systems. In cardiac myocytes, [Ca2+]SR regulates cardiac contractility. We directly measure [Ca2+]SR in intact cardiac myocytes dynamically and quantitatively during beats, with high spatial resolution. Diastolic [Ca2+]SR (1 to 1.5 mmol/L) is only partially depleted (24% to 63%) during contraction. There is little temporal delay in the decline in [Ca2+]SR at release junctions and between junctions, indicating rapid internal diffusion. The incomplete local Ca2+ release shows that the inherently positive feedback of Ca2+-induced Ca2+ release terminates, despite a large residual driving force. These findings place stringent novel constraints on how excitation-contraction coupling works in heart and also reveal a Ca2+ store reserve that could in principle be a therapeutic target to enhance cardiac function in heart failure.

  13. Expression profiles of muscle disease-associated genes and their isoforms during differentiation of cultured human skeletal muscle cells

    PubMed Central

    2012-01-01

    Background The formation of contractile myofibrils requires the stepwise onset of expression of muscle specific proteins. It is likely that elucidation of the expression patterns of muscle-specific sarcomeric proteins is important to understand muscle disorders originating from defects in contractile sarcomeric proteins. Methods We investigated the expression profile of a panel of sarcomeric components with a focus on proteins associated with a group of congenital disorders. The analyses were performed in cultured human skeletal muscle cells during myoblast proliferation and myotube development. Results Our culture technique resulted in the development of striated myotubes and the expression of adult isoforms of the sarcomeric proteins, such as fast TnI, fast TnT, adult fast and slow MyHC isoforms and predominantly skeletal muscle rather than cardiac actin. Many proteins involved in muscle diseases, such as beta tropomyosin, slow TnI, slow MyBPC and cardiac TnI were readily detected in the initial stages of muscle cell differentiation, suggesting the possibility of an early role for these proteins as constituent of the developing contractile apparatus during myofibrillogenesis. This suggests that in disease conditions the mechanisms of pathogenesis for each of the mutated sarcomeric proteins might be reflected by altered expression patterns, and disturbed assembly of cytoskeletal, myofibrillar structures and muscle development. Conclusions In conclusion, we here confirm that cell cultures of human skeletal muscle are an appropriate tool to study developmental stages of myofibrillogenesis. The expression of several disease-associated proteins indicates that they might be a useful model system for studying the pathogenesis of muscle diseases caused by defects in specific sarcomeric constituents. PMID:23273262

  14. Ultrastructural alterations in skeletal muscle of pigs with acute monensin myotoxicosis.

    PubMed Central

    Van Vleet, J. F.; Ferrans, V. J.

    1984-01-01

    Large doses of monensin, a Na+-selective carboxylic ionophore, produce polyfocal, monophasic necrosis of skeletal muscle, with Type I fiber selectivity, in swine. For a study of the sequential ultrastructural alterations in affected skeletal muscles, 14 weanling pigs were given 40 mg monensin/kg body weight and were euthanatized 1, 2, 4, 8, and 16 days later. Myotoxicosis and myoglobinuria were apparent clinically. At necropsy, white, dry areas of necrosis were present in the muscle masses of the anterior and posterior thigh, shoulder, and loin. Two patterns of skeletal muscle necrosis were observed on Day 1, especially in Type I fibers. In fibers exhibiting the first of these patterns, the contractile material was disrupted, forming dense amorphous and filamentous clumps scattered within the persistent sheaths of external lamina (sarcolemmal tubes); the mitochondria were swollen and contained flocculent matrix densities, and the nuclei were pyknotic. Fibers showing the second pattern were uniformly dense, but their sarcoplasm was not disrupted. Sublethally injured fibers were also observed and showed focal myofibrillar lysis. On Days 2 and 4, the necrotic muscle had marked infiltration of macrophages in the interstitium and within sarcolemmal tubes. Rapid resolution of the fiber necrosis occurred by phagocytosis of the sarcoplasmic debris. Regeneration of affected muscles developed early following injury and progressed rapidly to complete restoration of the necrotic muscles without residual fibrosis. Regeneration was initiated on Day 1 by activation of satellite cells to form presumptive myoblasts; on Days 4 and 8 these cells showed evidence of fusion, forming myotubes to restore the necrotic fibers. Images Figure 1 Figure 3 Figure 4 Figure 5 Figure 6 Figure 7 Figure 8 Figure 9 Figure 10 Figure 11 PMID:6696050

  15. Geologic continuous casting below continental and deep-sea detachment faults and at the striated extrusion of Sacsayhuaman, Peru

    USGS Publications Warehouse

    Spencer, J.E.

    1999-01-01

    In the common type of industrial continuous casting, partially molten metal is extruded from a vessel through a shaped orifice called a mold in which the metal assumes the cross-sectional form of the mold as it cools and solidifies. Continuous casting can be sustained as long as molten metal is supplied and thermal conditions are maintained. I propose that a similar process produced parallel sets of grooves in three geologic settings, as follows: (1) corrugated metamorphic core complexes where mylonized mid-crustal rocks were exhumed by movement along low-angle normal faults known as detachment faults; (2) corrugated submarine surfaces where ultramafic and mafic rocks were exhumed by normal faulting within oceanic spreading centers; and (3) striated magma extrusions exemplified by the famous grooved outcrops at the Inca fortress of Sacsayhuaman in Peru. In each case, rocks inferred to have overlain the corrugated surface during corrugation genesis molded and shaped a plastic to partially molten rock mass as it was extruded from a moderate- to high-temperature reservoir.

  16. Acting Out Muscle Contraction.

    ERIC Educational Resources Information Center

    Hudson, Margaret

    2003-01-01

    Describes a science activity that can be implemented into anatomy and physiology courses that demonstrates the interactions between action and myosin, the roles of sodium and calcium ions in the regulation of contraction, and the functions of the plasma membrane and the sarcoplasmic reticulum. (YDS)

  17. Muscle fibers in the central nervous system of nemerteans: spatial organization and functional role.

    PubMed

    Petrov, A A; Zaitseva, O V

    2012-08-01

    The system of muscle fibers associated with the brain and lateral nerve cords is present in all major groups of enoplan nemerteans. Unfortunately, very little is known about the functional role and spatial arrangement of these muscles of the central nervous system. This article examines the architecture of the musculature of the central nervous system in two species of monostiliferous nemerteans (Emplectonema gracile and Tetrastemma cf. candidum) using phalloidin staining and confocal microscopy. The article also briefly discusses the body-wall musculature and the muscles of the cephalic region. In both species, the lateral nerve cords possess two pairs of cardinal muscles that run the length of the nerve cords and pass through the ventral cerebral ganglia. A system of peripheral muscles forms a meshwork around the lateral nerve cords in E. gracile. The actin-rich processes that ramify within the nerve cords in E. gracile (transverse fibers) might represent a separate population of glia-like cells or sarcoplasmic projections of the peripheral muscles of the central nervous system. The lateral nerve cords in T. cf. candidum lack peripheral muscles but have muscles similar in their position and orientation to the transverse fibers. The musculature of the central nervous system is hypothesized to function as a support system for the lateral nerve cords and brain, preventing rupturing and herniation of the nervous tissue during locomotion. The occurrence of muscles of the central nervous system in nemerteans and other groups and their possible relevance in taxonomy are discussed.

  18. Ultrastructural features of degenerated cardiac muscle cells in patients with cardiac hypertrophy.

    PubMed Central

    Maron, B. J.; Ferrans, V. J.; Roberts, W. C.

    1975-01-01

    Degenerated cardiac muscle cells were present in hypertrophied ventricular muscle obtained at operation from 12 (38%) of 32 patients with asymmetric septal hypertrophy (hypertrophic cardiomyopathy) or aortic valvular disease. Degenerated cells demonstrated a wide variety of ultrastructural alterations. Mildly altered cells were normal-sized or hypertrophied and showed focal changes, including preferential loss of thick (myosin) filaments, streaming and clumping of Z band material, and proliferation of the tubules of sarcoplasmic reticulum. Moderately and severely degenerated cells were normal-sized or atrophic and showed additional changes, including extensive myofibrillar lysis and loss of T tubules. The appearance of the most severely degenerated cells usually reflected the cytoplasmic organelle (sarcoplasmic reticulum, glycogen, or mitochondria) which underwent proliferation and filled the myofibril-free areas of these cells. Moderately and severely degenerated cells were present in areas of fibrosis, had thickened basement membranes, and had lost their intercellular connections. These observations suggest that degenerated cardiac muscle cells have poor contractile function and may be responsible for impaired cardiac performance in some patients with chronic ventricular hypertrophy. Images Fig 1 Fig 2 Fig 3 Figs 4-6 Figs 7-8 Fig 9 Fig 10 Fig 11 Figs 12-15 Fig 16 Fig 17 Figs 18-21 Figs 22-23 Fig 24 Fig 25 Fig 26 Fig 27 Figs 28-29 Fig 30 Figs 31-32 Fig 33 PMID:124533

  19. Resonant Reflection Spectroscopy of Biomolecular Arrays in Muscle

    PubMed Central

    Young, Kevin W.; Radic, Stojan; Myslivets, Evgeny; O’Connor, Shawn M.; Lieber, Richard L.

    2014-01-01

    Sarcomeres, the functional units of contraction in striated muscle, are composed of an array of interdigitating protein filaments. Direct interaction between overlapping filaments generates muscular force, which produces animal movement. When filament length is known, sarcomere length successfully predicts potential force, even in whole muscles that contain billions of sarcomere units. Inability to perform in vivo sarcomere measurements with submicrometer resolution is a long-standing challenge in the muscle physiology field and has hampered studies of normal muscle function, adaptation, injury, aging, and disease, particularly in humans. Here, we develop theory and demonstrate the feasibility of to our knowledge a new technique that measures sarcomere length with submicrometer resolution. In this believed novel approach, we examine sarcomere structure by measuring the multiple resonant reflections that are uniquely defined by Fourier decomposition of the sarcomere protein spatial framework. Using a new supercontinuum spectroscopic system, we show close agreement between sarcomere lengths measured by resonant reflection spectroscopy and laser diffraction in an ensemble of 10 distinct muscles. PMID:25418304

  20. Biohybrid Thin Films for Measuring Contractility in Engineered Cardiovascular Muscle

    PubMed Central

    Alford, Patrick W.; Feinberg, Adam W.; Sheehy, Sean P.; Parker, Kevin Kit

    2010-01-01

    In vitro cardiovascular disease models need to recapitulate tissue-scale function in order to provide in vivo relevance. We have developed a new method for measuring the contractility of engineered cardiovascular smooth and striated muscle in vitro during electrical and pharmacological stimulation. We present a growth theory-based finite elasticity analysis for calculating the contractile stresses of a 2D anisotropic muscle tissue cultured on a flexible synthetic polymer thin film. Cardiac muscle engineered with neonatal rat ventricular myocytes and paced at 0.5 Hz generated stresses of 9.2 ± 3.5 kPa at peak systole, similar to measurements of the contractility of papillary muscle from adult rats. Vascular tissue engineered with human umbilical arterial smooth muscle cells maintained a basal contractile tone of 13.1 ± 2.1 kPa and generated another 5.1 ± 0.8 kPa when stimulated with endothelin-1. These data suggest that this method may be useful in assessing the efficacy and safety of pharmacological agents on cardiovascular tissue. PMID:20149449

  1. To understand muscle you must take it apart

    PubMed Central

    Batters, Christopher; Veigel, Claudia; Homsher, Earl; Sellers, James R.

    2014-01-01

    Striated muscle is an elegant system for study at many levels. Much has been learned about the mechanism of contraction from studying the mechanical properties of intact and permeabilized (or skinned) muscle fibers. Structural studies using electron microscopy, X-ray diffraction or spectroscopic probes attached to various contractile proteins were possible because of the highly ordered sarcomeric arrangement of actin and myosin. However, to understand the mechanism of force generation at a molecular level, it is necessary to take the system apart and study the interaction of myosin with actin using in vitro assays. This reductionist approach has lead to many fundamental insights into how myosin powers muscle contraction. In addition, nature has provided scientists with an array of muscles with different mechanical properties and with a superfamily of myosin molecules. Taking advantage of this diversity in myosin structure and function has lead to additional insights into common properties of force generation. This review will highlight the development of the major assays and methods that have allowed this combined reductionist and comparative approach to be so fruitful. This review highlights the history of biochemical and biophysical studies of myosin and demonstrates how a broad comparative approach combined with reductionist studies have led to a detailed understanding of how myosin interacts with actin and uses chemical energy to generate force and movement in muscle contraction and motility in general. PMID:24653704

  2. Altered cross-bridge properties in skeletal muscle dystrophies

    PubMed Central

    Guellich, Aziz; Negroni, Elisa; Decostre, Valérie; Demoule, Alexandre; Coirault, Catherine

    2014-01-01

    Force and motion generated by skeletal muscle ultimately depends on the cyclical interaction of actin with myosin. This mechanical process is regulated by intracellular Ca2+ through the thin filament-associated regulatory proteins i.e.; troponins and tropomyosin. Muscular dystrophies are a group of heterogeneous genetic affections characterized by progressive degeneration and weakness of the skeletal muscle as a consequence of loss of muscle tissue which directly reduces the number of potential myosin cross-bridges involved in force production. Mutations in genes responsible for skeletal muscle dystrophies (MDs) have been shown to modify the function of contractile proteins and cross-bridge interactions. Altered gene expression or RNA splicing or post-translational modifications of contractile proteins such as those related to oxidative stress, may affect cross-bridge function by modifying key proteins of the excitation-contraction coupling. Micro-architectural change in myofilament is another mechanism of altered cross-bridge performance. In this review, we provide an overview about changes in cross-bridge performance in skeletal MDs and discuss their ultimate impacts on striated muscle function. PMID:25352808

  3. Interactions between beta-enolase and creatine kinase in the cytosol of skeletal muscle cells.

    PubMed Central

    Foucault, G; Vacher, M; Cribier, S; Arrio-Dupont, M

    2000-01-01

    We studied interactions in vivo between the cytosolic muscle isoform of creatine kinase (M-CK) and the muscle isoform of 2-phospho-D-glycerate hydrolyase (beta-enolase) in muscle sarcoplasm by incubating glycerol-skinned fibres with FITC-labelled beta-enolase in the presence or absence of free CK. A small amount of bound beta-enolase was observed in the presence of large concentrations of CK. The mobility of enolase was measured in cultured satellite cells by modulated-fringe-pattern photobleaching. FITC-labelled beta-enolase was totally mobile in both the presence and the absence of CK but its diffusion coefficient was slightly lower in the presence of CK. This suggests a weak interaction in vivo between enolase and CK. PMID:10657248

  4. Titin stiffness modifies the force-generating region of muscle sarcomeres

    PubMed Central

    Li, Yong; Lang, Patrick; Linke, Wolfgang A.

    2016-01-01

    The contractile units of striated muscle, the sarcomeres, comprise the thick (myosin) and thin (actin) filaments mediating active contraction and the titin filaments determining “passive” elasticity. We hypothesized that titin may be more active in muscle contraction by directly modulating thick-filament properties. We used single-myofibril mechanical measurements and atomic force microscopy of individual sarcomeres to quantify the effects of sarcomere strain and titin spring length on both the inter-filament lattice spacing and the lateral stiffness of the actin-myosin overlap zone (A-band). We found that strain reduced the lattice spacing similarly in sarcomeres with stiff (rabbit psoas) or compliant titin (rabbit diaphragm), but increased A-band lateral stiffness much more in psoas than in diaphragm. The strain-induced alterations in A-band stiffness that occur independently of lattice spacing effects may be due to titin stiffness-sensing by A-band proteins. This mechanosensitivity could play a role in the physiologically important phenomenon of length-dependent activation of striated muscle. PMID:27079135

  5. Chemotherapy-Induced Weakness and Fatigue in Skeletal Muscle: The Role of Oxidative Stress

    PubMed Central

    St. Clair, Daret K.

    2011-01-01

    Abstract Significance Fatigue is one of the most common symptoms of cancer and its treatment, manifested in the clinic through weakness and exercise intolerance. These side effects not only compromise patient's quality of life (QOL), but also diminish physical activity, resulting in limited treatment and increased morbidity. Recent Advances Oxidative stress, mediated by cancer or chemotherapeutic agents, is an underlying mechanism of the drug-induced toxicity. Nontargeted tissues, such as striated muscle, are severely affected by oxidative stress during chemotherapy, leading to toxicity and dysfunction. Critical Issues These findings highlight the importance of investigating clinically applicable interventions to alleviate the debilitating side effects. This article discusses the clinically available chemotherapy drugs that cause fatigue and oxidative stress in cancer patients, with an in-depth focus on the anthracycline doxorubicin. Doxorubicin, an effective anticancer drug, is a primary example of how chemotherapeutic agents disrupt striated muscle function through oxidative stress. Future Directions Further research investigating antioxidants could provide relief for cancer patients from debilitating muscle weakness, leading to improved quality of life. Antioxid. Redox Signal. 15, 2543–2563. PMID:21457105

  6. Invertebrate muscles: thin and thick filament structure; molecular basis of contraction and its regulation, catch and asynchronous muscle

    PubMed Central

    Hooper, Scott L.; Hobbs, Kevin H.; Thuma, Jeffrey B.

    2008-01-01

    This is the second in a series of canonical reviews on invertebrate muscle. We cover here thin and thick filament structure, the molecular basis of force generation and its regulation, and two special properties of some invertebrate muscle, catch and asynchronous muscle. Invertebrate thin filaments resemble vertebrate thin filaments, although helix structure and tropomyosin arrangement show small differences. Invertebrate thick filaments, alternatively, are very different from vertebrate striated thick filaments and show great variation within invertebrates. Part of this diversity stems from variation in paramyosin content, which is greatly increased in very large diameter invertebrate thick filaments. Other of it arises from relatively small changes in filament backbone structure, which results in filaments with grossly similar myosin head placements (rotating crowns of heads every 14.5 nm) but large changes in detail (distances between heads in azimuthal registration varying from three to thousands of crowns). The lever arm basis of force generation is common to both vetebrates and invertebrates, and in some invertebrates this process is understood on the near atomic level. Invertebrate actomyosin is both thin (tropomyosin:troponin) and thick (primarily via direct Ca++ binding to myosin) filament regulated, and most invertebrate muscles are dually regulated. These mechanisms are well understood on the molecular level, but the behavioral utility of dual regulation is less so. The phosphorylation state of the thick filament associated giant protein, twitchin, has been recently shown to be the molecular basis of catch. The molecular basis of the stretch activation underlying asynchronous muscle activity, however, remains unresolved. PMID:18616971

  7. Differentiation of Sparidae species by DNA sequence analysis, PCR-SSCP and IEF of sarcoplasmic proteins.

    PubMed

    Schiefenhövel, Karin; Rehbein, Hartmut

    2013-05-01

    The increasing global trade of fishery and aquaculture products makes it necessary to develop methods for species identification in case of fish fillets or other highly processed seafood with external morphological characteristics (e.g. gills, fins) of the original fish being removed. Species identification methods based on DNA- or protein-analysis, like DNA sequencing of PCR products, PCR-SSCP (polymerase chain reaction - single strand conformation polymorphism), and isoelectric focusing (IEF) of water-soluble proteins of fish fillet were applied successfully to distinguish between various sparid species including Sparus aurata, Spondyliosoma cantharus, Boops boops, Acanthopagrus bifasciatus, Argyrops spinifer, Pagellus bogaraveo and Lithognathus mormyrus of the family Sparidae available on European markets. Indication of the presence of parvalbumins in light muscle of all of the studied species is presented by the location of heat-stable proteins in the anodic range of the IEF gels.

  8. The T-tubular network and its triads in the sole plate sarcoplasm of the motor end-plate of mammals.

    PubMed

    Dauber, W; Voigt, T; Härtel, X; Mayer, J

    2000-01-01

    The transition between the subsynaptic folds and the T-tubules demonstrated in a former paper was further investigated in the sole plate area by using the extracellular marker Lanthanum. A tubular network of the T-system of the sole plate area which is connected to the subsynaptic folds and to the t-tubular elements between the myofibrils is described for the first time. T-tubules of this network criss-cross through the sarcoplasm of the sole plate and lie in close proximity to sole plate nuclei and mitochondria. Cisterns of sarcoplasmic reticulum of the sole plate area make contact with these t-tubules forming triads. The possible physiological role of this sole plate network and its triads will be discussed with regard to a transport of substances in tubules with the dimension of nanotubes and Ca2+ activated processes in the sole plate area.

  9. An Altered Phenotype in a Conditional Knockout of Pitx2 in Extraocular Muscle

    PubMed Central

    Zhou, Yuefang; Cheng, Georgiana; Dieter, Lisa; Hjalt, Tord A.; Andrade, Francisco H.; Stahl, John S.; Kaminski, Henry J.

    2015-01-01

    Purpose To determine the temporal and spatial expression of Pitx2, a bicoid-like homeobox transcription factor, during postnatal development of mouse extraocular muscle and to evaluate its role in the growth and phenotypic maintenance of postnatal extraocular muscle. Methods Mouse extraocular muscles of different ages were examined for the expression of Pitx2 by RT-PCR, q-PCR, and immunostaining. A conditional mutant mouse strain, in which Pitx2 function is inactivated at postnatal day (P)0, was generated with a Cre-loxP strategy. Histology, immunostaining, realtime PCR, in vitro muscle contractility, and in vivo ocular motility were used to study the effect of Pitx2 depletion on extraocular muscle. Results All three Pitx2 isoforms were expressed by extraocular muscle and at higher levels than in other striated muscles. Immunostaining demonstrated the presence of Pitx2 mainly in extraocular muscle myonuclei. However, no obvious expression patterns were observed in terms of anatomic region (orbital versus global layer), innervation zone, or muscle fiber types. The mutant extraocular muscle had no obvious pathology but had altered muscle fiber sizes. Expression levels of myosin isoforms Myh1, Myh6, Myh7, and Myh13 were reduced, whereas Myh2, Myh3, Myh4, and Myh8 were not affected by postnatal loss of Pitx2. In vitro, Pitx2 loss made the extraocular muscles stronger, faster, and more fatigable. Eye movement recordings found saccades to have a lower peak velocity. Conclusions Pitx2 is important in maintaining the mature extraocular muscle phenotype and regulating the expression of critical contractile proteins. Modulation of Pitx2 expression can influence extraocular muscle function with long-term therapeutic implications. PMID:19407022

  10. Comparative transcriptome profiling of longissimus muscle tissues from Qianhua Mutton Merino and Small Tail Han sheep.

