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Sample records for striated muscle sarcoplasmic

  1. An invertebrate smooth muscle with striated muscle myosin filaments.

    PubMed

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

    2015-10-20

    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.

  2. 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

  3. Independent evolution of striated muscles in cnidarians and bilaterians.

    PubMed

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

    2012-07-12

    Striated muscles are present in bilaterian animals (for example, vertebrates, insects and annelids) and some non-bilaterian eumetazoans (that is, cnidarians and ctenophores). The considerable ultrastructural similarity of striated muscles between these animal groups is thought to reflect a common evolutionary origin. Here we show that a muscle protein core set, including a type II myosin heavy chain (MyHC) motor protein characteristic of striated muscles in vertebrates, was already present in unicellular organisms before the origin of multicellular animals. Furthermore, 'striated muscle' and 'non-muscle' myhc orthologues are expressed differentially in two sponges, compatible with a functional diversification before the origin of true muscles and the subsequent use of striated muscle MyHC in fast-contracting smooth and striated muscle. Cnidarians and ctenophores possess striated muscle myhc orthologues but lack crucial components of bilaterian striated muscles, such as genes that code for titin and the troponin complex, 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 new proteins to a pre-existing, ancestral contractile apparatus may serve as a model for the evolution of complex animal cell types.

  4. 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

  5. Caffeine-induced contracture in oesophageal striated muscle of normotensive and hypertensive rats.

    PubMed

    Sekiguchi, Fumiko; Kawata, Kyoko; Komori, Mayumi; Sunano, Satoru

    2003-03-28

    To elucidate whether properties of the sarcoplasmic reticulum are altered, not only in vascular smooth muscle, but also in visceral striated muscle of spontaneously hypertensive rats (SHR), caffeine-induced contractures in oesophageal striated muscle of Wistar Kyoto rats (WKY) and stroke-prone SHR (SHRSP) were compared. In both preparations, 30 mM caffeine induced a contracture with two components. The second component, which was diminished by extracellular Ca(2+) removal or Ni(2+) but not by verapamil, was much smaller in SHRSP. Both components and differences between WKY and SHRSP coincided with changes in intracellular Ca(2+). Although membrane potential was identical between these preparations, caffeine induced slight depolarization only in WKY preparations. Similar depolarization was observed with 10 mM K(+), which induced no contraction. It is suggested that the first and the second components of caffeine-induced contracture were induced by Ca(2+) released from sarcoplasmic reticulum and by Ca(2+) that entered through channels activated by sarcoplasmic reticulum Ca(2+) depletion, respectively. In SHRSP preparations, Ca(2+) from the latter pathway was clearly decreased, although this change is thought not to be related to the initiation of hypertension. These results suggest that Ca(2+) handling properties of cell membrane and sarcoplasmic reticulum are generally altered in muscles of SHRSP.

  6. 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.

  7. Tropomodulin Capping of Actin Filaments in Striated Muscle Development and Physiology

    PubMed Central

    Gokhin, David S.; Fowler, Velia M.

    2011-01-01

    Efficient striated muscle contraction requires precise assembly and regulation of diverse actin filament systems, most notably the sarcomeric thin filaments of the contractile apparatus. By capping the pointed ends of actin filaments, tropomodulins (Tmods) regulate actin filament assembly, lengths, and stability. Here, we explore the current understanding of the expression patterns, localizations, and functions of Tmods in both cardiac and skeletal muscle. We first describe the mechanisms by which Tmods regulate myofibril assembly and thin filament lengths, as well as the roles of closely related Tmod family variants, the leiomodins (Lmods), in these processes. We also discuss emerging functions for Tmods in the sarcoplasmic reticulum. This paper provides abundant evidence that Tmods are key structural regulators of striated muscle cytoarchitecture and physiology. PMID:22013379

  8. 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

  9. Mitochondria Are Linked to Calcium Stores in Striated Muscle by Developmentally Regulated Tethering Structures

    PubMed Central

    Boncompagni, Simona; Rossi, Ann E.; Micaroni, Massimo; Beznoussenko, Galina V.; Polishchuk, Roman S.; Dirksen, Robert T.

    2009-01-01

    Bi-directional calcium (Ca2+) signaling between mitochondria and intracellular stores (endoplasmic/sarcoplasmic reticulum) underlies important cellular functions, including oxidative ATP production. In striated muscle, this coupling is achieved by mitochondria being located adjacent to Ca2+ stores (sarcoplasmic reticulum [SR]) and in proximity of release sites (Ca2+ release units [CRUs]). However, limited information is available with regard to the mechanisms of mitochondrial-SR coupling. Using electron microscopy and electron tomography, we identified small bridges, or tethers, that link the outer mitochondrial membrane to the intracellular Ca2+ stores of muscle. This association is sufficiently strong that treatment with hypotonic solution results in stretching of the SR membrane in correspondence of tethers. We also show that the association of mitochondria to the SR is 1) developmentally regulated, 2) involves a progressive shift from a longitudinal clustering at birth to a specific CRU-coupled transversal orientation in adult, and 3) results in a change in the mitochondrial polarization state, as shown by confocal imaging after JC1 staining. Our results suggest that tethers 1) establish and maintain SR–mitochondrial association during postnatal maturation and in adult muscle and 2) likely provide a structural framework for bi-directional signaling between the two organelles in striated muscle. PMID:19037102

  10. Mitochondria are linked to calcium stores in striated muscle by developmentally regulated tethering structures.

    PubMed

    Boncompagni, Simona; Rossi, Ann E; Micaroni, Massimo; Beznoussenko, Galina V; Polishchuk, Roman S; Dirksen, Robert T; Protasi, Feliciano