    PubMed

    Sun, Limin; Bai, Man; Xiang, Lujie; Zhang, Guishan; Ma, Wei; Jiang, Huaizhi

    2016-01-01

    The Qianhua Mutton Merino (QHMM) is a new sheep (Ovis aries) variety with better meat performance compared with the traditional local variety Small Tail Han (STH) sheep. We aimed to evaluate the transcriptome regulators associated with muscle growth and development between the QHMM and STH. We used RNA-Seq to obtain the transcriptome profiles of the longissimus muscle from the QHMM and STH. The results showed that 960 genes were differentially expressed (405 were up-regulated and 555 were down-regulated). Among these, 463 differently expressed genes (DEGs) were probably associated with muscle growth and development and were involved in biological processes such as skeletal muscle tissue development and muscle cell differentiation; molecular functions such as catalytic activity and oxidoreductase activity; cellular components such as mitochondrion and sarcoplasmic reticulum; and pathways such as metabolic pathways and citrate cycle. From the potential genes, a gene-act-network and co-expression-network closely related to muscle growth and development were identified and established. Finally, the expressions of nine genes were validated by real-time PCR. The results suggested that some DEGs, including MRFs, GXP1 and STAC3, play crucial roles in muscle growth and development processes. This genome-wide transcriptome analysis of QHMM and STH muscle is reported for the first time. PMID:27645777

  11. Comparative transcriptome profiling of longissimus muscle tissues from Qianhua Mutton Merino and Small Tail Han sheep

    PubMed Central

    Sun, Limin; Bai, Man; Xiang, Lujie; Zhang, Guishan; Ma, Wei; Jiang, Huaizhi

    2016-01-01

    The Qianhua Mutton Merino (QHMM) is a new sheep (Ovis aries) variety with better meat performance compared with the traditional local variety Small Tail Han (STH) sheep. We aimed to evaluate the transcriptome regulators associated with muscle growth and development between the QHMM and STH. We used RNA-Seq to obtain the transcriptome profiles of the longissimus muscle from the QHMM and STH. The results showed that 960 genes were differentially expressed (405 were up-regulated and 555 were down-regulated). Among these, 463 differently expressed genes (DEGs) were probably associated with muscle growth and development and were involved in biological processes such as skeletal muscle tissue development and muscle cell differentiation; molecular functions such as catalytic activity and oxidoreductase activity; cellular components such as mitochondrion and sarcoplasmic reticulum; and pathways such as metabolic pathways and citrate cycle. From the potential genes, a gene-act-network and co-expression-network closely related to muscle growth and development were identified and established. Finally, the expressions of nine genes were validated by real-time PCR. The results suggested that some DEGs, including MRFs, GXP1 and STAC3, play crucial roles in muscle growth and development processes. This genome-wide transcriptome analysis of QHMM and STH muscle is reported for the first time. PMID:27645777

  12. Evidence that lipid lateral phase separation induces functionally significant structural changes in the Ca+2ATPase of the sarcoplasmic reticulum.

    PubMed Central

    Asturias, F J; Pascolini, D; Blasie, J K

    1990-01-01

    We have studied lipid lateral phase separation (LPS) in the intact sarcoplasmic reticulum (SR) membrane and in bilayers of isolated SR membrane lipids as a function of temperature, [Mg+2], and degree of hydration. Lipid LPS was observed in both the intact membrane and in the bilayers of isolated SR lipids, and the LPS behavior of both systems was found to be qualitatively similar. Namely, lipid LPS occurs only at relatively low temperature and water content, independently of the [Mg+2], and the upper characteristic temperature (th) for lipid LPS for both the membrane and bilayers of its isolated lipids coincide to within a few degrees. However, at similar temperatures, isolated lipids show more LPS than the lipids in the intact membrane. Lipid LPS in the intact membrane and in bilayers of the isolated lipids is fully reversible, and more extensive for samples partially dehydrated at temperatures below th. Our previous x-ray diffraction studies established the existence of a temperature-induced transition in the profile structure of the sarcoplasmic reticulum Ca+2ATPase which occurs at a temperature corresponding to the [Mg+2]-dependent upper characteristic temperature for lipid LPS in the SR membrane. Furthermore, the functionality of the ATPase, and in particular the lifetime of the first phosphorylated enzyme conformation (E1 approximately P) in the Ca+2 transport cycle, were also found to be linked to the occurrence of this structural transition. The hysterisis observed in lipid LPS behavior as a function of temperature and water content provides a possible explanation for the more efficient transient trapping of the enzyme in the E1 approximately P conformation observed in SR membranes partially dehydrated at temperatures below th. The observation that LPS behavior for the intact SR membrane and bilayers of isolated SR lipids (no protein present) are qualitatively similar strongly suggests that the LPS behavior of the SR membrane lipids is responsible for the

  13. Osmosensation in TRPV2 dominant negative expressing skeletal muscle fibres

    PubMed Central

    Zanou, Nadège; Mondin, Ludivine; Fuster, Clarisse; Seghers, François; Dufour, Inès; de Clippele, Marie; Schakman, Olivier; Tajeddine, Nicolas; Iwata, Yuko; Wakabayashi, Shigeo; Voets, Thomas; Allard, Bruno; Gailly, Philippe

    2015-01-01

    Abstract Increased plasma osmolarity induces intracellular water depletion and cell shrinkage followed by activation of a regulatory volume increase (RVI). In skeletal muscle, this is accompanied by transverse tubule (TT) dilatation and by a membrane depolarization responsible for a release of Ca2+ from intracellular pools. We observed that both hyperosmotic shock-induced Ca2+ transients and RVI were inhibited by Gd3+, ruthenium red and GsMTx4 toxin, three inhibitors of mechanosensitive ion channels. The response was also completely absent in muscle fibres overexpressing a non-permeant, dominant negative (DN) mutant of the transient receptor potential, V2 isoform (TRPV2) ion channel, suggesting the involvement of TRPV2 or of a TRP isoform susceptible to heterotetramerization with TRPV2. The release of Ca2+ induced by hyperosmotic shock was increased by cannabidiol, an activator of TRPV2, and decreased by tranilast, an inhibitor of TRPV2, suggesting a role for the TRPV2 channel itself. Hyperosmotic shock-induced membrane depolarization was impaired in TRPV2-DN fibres, suggesting that TRPV2 activation triggers the release of Ca2+ from the sarcoplasmic reticulum by depolarizing TTs. RVI requires the sequential activation of STE20/SPS1-related proline/alanine-rich kinase (SPAK) and NKCC1, a Na+–K+–Cl− cotransporter, allowing ion entry and driving osmotic water flow. In fibres overexpressing TRPV2-DN as well as in fibres in which Ca2+ transients were abolished by the Ca2+ chelator BAPTA, the level of P-SPAKSer373 in response to hyperosmotic shock was reduced, suggesting a modulation of SPAK phosphorylation by intracellular Ca2+. We conclude that TRPV2 is involved in osmosensation in skeletal muscle fibres, acting in concert with P-SPAK-activated NKCC1. Key points Increased plasma osmolarity induces intracellular water depletion and cell shrinkage (CS) followed by activation of a regulatory volume increase (RVI). In skeletal muscle, the hyperosmotic shock

  14. Spaceflight effects on adult rat muscle protein, nucleic acids, and amino acids

    NASA Technical Reports Server (NTRS)

    Steffen, J. M.; Musacchia, X. J.

    1986-01-01

    Exposure to conditions of weightlessness has been associated with decrements in muscle mass and strength. The present studies were undertaken to determine muscle responses at the cellular level. Male Sprague-Dawley rats (360-410 g) were exposed to 7 days of weightlessness during the Spacelab-3 shuttle flight (May 1985). Animals were killed 12 h postflight, and soleus (S), gastrocnemius (G), and extensor digitorum longus (EDL) muscles were excised. Muscle protein, RNA, and DNA were extracted and quantified. Differential muscle atrophy was accompanied by a significant (P less than 0.05) reduction in total protein only in S muscles. There were no significant changes in protein concentration (mg/g) in the muscles examined. In S muscles from flight animals, sarcoplasmic protein accounted for a significantly greater proportion of total protein that in ground controls (37.5 vs. 32.5%). Tissue concentrations (nmol/g) of asparagine-aspartate, glutamine-glutamate, glycine, histidine, and lysine were significantly reduced (from 17 to 63%) in S muscles from flight animals, but only glutamine-glutamate levels were decreased in the G and EDL. Muscle DNA content (microgram) was unchanged in the tissues examined, but S muscle DNA concentration (micrograms/mg) increased 27%. RNA content (micrograms) was significantly (P less than 0.025) reduced in S (-28%) and G(-22%) muscles following spaceflight. These results identify specific alterations in rat skeletal muscle during short term (7-day) exposure to weightlessness and compare favorably with observations previously obtained from ground-based suspension simulations.

  15. Proto-oncogene expression during terminal differentiation of cardiac and skeletal muscle

    SciTech Connect

    Claycomb, W.C.; Lanson, N.A. Jr.; Springhorn, J.P.

    1986-05-01

    The authors have examined the expression of 17 different protooncogenes in proliferating and terminally differentiating cardiac and skeletal muscle cells. Cardiac muscle cells at various periods during differentiation were obtained from the rat. The L6 skeletal muscle cell line and a primary culture of human skeletal muscle satellite cells were the source of skeletal muscle cells. Total cellular RNA was isolated by the quanidinium procedure and purified by CsCl. RNA was separated on 1.2% agarose-formaldehyde gels and blotted onto Zeta-Probe nylon membranes. DNA probes, labeled with /sup 32/P, were generated by nick translation of purified DNA fragments or recombinant plasmid DNA. Northern blots were hybridized with /sup 32/P-DNA in 50% formamide, 1 mM EDTA, 7% SDS, 0.5 M NaHPO/sub 4/, 0.5 mg/ml denatured herring testes DNA and washed in 1 mM EDTA, 40 mM NaHPO/sub 4/ and 5% SDS. As positive controls, to assess DNA synthesis and cell proliferation, human histone H/sub 4/ and thymidine kinase were used as probes; rat cardiac muscle myosin heavy chain and M creatine kinase served to assess muscle cell differentiation. Results of these studies indicate that several of these oncogenes may be involved with the regulation of cell proliferation and terminal cell differentiation in striated muscle.

  16. Transgenic expression and purification of myosin isoforms using the Drosophila melanogaster indirect flight muscle system.

    PubMed

    Caldwell, James T; Melkani, Girish C; Huxford, Tom; Bernstein, Sanford I

    2012-01-01

    Biophysical and structural studies on muscle myosin rely upon milligram quantities of extremely pure material. However, many biologically interesting myosin isoforms are expressed at levels that are too low for direct purification from primary tissues. Efforts aimed at recombinant expression of functional striated muscle myosin isoforms in bacterial or insect cell culture have largely met with failure, although high level expression in muscle cell culture has recently been achieved at significant expense. We report a novel method for the use of strains of the fruit fly Drosophila melanogaster genetically engineered to produce histidine-tagged recombinant muscle myosin isoforms. This method takes advantage of the single muscle myosin heavy chain gene within the Drosophila genome, the high level of expression of accessible myosin in the thoracic indirect flight muscles, the ability to knock out endogenous expression of myosin in this tissue and the relatively low cost of fruit fly colony production and maintenance. We illustrate this method by expressing and purifying a recombinant histidine-tagged variant of embryonic body wall skeletal muscle myosin II from an engineered fly strain. The recombinant protein shows the expected ATPase activity and is of sufficient purity and homogeneity for crystallization. This system may prove useful for the expression and isolation of mutant myosins associated with skeletal muscle diseases and cardiomyopathies for their biochemical and structural characterization.

  17. Effect of hypokinesia on contractile function of cardiac muscle

    NASA Technical Reports Server (NTRS)

    Meyerson, F. Z.; Kapelko, V. I.; Trikhpoyeva, A. M.; Gorina, M. S.

    1980-01-01

    Rats were subjected to hypokinesia for two months and the contractile function of isolated papillary muscle was studied. Hypokinesia reduced significantly the isotonic contraction rate which depended on the ATPase activity of the myofibrils; it also reduced the rate and index of relaxation which depended on the functional capacity of the Ca(++) pump of the sarcoplasmic reticulum. The maximum force of isometric contraction determined by the quantity of actomyosin bridges in the myofibrils did not change after hypokinesia. This complex of changes is contrary to that observed in adaptation to exercise when the rate of isotonic contraction and relaxation increases while the force of isometric contraction does not change. The possible mechanism of this stability of the contractile force during adaptation and readaptation of the heart is discussed.

  18. Giant muscle fibres in pigs with different Ryr1 genotype.

    PubMed

    Fazarinc, G; Candek-Potokar, M; Ursic, M; Vrecl, M; Pogacnik, A

    2002-12-01

    This study examined the frequency, morphological and immunohistochemical characteristics of the giant fibres in the longissimus muscle of local Krsko polje pigs with different Ryr1 genotypes. Giant fibres were round-shaped and had significantly increased cross-sectional area compared with normal muscle fibres. Only fast-twitch glycolytic fibres were affected, usually showing enhanced succinate dehydrogenase activity. On the ultrastructural level, the dilation of the sarcoplasmic reticulum, swelling of mitochondria and destruction of myofilaments was observed. The incidence of giant fibres was the highest in Ryr1 dimutant pigs (Ryr1 nn), which also exhibited lower muscle pH1 than heterozygous (Ryr1 Nn) or pigs with the wild Ryr1 gene (Ryr1 NN). However, the giant fibres were also present in pigs free of Ryr1 gene mutation. Our results suggest that the giant fibre syndrome depends mostly upon the rate and intensity of early post-mortem glycolysis, which results in acidity of muscle tissue. We suppose that the giant fibre formation is a result of excessive intracellular lactate accumulation in some fast-twitch glycolytic fibres. This process could also explain the ultrastructural alterations and the consequent changes in the oxidative enzymes and myofibrillar ATPase staining pattern observed in our and some previous studies.

  19. Excitation—contraction coupling in amphioxus muscle cells

    PubMed Central

    Hagiwara, S.; Henkart, Maryanna P.; Kidokoro, Y.

    1971-01-01

    1. Excitation-contraction coupling was studied in myotomal muscles of amphioxus, Branchiostoma californiense. 2. The action potential of a muscle cell produces a twitch with a rise time of 30-40 msec at 11° C and its Q10 is about 2·2. 3. The twitch increases in amplitude with increasing external Ca concentration and is abolished in Ca-free saline (1 mM-EGTA and 55·7 mM-MgCl2); the twitch amplitude is suppressed by Co or La ions. 4. Caffeine at concentrations above 1 mM in the external saline causes a prolongation of the action potential and a contracture which lasts several minutes. 5. After exposure to caffeine the responsiveness of the muscle to subsequent applications of caffeine recovers in normal saline in 20-30 minutes but not in Ca-free saline. 6. The amplitude of the caffeine contracture is independent of the external Ca concentration and is unaltered after the twitch is eliminated in Ca-free saline. 7. After exposure to caffeine a full-sized twitch can be obtained before the responsiveness to caffeine shows any significant recovery. 8. It is concluded that the twitch is produced by the Ca influx resulting from the increased permeability of the muscle cell membrane to Ca during the action potential and that the Ca mobilized by caffeine is not necessary to the initiation of the twitch. 9. Electronmicroscopy shows the existence of sarcoplasmic reticulum. Imagesabc and dabcd PMID:5158596

  20. The myogenic electric organ of Sternopygus macrurus: a non-contractile tissue with a skeletal muscle transcriptome

    PubMed Central

    Samanta, Manoj P.; Chaidez, Alexander

    2016-01-01

    In most electric fish species, the electric organ (EO) derives from striated muscle cells that suppress many muscle properties. In the gymnotiform Sternopygus macrurus, mature electrocytes, the current-producing cells of the EO, do not contain sarcomeres, yet they continue to make some cytoskeletal and sarcomeric proteins and the muscle transcription factors (MTFs) that induce their expression. In order to more comprehensively examine the transcriptional regulation of genes associated with the formation and maintenance of the contractile sarcomere complex, results from expression analysis using qRT-PCR were informed by deep RNA sequencing of transcriptomes and miRNA compositions of muscle and EO tissues from adult S. macrurus. Our data show that: (1) components associated with the homeostasis of the sarcomere and sarcomere-sarcolemma linkage were transcribed in EO at levels similar to those in muscle; (2) MTF families associated with activation of the skeletal muscle program were not differentially expressed between these tissues; and (3) a set of microRNAs that are implicated in regulation of the muscle phenotype are enriched in EO. These data support the development of a unique and highly specialized non-contractile electrogenic cell that emerges from a striated phenotype and further differentiates with little modification in its transcript composition. This comprehensive analysis of parallel mRNA and miRNA profiles is not only a foundation for functional studies aimed at identifying mechanisms underlying the transcription-independent myogenic program in S. macrurus EO, but also has important implications to many vertebrate cell types that independently activate or suppress specific features of the skeletal muscle program. PMID:27114860

  1. Some distinctive features of zebrafish myogenesis based on unexpected distributions of the muscle cytoskeletal proteins actin, myosin, desmin, alpha-actinin, troponin and titin.

    PubMed

    Costa, Manoel L; Escaleira, Roberta C; Rodrigues, Viviane B; Manasfi, Muhamed; Mermelstein, Claudia S

    2002-08-01

    The current myofibrillogenesis model is based mostly on in vitro cell cultures and on avian and mammalian embryos in situ. We followed the expression of actin, myosin, desmin, alpha-actinin, titin, and troponin using immunofluorescence microscopy of zebrafish (Danio rerio) embryos. We could see young mononucleated myoblasts with sharp striations. The striations were positive for all the sarcomeric proteins. Desmin distribution during muscle maturation changes from dispersed aggregates to a perinuclear concentration to striated afterwards. We could not observe desmin-positive, myofibrillar-proteins-negative cells, and we could not find any non-striated distribution of sarcomeric proteins, such as stress fiber-like structures. Some steps, like fusion before striation, seem to be different in the zebrafish when compared with the previously described myogenesis sequences.

  2. The nuclear receptor NOR-1 regulates the small muscle protein, X-linked (SMPX) and myotube differentiation

    PubMed Central

    Ferrán, Beatriz; Martí-Pàmies, Ingrid; Alonso, Judith; Rodríguez-Calvo, Ricardo; Aguiló, Silvia; Vidal, Francisco; Rodríguez, Cristina; Martínez-González, José

    2016-01-01

    Recent works have highlighted the role of NOR-1 in both smooth and skeletal muscle, and have proposed this nuclear receptor as a nexus that coordinates muscle performance and metabolic capacity. However, no muscle specific genes regulated by NOR-1 have been identified so far. To identify NOR-1 target genes, we over-expressed NOR-1 in human vascular smooth muscle cells (VSMC). These cells subjected to sustained over-expression of supraphysiological levels of NOR-1 experienced marked phenotypic changes and up-regulated the skeletal muscle protein X-linked (SMPX), a protein typically expressed in striated muscle and associated to cell shape. By transcriptional studies and DNA-protein binding assays, we identified a non-consensus NBRE site in human SMPX promoter, critical for NOR-1 responsiveness. The expression of SMPX was higher in human skeletal muscle myoblasts (HSMM) than in human VSMC, and further increased in HSMM differentiated to myotubes. NOR-1 silencing prevented SMPX expression in HSMM, as well as their differentiation to myotubes, but the up-regulation of SMPX was dispensable for HSMM differentiation. Our results indicate that NOR-1 regulate SMPX in human muscle cells and acts as a muscle regulatory factor, but further studies are required to unravel its role in muscle differentiation and hypertrophy. PMID:27181368

  3. Fatty acid composition, sarcoplasmic reticular lipid oxidation, and immunity of hard clam (Meretrix lusoria) fed different dietary microalgae.

    PubMed

    Chen, Shu-Mei; Tseng, Kai-Yi; Huang, Chen-Huei

    2015-07-01

    Fatty acid profiles, activities of biomembrane lipid peroxidation, and immunity of a seawater clam (Meretrix lusoria) fed three species of dietary microalgae were investigated. Clams of a marketable size (25 g mean weight) were fed Tetraselmis chui, Chaetoceros muelleri, or Isochrysis galbana for 8 weeks. Fatty acids, particularly eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA), in the polar lipid fractions of clams reflected those of the dietary algae species. Clams fed with T. chui and C. muelleri contained higher proportion of non-methylene interrupted (NMI) fatty acids than those fed I. galbana. Proportion of DHA in lipids of the clams fed with I. galbana was the highest among test groups. The NADH-dependent sarcoplasmic reticular lipid peroxidation activity of clams fed I. galbana was significantly greater (p < 0.05) than that of clams fed T. chui or C. muelleri. The hemocyte adhesion capacity of clams fed C. muelleri or I. galbana was significantly higher (p < 0.05) than that of clams fed T. chui. No significant differences (p ≥ 0.05) in total hemocyte count, phenoloxidase activity, clearance efficiency hemocyte and phagocytosis were detected among clams fed different microalgae.

  4. A new method of vesicle formation by salting-out and its application to the reconstitution of sarcoplasmic reticulum.

    PubMed

    Taguchi, T; Kasai, M

    1983-04-01

    This paper describes a new method of forming membrane vesicles. It was found that the addition of salt such as KCl into a solution containing lipid (asolectin) and a non-ionic surfactant, Triton X-114, led to the formation of closed membrane vesicles. The vesicles were separated from Triton X-114 by hydrophobic interaction chromatography. Electron microscopy revealed that the mean diameter of the vesicles was 110 nm +/- 69 nm (S.D.). Measurement of osmotic volume change showed that the permeability of the vesicle was very low to salts, sugar (glucose) and amphoteric ion (glycine), but very high to glycerol, ethylene glycol and water. Vesicle formation by this 'salting-out' method is very useful for reconstitution of transport systems in biomembranes because of its advantages: completion within a short time; high yield; and the possibility of utilizing samples in non-ionic surfactant solution. When we applied the method to the reconstitution of sarcoplasmic reticulum, Ca2+-ATPase was incorporated into the reconstituted vesicles and was enzymatically active in the membrane. PMID:6830789

  5. Hydrogen sulfide preconditioning protects against myocardial ischemia/reperfusion injury in rats through inhibition of endo/sarcoplasmic reticulum stress

    PubMed Central

    Li, Changyong; Hu, Min; Wang, Yuan; Lu, Huan; Deng, Jing; Yan, Xiaohong

    2015-01-01

    Ischemia reperfusion (I/R) injury is a major cause of myocardial damage. Hydrogen sulfide (H2S), a gaseous signal molecule, has drawn considerable attention for its role in various pathophysiological processes. Multiple lines of evidence reveal the protective effects of H2S in various models of cardiac injury, however, the exact mechanism underlying this protective effect of H2S against myocardial I/R injury is not fully understood. The present study was designed to investigate whether H2S preconditioning attenuates myocardial I/R injury in rats and whether the observed protection is associated with reduced endo/sarcoplasmic reticulum (ER/SR) stress. We found that H2S preconditioning significantly reduced myocardial infarct size, preserved left ventricular function, and inhibited I/R-induced cardiomyocyte apoptosis in vivo. Furthermore, H2S preconditioning significantly attenuated I/R-induced ER/SR stress responses, including the increased expression of glucose-regulated protein 78, C/EBP homologous protein, and activate transcription factor in myocardium. Additionally, we demonstrate that H2S preconditioning attenuates ER/SR stress and inhibits cardiomyocyte apoptosis in an in vitro model of hypoxia/reoxygenation in rat H9c2 cardiac myocytes. In conclusion, these results suggest that H2S-attenuated ER/SR stress plays an important role in its protective effects against I/R-induced myocardial injury. PMID:26339339

  6. Short and long range functions of amino acids in the transmembrane region of the sarcoplasmic reticulum ATPase. A mutational study.

    PubMed

    Chen, L; Sumbilla, C; Lewis, D; Zhong, L; Strock, C; Kirtley, M E; Inesi, G

    1996-05-01

    Mutational analysis of several amino acids in the transmembrane region of the sarcoplasmic reticulum ATPase was performed by expressing wild type ATPase and 32 site-directed mutants in COS-1 cells followed by functional characterization of the microsomal fraction. Four different phenotype characteristics were observed in the mutants: (a) functions similar to those sustained by the wild type ATPase; (b) Ca2+ transport inhibited to a greater extent than ATPase hydrolytic activity; (c) inhibition of transport and hydrolytic activity in the presence of high levels of phosphorylated enzyme intermediate; and (d) total inhibition of ATP utilization by the enzyme while retaining the ability to form phosphoenzyme by utilization of P(i). Analysis of experimental observations and molecular models revealed short and long range functions of several amino acids within the transmembrane region. Short range functions include: (a) direct involvement of five amino acids in Ca2+ binding within a channel formed by clustered transmembrane helices M4, M5, M6, and M8; (b) roles of several amino acids in structural stabilization of the helical cluster for optimal channel function; and (c) a specific role of Lys297 in sealing the distal end of the channel, suggesting that the M4 helix rotates to allow vectorial flux of Ca2+ upon enzyme phosphorylation. Long range functions are related to the influence of several transmembrane amino acids on phosphorylation reactions with ATP or P(i), transmitted to the extramembranous region of the ATPase in the presence or in the absence of Ca2+.

  7. Luteolin Exerts Cardioprotective Effects through Improving Sarcoplasmic Reticulum Ca2+-ATPase Activity in Rats during Ischemia/Reperfusion In Vivo

    PubMed Central

    Nai, Changsheng; Xuan, Haochen; Zhang, Yingying; Shen, Mengxiao; Xu, Tongda; Pan, Defeng; Zhang, Congwei; Zhang, Yanbin; Li, Dongye

    2015-01-01

    The flavonoid luteolin exists in many types of fruits, vegetables, and medicinal herbs. Our previous studies have demonstrated that luteolin reduced ischemia/reperfusion (I/R) injury in vitro, which was related with sarcoplasmic reticulum Ca2+-ATPase (SERCA2a) activity. However, the effects of luteolin on SERCA2a activity during I/R in vivo remain unclear. To investigate whether luteolin exerts cardioprotective effects and to monitor changes in SERCA2a expression and activity levels in vivo during I/R, we created a myocardial I/R rat model by ligating the coronary artery. We demonstrated that luteolin could reduce the myocardial infarct size, lactate dehydrogenase release, and apoptosis during I/R injury in vivo. Furthermore, we found that luteolin inhibited the I/R-induced decrease in SERCA2a activity in vivo. However, neither I/R nor luteolin altered SERCA2a expression levels in myocardiocytes. Moreover, the PI3K/Akt signaling pathway played a vital role in this mechanism. In conclusion, the present study has confirmed for the first time that luteolin yields cardioprotective effects against I/R injury by inhibiting the I/R-induced decrease in SERCA2a activity partially via the PI3K/Akt signaling pathway in vivo, independent of SERCA2a protein level regulation. SERCA2a activity presents a novel biomarker to assess the progress of I/R injury in experimental research and clinical applications. PMID:26681967

  8. Reactive sulfhydryl groups of sarcoplasmic reticulum ATPase. II. Site of labeling with iodoacetamide and its fluorescent derivative.

    PubMed

    Yamashita, T; Kawakita, M

    1987-02-01

    Iodoacetamide (IAA) and its fluorescent derivative, 5-(2-iodoacetamidoethyl) amino-naphthalene-1-sulfonate (IAEDANS) specifically bind to a site on the C-terminal half of sarcoplasmic reticulum (SR) Ca2+,Mg2+-ATPase. The location of this specific binding site was identified. SR membranes were treated with 150 microM [14C]IAA at pH 7.0 and 30 degrees C. One mole of IAA per mole of ATPase was bound in 6 h without affecting the Ca2+-transport activity. [14C]IAA-labeled SR membranes were cleaved with BrCN, and 14C-labeled peptide fragments were separated by Sephadex LH-60 chromatography and then digested further with trypsin. A radioactive peptide (Ala-Cys 674-Cys-Phe-Ala-Arg) was purified by Sephadex LH-20 chromatography and C18 reversed phase HPLC (Cys denotes the [14C]IAA-binding site). IAEDANS-labeling was carried out by reacting SR membranes with 50 microM IAEDANS for 5 h, at pH 7.0 and 30 degrees C. A fluorescent peptide was successfully purified by the same procedures as for the IAA-labeled peptide, and the amino acid sequence analysis of this peptide revealed that the IAEDANS labeling site was identical with the IAA binding site.

  9. Calcium translocation mechanism in sarcoplasmic reticulum vesicles, deduced from location studies of protein-bound spin labels.

    PubMed Central

    Champeil, P; Rigaud, J L; Gary-Bobo, C M

    1980-01-01

    Sarcoplasmic reticulum vesicles were exposed to various thiol-directed spin labels, and the position of the label on the inner or outer vesicle surface was investigated as a function of the ATPase (adenosinetriphosphatase; ATP phosphohydrolase, EC 3.6.1.3) chemical state. Previous measurements of label accessibility to externally added ascorbate had been considered to suggest an external-internal transition of protein-bound labels, coupled with ion translocation [Tonomura, Y. & Morales, M.F. (1974) Proc. Natl. Acad. Sci. USA 71, 3687-3691]. We show that these ascorbate studies do not lead to convincing conclusions. We demonstrate, on the contrary, that transition ions (nickel and ferricyanide) can be used as selective line-broadening agents for the signals arising from external labels. No significant difference in nickel- or ferricyanide-label interaction can be attributed to a different orientation of the label in any of the enzyme chemical states tested. Our results therefore contradict the current interpretation of ascorbate quenching experiments in terms of calcium ATPase rotatory motion; rather they are consistent with ion transport models involving only limited conformational rearrangements of the pump. PMID:6446710

  10. Hydralazine-induced promoter demethylation enhances sarcoplasmic reticulum Ca2+ -ATPase and calcium homeostasis in cardiac myocytes.

    PubMed

    Kao, Yu-Hsun; Cheng, Chen-Chuan; Chen, Yao-Chang; Chung, Cheng-Chih; Lee, Ting-I; Chen, Shih-Ann; Chen, Yi-Jen

    2011-09-01

    Sarcoplasmic reticulum (SR) Ca(2+)-ATPase (SERCA2a) plays an essential role in Ca(2+) homeostasis and cardiac functions. The promoter region of SERCA2a has a high content of CpG islands; thus, epigenetic modification by inhibiting methylation can enhance SERCA2a expression in cardiomyocytes. Hydralazine, a drug frequently used in heart failure, is a potential DNA methylation inhibitor. We evaluated whether hydralazine can modulate Ca(2+) handling through an increase in SERCA2a expression via regulating methylation. We used indo-1 fluorescence, real-time RT-PCR, immunoblotting, and methylation-specific PCR to investigate intracellular Ca(2+), the expressions of RNA and protein, and methylation of SERCA2a in HL-1 cardiomyocytes with and without (control) the administration of hydralazine (1, 10, and 30 μM) for 72 h. Hydralazine (10 and 30 μM) increased the intracellular Ca(2+) transients and SR Ca(2+) contents. Hydralazine (10 and 30 μM) decreased methylation in the SERCA2a promoter region and increased the RNA and protein expressions of SERCA2a. Additionally, hydralazine (10 and 30 μM) decreased the expression of DNA methyltransferase 1. Moreover, treatment with hydralazine in isoproterenol-induced heart failure rats decreased the promoter methylation of SERCA2a and increased SERCA2a RNA expression. In conclusion, hydralazine-induced promoter demethylation may improve cardiac function through increasing SERCA2a and modulating calcium homeostasis in cardiomyocytes.

  11. Heterogeneity of neuromuscular junctions in striated muscle of human esophagus demonstrated by triple staining for the vesicular acetylcholine transporter, alpha-bungarotoxin, and acetylcholinesterase.

    PubMed

    Kallmünzer, Bernd; Sörensen, Björn; Neuhuber, Winfried L; Wörl, Jürgen

    2006-05-01

    During studies on enteric co-innervation in the human esophagus, we found that not all acetylcholinesterase (AChE)-positive motor endplates stained for alpha-bungarotoxin (alpha-BT) and the vesicular acetylcholine transporter (VAChT), respectively. Therefore, we probed for differences in neuromuscular junctions in human esophagus by using triple staining for VAChT, alpha-BT, and AChE followed by qualitative and quantitative analysis. To exclude that the results were caused by processing artifacts, we additionally examined the influence of a number of factors including post-mortem changes and the type and duration of fixation on the staining results. Four types of neuromuscular junction could be distinguished in human esophagus: type I with VAChT-positive and type II with VAChT-negative nerve terminals on a alpha-BT-positive and AChE-positive endplate area, type III with VAChT-positive nerve terminals on a alpha-BT-negative but AChE-positive endplate area, and type IV with VAChT-negative nerve terminals on a alpha-BT-negative but AChE-positive endplate area. On average, 32% of evaluated AChE-positive motor endplates were type I, 6% type II, 24% type III, and 38% type IV. Based on these results, we suggest that, in human esophagus, (1) the most reliable method for staining motor endplates is presently AChE histochemistry, (2) alpha-BT-sensitive and alpha-BT-resistant nicotinic acetylcholine receptors exist in neuromuscular junctions, and (3) different types of VAChT or transport mechanisms for acetylcholine probably exist in neuromuscular junctions.

  12. Histotopographical study of human periocular elastic fibers using aldehyde-fuchsin staining with special reference to the sleeve and pulley system for extraocular rectus muscles.

    PubMed

    Osanai, Hajime; Murakami, Gen; Ohtsuka, Aiji; Suzuki, Daisuke; Nakagawa, Takashi; Tatsumi, Haruyuki

    2009-09-01

    The aim of this study was to investigate the detailed configuration of periocular elastic fibers. Semiserial paraffin sections were made using 40 whole orbital contents from 27 elderly cadavers and stained by the aldehyde-fuchsin method. Periocular tissues were classified into three types according to directions of the elastic fibers, i.e., tissues containing anteroposteriorly running elastic fibers, those with mediolateral fibers, and those with meshwork of fibers. Anteroposterior elastic fiber-dominant tissue was seen in the upper eyelid and newly defined pulley plate for the medial and lateral recti (MR, LR). Mediolateral fibers were predominant in the central part of the inferior rectus pulley. In the pulley plates for the MR and LR, anteroposteriorly running fibers encased the striated muscle. Tenon's capsule and the epimysium of the recti were mediolateral fiber-dominant. However, at the entrance of the muscle terminal where Tenon's capsule reflects and continues to the epimysium, composite elastic fibers provided a meshwork-like skeleton. The elastic mesh was also seen around the lacrimal canaliculi. The pulley for the recti seemed to be composed of two parts--a connective tissue plate encasing the recti and specialized Tenon's capsule at an entrance or porta of the muscle. For both parts, elastic fibers were major functional components. The anteroposterior elastic fibers in the MR and LR pulley plates, especially, seemed to receive anteroposteriorly directed stress and tension from these striated muscles. The elastic interfaces seemed to prevent any concentration of stress that would interfere with periocular striated muscle functions, including hypothetical active pulleys.

  13. SERCA1 overexpression minimizes skeletal muscle damage in dystrophic mouse models

    PubMed Central

    Mázala, Davi A. G.; Pratt, Stephen J. P.; Chen, Dapeng; Molkentin, Jeffery D.; Lovering, Richard M.

    2015-01-01

    Duchenne muscular dystrophy (DMD) is characterized by progressive muscle wasting secondary to repeated muscle damage and inadequate repair. Elevations in intracellular free Ca2+ have been implicated in disease progression, and sarcoplasmic/endoplasmic reticulum Ca2+-ATPase 1 (SERCA1) overexpression has been shown to ameliorate the dystrophic phenotype in mdx mice. The purpose of this study was to assess the effects of SERCA1 overexpression in the more severe mdx/Utr−/− mouse model of DMD. Mice overexpressing SERCA1 were crossed with mdx/Utr+/− mice to generate mdx/Utr−/−/+SERCA1 mice and compared with wild-type (WT), WT/+SERCA1, mdx/+SERCA1, and genotype controls. Mice were assessed at ∼12 wk of age for changes in Ca2+ handling, muscle mass, quadriceps torque, markers of muscle damage, and response to repeated eccentric contractions. SERCA1-overexpressing mice had a two- to threefold increase in maximal sarcoplasmic reticulum Ca2+-ATPase activity compared with WT which was associated with normalization in body mass for both mdx/+SERCA1 and mdx/Utr−/−/+SERCA1. Torque deficit in the quadriceps after eccentric injury was 2.7-fold greater in mdx/Utr−/− vs. WT mice, but only 1.5-fold greater in mdx/Utr−/−/+SERCA1 vs. WT mice, an attenuation of 44%. Markers of muscle damage (% centrally nucleated fibers, necrotic area, and serum creatine kinase levels) were higher in both mdx and mdx/Utr−/− vs. WT, and all were attenuated by overexpression of SERCA1. These data indicate that SERCA1 overexpression ameliorates functional impairments and cellular markers of damage in a more severe mouse model of DMD. These findings support targeting intracellular Ca2+ control as a therapeutic approach for DMD. PMID:25652448

  14. Characterization of Post-Translational Modifications to Calsequestrins of Cardiac and Skeletal Muscle

    PubMed Central

    Lewis, Kevin M.; Munske, Gerhard R.; Byrd, Samuel S.; Kang, Jeehoon; Cho, Hyun-Jai; Ríos, Eduardo; Kang, ChulHee

    2016-01-01

    Calsequestrin is glycosylated and phosphorylated during its transit to its final destination in the junctional sarcoplasmic reticulum. To determine the significance and universal profile of these post-translational modifications to mammalian calsequestrin, we characterized, via mass spectrometry, the glycosylation and phosphorylation of skeletal muscle calsequestrin from cattle (B. taurus), lab mice (M. musculus) and lab rats (R. norvegicus) and cardiac muscle calsequestrin from cattle, lab rats and humans. On average, glycosylation of skeletal calsequestrin consisted of two N-acetylglucosamines and one mannose (GlcNAc2Man1), while cardiac calsequestrin had five additional mannoses (GlcNAc2Man6). Skeletal calsequestrin was not phosphorylated, while the C-terminal tails of cardiac calsequestrin contained between zero to two phosphoryls, indicating that phosphorylation of cardiac calsequestrin may be heterogeneous in vivo. Static light scattering experiments showed that the Ca2+-dependent polymerization capabilities of native bovine skeletal calsequestrin are enhanced, relative to the non-glycosylated, recombinant isoform, which our crystallographic studies suggest may be due to glycosylation providing a dynamic “guiderail”-like scaffold for calsequestrin polymerization. Glycosylation likely increases a polymerization/depolymerization response to changing Ca2+ concentrations, and proper glycosylation, in turn, guarantees both effective Ca2+ storage/buffering of the sarcoplasmic reticulum and localization of calsequestrin (Casq) at its target site. PMID:27649144

  15. Electrical models of excitation-contraction coupling and charge movement in skeletal muscle.

    PubMed

    Mathias, R T; Levis, R A; Eisenberg, R S

    1980-07-01

    The consequences of ionic current flow from the T system to the sarcoplasmic reticulum (SR) of skeletal muscle are examined. The Appendix analyzes a simple model in which the conductance gx, linking T system and SR, is in series with a parallel resistor and capacitor having fixed values. The conductance gx is supposed to increase rapidly with depolarization and to decrease slowly with repolarization. Nonlinear transient currents computed from this model have some of the properties of gating currents produced by intramembrane charge movement. In particular, the integral of the transient current upon depolarization approximates that upon repolarization. Thus, equality of nonlinear charge movement can occur without intramembrane charge movement. A more complicated model is used in the text to fit the structure of skeletal muscle and other properties of its charge movement. Rectification is introduced into gx and the membrane conductance of the terminal cisternae to give asymmetry in the time-course of the transient currents and saturation in the curve relating charge movement to depolarization, respectively. The more complex model fits experimental data quite well if the longitudinal tubules of the sarcoplasmic reticulum are isolated from the terminal cisternae by a substantial resistance and if calcium release from the terminal cisternae is, for the most part, electrically silent. Specific experimental tests of the model are proposed, and the implications for excitation-contraction coupling are discussed.

  16. Characterization of Post-Translational Modifications to Calsequestrins of Cardiac and Skeletal Muscle.

    PubMed

    Lewis, Kevin M; Munske, Gerhard R; Byrd, Samuel S; Kang, Jeehoon; Cho, Hyun-Jai; Ríos, Eduardo; Kang, ChulHee

    2016-01-01

    Calsequestrin is glycosylated and phosphorylated during its transit to its final destination in the junctional sarcoplasmic reticulum. To determine the significance and universal profile of these post-translational modifications to mammalian calsequestrin, we characterized, via mass spectrometry, the glycosylation and phosphorylation of skeletal muscle calsequestrin from cattle (B. taurus), lab mice (M. musculus) and lab rats (R. norvegicus) and cardiac muscle calsequestrin from cattle, lab rats and humans. On average, glycosylation of skeletal calsequestrin consisted of two N-acetylglucosamines and one mannose (GlcNAc₂Man₁), while cardiac calsequestrin had five additional mannoses (GlcNAc₂Man₆). Skeletal calsequestrin was not phosphorylated, while the C-terminal tails of cardiac calsequestrin contained between zero to two phosphoryls, indicating that phosphorylation of cardiac calsequestrin may be heterogeneous in vivo. Static light scattering experiments showed that the Ca(2+)-dependent polymerization capabilities of native bovine skeletal calsequestrin are enhanced, relative to the non-glycosylated, recombinant isoform, which our crystallographic studies suggest may be due to glycosylation providing a dynamic "guiderail"-like scaffold for calsequestrin polymerization. Glycosylation likely increases a polymerization/depolymerization response to changing Ca(2+) concentrations, and proper glycosylation, in turn, guarantees both effective Ca(2+) storage/buffering of the sarcoplasmic reticulum and localization of calsequestrin (Casq) at its target site. PMID:27649144

  17. Cellular Trafficking of Phospholamban and Formation of Functional Sarcoplasmic Reticulum During Myocyte DIfferentiation

    SciTech Connect

    Stenoien, David L.; Knyushko, Tatyana V.; Londono, Monica P.; Opresko, Lee; Mayer, M. Uljana; Brady, Scott T.; Squier, Thomas C.; Bigelow, Diana J.

    2007-06-01

    The sarco/endoplasmic reticulum Ca-ATPase (SERCA) family members are transmembrane proteins that play an essential role in regulating intracellular calcium levels. Phospholamban (PLB), a 52 amino acid phosphoprotein, regulates SERCA activity in adult heart and skeletal muscle. Using the C2C12 myocyte cell line, we find endogenous PLB constitutively expressed in both myoblasts and myotubes, whereas SERCA expression coincides with activation of the differentiation program. PLB has a punctuate distribution in myoblasts changing to a reticular distribution in myotubes where it colocalizes with SERCAs. To examine the distribution and dynamics of PLB and SERCA, we expressed fluorescent fusion proteins (GFP, CFP, and YFP) of PLB and SERCA in myoblasts. Coexpressed PLB and SERCA localize to distinct cellular compartments in myoblasts but begin to colocalize as cells differentiate. Fluorescence Recovery After Photobleaching (FRAP) studies show different recovery patterns for each protein in myoblasts confirming their localization to distinct compartments. To extend these studies, we created stable cell lines expressing O6-alkylguanine-DNA alkyltransferase (AGT) fusions with PLB or SERCA to track their localization as myocytes differentiate. These experiments demonstrate that PLB localizes to punctate vesicles in myoblasts and adopts a reticular distribution that coincides with SERCA distribution after differentiation. Colocalization experiments indicate that a subset of PLB in myoblasts colocalizes with endosomes, Golgi, and the plasma membrane however PLB also localizes to other, as yet unidentified vesicles. Our results indicate that differentiation plays a critical role in regulating PLB distribution to ensure its colocalization within the same cellular compartment as SERCA in differentiated cells. The presence and altered distribution of PLB in undifferentiated myoblasts raises the possibility that this protein has additional functions distinct from SERCA regulation.

  18. Quantifying bond distortions in transient enzyme species by a combination of density functional theory calculations and time-resolved infrared difference spectroscopy. Implications for the mechanism of dephosphorylation of the sarcoplasmic reticulum Ca(2+)-ATPase (SERCA1a).

    PubMed

    Barth, Andreas

    2015-10-01

    The sarcoplasmic Ca(2+)-ATPase (SERCA1a) forms two phosphoenzyme intermediates during Ca(2+) pumping. The second intermediate E2P hydrolyzes rapidly, which is essential for the rapid removal of Ca(2+) from the cytosol of muscle cells. The present work studies whether a weakening of the scissile PO bond in the E2P ground state facilitates dephosphorylation. To this end, the experimentally known vibrational spectrum of the E2P phosphate group was calculated with density functional theory (DFT) using structural models at two levels of structural complexity: (i) Models of acetyl phosphate in simple environments and (ii) ~150 atom models of the catalytic site. It was found that the enzyme environment distorts the structure of the phosphate group: one of the terminal PO bonds is shorter in the catalytic site indicating weaker interactions than in water. However, the bond that bridges phosphate and Asp351 is unaffected. This indicates that the scissile PO bond is not weakened by the enzyme environment of E2P. A second finding was that the catalytic site of the E2P state in aqueous solution appears to adopt a structure as in the crystals with BeF3(-), where the ATPase is in a non-reactive conformation. The reactant state of the dephosphorylation reaction differs from the E2P ground state: Glu183 faces Asp351 and positions the attacking water molecule. This state has a 0.04Å longer, and thus weaker, bridging PO bond. The reactant state is not detected in our experiments, indicating that its energy is at least 1kcal/mol higher than that of the E2P ground state.

  19. The RNA-binding protein Rbfox1 regulates splicing required for skeletal muscle structure and function

    PubMed Central

    Pedrotti, Simona; Giudice, Jimena; Dagnino-Acosta, Adan; Knoblauch, Mark; Singh, Ravi K.; Hanna, Amy; Mo, Qianxing; Hicks, John; Hamilton, Susan; Cooper, Thomas A.

    2015-01-01

    The Rbfox family of RNA-binding proteins is highly conserved with established roles in alternative splicing (AS) regulation. High-throughput studies aimed at understanding transcriptome remodeling have revealed skeletal muscle as displaying one of the largest number of AS events. This finding is consistent with requirements for tissue-specific protein isoforms needed to sustain muscle-specific functions. Rbfox1 is abundant in vertebrate brain, heart and skeletal muscle. Genome-wide genetic approaches have linked the Rbfox1 gene to autism, and a brain-specific knockout mouse revealed a critical role for this splicing regulator in neuronal function. Moreover, a Caenorhabditis elegans Rbfox1 homolog regulates muscle-specific splicing. To determine the role of Rbfox1 in muscle function, we developed a conditional knockout mouse model to specifically delete Rbfox1 in adult tissue. We show that Rbfox1 is required for muscle function but a >70% loss of Rbfox1 in satellite cells does not disrupt muscle regeneration. Deep sequencing identified aberrant splicing of multiple genes including those encoding myofibrillar and cytoskeletal proteins, and proteins that regulate calcium handling. Ultrastructure analysis of Rbfox1−/− muscle by electron microscopy revealed abundant tubular aggregates. Immunostaining showed mislocalization of the sarcoplasmic reticulum proteins Serca1 and Ryr1 in a pattern indicative of colocalization with the tubular aggregates. Consistent with mislocalization of Serca1 and Ryr1, calcium handling was drastically altered in Rbfox1−/− muscle. Moreover, muscle function was significantly impaired in Rbfox1−/− muscle as indicated by decreased force generation. These results demonstrate that Rbfox1 regulates a network of AS events required to maintain multiple aspects of muscle physiology. PMID:25575511

  20. Murine Muscle Engineered from Dermal Precursors: An In Vitro Model for Skeletal Muscle Generation, Degeneration, and Fatty Infiltration

    PubMed Central

    García-Parra, Patricia; Naldaiz-Gastesi, Neia; Maroto, Marcos; Padín, Juan Fernando; Goicoechea, María; Aiastui, Ana; Fernández-Morales, José Carlos; García-Belda, Paula; Lacalle, Jaione; Álava, Jose Iñaki; García-Verdugo, José Manuel; García, Antonio G.

    2014-01-01

    Skeletal muscle can be engineered by converting dermal precursors into muscle progenitors and differentiated myocytes. However, the efficiency of muscle development remains relatively low and it is currently unclear if this is due to poor characterization of the myogenic precursors, the protocols used for cell differentiation, or a combination of both. In this study, we characterized myogenic precursors present in murine dermospheres, and evaluated mature myotubes grown in a novel three-dimensional culture system. After 5–7 days of differentiation, we observed isolated, twitching myotubes followed by spontaneous contractions of the entire tissue-engineered muscle construct on an extracellular matrix (ECM). In vitro engineered myofibers expressed canonical muscle markers and exhibited a skeletal (not cardiac) muscle ultrastructure, with numerous striations and the presence of aligned, enlarged mitochondria, intertwined with sarcoplasmic reticula (SR). Engineered myofibers exhibited Na+- and Ca2+-dependent inward currents upon acetylcholine (ACh) stimulation and tetrodotoxin-sensitive spontaneous action potentials. Moreover, ACh, nicotine, and caffeine elicited cytosolic Ca2+ transients; fiber contractions coupled to these Ca2+ transients suggest that Ca2+ entry is activating calcium-induced calcium release from the SR. Blockade by d-tubocurarine of ACh-elicited inward currents and Ca2+ transients suggests nicotinic receptor involvement. Interestingly, after 1 month, engineered muscle constructs showed progressive degradation of the myofibers concomitant with fatty infiltration, paralleling the natural course of muscular degeneration. We conclude that mature myofibers may be differentiated on the ECM from myogenic precursor cells present in murine dermospheres, in an in vitro system that mimics some characteristics found in aging and muscular degeneration. PMID:23631552

  1. Morphological and functional maturation of a skeletal muscle regulated by juvenile hormone.

    PubMed

    Rose, Uwe

    2004-01-01

    Reproductive behaviour of animals requires a well-adapted muscular system. This study examines the structural and functional development of ovipositor muscle properties in female locusts during reproductive development. A possible regulation by juvenile hormone (JH) was assessed by comparing muscle properties in immature and mature females and with those whose JH production was inhibited by allatectomy early in adult life. The results are related to the reproductive behaviour of locusts. Histological and ultrastructural comparison of muscle fibres and their associated cuticular structures (apodemes) revealed dramatic growth during the first 2 weeks of reproductive development. The cross-sectional area of muscle fibres increased sevenfold, and their mass-per-length 5.3-fold. Ultrastructural examination showed growth of mitochondria, development of sarcoplasmic reticulum and increasing levels of structural organisation of myofibrils. Muscles of mature females displayed pronounced fatigue resistance, contracted more powerfully (twitch, 33.22+/-10.8 mN; 50 Hz, 623.66+/-115.77 mN) and had almost two times faster kinetics than those of immature females (twitch, 6.5+/-2.6 mN; 50 Hz, 14.19+/-2.58 mN). Together with muscular maturation, cuticular apodemes, which serve as attachment sides for ovipositor muscles, grow considerably in length and width and assume a complex surface structure. Most of the described changes were suppressed in females deprived of JH (allatectomised). The results demonstrate an adaptation of muscle properties to the requirements of reproductive behaviour that is largely regulated by juvenile hormone.

  2. Stimulus rate dependence of regional cerebral blood flow in human striate cortex, demonstrated by positron emission tomography

    SciTech Connect

    Fox, P.T.; Raichle, M.E.

    1984-05-01

    The purpose of this investigation was to determine the relationship between the repetition rate of a simple sensory stimulus and regional cerebral blood flow (rCBF) in the human brain. Positron emission tomography (PET), using intravenously administered H/sub 2/(/sup 15/)O as the diffusible blood-flow tracer, was employed for all CBF measurements. The use of H/sub 2/(/sup 15/)O with PET allowed eight CBF measurements to be made in rapid sequence under multiple stimulation conditions without removing the subject from the tomograph. Nine normal volunteers each underwent a series of eight H2(/sup 15/)O PET measurements of CBF. Initial and final scans were made during visual deprivation. The six intervening scans were made during visual activation with patterned-flash stimuli given in random order at 1.0-, 3.9-, 7.8-, 15.5-, 33.1-, and 61-Hz repetition rates. The region of greatest rCBF increase was determined. Within this region the rCBF was determined for every test condition and then expressed as the percentage change from the value of the initial unstimulated scan (rCBF% delta). In every subject, striate cortex rCBF% delta varied systematically with stimulus rate. Between 0 and 7.8 Hz, rCBF% delta was a linear function of stimulus repetition rate. The rCBF response peaked at 7.8 Hz and then declined. The rCBF% delta during visual stimulation was significantly greater than that during visual deprivation for every stimulus rate except 1.0 Hz. The anatomical localization of the region of peak rCBF response was determined for every subject to be the mesial occipital lobes along the calcarine fissure, primary visual cortex. Stimulus rate is a significant determinant of rCBF response in the visual cortex. Investigators of brain responses to selective activation procedures should be aware of the potential effects of stimulus rate on rCBF and other measurements of cerebral metabolism.

  3. Changes in Late Cretaceous-Quaternary Caribbean plate motion directions inferred from paleostress measurements from striated fault planes

    NASA Astrophysics Data System (ADS)

    Batbayar, K.; Mann, P.; Hippolyte, J.

    2013-12-01

    We compiled paleostress analyses from previous research works collected at 591 localities of striated fault planes in rocks ranging in age from Late Cretaceous to Quaternary in the circum-Caribbean and Mexico. The purpose of the study is to quantify a progressive clockwise rotation of the Caribbean plate during its Late Cretaceous to recent subduction of the Proto-Caribbean seaway. Paleostress analysis is based on the assumption that slickenside lineations indicate both the direction and sense of maximum resolved shear stress on that fault plane. We have plotted directions of maximum horizontal stress onto plate tectonic reconstructions of the circum-Caribbean plate boundaries and infer that these directions are proxies for paleo-plate motion directions of the Caribbean plate. Plotting these stress directions onto reconstructions provided a better visualization of the relation of stress directions to blocks at their time of Late Cretaceous to recent deformation. Older, more deformed rocks of Late Cretaceous to Eocene ages yield a greater scatter in derived paleostress directions as these rocks have steeper dips, more pervasive faulting, and were likely affected by large rotations as known from previous paleomagnetic studies of Caribbean plate margins. Despite more scatter in measurements from older rock units, four major events that affected the Caribbean plate and the Great Arc of the Caribbean (GAC) are recognizable from changing orientations of stress directions: 1) Late Cretaceous collision of the GAC with southern Mexico and Colombia is consistent with NE directions of maximum compression in rocks of this age range in southern Mexico and EW directions in Colombia as the GAC approached the Proto-Caribbean seaway; 2) Paleocene-Eocene collision of the GAC with the Bahamas platform in Cuba and Hispaniola and with the South American plate in Venezuela is consistent with CW rotations of stress directions in rocks of these ages in the northern Caribbean and CCW

  4. Towards muscle-specific meat color stability of Chinese Luxi yellow cattle: A proteomic insight into post-mortem storage.

    PubMed

    Wu, Wei; Yu, Qian-Qian; Fu, Yu; Tian, Xiao-Jing; Jia, Fei; Li, Xing-Min; Dai, Rui-Tong

    2016-09-16

    Searching for potential predictors of meat color is a challenging task for the meat industry. In this study, the relationship between meat color parameters and the sarcoplasmic proteome of M. longissimuss lumborum (LL) and M. psoas major (PM) from Chinese Luxi yellow cattle during post-mortem storage (0, 5, 10 and 15days) were explored with the aid of the integrated proteomics and bioinformatics approaches. Meat color attributes revealed that LL displayed better color stability than PM during storage. Furthermore, sarcoplasmic proteins of these two muscles were compared between days 5, 10, 15 and day 0. Several proteins were closely correlated with meat color attributes and they were muscle-specific and responsible for the meat color stability at different storage periods. Glycerol-3-phosphate dehydrogenase, fructose-bisphosphate aldolase A isoform, glycogen phosphorylase, peroxiredoxin-2, phosphoglucomutase-1, superoxide dismutase [Cu-Zn], heat shock cognate protein (71kDa) might serve as the candidate predictors of meat color stability during post-mortem storage. In addition, bioinformatics analyses indicated that more proteins were involved in glycolytic metabolism of LL, which contributed to better meat color stability of LL than PM. The present results could provide a proteomic insight into muscle-specific meat color stability of Chinese Luxi yellow cattle during post-mortem storage.

  5. Cavin4b/Murcb Is Required for Skeletal Muscle Development and Function in Zebrafish

    PubMed Central

    Housley, Michael P.; Njaine, Brian; Ricciardi, Filomena; Stone, Oliver A.; Hölper, Soraya; Krüger, Marcus; Kostin, Sawa; Stainier, Didier Y. R.

    2016-01-01

    Skeletal muscles provide metazoans with the ability to feed, reproduce and avoid predators. In humans, a heterogeneous group of genetic diseases, termed muscular dystrophies (MD), lead to skeletal muscle dysfunction. Mutations in the gene encoding Caveolin-3, a principal component of the membrane micro-domains known as caveolae, cause defects in muscle maintenance and function; however it remains unclear how caveolae dysfunction underlies MD pathology. The Cavin family of caveolar proteins can form membrane remodeling oligomers and thus may also impact skeletal muscle function. Changes in the distribution and function of Cavin4/Murc, which is predominantly expressed in striated muscles, have been reported to alter caveolae structure through interaction with Caveolin-3. Here, we report the generation and phenotypic analysis of murcb mutant zebrafish, which display impaired swimming capacity, skeletal muscle fibrosis and T-tubule abnormalities during development. To understand the mechanistic importance of Murc loss of function, we assessed Caveolin-1 and 3 localization and found it to be abnormal. We further identified an in vivo function for Murc in Erk signaling. These data link Murc with developmental defects in T-tubule formation and progressive muscle dysfunction, thereby providing a new candidate for the etiology of muscular dystrophy. PMID:27294373

  6. Selenoprotein-deficient transgenic mice exhibit enhanced exercise-induced muscle growth.

    PubMed

    Hornberger, Troy A; McLoughlin, Thomas J; Leszczynski, Jori K; Armstrong, Dustin D; Jameson, Ruth R; Bowen, Phyllis E; Hwang, Eun-Sun; Hou, Honglin; Moustafa, Mohamed E; Carlson, Bradley A; Hatfield, Dolph L; Diamond, Alan M; Esser, Karyn A

    2003-10-01

    Dietary intake of selenium has been implicated in a wide range of health issues, including aging, heart disease and cancer. Selenium deficiency, which can reduce selenoprotein levels, has been associated with several striated muscle pathologies. To investigate the role of selenoproteins in skeletal muscle biology, we used a transgenic mouse (referred to as i6A-) that has reduced levels of selenoproteins due to the introduction and expression of a dominantly acting mutant form of selenocysteine transfer RNA (tRNA[Ser]Sec). As a consequence, each organ contains reduced levels of most selenoproteins, yet these mice are normal with regard to fertility, overall health, behavior and blood chemistries. In the present study, although skeletal muscles from i6A- mice were phenotypically indistinguishable from those of wild-type mice, plantaris muscles were approximately 50% heavier after synergist ablation, a model of exercise overload. Like muscle in wild-type mice, the enhanced growth in the i6A- mice was completely blocked by inhibition of the mammalian target of rapamycin (mTOR) pathway. Muscles of transgenic mice exhibited increased site-specific phosphorylation on both Akt and p70 ribosomal S6 kinase (p70S6k) (P < 0.05) before ablation, perhaps accounting for the enhanced response to synergist ablation. Thus, a single genetic alteration resulted in enhanced skeletal muscle adaptation after exercise, and this is likely through subtle changes in the resting phosphorylation state of growth-related kinases.

  7. Polarization gating enables sarcomere length measurements by laser diffraction in fibrotic muscle

    NASA Astrophysics Data System (ADS)

    Young, Kevin W.; Dayanidhi, Sudarshan; Lieber, Richard L.

    2014-11-01

    Sarcomere length is a key parameter commonly measured in muscle physiology since it dictates striated muscle active force. Laser diffraction (LD)-based measurements of sarcomere length are time-efficient and sample a greater number of sarcomeres compared with traditional microscopy-based techniques. However, a limitation to LD techniques is that signal quality is severely degraded by scattering events as photons propagate through tissue. Consequently, sarcomere length measurements are unattainable when the number of scattering events is sufficiently large in muscle tissue with a high scattering probability. This occurs in fibrotic skeletal muscle seen in muscular dystrophies and secondary to tissue trauma, thus eliminating the use of LD to study these skeletal muscle ailments. Here, we utilize polarization gating to extract diffracted signals that are buried in noise created by scattering. Importantly, we demonstrate that polarization-gated laser diffraction (PGLD) enables sarcomere length measurements in muscles from chronically immobilized mice hind limbs; these muscles have a substantial increase of intramuscular connective tissue that scatter light and disable sarcomere length measurements by traditional LD. Further, we compare PGLD sarcomere lengths to those measured by bright field (BF) and confocal microscopy as positive controls and reveal a significant bias of BF but not of confocal microscopy.

  8. Identification of a putative pathway for the muscle homing of stem cells in a muscular dystrophy model.

    PubMed

    Torrente, Yvan; Camirand, Geoffrey; Pisati, Federica; Belicchi, Marzia; Rossi, Barbara; Colombo, Fabio; El Fahime, Mosthapha; Caron, Nicolas J; Issekutz, Andrew C; Constantin, Gabriela; Tremblay, Jacques P; Bresolin, Nereo

    2003-08-01

    Attempts to repair muscle damage in Duchenne muscular dystrophy (DMD) by transplanting skeletal myoblasts directly into muscles are faced with the problem of the limited migration of these cells in the muscles. The delivery of myogenic stem cells to the sites of muscle lesions via the systemic circulation is a potential alternative approach to treat this disease. Muscle-derived stem cells (MDSCs) were obtained by a MACS(R) multisort method. Clones of MDSCs, which were Sca-1+/CD34-/L-selectin+, were found to adhere firmly to the endothelium of mdx dystrophic muscles after i.v. or i.m. injections. The subpopulation of Sca-1+/CD34- MDSCs expressing L-selectin was called homing MDSCs (HMDSCs). Treatment of HMDSCs with antibodies against L-selectin prevented adhesion to the muscle endothelium. Importantly, we found that vascular endothelium from striate muscle of young mdx mice expresses mucosal addressin cell adhesion molecule-1 (MAdCAM-1), a ligand for L-selectin. Our results showed for the first time that the expression of the adhesion molecule L-selectin is important for muscle homing of MDSCs. This discovery will aid in the improvement of a potential therapy for muscular dystrophy based on the systemic delivery of MDSCs.

  9. [Receptive fields of cat striate neurons: dynamics of weight and topographical characteristics of the excitatory zone of the neuron receptive fields in the visual cortex].

    PubMed

    Lazareva, N A; Kozhukhov, S A; Sharaev, G A; Novikova, R V; Tikhomirov, A S; Tsutskiridze, D Iu; Shevelev, I A

    2010-01-01

    In 22 acute experiments on anesthetized and immobilized adult cats, the dynamics of 83 on- and/or off-receptive fields (RF) was studied in 47 striate neurons with the method of temporal slices by mapping with single-flash stimuli. The latency of the RF appearance was 88 +/- 5 ms, and its duration was 192 +/- 12 ms. Dynamical wavy changes in the RF sizes and weights repeating up to three times were revealed. The mean duration of one wave was 95 +/- 4 ms. In 99% of cases, the RF discharge center defined for one temporal slice also displaced in an undulatory way relatively the RF's center defined for whole analysis period. The mean duration of this undulatory cycle was 67.3 +/- 3 ms. In 72.5% of cases, the displacement of the discharge center followed different trajectories inscribed in an ellipse. The functional significance of such changes in the RFs of striate neurons is discussed with regard to the dynamics of their detector features and underlying mechanisms.

  10. Azumolene inhibits a component of store-operated calcium entry coupled to the skeletal muscle ryanodine receptor.

    PubMed

    Zhao, Xiaoli; Weisleder, Noah; Han, Xuehai; Pan, Zui; Parness, Jerome; Brotto, Marco; Ma, Jianjie

    2006-11-01

    Dantrolene reduces the elevated myoplasmic Ca(2+) generated during malignant hyperthermia, a pharmacogenetic crisis triggered by volatile anesthetics. Although specific binding of dantrolene to the type 1 ryanodine receptor (RyR1), the Ca(2+) release channel of skeletal muscle sarcoplasmic reticulum, has been demonstrated, there is little evidence for direct dantrolene inhibition of RyR1 channel function. Recent studies suggest store-operated Ca(2+) entry (SOCE) contributes to skeletal muscle function, but the effect of dantrolene on this pathway has not been examined. Here we show that azumolene, an equipotent dantrolene analog, inhibits a component of SOCE coupled to activation of RyR1 by caffeine and ryanodine, whereas the SOCE component induced by thapsigargin is not affected. Our data suggest that azumolene distinguishes between two mechanisms of cellular signaling to SOCE in skeletal muscle, one that is coupled to and one independent from RyR1. PMID:16945924

  11. The effects of sarcolipin over-expression in mouse skeletal muscle on metabolic activity.

    PubMed

    Butler, John; Smyth, Neil; Broadbridge, Robert; Council, Claire E; Lee, Anthony G; Stocker, Claire J; Hislop, David C; Arch, Jonathan R S; Cawthorne, Michael A; Malcolm East, J

    2015-03-01

    Studies in sarcolipin knockout mice have led to the suggestion that skeletal muscle sarcolipin plays a role in thermogenesis. The mechanism proposed is uncoupling of the sarcoplasmic reticulum calcium pump. However, in other work sarcolipin was not detected in mouse skeletal tissue. We have therefore measured sarcolipin levels in mouse skeletal muscle using semi-quantitative western blotting and synthetic mouse sarcolipin. Sarcolipin levels were so low that it is unlikely that knocking out sarcolipin would have a measurable effect on thermogenesis by SERCA. In addition, overexpression of neither wild type nor FLAG-tagged variants of mouse sarcolipin in transgenic mice had any major significant effects on body mass, energy expenditure, even when mice were fed on a high fat diet. PMID:25660043

  12. Changes in qualitative composition of white muscle with nutritional status of Atlantic cod, Gadus morhua.

    PubMed

    Beaulieu, M A; Guderley, H

    1998-10-01

    Proteins from white muscle are mobilized to cover energy requirements during long-term starvation in fish. Using SDS-polyacrylamide gel electrophoresis, we compared the soluble and insoluble fractions of white muscle proteins from fed and starved Atlantic cod, Gadus morhua, to establish whether preferential preservation or degradation of specific proteins occurred during starvation. While starvation induced no qualitative changes in the electrophoretic pattern of the myofibrillar fraction, our results document differential decreases in the levels of soluble proteins during fasting. Moreover, immunoblot analysis using a monoclonal antibody directed against actin, showed a marked accumulation of this protein in the sarcoplasmic fraction of starved individuals, most likely due to myofibrillar degradation. PMID:9883575

  13. The effects of sarcolipin over-expression in mouse skeletal muscle on metabolic activity.

    PubMed

    Butler, John; Smyth, Neil; Broadbridge, Robert; Council, Claire E; Lee, Anthony G; Stocker, Claire J; Hislop, David C; Arch, Jonathan R S; Cawthorne, Michael A; Malcolm East, J

    2015-03-01

    Studies in sarcolipin knockout mice have led to the suggestion that skeletal muscle sarcolipin plays a role in thermogenesis. The mechanism proposed is uncoupling of the sarcoplasmic reticulum calcium pump. However, in other work sarcolipin was not detected in mouse skeletal tissue. We have therefore measured sarcolipin levels in mouse skeletal muscle using semi-quantitative western blotting and synthetic mouse sarcolipin. Sarcolipin levels were so low that it is unlikely that knocking out sarcolipin would have a measurable effect on thermogenesis by SERCA. In addition, overexpression of neither wild type nor FLAG-tagged variants of mouse sarcolipin in transgenic mice had any major significant effects on body mass, energy expenditure, even when mice were fed on a high fat diet.

  14. Comparison of Ca2+ release and uptake characteristics of the sarcoplasmic reticulum in isolated horse and rabbit cardiomyocytes.

    PubMed

    Loughrey, C M; Smith, G L; MacEachern, K E

    2004-09-01

    Both the cardiac action potential duration (APD) (0.6-1 s) and resting heart rate (30-40 beats/min) in the horse are significantly different from humans and smaller mammals, including the rabbit. This would be anticipated to have consequences for excitation-contraction (EC) coupling and require adaptation of the individual processes involved. The sarcoplasmic reticulum (SR) is one of the main components involved in EC coupling. This study examines and compares the activity of this organelle in the horse with that of the rabbit. In particular, the study focuses on SR Ca2+ release via the Ca2+ release channel/ryanodine receptor (RyR2) and Ca2+ uptake via the sarco(endo)plasmic reticulum Ca(2+)-ATPase (SERCA) pump. Isolated cardiomyocytes from both horse and rabbit hearts were permeabilized, bathed in a mock intracellular solution, and exposed to a specified [Ca2+]. Rabbit cardiomyocytes exposed to 260 nM [Ca2+] produced spontaneous Ca2+ release and propagated Ca2+ waves. Horse cells failed to produce Ca2+ waves; instead, only local release in the form of Ca2+ sparks was evident. However, at 550 nM [Ca2+], Ca2+ waves were produced in both species. Ca2+ waves were four times less frequent yet approximately 1.5 times greater in amplitude in the horse compared with the rabbit. Ca2+ wave velocity was comparable between the species. The reason for this disparity in Ca2+ wave characteristics is unknown. Separate measurements of oxalate-supported Ca2+ uptake into the SR suggest that both horse and rabbit cardiomyocytes have comparable levels SERCA activity. The possible reasons for the observed differences in SR Ca2+ release between the horse and rabbit are discussed.

  15. Dissociation of mitochondrial from sarcoplasmic reticular stress in Drosophila cardiomyopathy induced by molecularly distinct mitochondrial fusion defects

    PubMed Central

    Bhandari, Poonam; Song, Moshi; Dorn, Gerald W

    2015-01-01

    Mitochondrial dynamism (fusion and fission) is responsible for remodeling interconnected mitochondrial networks in some cell types. Adult cardiac myocytes lack mitochondrial networks, and their mitochondria are inherently “fragmented”. Mitochondrial fusion/fission is so infrequent in cardiomyocytes as to not be observable under normal conditions, suggesting that mitochondrial dynamism may be dispensable in this cell type. However, we previously observed that cardiomyocyte-specific genetic suppression of mitochondrial fusion factors optic atrophy 1 (Opa1) and mitofusin/MARF evokes cardiomyopathy in Drosophila hearts. We posited that fusion-mediated remodeling of mitochondria may be critical for cardiac homeostasis, although never directly observed. Alternately, we considered that inner membrane Opa1 and outer membrane mitofusin/MARF might have other as-yet poorly described roles that affect mitochondrial and cardiac function. Here we compared heart tube function in three models of mitochondrial fragmentation in Drosophila cardiomyocytes: Drp1 expression, Opa1 RNAi, and mitofusin MARF RNA1. Mitochondrial fragmentation evoked by enhanced Drp1-mediated fission did not adversely impact heart tube function. In contrast, RNAi-mediated suppression of either Opa1 or mitofusin/MARF induced cardiac dysfunction associated with mitochondrial depolarization and ROS production. Inhibiting ROS by overexpressing superoxide dismutase (SOD) or suppressing ROMO1 prevented mitochondrial and heart tube dysfunction provoked by Opa1 RNAi, but not by mitofusin/MARF RNAi. In contrast, enhancing the ability of endoplasmic/sarcoplasmic reticulum to handle stress by expressing Xbp1 rescued the cardiomyopathy of mitofusin/MARF insufficiency without improving that caused by Opa1 deficiency. We conclude that decreased mitochondrial size is not inherently detrimental to cardiomyocytes. Rather, preservation of mitochondrial function by Opa1 located on the inner mitochondrial membrane, and

  16. Facilitation of cytosolic calcium wave propagation by local calcium uptake into the sarcoplasmic reticulum in cardiac myocytes.

    PubMed

    Maxwell, Joshua T; Blatter, Lothar A

    2012-12-01

    The widely accepted paradigm for cytosolic Ca(2+) wave propagation postulates a 'fire-diffuse-fire' mechanism where local Ca(2+)-induced Ca(2+) release (CICR) from the sarcoplasmic reticulum (SR) via ryanodine receptor (RyR) Ca(2+) release channels diffuses towards and activates neighbouring release sites, resulting in a propagating Ca(2+) wave. A recent challenge to this paradigm proposed the requirement for an intra-SR 'sensitization' Ca(2+) wave that precedes the cytosolic Ca(2+) wave and primes RyRs from the luminal side to CICR. Here, we tested this hypothesis experimentally with direct simultaneous measurements of cytosolic ([Ca(2+)](i); rhod-2) and intra-SR ([Ca(2+)](SR); fluo-5N) calcium signals during wave propagation in rabbit ventricular myocytes, using high resolution fluorescence confocal imaging. The increase in [Ca(2+)](i) at the wave front preceded depletion of the SR at each point along the calcium wave front, while during this latency period a transient increase of [Ca(2+)](SR) was observed. This transient elevation of [Ca(2+)](SR) could be identified at individual release junctions and depended on the activity of the sarco-endoplasmic reticulum Ca(2+)-ATPase (SERCA). Increased SERCA activity (β-adrenergic stimulation with 1 μM isoproterenol (isoprenaline)) decreased the latency period and increased the amplitude of the transient elevation of [Ca(2+)](SR), whereas inhibition of SERCA (3 μM cyclopiazonic acid) had the opposite effect. In conclusion, the data provide experimental evidence that local Ca(2+) uptake by SERCA into the SR facilitates the propagation of cytosolic Ca(2+) waves via luminal sensitization of the RyR, and supports a novel paradigm of a 'fire-diffuse-uptake-fire' mechanism for Ca(2+) wave propagation in cardiac myocytes.

  17. Two alternate kinetic routes for the decomposition of the phosphorylated intermediate of sarcoplasmic reticulum Ca2+-ATPase.

    PubMed

    Nakamura, Y

    1984-07-10

    The decomposition of the phosphorylated intermediate (EP) of sarcoplasmic reticulum ATPase, purified by the method of deoxycholic acid extraction, was studied by first phosphorylating with [gamma-32P]ATP, then diluting the reaction mixture with 20 volumes of medium containing nonradioactive ATP, and finally quenching serial samples with acid for determination of residual [32P]EP. The time course of [32P]EP decomposition consists of an initial fast phase followed by a slow phase. The two components of EP, EPfast (1.1 nmol/mg) and EPslow (2.8 nmol/mg), decomposed with the rate constants of 6 and 0.8 min-1, respectively, in the presence of 0.5 mM CaCl2, 5mM MgCl2, and 90 mM KCl at pH 7.0 and O degrees C. The sum of the hydrolytic activities corresponding to the two components accounts for the steady state velocity of the Pi production under the same conditions, indicating that the two components represent simultaneous pathways, rather than sequential steps of EP decomposition. As the time of phosphorylation with [gamma-32P]ATP is increased from 2 to 15 s, the fraction of EPfast decreases in favor of EPslow. This conversion decreases the rate of total Pi production by the enzyme following an initial Pi burst. Conversion of EPfast to EPslow is favored by millimolar concentrations of Ca2+. On the other hand, conversion of EPslow to EPfast is obtained by reducing Ca2+ or raising Mg2+ concentration, but is prevented by removal of ADP. The EPslow fraction decreases in favor of EPfast as the temperature is increased from 0 to 22 degrees C. PMID:6234309

  18. Resveratrol, an activator of SIRT1, upregulates sarcoplasmic calcium ATPase and improves cardiac function in diabetic cardiomyopathy

    PubMed Central

    Sulaiman, M.; Matta, M. J.; Sunderesan, N. R.; Periasamy, M.; Gupta, M.

    2010-01-01

    Reduced sarcoplasmic calcium ATPase (SERCA2a) expression has been shown to play a significant role in the cardiac dysfunction in diabetic cardiomyopathy. The mechanism of SERCA2a repression is, however, not known. This study was designed to examine the effect of resveratrol (RSV), a potent activator of SIRT1, on cardiac function and SERCA2a expression in chronic type 1 diabetes. Adult male mice were injected with streptozotocin (STZ) and fed with either a regular diet or a diet enriched with RSV. STZ administration produced progressive decline in cardiac function, associated with markedly reduced SERCA2a and SIRT1 protein levels and increased collagen deposition; RSV treatment to these mice had a tremendous beneficial effect both in terms of improving SERCA2a expression and on cardiac function. In cultured cardiomyocytes, RSV restored SERCA2 promoter activity, which was otherwise highly repressed in high-glucose media. Protective effects of RSV were found to be dependent on its ability to activate Silent information regulator (SIRT) 1. In cardiomyocytes, overexpression of SIRT1 was found sufficient to activate SERCA2 promoter in a dose-dependent manner. In contrast, pretreatment of cardiomyocytes with SIRT1 antagonist, splitomycin, blocked these beneficial effects of RSV. In addition, SIRT1 knockout (+/−) mice were also found to be more sensitive to STZ-induced decline in SERCA2a mRNA. The data demonstrate that, in chronic diabetes, 1) the enzymatic activity of cardiac SIRT1 is reduced, which contributes to reduced expression of SERCA2a and 2) through activation of SIRT1, RSV enhances expression of SERCA2a and improves cardiac function. PMID:20008278

  19. R-CEPIA1er as a new tool to directly measure sarcoplasmic reticulum [Ca] in ventricular myocytes.

    PubMed

    Bovo, Elisa; Martin, Jody L; Tyryfter, Jollyn; de Tombe, Pieter P; Zima, Aleksey V

    2016-07-01

    In cardiomyocytes, [Ca] within the sarcoplasmic reticulum (SR; [Ca]SR) partially determines the amplitude of cytosolic Ca transient that, in turn, governs myocardial contraction. Therefore, it is critical to understand the molecular mechanisms that regulate [Ca]SR handling. Until recently, the best approach available to directly measure [Ca]SR was to use low-affinity Ca indicators (e.g., Fluo-5N). However, this approach presents several limitations, including nonspecific cellular localization, dye extrusion, and species limitation. Recently a new genetically encoded family of Ca indicators has been generated, named Ca-measuring organelle-entrapped protein indicators (CEPIA). Here, we tested the red fluorescence SR-targeted Ca sensor (R-CEPIA1er) as a tool to directly measure [Ca]SR dynamics in ventricular myocytes. Infection of rabbit and rat ventricular myocytes with an adenovirus expressing the R-CEPIA1er gene displayed prominent localization in the SR and nuclear envelope. Calibration of R-CEPIA1er in myocytes resulted in a Kd of 609 μM, suggesting that this sensor is sensitive in the whole physiological range of [Ca]SR [Ca]SR dynamics measured with R-CEPIA1er were compared with [Ca]SR measured with Fluo5-N. We found that both the time course of the [Ca]SR depletion and fractional SR Ca release induced by an action potential were similar between these two Ca sensors. R-CEPIA1er fluorescence did not decline during experiments, indicating lack of dye extrusion or photobleaching. Furthermore, measurement of [Ca]SR with R-CEPIA1er can be combined with cytosolic [Ca] measurements (with Fluo-4) to obtain more detailed information regarding Ca handling in cardiac myocytes. In conclusion, R-CEPIA1er is a promising tool that can be used to measure [Ca]SR dynamics in myocytes from different animal species. PMID:27233762

  20. Localization of site-specific probes on the Ca-ATPase of sarcoplasmic reticulum using fluorescence energy transfer.

    PubMed

    Squier, T C; Bigelow, D J; Garcia de Ancos, J; Inesi, G

    1987-04-01

    Highly reactive sulfhydryls, previously labeled with an iodoacetamide spin label on the Ca-ATPase of sarcoplasmic reticulum, were labeled with the fluorescent probe, 5-(2-[iodoacetyl)amino)ethyl)aminonaphthalene-1-sulfonic acid (IAEDANS), without loss of enzymatic activity. We have selectively measured the apparent distance of the more reactive site, relative to other site-specific probes at both the nucleotide and the high affinity calcium binding sites. Fluorescence energy transfer efficiencies from the donor IAEDANS to two acceptors: fluorescein 5'-isothiocyanate or 2',3'-O-(2,4,3-trinitrophenyl)adenosine monophosphate, situated at or near the nucleotide site, were measured using fluorescence lifetimes and yields. Fluorescence on polyacrylamide gels shows that the IAEDANS and fluorescein 5'-isothiocyanate labels are both associated with the B tryptic fragment. The energy transfer measurements are consistent with distances of 56 and 68 A between IAEDANS and these respective binding sites. On the other hand, energy transfer measurements using the lanthanide, praseodymium (Pr3+), as an acceptor indicate that IAEDANS is located 16-18 A from the binding site(s) of this calcium analog. Pr3+ is shown to be a good analog for calcium binding to the high affinity sites on the enzyme since it competitively displaces calcium, as evidenced by the reversal of the specific calcium-dependent intrinsic fluorescent signal and inactivation of ATPase activity, over the same narrow range in Pr3+ concentration where energy transfer is observed. Our observations suggest that the portion of the B fragment spanning the cytoplasmic portion of the ATPase is folded onto the A fragment, bringing the IAEDANS label in close proximity to the high affinity calcium binding domain.

  1. Structural dynamics of the Ca2(+)-ATPase of sarcoplasmic reticulum. Temperature profiles of fluorescence polarization and intramolecular energy transfer.

    PubMed

    Jona, I; Matko, J; Martonosi, A

    1990-10-01

    The temperature dependence of fluorescence polarization and Förster-type resonance energy transfer (FRET) was analyzed in the Ca2(+)-ATPase of sarcoplasmic reticulum using protein tryptophan and site-specific fluorescence indicators such as 5-[2-[iodoacetyl)amino)ethyl]aminonaphthalene-1-sulfonic acid (IAEDANS), fluorescein 5'-isothiocyanate (FITC), 2',3'-O-(2,4,3-trinitrophenyl)adenosine monophosphate (TNP-AMP) or lanthanides (Pr3+, Nd3+) as probes. The normalized energy transfer efficiency between AEDANS bound at cysteine-670 and -674 and FITC bound at lysine-515 increases with increasing temperature in the range of 10-37 degrees C, indicating the existence of a relatively flexible structure in the region of the ATPase molecule that links the AEDANS to the FITC site. These observations are consistent with the theory of Somogyi, Matko, Papp, Hevessy, Welch and Damjanovich (Biochemistry 23 (1984) 3403-3411) that thermally induced structural fluctuations increase the energy transfer. Structural fluctuations were also evident in the energy transfer between FITC linked to the nucleotide-binding domain and Nd3+ bound at the putative Ca2+ sites. By contrast the normalized energy transfer efficiency between AEDANS and Pr3+ was relatively insensitive to temperature, suggesting that the region between cysteine-670 and the putative Ca2+ site monitored by the AEDANS-Pr3+ pair is relatively rigid. A combination of the energy transfer data with the structural information derived from analysis of Ca2(+)-ATPase crystals yields a structural model, in which the location of the AEDANS-, FITC- and Ca2+ sites are tentatively identified.

  2. Sarcoplasmic reticulum Ca²⁺ release is both necessary and sufficient for SK channel activation in ventricular myocytes.

    PubMed

    Terentyev, Dmitry; Rochira, Jennifer A; Terentyeva, Radmila; Roder, Karim; Koren, Gideon; Li, Weiyan

    2014-03-01

    SK channels are upregulated in human patients and animal models of heart failure (HF). However, their activation mechanism and function in ventricular myocytes remain poorly understood. We aim to test the hypotheses that activation of SK channels in ventricular myocytes requires Ca(2+) release from sarcoplasmic reticulum (SR) and that SK currents contribute to reducing triggered activity. SK2 channels were overexpressed in adult rat ventricular myocytes using adenovirus gene transfer. Simultaneous patch clamp and confocal Ca(2+) imaging experiments in SK2-overexpressing cells demonstrated that depolarizations resulted in Ca(2+)-dependent outward currents sensitive to SK inhibitor apamin. SR Ca(2+) release induced by rapid application of 10 mM caffeine evoked repolarizing SK currents, whereas complete depletion of SR Ca(2+) content eliminated SK currents in response to depolarizations, despite intact Ca(2+) influx through L-type Ca(2+) channels. Furthermore, voltage-clamp experiments showed that SK channels can be activated by global spontaneous SR Ca(2+) release events Ca(2+) waves (SCWs). Current-clamp experiments revealed that SK overexpression reduces the amplitude of delayed afterdepolarizations (DADs) resulting from SCWs and shortens action potential duration. Immunolocalization studies showed that overexpressed SK channels are distributed both at external sarcolemmal membranes and along the Z-lines, resembling the distribution of endogenous SK channels. In summary, SR Ca(2+) release is both necessary and sufficient for the activation of SK channels in rat ventricular myocytes. SK currents contribute to repolarization during action potentials and attenuate DADs driven by SCWs. Thus SK upregulation in HF may have an anti-arrhythmic effect by reducing triggered activity.

  3. Phospholamban Modulates the Functional Coupling between Nucleotide Domains in Ca-ATPase Oligomeric Complexes in Cardiac Sarcoplasmic Reticulum

    SciTech Connect

    Chen, L.; Yao, Qing; Soares, Thereza A.; Squier, Thomas C.; Bigelow, Diana J.

    2009-03-24

    Oligomeric interactions between Ca-ATPase polypeptide chains and their modulation by phospholamban (PLB) were measured in native cardiac sarcoplasmic reticulum (SR) microsomes. Progressive modification of Lys514 with fluorescein-5-isothiocyanate (FITC), which physically blocks access to the nucleotide binding site by ATP, demonstrates that Ca-ATPase active sites function independently of one another prior to the phosphorylation of PLB. However, upon PKA-dependent phosphorylation of PLB, a second-order dependence between enzyme activity and the fraction of active sites is observed, consistent with a dimeric functional complex. Complementary distance measurements were made using FITC or 5-iodoacetamido-fluorescein (IAF) bound to Cys674 within the N- or P-domains respectively, to detect structural coupling within oligomeric complexes. Accompanying the phosphorylation of PLB, neighboring Ca-ATPase polypeptide chains exhibit a 4 ± 2 Å decrease in the proximity between FITC sites within the N-domain and a 9 ± 3 Å increase in the proximity between IAF sites within P-domains. Thus, the phosphorylation of PLB induces spatial rearrangements between the N- and P-domain elements of proximal Ca-ATPase polypeptide chains which restore functional interactions between neighboring polypeptide chains and, in turn, result in increased rates of catalytic turnover. These results are interpreted in terms of a structural model, calculated through optimization of shape complementarity, desolvation, and electrostatic energies, which suggests a dimeric arrangement of Ca-ATPase polypeptide chains through the proximal association of N-domains. We suggest that the phosphorylation of PLB acts to release constraints involving interdomain subunit interactions that enhance catalytically important N-domain motions.

  4. Phospholamban Modulates the Functional Coupling between Nucleotide Domains in Ca-ATPase Oligomeric Complexes in Cardiac Sarcoplasmic Reticulum

    PubMed Central

    Chen, Linda T.L.; Yao, Qing; Soares, Thereza A.; Squier, Thomas C.; Bigelow, Diana J.

    2009-01-01

    Oligomeric interactions between Ca-ATPase polypeptide chains and their modulation by phospholamban (PLB) were measured in native cardiac sarcoplasmic reticulum (SR) microsomes. Progressive modification of Lys514 with fluorescein-5-isothiocyanate (FITC), which physically blocks access to the nucleotide binding site by ATP, demonstrates that Ca-ATPase active sites function independently of one another prior to the phosphorylation of PLB. However, upon PKA-dependent phosphorylation of PLB, a second-order dependence between residual enzyme activity and the fraction of active sites is observed, consistent with a dimeric functional complex. Complementary distance measurements were made using FITC or 5-iodoacetamido-fluorescein (IAF) bound to Cys674 within the N- or P-domains respectively, to detect structural coupling within oligomeric complexes. Accompanying the phosphorylation of PLB, neighboring Ca-ATPase polypeptide chains exhibit a 4 ± 2 Å decrease in the proximity between FITC sites within the N-domain and a 9 ± 3 Å increase in the proximity between IAF sites within P-domains. Thus, the phosphorylation of PLB induces spatial rearrangements between the N- and P-domain elements of proximal Ca-ATPase polypeptide chains which restore functional interactions between neighboring polypeptide chains and, in turn, result in increased rates of catalytic turnover. These results are interpreted in terms of a structural model, calculated through optimization of shape complementarity, desolvation, and electrostatic energies, which suggests a dimeric arrangement of Ca-ATPase polypeptide chains through the proximal association of N-domains that accommodates interaction with PLB. We suggest that the phosphorylation of PLB acts to release constraints involving interdomain subunit interactions that enhance catalytically important N-domain motions. PMID:19191503

  5. Peptide fragments of the dihydropyridine receptor can modulate cardiac ryanodine receptor channel activity and sarcoplasmic reticulum Ca2+ release.

    PubMed Central

    Dulhunty, Angela F; Curtis, Suzanne M; Cengia, Louise; Sakowska, Magdalena; Casarotto, Marco G

    2004-01-01

    We show that peptide fragments of the dihydropyridine receptor II-III loop alter cardiac RyR (ryanodine receptor) channel activity in a cytoplasmic Ca2+-dependent manner. The peptides were AC (Thr-793-Ala-812 of the cardiac dihydropyridine receptor), AS (Thr-671-Leu-690 of the skeletal dihydropyridine receptor), and a modified AS peptide [AS(D-R18)], with an extended helical structure. The peptides added to the cytoplasmic side of channels in lipid bilayers at > or = 10 nM activated channels when the cytoplasmic [Ca2+] was 100 nM, but either inhibited or did not affect channel activity when the cytoplasmic [Ca2+] was 10 or 100 microM. Both activation and inhibition were independent of bilayer potential. Activation by AS, but not by AC or AS(D-R18), was reduced at peptide concentrations >1 mM in a voltage-dependent manner (at +40 mV). In control experiments, channels were not activated by the scrambled AS sequence (ASS) or skeletal II-III loop peptide (NB). Resting Ca2+ release from cardiac sarcoplasmic reticulum was not altered by peptide AC, but Ca2+-induced Ca2+ release was depressed. Resting and Ca2+-induced Ca2+ release were enhanced by both the native and modified AS peptides. NMR revealed (i) that the structure of peptide AS(D-R18) is not influenced by [Ca2+] and (ii) that peptide AC adopts a helical structure, particularly in the region containing positively charged residues. This is the first report of specific functional interactions between dihydropyridine receptor A region peptides and cardiac RyR ion channels in lipid bilayers. PMID:14678014

  6. Interaction of nucleotides with Asp(351) and the conserved phosphorylation loop of sarcoplasmic reticulum Ca(2+)-ATPase.

    PubMed

    McIntosh, D B; Woolley, D G; MacLennan, D H; Vilsen, B; Andersen, J P

    1999-09-01

    The nucleotide binding properties of mutants with alterations to Asp(351) and four of the other residues in the conserved phosphorylation loop, (351)DKTGTLT(357), of sarcoplasmic reticulum Ca(2+)-ATPase were investigated using an assay based on the 2', 3'-O-(2,4,6-trinitrophenyl)-8-azidoadenosine triphosphate (TNP-8N(3)-ATP) photolabeling of Lys(492) and competition with ATP. In selected cases where the competition assay showed extremely high affinity, ATP binding was also measured by a direct filtration assay. At pH 8.5 in the absence of Ca(2+), mutations removing the negative charge of Asp(351) (D351N, D351A, and D351T) produced pumps that bound MgTNP-8N(3)-ATP and MgATP with affinities 20-156-fold higher than wild type (K(D) as low as 0.006 microM), whereas the affinity of mutant D351E was comparable with wild type. Mutations K352R, K352Q, T355A, and T357A lowered the affinity for MgATP and MgTNP-8N(3)-ATP 2-1000- and 1-6-fold, respectively, and mutation L356T completely prevented photolabeling of Lys(492). In the absence of Ca(2+), mutants D351N and D351A exhibited the highest nucleotide affinities in the presence of Mg(2+) and at alkaline pH (E1 state). The affinity of mutant D351A for MgATP was extraordinarily high in the presence of Ca(2+) (K(D) = 0.001 microM), suggesting a transition state like configuration at the active site under these conditions. The mutants with reduced ATP affinity, as well as mutants D351N and D351A, exhibited reduced or zero CrATP-induced Ca(2+) occlusion due to defective CrATP binding.

  7. Effects of sex hormones on genioglossal muscle contractility and SR Ca2+-ATPase activity in aged rat.

    PubMed

    Liu, Yue-Hua; Qi, Juan; Hou, Yu-Xia; Wang, Fei

    2008-04-01

    This investigation was designed to examine whether short-term administration of sex hormones could produce changes in contractile properties and the sarcoplasmic reticulum Ca(2+)-ATPase (SERCA) function in genioglossal muscle (GG) of aged male rats. Twenty-four aged male rats were randomly divided into three groups to receive an intramuscular injection of either 0.1mg/kg oestrogen (group A), 2.5 mg/kg testosterone (group B), or 0.2 ml sterile peanut oil (group C, control), twice a week, during 4 weeks. After hormone treatment, in vivo isometric contractile properties were determined using surgically prepared GG muscles with platinum electrodes for stimulation of the medial branch of the hypoglossus nerve. Sarcoplasmic reticulum Ca(2+)-ATPase activity was measured in muscle homogenates by detecting the amount of inorganic phosphorus ion released in a standard coupled enzyme assay. SERCA1 mRNA level was observed using a real-time quantitative reverse transcription-polymerase chain reaction (Q-RT-PCR). It was found that, compared with group C, testosterone treatment decreased the fatigue resistance in GG muscles (p<.05), whereas no change was observed in the isometric twitch (P(t)) and tetanic tension (P(0)) (p>.05). By contrast, in oestrogen treated GG muscles, no significant modification was found either in the contractile or in endurance properties (p>.05). The change in GG fatigue resistance of group B was associated with a marked decrease in SR Ca(2+)-ATPase activity when compared with that of the control group (p<.01). Furthermore, SERCA1 mRNA level was also down regulated in group B (p<.05). No prominent differences in SR Ca(2+)-ATPase activity and SERCA1 mRNA expression existed between group A and group C (p>.05). The present results show that exogenous testosterone produces more pronounced changes in GG muscle fatigue resistance than oestrogen does by acting at SR Ca(2+)-ATPase activity and SERCA gene expression.

  8. Functional ryanodine receptor channels in flatworm muscle fibres.

    PubMed

    Day, T A; Haithcock, J; Kimber, M; Maule, A G

    2000-04-01

    Caffeine, which stimulates intracellular Ca2+ release channels known as ryanodine receptor (RyR) channels, induces contraction of individual muscle fibres dissociated from the trematode Schistosoma mansoni, and the turbellarians Dugesia tigrina and Procerodes littoralis. Caffeine is much more potent on S. mansoni fibres (EC50 0.7 mM) than those from D. tigrina or P. littoralis (3.2 mM and 4.6 mM, respectively). These caffeine-induced contractions are blocked by ryanodine, confirming the presence of functional RyR channels in these flatworm muscles. However, the contractions are not blocked by typical RyR channel blockers ruthenium red or neomycin, indicating that there may be important pharmacological differences between the RyR channels in this early-diverging phylum and those of later animals. These studies demonstrate that RyR channels are present in the muscle of these flatworms, and that the sarcoplasmic reticulum stores sufficient Ca2+ to support contraction.

  9. Muscle ultrastructural changes from exhaustive exercise performed after prolonged restricted activity and retraining in dogs

    NASA Technical Reports Server (NTRS)

    Nazar, K.; Greenleaf, J. E.; Philpott, D.; Pohoska, E.; Olszewska, K.; Kaciuba-Uscilko, H.

    1991-01-01

    The effect of exhaustive treadmill exercise on ultrastructural changes in the quadriceps femoris muscle was studied in 7 normal, healthy dogs, before and after restricted activity (RA), and following a subsequent 2 month treadmill exercise retraining period for the 5 mo group. Mean time to exhaustion in the 2 mo group decreased from 177 + or - 22 min before to 90 + or - 32 min after RA. Retraining increased tolerance to 219 + or - 73 min; 24 pct. above the before RA and 143 pct. above the after RA time. After RA exhaustion time in the 5 mo group was 25 and 45 min. Before RA, pre-exercise muscle structure was normal and post exercise there was only slight swelling of mitochondria. After RA, pre-exercise, numerous glycogen granules and lipid droplets appeared in the muscle fibers, mitochondria were smaller, and sarcoplasmic reticulum channels widened; post exercise these changes were accentuated and some areas were devoid of glycogen, and there was fiber degradation. After 5 mo RA pre-exercise there were more pronounced changes; mitochondria were very small and dense, there were many lipid droplets, myofibrils were often separated, and the fibers appeared edematous and degenerating; post exercise the sarcoplasmic reticulum was swollen, no glycogen was present, and there was marked swelling and deformation of mitochondria. After retraining, both pre-exercise and post exercise there was still evidence of fiber degeneration. Thus, susceptibility of active skeletal muscle structures and subcellular elements, e.g., mitochondria, to the action of damaging factors occurring during exhaustive exercise is enhanced considerably by prolonged disuse.

  10. Muscle strain (image)

    MedlinePlus

    A muscle strain is the stretching or tearing of muscle fibers. A muscle strain can be caused by sports, exercise, a ... something that is too heavy. Symptoms of a muscle strain include pain, tightness, swelling, tenderness, and the ...

  11. Survey of studies on how spaceflight affects rodent skeletal muscle.

    PubMed

    Fejtek, M B; Wassersug, R J

    1999-01-01

    Rodent muscles have been examined in more than 89 spaceflight studies over the last 25 years with much variation in the procedures and results. Mission duration ranged from four days to three weeks, postflight data collection ranged from a few hours to two days after landing, and there is great diversity in the number, size, and age of the rats that have flown. Several different types and sizes of animal enclosures have also been used--a significant factor because cage design affects animal activity and muscle loading. Only a small percentage (approximately 16%) of the total number of striated muscles in the rat have been examined. We have identified both substantial redundancy and inconsistencies in the results from studies to date. However, many of these appear unavoidable due to the great variation in experimental protocol of the different missions. Nevertheless these studies repeatedly confirm that exposure to spaceflight decreases the mass of limb muscles and leads to muscle atrophy. The majority of missions were flown by the former Soviet Union, but the majority of papers have been published by U.S. researchers. A relatively small number of investigators (about 50) clustered into fewer than 15 identifiable research groups worldwide account for most of the results to date. These groups have had access to rodent muscle tissue from two to seven spaceflights each. International cooperation in the post-cold war era and the publication of future work in peer-reviewed international journals should help greatly in reducing redundancy and enriching our knowledge of how gravity affects biological systems.

  12. Differentiation of pluripotent stem cells to muscle fiber to model Duchenne muscular dystrophy.

    PubMed

    Chal, Jérome; Oginuma, Masayuki; Al Tanoury, Ziad; Gobert, Bénédicte; Sumara, Olga; Hick, Aurore; Bousson, Fanny; Zidouni, Yasmine; Mursch, Caroline; Moncuquet, Philippe; Tassy, Olivier; Vincent, Stéphane; Miyanari, Ayako; Bera, Agata; Garnier, Jean-Marie; Guevara, Getzabel; Hestin, Marie; Kennedy, Leif; Hayashi, Shinichiro; Drayton, Bernadette; Cherrier, Thomas; Gayraud-Morel, Barbara; Gussoni, Emanuela; Relaix, Frédéric; Tajbakhsh, Shahragim; Pourquié, Olivier

    2015-09-01

    During embryonic development, skeletal muscles arise from somites, which derive from the presomitic mesoderm (PSM). Using PSM development as a guide, we establish conditions for the differentiation of monolayer cultures of mouse embryonic stem (ES) cells into PSM-like cells without the introduction of transgenes or cell sorting. We show that primary and secondary skeletal myogenesis can be recapitulated in vitro from the PSM-like cells, providing an efficient, serum-free protocol for the generation of striated, contractile fibers from mouse and human pluripotent cells. The mouse ES cells also differentiate into Pax7(+) cells with satellite cell characteristics, including the ability to form dystrophin(+) fibers when grafted into muscles of dystrophin-deficient mdx mice, a model of Duchenne muscular dystrophy (DMD). Fibers derived from ES cells of mdx mice exhibit an abnormal branched phenotype resembling that described in vivo, thus providing an attractive model to study the origin of the pathological defects associated with DMD. PMID:26237517

  13. Muscle intermediate filaments and their links to membranes and membranous organelles

    SciTech Connect

    Capetanaki, Yassemi . E-mail: ycapetanaki@bioacademy.gr; Bloch, Robert J.; Kouloumenta, Asimina; Mavroidis, Manolis; Psarras, Stelios

    2007-06-10

    Intermediate filaments (IFs) play a key role in the integration of structure and function of striated muscle, primarily by mediating mechanochemical links between the contractile apparatus and mitochondria, myonuclei, the sarcolemma and potentially the vesicle trafficking apparatus. Linkage of all these membranous structures to the contractile apparatus, mainly through the Z-disks, supports the integration and coordination of growth and energy demands of the working myocyte, not only with force transmission, but also with de novo gene expression, energy production and efficient protein and lipid trafficking and targeting. Desmin, the most abundant and intensively studied muscle intermediate filament protein, is linked to proper costamere organization, myoblast and stem cell fusion and differentiation, nuclear shape and positioning, as well as mitochondrial shape, structure, positioning and function. Similar links have been established for lysosomes and lysosome-related organelles, consistent with the presence of widespread links between IFs and membranous structures and the regulation of their fusion, morphology and stabilization necessary for cell survival.

  14. Isolation and characterization of the inositol trisphosphate receptor from smooth muscle

    SciTech Connect

    Chadwick, C.C.; Saito, A.; Fleischer, S. )

    1990-03-01

    The release of Ca{sup 2+} from internal stores is requisite to muscle contraction. In skeletal muscle and heart, the Ca{sup 2+} release channels (ryanodine receptor) of sarcoplasmic reticulum, involved in excitation-contraction coupling, have recently been isolated and characterized. In smooth muscle, inositol 1,4,5-trisphosphate (IP{sub 3}) is believed to mobilize Ca{sup 2+} from internal stores and thereby modulate contraction. The authors describe the isolation of an IP{sub 3} receptor from smooth muscle. Bovine aorta smooth muscle microsomes were solubilized with 3-((3-cholamidopropyl) dimethylammonio)-1-propanesulfonate, and the IP{sub 3} receptor was purified by sucrose gradient centrifugation and column chromatography with heparin-agarose and wheat germ agglutinin-agarose. The receptor is an oligomer of a single polypeptide with a M{sub r} of 224,000 as determined by SDS/PAGE. Negative-staining electron microscopy reveals that the receptor is a large pinwheel-like structure having surface dimensions of {approx}250 {times} 250 {angstrom} with fourfold symmetry. The IP{sub 3} receptor from smooth muscle is similar to the ryanodine receptor with regard to its large size and fourfold symmetry, albeit distinct with regard to appearance, protomer size, and ligand binding.

  15. [Congenital myopathies - skeletal muscle diseases related to disorder of actin filament structure and functions].

    PubMed

    Robaszkiewicz, Katarzyna; Moraczewska, Joanna

    2011-01-01

    Congenital myopathies are clinically and genetically heterogeneous disorders characterized by muscle structural abnormalities, muscle weakness and deformities. The clinical spectrum of the disease ranges from severe cases with early death to adult-onset cases with slow progression. In the skeletal muscle fibers, the specific structural changes are rod-shaped structures present in the sarcoplasm (nemaline myopathy – NM) or nuclei (intranuclear rod myopathy – IRM), cap-like structures peripherally located within muscle fibers (cap disease – CD), accumulations of actin filaments (actin myopathy – AM), changes in the fiber type proportion and size (congenital fiber type disproportion – CFTD), irregularity of Z-lines and abnormal localization of myofiber nuclei. Mutations in several genes encoding muscle proteins have been linked to congenital myopathy. These genes include a-skeletal actin (ACTA1), tropomyosin (TPM2 and TPM3), troponin (TNNT1) and nebulin (NEB). In vitro and in vivo studies show that mutations identified within these genes have varying impacts on thin filament protein structure, which affect polymerization and stabilization of actin filament, actin cellular localization and regulation of actin-myosin activity. Many lines of evidence suggest that mutated proteins have "toxic" effects. Unfortunately, there is no existing simple correlation between the degree of protein disruption, muscle pathologies and disease severity. PMID:21677359

  16. Effect of spaceflight on the functional, biochemical, and metabolic properties of skeletal muscle

    NASA Technical Reports Server (NTRS)

    Baldwin, K. M.

    1996-01-01

    This paper summarizes the effects of spaceflight on the functional, morphological, and biochemical properties of human and rodent skeletal muscle. The findings suggest that following as little as 5-6 in space there are deficits in both human and rodent motor capacity, strength, and endurance properties of skeletal muscle. The reduced strength is associated, in part, with a reduction in muscle mass as reflected in smaller cross-sectional areas of both fast- and slow-twitch fibers. Available evidence in animal models suggests that slow-twitch fibers are more sensitive to the atrophying process. Accompanying the atrophy is a transformation of slow to fast protein phenotype involving myosin heavy chain and sarcoplasmic reticulum protein isoforms. These transformations appear to be regulated, in part, by pretranslational processes. Data on the oxidative capacity of rodent skeletal muscle suggest a bias toward preferential utilization of carbohydrate as the primary substrate. These collective findings suggest that skeletal muscles comprised chiefly of slow fibers are highly dependent on gravity for the normal expression of protein mass and slow phenotype. Future studies need to focus on elucidating the mechanisms associated with the atrophy response, as well as identifying suitable exercise and other countermeasures capable of preserving the structural and functional integrity of skeletal muscle.

  17. Beta-Adrenoceptor Stimulation Reveals Ca2+ Waves and Sarcoplasmic Reticulum Ca2+ Depletion in Left Ventricular Cardiomyocytes from Post-Infarction Rats with and without Heart Failure.

    PubMed

    Sadredini, Mani; Danielsen, Tore Kristian; Aronsen, Jan Magnus; Manotheepan, Ravinea; Hougen, Karina; Sjaastad, Ivar; Stokke, Mathis Korseberg

    2016-01-01

    Abnormal cellular Ca2+ handling contributes to both contractile dysfunction and arrhythmias in heart failure. Reduced Ca2+ transient amplitude due to decreased sarcoplasmic reticulum Ca2+ content is a common finding in heart failure models. However, heart failure models also show increased propensity for diastolic Ca2+ release events which occur when sarcoplasmic reticulum Ca2+ content exceeds a certain threshold level. Such Ca2+ release events can initiate arrhythmias. In this study we aimed to investigate if both of these aspects of altered Ca2+ homeostasis could be found in left ventricular cardiomyocytes from rats with different states of cardiac function six weeks after myocardial infarction when compared to sham-operated controls. Video edge-detection, whole-cell Ca2+ imaging and confocal line-scan imaging were used to investigate cardiomyocyte contractile properties, Ca2+ transients and Ca2+ waves. In baseline conditions, i.e. without beta-adrenoceptor stimulation, cardiomyocytes from rats with large myocardial infarction, but without heart failure, did not differ from sham-operated animals in any of these aspects of cellular function. However, when exposed to beta-adrenoceptor stimulation, cardiomyocytes from both non-failing and failing rat hearts showed decreased sarcoplasmic reticulum Ca2+ content, decreased Ca2+ transient amplitude, and increased frequency of Ca2+ waves. These results are in line with a decreased threshold for diastolic Ca2+ release established by other studies. In the present study, factors that might contribute to a lower threshold for diastolic Ca2+ release were increased THR286 phosphorylation of Ca2+/calmodulin-dependent protein kinase II and increased protein phosphatase 1 abundance. In conclusion, this study demonstrates both decreased sarcoplasmic reticulum Ca2+ content and increased propensity for diastolic Ca2+ release events in ventricular cardiomyocytes from rats with heart failure after myocardial infarction, and that these

  18. Excitation-contraction coupling in zebrafish ventricular myocardium is regulated by trans-sarcolemmal Ca2+ influx and sarcoplasmic reticulum Ca2+ release.

    PubMed

    Haustein, Moritz; Hannes, Tobias; Trieschmann, Jan; Verhaegh, Rabea; Köster, Annette; Hescheler, Jürgen; Brockmeier, Konrad; Adelmann, Roland; Khalil, Markus

    2015-01-01

    Zebrafish (Danio rerio) have become a popular model in cardiovascular research mainly due to identification of a large number of mutants with structural defects. In recent years, cardiomyopathies and other diseases influencing contractility of the heart have been studied in zebrafish mutants. However, little is known about the regulation of contractility of the zebrafish heart on a tissue level. The aim of the present study was to elucidate the role of trans-sarcolemmal Ca(2+)-flux and sarcoplasmic reticulum Ca(2+)-release in zebrafish myocardium. Using isometric force measurements of fresh heart slices, we characterised the effects of changes of the extracellular Ca(2+)-concentration, trans-sarcolemmal Ca(2+)-flux via L-type Ca(2+)-channels and Na(+)-Ca(2+)-exchanger, and Ca(2+)-release from the sarcoplasmic reticulum as well as beating frequency and β-adrenergic stimulation on contractility of adult zebrafish myocardium. We found an overall negative force-frequency relationship (FFR). Inhibition of L-type Ca(2+)-channels by verapamil (1 μM) decreased force of contraction to 22 ± 7% compared to baseline (n=4, p<0.05). Ni(2+) was the only substance to prolong relaxation (5 mM, time after peak to 50% relaxation: 73 ± 3 ms vs. 101 ± 8 ms, n=5, p<0.05). Surprisingly though, inhibition of the sarcoplasmic Ca(2+)-release decreased force development to 54 ± 3% in ventricular (n=13, p<0.05) and to 52 ± 8% in atrial myocardium (n=5, p<0.05) suggesting a substantial role of SR Ca(2+)-release in force generation. In line with this finding, we observed significant post pause potentiation after pauses of 5 s (169 ± 7% force compared to baseline, n=8, p<0.05) and 10 s (198 ± 9% force compared to baseline, n=5, p<0.05) and mildly positive lusitropy after β-adrenergic stimulation. In conclusion, force development in adult zebrafish ventricular myocardium requires not only trans-sarcolemmal Ca2+-flux, but also intact sarcoplasmic reticulum Ca(2+)-cycling. In contrast to

  19. Beta-Adrenoceptor Stimulation Reveals Ca2+ Waves and Sarcoplasmic Reticulum Ca2+ Depletion in Left Ventricular Cardiomyocytes from Post-Infarction Rats with and without Heart Failure.

    PubMed

    Sadredini, Mani; Danielsen, Tore Kristian; Aronsen, Jan Magnus; Manotheepan, Ravinea; Hougen, Karina; Sjaastad, Ivar; Stokke, Mathis Korseberg

    2016-01-01

    Abnormal cellular Ca2+ handling contributes to both contractile dysfunction and arrhythmias in heart failure. Reduced Ca2+ transient amplitude due to decreased sarcoplasmic reticulum Ca2+ content is a common finding in heart failure models. However, heart failure models also show increased propensity for diastolic Ca2+ release events which occur when sarcoplasmic reticulum Ca2+ content exceeds a certain threshold level. Such Ca2+ release events can initiate arrhythmias. In this study we aimed to investigate if both of these aspects of altered Ca2+ homeostasis could be found in left ventricular cardiomyocytes from rats with different states of cardiac function six weeks after myocardial infarction when compared to sham-operated controls. Video edge-detection, whole-cell Ca2+ imaging and confocal line-scan imaging were used to investigate cardiomyocyte contractile properties, Ca2+ transients and Ca2+ waves. In baseline conditions, i.e. without beta-adrenoceptor stimulation, cardiomyocytes from rats with large myocardial infarction, but without heart failure, did not differ from sham-operated animals in any of these aspects of cellular function. However, when exposed to beta-adrenoceptor stimulation, cardiomyocytes from both non-failing and failing rat hearts showed decreased sarcoplasmic reticulum Ca2+ content, decreased Ca2+ transient amplitude, and increased frequency of Ca2+ waves. These results are in line with a decreased threshold for diastolic Ca2+ release established by other studies. In the present study, factors that might contribute to a lower threshold for diastolic Ca2+ release were increased THR286 phosphorylation of Ca2+/calmodulin-dependent protein kinase II and increased protein phosphatase 1 abundance. In conclusion, this study demonstrates both decreased sarcoplasmic reticulum Ca2+ content and increased propensity for diastolic Ca2+ release events in ventricular cardiomyocytes from rats with heart failure after myocardial infarction, and that these

  20. Beta-Adrenoceptor Stimulation Reveals Ca2+ Waves and Sarcoplasmic Reticulum Ca2+ Depletion in Left Ventricular Cardiomyocytes from Post-Infarction Rats with and without Heart Failure

    PubMed Central

    Danielsen, Tore Kristian; Aronsen, Jan Magnus; Manotheepan, Ravinea; Hougen, Karina; Sjaastad, Ivar; Stokke, Mathis Korseberg

    2016-01-01

    Abnormal cellular Ca2+ handling contributes to both contractile dysfunction and arrhythmias in heart failure. Reduced Ca2+ transient amplitude due to decreased sarcoplasmic reticulum Ca2+ content is a common finding in heart failure models. However, heart failure models also show increased propensity for diastolic Ca2+ release events which occur when sarcoplasmic reticulum Ca2+ content exceeds a certain threshold level. Such Ca2+ release events can initiate arrhythmias. In this study we aimed to investigate if both of these aspects of altered Ca2+ homeostasis could be found in left ventricular cardiomyocytes from rats with different states of cardiac function six weeks after myocardial infarction when compared to sham-operated controls. Video edge-detection, whole-cell Ca2+ imaging and confocal line-scan imaging were used to investigate cardiomyocyte contractile properties, Ca2+ transients and Ca2+ waves. In baseline conditions, i.e. without beta-adrenoceptor stimulation, cardiomyocytes from rats with large myocardial infarction, but without heart failure, did not differ from sham-operated animals in any of these aspects of cellular function. However, when exposed to beta-adrenoceptor stimulation, cardiomyocytes from both non-failing and failing rat hearts showed decreased sarcoplasmic reticulum Ca2+ content, decreased Ca2+ transient amplitude, and increased frequency of Ca2+ waves. These results are in line with a decreased threshold for diastolic Ca2+ release established by other studies. In the present study, factors that might contribute to a lower threshold for diastolic Ca2+ release were increased THR286 phosphorylation of Ca2+/calmodulin-dependent protein kinase II and increased protein phosphatase 1 abundance. In conclusion, this study demonstrates both decreased sarcoplasmic reticulum Ca2+ content and increased propensity for diastolic Ca2+ release events in ventricular cardiomyocytes from rats with heart failure after myocardial infarction, and that these

  1. Excitation-Contraction Coupling in Zebrafish Ventricular Myocardium Is Regulated by Trans-Sarcolemmal Ca2+ Influx and Sarcoplasmic Reticulum Ca2+ Release

    PubMed Central

    Trieschmann, Jan; Verhaegh, Rabea; Köster, Annette; Hescheler, Jürgen; Brockmeier, Konrad; Adelmann, Roland; Khalil, Markus

    2015-01-01

    Zebrafish (Danio rerio) have become a popular model in cardiovascular research mainly due to identification of a large number of mutants with structural defects. In recent years, cardiomyopathies and other diseases influencing contractility of the heart have been studied in zebrafish mutants. However, little is known about the regulation of contractility of the zebrafish heart on a tissue level. The aim of the present study was to elucidate the role of trans-sarcolemmal Ca2+-flux and sarcoplasmic reticulum Ca2+-release in zebrafish myocardium. Using isometric force measurements of fresh heart slices, we characterised the effects of changes of the extracellular Ca2+-concentration, trans-sarcolemmal Ca2+-flux via L-type Ca2+-channels and Na+-Ca2+-exchanger, and Ca2+-release from the sarcoplasmic reticulum as well as beating frequency and β-adrenergic stimulation on contractility of adult zebrafish myocardium. We found an overall negative force-frequency relationship (FFR). Inhibition of L-type Ca2+-channels by verapamil (1 μM) decreased force of contraction to 22±7% compared to baseline (n=4, p<0.05). Ni2+ was the only substance to prolong relaxation (5 mM, time after peak to 50% relaxation: 73±3 ms vs. 101±8 ms, n=5, p<0.05). Surprisingly though, inhibition of the sarcoplasmic Ca2+-release decreased force development to 54±3% in ventricular (n=13, p<0.05) and to 52±8% in atrial myocardium (n=5, p<0.05) suggesting a substantial role of SR Ca2+-release in force generation. In line with this finding, we observed significant post pause potentiation after pauses of 5 s (169±7% force compared to baseline, n=8, p<0.05) and 10 s (198±9% force compared to baseline, n=5, p<0.05) and mildly positive lusitropy after β-adrenergic stimulation. In conclusion, force development in adult zebrafish ventricular myocardium requires not only trans-sarcolemmal Ca2+-flux, but also intact sarcoplasmic reticulum Ca2+-cycling. In contrast to mammals, FFR is strongly negative in the

  2. Mutations in either the essential or regulatory light chains of myosin are associated with a rare myopathy in human heart and skeletal muscle.

    PubMed

    Poetter, K; Jiang, H; Hassanzadeh, S; Master, S R; Chang, A; Dalakas, M C; Rayment, I; Sellers, J R; Fananapazir, L; Epstein, N D

    1996-05-01

    The muscle myosins and hexomeric proteins consisting of two heavy chains and two pairs of light chains, the latter called essential (ELC) and regulatory (RLC). The light chains stabilize the long alpha helical neck of the myosin head. Their function in striated muscle, however, is only partially understood. We report here the identification of distinct missense mutations in a skeletal/ventricular ELC and RLC, each of which are associated with a rare variant of cardiac hypertrophy as well as abnormal skeletal muscle. We show that myosin containing the mutant ELC has abnormal function, map the mutant residues on the three-dimensional structure of myosin and suggest that the mutations disrupt the stretch activation response of the cardiac papillary muscles.

  3. 2,3,6-triaminopyridine, a metabolite of the urinary tract analgesic phenazopyridine, causes muscle necrosis and renal damage in rats.

    PubMed

    Munday, R; Manns, E

    1998-01-01

    Some aromatic polyamines form very stable free radicals and readily undergo autoxidation with concomitant formation of 'active oxygen' species. These substances cause necrosis of striated muscle in rats, and it has been suggested that this is due to free radical formation and disruption of energy production through their oxidation via the cytochrome c/cytochrome oxidase system of mitochondria. 2,3,6-Triaminopyridine, which is structurally related to the myotoxic amines and likewise undergoes autoxidation and disrupts mitochondrial metabolism, is a metabolite of the widely used urinary analgesic phenazopyridine. When administered to rats, triaminopyridine caused extensive necrosis of skeletal muscle and a lesser degree of damage to heart muscle. It also induced vacuolation and necrosis of distal tubules of the kidney, associated with tubular dilatation and cast formation. Both muscle damage and renal tubular necrosis have been reported following use or abuse of phenazopyridine, and it is likely that triaminopyridine is responsible for both of these effects.

  4. Molecular analysis of the muscle protein projectin in Lepidoptera.

    PubMed

    Ayme-Southgate, A J; Turner, L; Southgate, R J

    2013-01-01

    Striated muscles of both vertebrates and insects contain a third filament composed of the giant proteins, namely kettin and projectin (insects) and titin (vertebrates). All three proteins have been shown to contain several domains implicated in conferring elasticity, in particular a PEVK segment. In this study, the characterization of the projectin protein in the silkmoth, Bombyx mori L. (Lepidoptera: Bombycidae), and the monarch butterfly, Danaus plexippus L. (Lepidoptera: Nymphalidae), as well as a partial characterization in the Carolina sphinx, Manduca sexta L. (Lepidoptera: Sphingidae), are presented. This study showed that, similar to other insects, projectin's overall modular organization was conserved, but in contrast, the PEVK region had a highly divergent sequence. The analysis of alternative splicing in the PEVK region revealed a small number of possible isoforms and the lack of a flight-muscle specific variant, both characteristics being in sharp contrast with findings from other insects. The possible correlation with difference in flight muscle stiffness and physiology between Lepidoptera and other insect orders is discussed.

  5. Molecular analysis of the muscle protein projectin in Lepidoptera.

    PubMed

    Ayme-Southgate, A J; Turner, L; Southgate, R J

    2013-01-01

    Striated muscles of both vertebrates and insects contain a third filament composed of the giant proteins, namely kettin and projectin (insects) and titin (vertebrates). All three proteins have been shown to contain several domains implicated in conferring elasticity, in particular a PEVK segment. In this study, the characterization of the projectin protein in the silkmoth, Bombyx mori L. (Lepidoptera: Bombycidae), and the monarch butterfly, Danaus plexippus L. (Lepidoptera: Nymphalidae), as well as a partial characterization in the Carolina sphinx, Manduca sexta L. (Lepidoptera: Sphingidae), are presented. This study showed that, similar to other insects, projectin's overall modular organization was conserved, but in contrast, the PEVK region had a highly divergent sequence. The analysis of alternative splicing in the PEVK region revealed a small number of possible isoforms and the lack of a flight-muscle specific variant, both characteristics being in sharp contrast with findings from other insects. The possible correlation with difference in flight muscle stiffness and physiology between Lepidoptera and other insect orders is discussed. PMID:24206568

  6. Motor activity and muscle properties in the hemidecerebellate cat.

    PubMed

    Stenvers, J W; Eerbeek, O; de Jong, J M; Meijer, A E

    1983-09-01

    Luciani's ipsilaterally acting 'trophic' cerebellar influence on striated muscle was reinvestigated in hemidecerebellate preparations of varying extent. Cats with hindlimb postural asymmetries for 4 or more days after the lesion developed a bilateral reduction of maximum tetanic tension and increased twitch/tetanus ratios of soleus. In addition, soleus on the side of the lesion lost force and weight, showed decreased twitch contraction and half-relaxation times, elevated myosin ATPase activity in part of its fibres, occasional fibre necrosis and a few snake coils. Protracted postural asymmetry occurred only if complete hemicerebellectomy included ablation of the lateral vestibular nucleus and extended for at least 3.0 mm across the midline into the contralateral vermal and intermediate cortex, especially of Larsell's lobuli IV and V. Most simply, the cerebellar effect on muscle is explained as the result of altered motoneuronal activation patterns. Comparison of the experimental soleus changes with Holmes's clinical findings in cases of cerebellar injury suggests that muscle itself participates in experimental and human cerebellar asthenia.

  7. Severe protein aggregate myopathy in a knockout mouse model points to an essential role of cofilin2 in sarcomeric actin exchange and muscle maintenance.

    PubMed

    Gurniak, Christine B; Chevessier, Frédéric; Jokwitz, Melanie; Jönsson, Friederike; Perlas, Emerald; Richter, Hendrik; Matern, Gabi; Boyl, Pietro Pilo; Chaponnier, Christine; Fürst, Dieter; Schröder, Rolf; Witke, Walter

    2014-01-01

    Mutations in the human actin depolymerizing factor cofilin2 result in an autosomal dominant form of nemaline myopathy. Here, we report on the targeted ablation of murine cofilin2, which leads to a severe skeletal muscle specific phenotype within the first two weeks after birth. Apart from skeletal muscle, cofilin2 is also expressed in heart and CNS, however the pathology was restricted to skeletal muscle. The two close family members of cofilin2 - ADF and cofilin1 - were co-expressed in muscle, but unable to compensate for the loss of cofilin2. While primary myofibril assembly and muscle development were unaffected in cofilin2 mutant mice, progressive muscle degeneration was observed between postnatal days 3 and 7. Muscle pathology was characterized by sarcoplasmic protein aggregates, fiber size disproportion, mitochondrial abnormalities and internal nuclei. The observed muscle pathology differed from nemaline myopathy, but showed combined features of actin-associated myopathy and myofibrillar myopathy. In cofilin2 mutant mice, the postnatal expression pattern and turnover of sarcomeric α-actin isoforms were altered. Levels of smooth muscle α-actin were increased and remained high in developing muscles, suggesting that cofilin2 plays a crucial role during the exchange of α-actin isoforms during the early postnatal remodeling of the sarcomere. PMID:24598388

  8. cap alpha. -skeletal and. cap alpha. -cardiac actin genes are coexpressed in adult human skeletal muscle and heart

    SciTech Connect

    Gunning, P.; Ponte, P.; Blau, H.; Kedes, L.

    1983-11-01

    The authors determined the actin isotypes encoded by 30 actin cDNA clones previously isolated from an adult human muscle cDNA library. Using 3' untranslated region probes, derived from ..cap alpha.. skeletal, ..beta..- and ..gamma..-actin cDNAs and from an ..cap alpha..-cardiac actin genomic clone, they showed that 28 of the cDNAs correspond to ..cap alpha..-skeletal actin transcripts. Unexpectedly, however, the remaining two cDNA clones proved to derive from ..cap alpha..-cardiac actin mRNA. Sequence analysis confirmed that the two skeletal muscle ..cap alpha..-cardiac actin cDNAs are derived from transcripts of the cloned ..cap alpha..-cardiac actin gene. Comparison of total actin mRNA levels in adult skeletal muscle and adult heart revealed that the steady-state levels in skeletal muscle are about twofold greater, per microgram of total cellular RNA, than those in heart. Thus, in skeletal muscle and in heart, both of the sarcomeric actin mRNA isotypes are quite abundant transcripts. They conclude that ..cap alpha..-skeletal and ..cap alpha..-cardiac actin genes are coexpressed as an actin pair in human adult striated muscles. Since the smooth-muscle actins (aortic and stomach) and the cytoplasmic actins (..beta.. and ..gamma..) are known to be coexpressed in smooth muscle and nonmuscle cells, respectively, they postulate that coexpression of actin pairs may be a common feature of mammalian actin gene expression in all tissues.

  9. Pathogenesis of coxsackie virus infection; multiplication of virus and evolution of the muscle lesion in mice.

    PubMed

    MELNICK, J L; GODMAN, G C

    1951-03-01

    The quantitative distribution of the Conn.-5 strain of Coxsackie virus in different tissues was determined by serial titration at intervals after inoculation of 4 to 5 day old mice. High titers were reached by the 2nd day in blood, heart, liver, muscle, intestine, and its contents, and these were maintained through the 8th day, except for the blood, in which the virus level fell earlier. In paralyzed mice, muscle and brain attained the highest titers and it was in these tissues alone that virus persisted through the 9th day of illness. The pathology of the infection has been briefly described. In particular, the evolution of morbid changes in striated muscle was correlated with the concentrations of virus in muscle. Acute muscle necrosis first occurred when there was a peak viral concentration (4th day), and reached maximal intensity on the 8th day. Scattered acute lesions continued to appear while the virus titer remained above 10(-4), from the 9th to 12th day. With the decrease in the myositis, there was a concomitant decrease in the incidence of perceptible disease. Inflammation was found to follow upon the development of necrosis, and subsided slowly. Regeneration began very early, became exuberant, and led finally to restitution of the muscle.

  10. On the footsteps of Triadin and its role in skeletal muscle

    PubMed Central

    Perez, Claudio F

    2011-01-01

    Calcium is a crucial element for striated muscle function. As such, myoplasmic free Ca2+ concentration is delicately regulated through the concerted action of multiple Ca2+ pathways that relay excitation of the plasma membrane to the intracellular contractile machinery. In skeletal muscle, one of these major Ca2+ pathways is Ca2+ release from intracellular Ca2+ stores through type-1 ryanodine receptor/Ca2+ release channels (RyR1), which positions RyR1 in a strategic cross point to regulate Ca2+ homeostasis. This major Ca2+ traffic point appears to be highly sensitive to the intracellular environment, which senses through a plethora of chemical and protein-protein interactions. Among these modulators, perhaps one of the most elusive is Triadin, a muscle-specific protein that is involved in many crucial aspect of muscle function. This family of proteins mediates complex interactions with various Ca2+ modulators and seems poised to be a relevant modulator of Ca2+ signaling in cardiac and skeletal muscles. The purpose of this review is to examine the most recent evidence and current understanding of the role of Triadin in muscle function, in general, with particular emphasis on its contribution to Ca2+ homeostasis. PMID:21909459

  11. Pleomorphic rhabdomyosarcoma showing smooth-muscle and fibrohistiocytic differentiation: a single case report.

    PubMed

    Eyden, Brian

    2010-02-01

    Rhabdomyosarcoma has traditionally been subclassified into alveolar, embryonal, and pleomorphic variants. Less commonly, spindle-cell, neuroendocrine, sclerosing, and lipid-rich or clear-cell subtypes are seen. The author recently encountered a myogenic sarcoma, with all the common markers of rhabdomyosarcoma, but expressing the unusual features of alpha-smooth-muscle actin and abundant rough endoplasmic reticulum (rER). This myogenic sarcoma, therefore, exhibited four lines of differentiation, and is documented here. The patient was a 65-year-old man with an inguinal soft tissue mass. Following surgical excision, the patient was given radiotherapy and was well without disease after 6 years. The tumor was positive for vimentin, desmin, alpha-smooth-muscle actin, alpha-sarcomeric actin, myogenin, MyoD1, and CD68. Cytoplasm was dominated by abundant rER intermingled with lipid droplets and lysosomes. Cell surfaces exhibited microvillous processes and focal adhesions, but no lamina. Subplasmalemmal smooth-muscle-type myofilaments with focal densities and rare sarcomeric filaments were seen. The low level of expression of some markers was interpreted as consistent with a poorly differentiated tumor. Given the four lines of differentiation--striated muscle, smooth muscle, fibroblastic, and histiocytic--a name reflecting its phenotype would be pleomorphic rhabdomyosarcoma showing smooth-muscle and fibrohistiocytic differentiation. PMID:20070153

  12. Differences in the Expression and Distribution of Flotillin-2 in Chick, Mice and Human Muscle Cells

    PubMed Central

    Possidonio, Ana Claudia Batista; Soares, Carolina Pontes; Portilho, Débora Morueco; Midlej, Victor; Benchimol, Marlene; Butler-Browne, Gillian; Costa, Manoel Luis; Mermelstein, Claudia

    2014-01-01

    Myoblasts undergo a series of changes in the composition and dynamics of their plasma membranes during the initial steps of skeletal muscle differentiation. These changes are crucial requirements for myoblast fusion and allow the formation of striated muscle fibers. Membrane microdomains, or lipid rafts, have been implicated in myoblast fusion. Flotillins are scaffold proteins that are essential for the formation and dynamics of lipid rafts. Flotillins have been widely studied over the last few years, but still little is known about their role during skeletal muscle differentiation. In the present study, we analyzed the expression and distribution of flotillin-2 in chick, mice and human muscle cells grown in vitro. Primary cultures of chick myogenic cells showed a decrease in the expression of flotillin-2 during the first 72 hours of muscle differentiation. Interestingly, flotillin-2 was found to be highly expressed in chick myogenic fibroblasts and weakly expressed in chick myoblasts and multinucleated myotubes. Flotillin-2 was distributed in vesicle-like structures within the cytoplasm of chick myogenic fibroblasts, in the mouse C2C12 myogenic cell line, and in neonatal human muscle cells. Cryo-immunogold labeling revealed the presence of flotillin-2 in vesicles and in Golgi stacks in chick myogenic fibroblasts. Further, brefeldin A induced a major reduction in the number of flotillin-2 containing vesicles which correlates to a decrease in myoblast fusion. These results suggest the involvement of flotillin-2 during the initial steps of skeletal myogenesis. PMID:25105415

  13. Differences in the expression and distribution of flotillin-2 in chick, mice and human muscle cells.

    PubMed

    Possidonio, Ana Claudia Batista; Soares, Carolina Pontes; Portilho, Débora Morueco; Midlej, Victor; Benchimol, Marlene; Butler-Browne, Gillian; Costa, Manoel Luis; Mermelstein, Claudia

    2014-01-01

    Myoblasts undergo a series of changes in the composition and dynamics of their plasma membranes during the initial steps of skeletal muscle differentiation. These changes are crucial requirements for myoblast fusion and allow the formation of striated muscle fibers. Membrane microdomains, or lipid rafts, have been implicated in myoblast fusion. Flotillins are scaffold proteins that are essential for the formation and dynamics of lipid rafts. Flotillins have been widely studied over the last few years, but still little is known about their role during skeletal muscle differentiation. In the present study, we analyzed the expression and distribution of flotillin-2 in chick, mice and human muscle cells grown in vitro. Primary cultures of chick myogenic cells showed a decrease in the expression of flotillin-2 during the first 72 hours of muscle differentiation. Interestingly, flotillin-2 was found to be highly expressed in chick myogenic fibroblasts and weakly expressed in chick myoblasts and multinucleated myotubes. Flotillin-2 was distributed in vesicle-like structures within the cytoplasm of chick myogenic fibroblasts, in the mouse C2C12 myogenic cell line, and in neonatal human muscle cells. Cryo-immunogold labeling revealed the presence of flotillin-2 in vesicles and in Golgi stacks in chick myogenic fibroblasts. Further, brefeldin A induced a major reduction in the number of flotillin-2 containing vesicles which correlates to a decrease in myoblast fusion. These results suggest the involvement of flotillin-2 during the initial steps of skeletal myogenesis.

  14. Neural control of the female urethral and anal rhabdosphincters and pelvic floor muscles

    PubMed Central

    de Groat, William C.

    2010-01-01

    The urethral rhabdosphincter and pelvic floor muscles are important in maintenance of urinary continence and in preventing descent of pelvic organs [i.e., pelvic organ prolapse (POP)]. Despite its clinical importance and complexity, a comprehensive review of neural control of the rhabdosphincter and pelvic floor muscles is lacking. The present review places historical and recent basic science findings on neural control into the context of functional anatomy of the pelvic muscles and their coordination with visceral function and correlates basic science findings with clinical findings when possible. This review briefly describes the striated muscles of the pelvis and then provides details on the peripheral innervation and, in particular, the contributions of the pudendal and levator ani nerves to the function of the various pelvic muscles. The locations and unique phenotypic characteristics of rhabdosphincter motor neurons located in Onuf's nucleus, and levator ani motor neurons located diffusely in the sacral ventral horn, are provided along with the locations and phenotypes of primary afferent neurons that convey sensory information from these muscles. Spinal and supraspinal pathways mediating excitatory and inhibitory inputs to the motor neurons are described; the relative contributions of the nerves to urethral function and their involvement in POP and incontinence are discussed. Finally, a detailed summary of the neurochemical anatomy of Onuf's nucleus and the pharmacological control of the rhabdosphincter are provided. PMID:20484700

  15. Manganese depresses rat heart muscle respiration.

    PubMed

    Miller, Kevin B; Caton, Joel S; Finley, John W

    2006-01-01

    It has previously been reported that moderately high dietary manganese (Mn) in combination with marginal magnesium (Mg) resulted in ultrastructural damage to heart mitochondria. Manganese may replace Mg in biological functions, including the role of enzyme cofactor. Manganese may accumulate and substitute for Mg during the condition of Mg-deficiency. The objective of the current study was to determine whether high Mn alters heart muscle respiration and Mg-enzyme activity as well as whole body Mn retention under marginal Mg. An additional objective was to determine whether high Mn results in increased oxidative stress. In experiment 1: forty-eight rats were fed a 2 x 3 factorial arrangement of Mn (10, 100, or 1000 mg/kg) and Mg (200 or 500 mg/kg). In experiment 2: thirty-two rats were fed one of four diets in a 2 x 2 factorial arrangement of Mn (10 or 250 mg/kg) and Mg (200 or 500 mg/kg). In experiment 3: thirty-two rats were fed one of four diets in a 2 x 2 factorial arrangement of Mn (10 or 650 mg/kg) and Mg (200 or 500 mg/kg). In experiment 2, high Mn and marginal Mg reduced (P<0.05) oxygen consumption of left ventricle muscle. Marginal Mg, but not Mn, reduced (P<0.05) activity of sarcoplasmic reticulum calcium-ATPase enzyme. Dietary Mg had no affect on (54)Mn kinetics, but high dietary Mn decreased (P<0.01) absorption, retention, and rate of excretion of (54)Mn. Neither cellular stress, measured by Comet assay, nor antioxidant activities were increased by high Mn. A strong interaction (P<0.001) between increasing Mn and adequate Mg on hematology was observed. These results confirm previous research in swine that high Mn alters myocardial integrity as well as function, but not as a result of altered calcium transport or oxidative stress.

  16. Capillary muscle

    PubMed Central

    Cohen, Caroline; Mouterde, Timothée; Quéré, David; Clanet, Christophe

    2015-01-01

    The contraction of a muscle generates a force that decreases when increasing the contraction velocity. This “hyperbolic” force–velocity relationship has been known since the seminal work of A. V. Hill in 1938 [Hill AV (1938) Proc R Soc Lond B Biol Sci 126(843):136–195]. Hill’s heuristic equation is still used, and the sliding-filament theory for the sarcomere [Huxley H, Hanson J (1954) Nature 173(4412):973–976; Huxley AF, Niedergerke R (1954) Nature 173(4412):971–973] suggested how its different parameters can be related to the molecular origin of the force generator [Huxley AF (1957) Prog Biophys Biophys Chem 7:255–318; Deshcherevskiĭ VI (1968) Biofizika 13(5):928–935]. Here, we develop a capillary analog of the sarcomere obeying Hill’s equation and discuss its analogy with muscles. PMID:25944938

  17. Regulation of phosphorylase kinase by low concentrations of Ca ions upon muscle contraction: the connection between metabolism and muscle contraction and the connection between muscle physiology and Ca-dependent signal transduction

    PubMed Central

    OZAWA, Eijiro

    2011-01-01

    It had long been one of the crucial questions in muscle physiology how glycogenolysis is regulated in connection with muscle contraction, when we found the answer to this question in the last half of the 1960s. By that time, the two principal currents of muscle physiology, namely, the metabolic flow starting from glycogen and the mechanisms of muscle contraction, had already been clarified at the molecular level thanks to our senior researchers. Thus, the final question we had to answer was how to connect these two currents. We found that low concentrations of Ca ions (10−7–10−4 M) released from the sarcoplasmic reticulum for the regulation of muscle contraction simultaneously reversibly activate phosphorylase kinase, the enzyme regulating glycogenolysis. Moreover, we found that adenosine 3′,5′-monophosphate (cyclic AMP), which is already known to activate muscle phosphorylase kinase, is not effective in the absence of such concentrations of Ca ions. Thus, cyclic AMP is not effective by itself alone and only modifies the activation process in the presence of Ca ions (at that time, cyclic AMP-dependent protein kinase had not yet been identified). After a while, it turned out that our works have not only provided the solution to the above problem on muscle physiology, but have also been considered as the first report of Ca-dependent protein phosphorylation, which is one of the central problems in current cell biology. Phosphorylase kinase is the first protein kinase to phosphorylate a protein resulting in the change in the function of the phosphorylated protein, as shown by Krebs and Fischer. Our works further showed that this protein kinase is regulated in a Ca-dependent manner. Accordingly, our works introduced the concept of low concentrations of Ca ions, which were first identified as the regulatory substance of muscle contraction, to the vast field of Ca biology including signal transduction. PMID:21986313

  18. Alterations of cAMP-dependent signaling in dystrophic skeletal muscle

    PubMed Central

    Rudolf, Rüdiger; Khan, Muzamil M.; Lustrino, Danilo; Labeit, Siegfried; Kettelhut, Ísis C.; Navegantes, Luiz C. C.

    2013-01-01

    Autonomic regulation processes in striated muscles are largely mediated by cAMP/PKA-signaling. In order to achieve specificity of signaling its spatial-temporal compartmentation plays a critical role. We discuss here how specificity of cAMP/PKA-signaling can be achieved in skeletal muscle by spatio-temporal compartmentation. While a microdomain containing PKA type I in the region of the neuromuscular junction (NMJ) is important for postsynaptic, activity-dependent stabilization of the nicotinic acetylcholine receptor (AChR), PKA type I and II microdomains in the sarcomeric part of skeletal muscle are likely to play different roles, including the regulation of muscle homeostasis. These microdomains are due to specific A-kinase anchoring proteins, like rapsyn and myospryn. Importantly, recent evidence indicates that compartmentation of the cAMP/PKA-dependent signaling pathway and pharmacological activation of cAMP production are aberrant in different skeletal muscles disorders. Thus, we discuss here their potential as targets for palliative treatment of certain forms of dystrophy and myasthenia. Under physiological conditions, the neuropeptide, α-calcitonin-related peptide, as well as catecholamines are the most-mentioned natural triggers for activating cAMP/PKA signaling in skeletal muscle. While the precise domains and functions of these first messengers are still under investigation, agonists of β2-adrenoceptors clearly exhibit anabolic activity under normal conditions and reduce protein degradation during atrophic periods. Past and recent studies suggest direct sympathetic innervation of skeletal muscle fibers. In summary, the organization and roles of cAMP-dependent signaling in skeletal muscle are increasingly understood, revealing crucial functions in processes like nerve-muscle interaction and muscle trophicity. PMID:24146652

  19. Phosphorylation-dependent changes in the spatial relationship between Ca-ATPase polypeptide chains in sarcoplasmic reticulum membranes.

    PubMed

    Bigelow, D J; Squier, T C; Inesi, G

    1992-04-01

    In order to investigate possible structural changes associated with the coupling mechanisms of the Ca-ATPase in sarcoplasmic reticulum membranes, we have utilized fluorescence resonance energy transfer between spectroscopic probes covalently bound to different domains of the ATPase. Using time-correlated single photon counting, we have directly measured the energy transfer efficiency between 5-[2-[(iodoacetyl)amino]ethyl]aminonaphthalene-1-sulfonic acid (IAEDANS), that is specifically bound to the B trypic fragment at cysteines 670 and 674 and acceptors covalently bound either near the nucleotide binding site, i.e. fluorescein 5-isothiocyanate at lysine 515, also on the B fragment, or maleimide-directed probes specifically located on the A1, tryptic fragment, i.e. 4-dimethylaminoazobenzene-4'-maleimide (DABmal) or fluorescein-5-maleimide (Fmal), probably at cysteines 344 and 364. All of these donor-acceptor pairs exhibit energy transfer both within and between Ca-ATPase molecules allowing us to investigate spatial relationships between the A1 and B domains and between different ATPase polypeptide chains. Differentiation between the intra- and intermolecular components of energy transfer was accomplished in two ways: 1) by comparing the transfer efficiencies in native membranes before and after detergent solubilization and 2) by reconstituting ATPase chains that have already been labeled with either the donor or acceptor chromophores. Using this approach, we find no significant change in the intramolecular transfer efficiency between any of these donor-acceptor pairs either upon binding of calcium to the high affinity sites or upon stabilization of the phosphoenzyme intermediate, indicating that there are no large structural changes within the B tryptic fragment or, alternatively, between the A1 and B fragments. With respect to intermolecular energy transfer, we observe no effect of calcium binding on the unliganded enzyme with either donor-acceptor pair. However

  20. Reticulated, Hyperchromic Rash in a Striated Pattern Mimicking Atopic Dermatitis and Fungal Infection in a 2-Month-Old Female: A Case of Incontinentia Pigmenti

    PubMed Central

    Poliak, Nina; Le, Alexandre; Rainey, Anthony

    2016-01-01

    We present a 12-month-old Hispanic female with a reticulated, hyperchromic rash in a striated pattern appearing on upper and lower extremities and trunk and back since the age of 6 weeks. Over the next 10 months, the rash persisted. The rash did not respond to treatment with antifungals and steroids. During her 6-month wellness visit, the patient was diagnosed with incontinentia pigmenti (IP), a rare X-linked dominant disorder, fatal to male fetuses in utero. IP can lead to serious neurological and ophthalmologic consequences. Early diagnosis by primary care physicians and parental education about the condition are essential for prevention of retinal detachment, developmental delay, and dental abnormalities. PMID:27195166

  1. Segregation of short-wavelength sensitive ("blue") cone signals among neurons in the lateral geniculate nucleus and striate cortex of marmosets.

    PubMed

    Hashemi-Nezhad, Maziar; Blessing, Esther M; Dreher, Bogdan; Martin, Paul R

    2008-11-01

    We measured functional input from short-wavelength selective (S) cones to neurons in the dorsal lateral geniculate nucleus (LGN) and striate cortex (area V1) in anaesthetized marmosets. We found that most magnocellular (MC) and parvocellular (PC) cells receive very little (<5%) functional input from S cones, whereas blue-on cells of the koniocellular (KC) pathway receive dominant input from S cones. Cells dominated by S cone input were not encountered in V1, but V1 cells received more S cone input than PC or MC cells. This suggests that S cone inputs are distributed broadly among neurons in V1. No differences in strength of S cone inputs were seen on comparing dichromatic and trichromatic marmosets, suggesting that the addition of a medium-long wavelength selective cone-opponent ("red-green") channel to a dichromatic visual system does not detectably affect the chromatic properties of the S cone pathways. PMID:18397798

  2. Modification of intracellular free calcium in cultured A10 vascular smooth muscle cells by exogenous phosphatidic acid.

    PubMed

    Bhugra, Praveen; Xu, Yan-Jun; Rathi, Satyajeet; Dhalla, Naranjan S

    2003-06-15

    Exogenous phosphatidic acid (PA) was observed to produce a concentration-dependent increase in [Ca(2+)](i) in cultured A10 vascular smooth muscle cells. Preincubation of cells with sarcoplasmic reticulum Ca(2+)-ATPase inhibitors (cyclopiazonic acid and thapsigargin), a phospholipase C inhibitor (2-nitro-4-carboxyphenyl-N,N-diphenylcarbamate), inositol 1,4,5-trisphosphate receptor antagonists (2-aminoethoxydiphenyl borate and xestospongin), and an activator of protein kinase C (PKC) (phorbol 12-myristate 13-acetate) depressed the PA-evoked increase in [Ca(2+)](i). Although EGTA, an extracellular Ca(2+) chelator, decreased the PA-induced increase in [Ca(2+)](i), sarcolemmal Ca(2+)-channel blockers (verapamil or diltiazem) did not alter the action of PA. On the other hand, inhibitors of PKC (bisindolylmaleimide I) and G(i)-protein (pertussis toxin) potentiated the increase in [Ca(2+)](i) evoked by PA significantly. These results suggest that the PA-induced increase in [Ca(2+)](i) in vascular smooth muscle cells may occur upon the activation of phospholipase C and the subsequent release of Ca(2+) from the inositol 1,4,5-trisphosphate-sensitive Ca(2+) pool in the sarcoplasmic reticulum. This action of PA may be mediated through the involvement of PKC. PMID:12787890

  3. Modification of intracellular free calcium in cultured A10 vascular smooth muscle cells by exogenous phosphatidic acid.

    PubMed

    Bhugra, Praveen; Xu, Yan-Jun; Rathi, Satyajeet; Dhalla, Naranjan S

    2003-06-15

    Exogenous phosphatidic acid (PA) was observed to produce a concentration-dependent increase in [Ca(2+)](i) in cultured A10 vascular smooth muscle cells. Preincubation of cells with sarcoplasmic reticulum Ca(2+)-ATPase inhibitors (cyclopiazonic acid and thapsigargin), a phospholipase C inhibitor (2-nitro-4-carboxyphenyl-N,N-diphenylcarbamate), inositol 1,4,5-trisphosphate receptor antagonists (2-aminoethoxydiphenyl borate and xestospongin), and an activator of protein kinase C (PKC) (phorbol 12-myristate 13-acetate) depressed the PA-evoked increase in [Ca(2+)](i). Although EGTA, an extracellular Ca(2+) chelator, decreased the PA-induced increase in [Ca(2+)](i), sarcolemmal Ca(2+)-channel blockers (verapamil or diltiazem) did not alter the action of PA. On the other hand, inhibitors of PKC (bisindolylmaleimide I) and G(i)-protein (pertussis toxin) potentiated the increase in [Ca(2+)](i) evoked by PA significantly. These results suggest that the PA-induced increase in [Ca(2+)](i) in vascular smooth muscle cells may occur upon the activation of phospholipase C and the subsequent release of Ca(2+) from the inositol 1,4,5-trisphosphate-sensitive Ca(2+) pool in the sarcoplasmic reticulum. This action of PA may be mediated through the involvement of PKC.

  4. Cross-bridge elasticity in single smooth muscle cells

    PubMed Central

    1983-01-01

    In smooth muscle, a cross-bridge mechanism is believed to be responsible for active force generation and fiber shortening. In the present studies, the viscoelastic and kinetic properties of the cross- bridge were probed by eliciting tension transients in response to small, rapid, step length changes (delta L = 0.3-1.0% Lcell in 2 ms). Tension transients were obtained in a single smooth muscle cell isolated from the toad (Bufo marinus) stomach muscularis, which was tied between a force transducer and a displacement device. To record the transients, which were of extremely small magnitude (0.1 microN), a high-frequency (400 Hz), ultrasensitive force transducer (18 mV/microN) was designed and built. The transients obtained during maximal force generation (Fmax = 2.26 microN) were characterized by a linear elastic response (Emax = 1.26 X 10(4) mN/mm2) coincident with the length step, which was followed by a biphasic tension recovery made up of two exponentials (tau fast = 5-20 ms, tau slow = 50-300 ms). During the development of force upon activation, transients were elicited. The relationship between stiffness and force was linear, which suggests that the transients originate within the cross-bridge and reflect the cross-bridge's viscoelastic and kinetic properties. The observed fiber elasticity suggests that the smooth muscle cross-bridge is considerably more compliant than in fast striated muscle. A thermodynamic model is presented that allows for an analysis of the factors contributing to the increased compliance of the smooth muscle cross-bridge. PMID:6413640

  5. High pressure induced changes in beef muscle proteome: correlation with quality parameters.

    PubMed

    Marcos, Begonya; Mullen, Anne Maria

    2014-05-01

    The relationship between pressure induced changes on individual proteins and selected quality parameters in bovine longissimus thoracis et lumborum (LTL) muscle was studied. Pressures ranging from 200 to 600 MPa at 20°C were used. High pressure processing (HPP) at pressures above 200 MPa induced strong modifications of protein solubility, meat colour and water holding capacity (WHC). The protein profiles of non-treated and pressure treated meat were observed using two dimensional electrophoresis. Proteins showing significant differences in abundance among treatments were identified by mass spectrometry. Pressure levels above 200 MPa strongly modified bovine LTL proteome with main effects being insolubilisation of sarcoplasmic proteins and solubilisation of myofibrillar proteins. Sarcoplasmic proteins were more susceptible to HPP effects than myofibrillar. Individual protein changes were significantly correlated with protein solubility, L, b and WHC, providing further insights into the mechanistic processes underlying HPP influence on quality and providing the basis for the future development of protein markers to assess the quality of processed meats.

  6. Intracellular energetic units in red muscle cells.

    PubMed Central

    Saks, V A; Kaambre, T; Sikk, P; Eimre, M; Orlova, E; Paju, K; Piirsoo, A; Appaix, F; Kay, L; Regitz-Zagrosek, V; Fleck, E; Seppet, E

    2001-01-01

    The kinetics of regulation of mitochondrial respiration by endogenous and exogenous ADP in muscle cells in situ was studied in skinned cardiac and skeletal muscle fibres. Endogenous ADP production was initiated by addition of MgATP; under these conditions the respiration rate and ADP concentration in the medium were dependent on the calcium concentration, and 70-80% of maximal rate of respiration was achieved at ADP concentration below 20 microM in the medium. In contrast, when exogenous ADP was added, maximal respiration rate was observed only at millimolar concentrations. An exogenous ADP-consuming system consisting of pyruvate kinase (PK; 20-40 units/ml) and phosphoenolpyruvate (PEP; 5 mM), totally suppressed respiration activated by exogenous ADP, but the respiration maintained by endogenous ADP was not suppressed by more than 20-40%. Creatine (20 mM) further activated respiration in the presence of ATP and PK+PEP. Short treatment with trypsin (50-500 nM for 5 min) decreased the apparent K(m) for exogenous ADP from 300-350 microM to 50-60 microM, increased inhibition of respiration by PK+PEP system up to 70-80%, with no changes in MgATPase activity and maximal respiration rates. Electron-microscopic observations showed detachment of mitochondria and disordering of the regular structure of the sarcomere after trypsin treatment. Two-dimensional electrophoresis revealed a group of at least seven low-molecular-mass proteins in cardiac skinned fibres which were very sensitive to trypsin and not present in glycolytic fibres, which have low apparent K(m) for exogenous ADP. It is concluded that, in oxidative muscle cells, mitochondria are incorporated into functional complexes ('intracellular energetic units') with adjacent ADP-producing systems in myofibrils and in sarcoplasmic reticulum, probably due to specific interaction with cytoskeletal elements responsible for mitochondrial distribution in the cell. It is suggested that these complexes represent the basic

  7. Alcohol differentially alters extracellular matrix and adhesion molecule expression in skeletal muscle and heart

    PubMed Central

    Steiner, Jennifer L.; Pruznak, Anne M.; Navaratnarajah, Maithili; Lang, Charles H.

    2015-01-01

    Background The production of fibrosis in response to chronic alcohol abuse is well recognized in liver but has not been fully characterized in striated muscle and may contribute to functional impairment. Therefore, the purpose of this study was to use an unbiased discovery-based approach to determine the effect of chronic alcohol consumption on the expression profile of genes important for cell-cell and cell-extracellular matrix (ECM) interactions in both skeletal and cardiac muscle. Methods Adult male rats were pair-fed an alcohol-containing liquid diet or control diet for 24 wks, and skeletal muscle (gastrocnemius) and heart collected in the freely fed state. A pathway-focused gene expression PCR array was performed on these tissues to assess mRNA content for 84 ECM proteins, and selected proteins were confirmed by Western analysis. Results In gastrocnemius, alcohol feeding up-regulated expression of 11 genes and down-regulated expression of 1 gene. Alcohol increased fibrosis as indicated by increased mRNA and/or protein for collagen α1(I), α2(I), α1(III) and α2(IV) as well as hydroxyproline. Alcohol also increased α-smooth muscle actin protein, an index of myofibroblast activation, but no concomitant change in TGF-β was detected. The mRNA and protein content for other ECM components, such as integrin α-5, L-selectin, PECAM, Sparc and Adamts2 was also increased by alcohol. Only laminin α-3 mRNA was decreased in gastrocnemius from alcohol-fed rats, while 66 ECM- or cell adhesion-related mRNAs were unchanged by alcohol. For heart, expression of 16 genes was up-regulated, expression of 3 genes was down-regulated, and 65 mRNAs were unchanged by alcohol; there were no common alcohol-induced gene expression changes between heart and skeletal muscle. Finally, alcohol increased TNFα and IL-12 mRNA in both skeletal and cardiac muscle, but IL-6 mRNA was increased and IL-10 mRNA decreased only in skeletal muscle. Conclusions These data demonstrate a fibrotic

  8. Increased Reliance on Muscle-based Thermogenesis upon Acute Minimization of Brown Adipose Tissue Function.

    PubMed

    Bal, Naresh C; Maurya, Santosh K; Singh, Sushant; Wehrens, Xander H T; Periasamy, Muthu

    2016-08-12

    Skeletal muscle has been suggested as a site of nonshivering thermogenesis (NST) besides brown adipose tissue (BAT). Studies in birds, which do not contain BAT, have demonstrated the importance of skeletal muscle-based NST. However, muscle-based NST in mammals remains poorly characterized. We recently reported that sarco/endoplasmic reticulum Ca(2+) cycling and that its regulation by SLN can be the basis for muscle NST. Because of the dominant role of BAT-mediated thermogenesis in rodents, the role of muscle-based NST is less obvious. In this study, we investigated whether muscle will become an important site of NST when BAT function is conditionally minimized in mice. We surgically removed interscapular BAT (iBAT, which constitutes ∼70% of total BAT) and exposed the mice to prolonged cold (4 °C) for 9 days. The iBAT-ablated mice were able to maintain optimal body temperature (∼35-37 °C) during the entire period of cold exposure. After 4 days in the cold, both sham controls and iBAT-ablated mice stopped shivering and resumed routine physical activity, indicating that they are cold-adapted. The iBAT-ablated mice showed higher oxygen consumption and decreased body weight and fat mass, suggesting an increased energy cost of cold adaptation. The skeletal muscles in these mice underwent extensive remodeling of both the sarcoplasmic reticulum and mitochondria, including alteration in the expression of key components of Ca(2+) handling and mitochondrial metabolism. These changes, along with increased sarcolipin expression, provide evidence for the recruitment of NST in skeletal muscle. These studies collectively suggest that skeletal muscle becomes the major site of NST when BAT activity is minimized.

  9. Increased Reliance on Muscle-based Thermogenesis upon Acute Minimization of Brown Adipose Tissue Function.

    PubMed

    Bal, Naresh C; Maurya, Santosh K; Singh, Sushant; Wehrens, Xander H T; Periasamy, Muthu

    2016-08-12

    Skeletal muscle has been suggested as a site of nonshivering thermogenesis (NST) besides brown adipose tissue (BAT). Studies in birds, which do not contain BAT, have demonstrated the importance of skeletal muscle-based NST. However, muscle-based NST in mammals remains poorly characterized. We recently reported that sarco/endoplasmic reticulum Ca(2+) cycling and that its regulation by SLN can be the basis for muscle NST. Because of the dominant role of BAT-mediated thermogenesis in rodents, the role of muscle-based NST is less obvious. In this study, we investigated whether muscle will become an important site of NST when BAT function is conditionally minimized in mice. We surgically removed interscapular BAT (iBAT, which constitutes ∼70% of total BAT) and exposed the mice to prolonged cold (4 °C) for 9 days. The iBAT-ablated mice were able to maintain optimal body temperature (∼35-37 °C) during the entire period of cold exposure. After 4 days in the cold, both sham controls and iBAT-ablated mice stopped shivering and resumed routine physical activity, indicating that they are cold-adapted. The iBAT-ablated mice showed higher oxygen consumption and decreased body weight and fat mass, suggesting an increased energy cost of cold adaptation. The skeletal muscles in these mice underwent extensive remodeling of both the sarcoplasmic reticulum and mitochondria, including alteration in the expression of key components of Ca(2+) handling and mitochondrial metabolism. These changes, along with increased sarcolipin expression, provide evidence for the recruitment of NST in skeletal muscle. These studies collectively suggest that skeletal muscle becomes the major site of NST when BAT activity is minimized. PMID:27298322

  10. Mercury distribution and lipid oxidation in fish muscle: Effects of washing and isoelectric protein precipitation

    USGS Publications Warehouse

    Gong, Y.; Krabbenhoft, D.P.; Ren, L.; Egelandsdal, B.; Richards, M.P.

    2011-01-01

    Nearly all the mercury (Hg) in whole muscle from whitefish (Coregonus clupeaformis) and walleye (Sander vitreus) was present as methyl mercury (MeHg). The Hg content in whole muscle from whitefish and walleye was 0.04-0.09 and 0.14-0.81 ppm, respectively. The myofibril fraction contained approximately three-fourths of the Hg in whitefish and walleye whole muscle. The sarcoplasmic protein fraction (e.g., press juice) was the next most abundant source of Hg. Isolated myosin, triacylglycerols, and cellular membranes contained the least Hg. Protein isolates prepared by pH shifting in the presence of citric acid did not decrease Hg levels. Addition of cysteine during washing decreased the Hg content in washed muscle probably through