    2009-02-01

    Bi-directional calcium (Ca(2+)) signaling between mitochondria and intracellular stores (endoplasmic/sarcoplasmic reticulum) underlies important cellular functions, including oxidative ATP production. In striated muscle, this coupling is achieved by mitochondria being located adjacent to Ca(2+) stores (sarcoplasmic reticulum [SR]) and in proximity of release sites (Ca(2+) release units [CRUs]). However, limited information is available with regard to the mechanisms of mitochondrial-SR coupling. Using electron microscopy and electron tomography, we identified small bridges, or tethers, that link the outer mitochondrial membrane to the intracellular Ca(2+) stores of muscle. This association is sufficiently strong that treatment with hypotonic solution results in stretching of the SR membrane in correspondence of tethers. We also show that the association of mitochondria to the SR is 1) developmentally regulated, 2) involves a progressive shift from a longitudinal clustering at birth to a specific CRU-coupled transversal orientation in adult, and 3) results in a change in the mitochondrial polarization state, as shown by confocal imaging after JC1 staining. Our results suggest that tethers 1) establish and maintain SR-mitochondrial association during postnatal maturation and in adult muscle and 2) likely provide a structural framework for bi-directional signaling between the two organelles in striated muscle.

  11. Evolution of striated muscle: jellyfish and the origin of triploblasty.

    PubMed

    Seipel, Katja; Schmid, Volker

    2005-06-01

    The larval and polyp stages of extant Cnidaria are bi-layered with an absence of mesoderm and its differentiation products. This anatomy originally prompted the diploblast classification of the cnidarian phylum. The medusa stage, or jellyfish, however, has a more complex anatomy characterized by a swimming bell with a well-developed striated muscle layer. Based on developmental histology of the hydrozoan medusa this muscle derives from the entocodon, a mesoderm-like third cell layer established at the onset of medusa formation. According to recent molecular studies cnidarian homologs to bilaterian mesoderm and myogenic regulators are expressed in the larval and polyp stages as well as in the entocodon and derived striated muscle. Moreover striated and smooth muscle cells may have evolved directly and independently from non-muscle cells as indicated by phylogenetic analysis of myosin heavy chain genes (MHC class II). To accommodate all evidences we propose that striated muscle-based locomotion coevolved with the nervous and digestive systems in a basic metazoan Bauplan from which the ancestors of the Ctenophora (comb jellyfish), Cnidaria (jellyfish and polyps), as well as the Bilateria are derived. We argue for a motile tri-layered cnidarian ancestor and a monophyletic descent of striated muscle in Cnidaria and Bilateria. As a consequence, diploblasty evolved secondarily in cnidarian larvae and polyps.

  12. 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

  13. Striated Muscle Regulation of Isometric Tension by Multiple Equilibria

    PubMed Central

    Zot, Henry G.; Hasbun, Javier E.; Van Minh, Nguyen

    2009-01-01

    Cooperative activation of striated muscle by calcium is based on the movement of tropomyosin described by the steric blocking theory of muscle contraction. Presently, the Hill model stands alone in reproducing both myosin binding data and a sigmoidal-shaped curve characteristic of calcium activation (Hill TL (1983) Two elementary models for the regulation of skeletal muscle contraction by calcium. Biophys J 44: 383–396.). However, the free myosin is assumed to be fixed by the muscle lattice and the cooperative mechanism is based on calcium-dependent interactions between nearest neighbor tropomyosin subunits, which has yet to be validated. As a result, no comprehensive model has been shown capable of fitting actual tension data from striated muscle. We show how variable free myosin is a selective advantage for activating the muscle and describe a mechanism by which a conformational change in tropomyosin propagates free myosin given constant total myosin. This mechanism requires actin, tropomyosin, and filamentous myosin but is independent of troponin. Hence, it will work equally well with striated, smooth and non-muscle contractile systems. Results of simulations with and without data are consistent with a strand of tropomyosin composed of ∼20 subunits being moved by the concerted action of 3–5 myosin heads, which compares favorably with the predicted length of tropomyosin in the overlap region of thick and thin filaments. We demonstrate that our model fits both equilibrium myosin binding data and steady-state calcium-dependent tension data and show how both the steepness of the response and the sensitivity to calcium can be regulated by the actin-troponin interaction. The model simulates non-cooperative calcium binding both in the presence and absence of strong binding myosin as has been observed. Thus, a comprehensive model based on three well-described interactions with actin, namely, actin-troponin, actin-tropomyosin, and actin-myosin can explain the

  14. 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.

  15. Transplantation and regeneration of striated muscle.

    PubMed Central

    Allbrook, D.

    1975-01-01

    This lecture explores the factors controlling regeneration and reconstitution of skeletal muscle following vascular and neural injury by giving an account of some experimental work in this field, which is then linked to the problem of the use of whole-muscle transplants in clinical surgery. Images Fig. 2 Fig. 4 Fig. 7 Fig. 8 Fig. 9 PMID:1147539

  16. Intracellular mechanisms of verapamil and diltiazem action on striated muscle of the rabbit.

    PubMed

    Su, J Y

    1988-09-01

    Skinned fibers from striated muscle were used to study the intracellular mechanisms (contractile proteins and sarcoplasmic reticulum [SR]) of action of diltiazem (DT) and verapamil (VP) on muscle contraction. Rabbit papillary muscle (PM), and the skeletal muscles adductor magnus (AM, fast-twitch) and soleus (SL, slow-twitch) were used. The muscles were skinned by homogenization and fibre bundles for PM and single fibres for AM and SL were dissected from the homogenate and mounted on photodiode force transducers. VP (0.1-3.0 mmol/l) (and to a lesser degree DT) increased Ca2+-activated tension development of the contractile protains in PM and SL and decreased it in AM (+[4-20]%, +4%, -[14-28]%, respectively). Both drugs increased the submaximal Ca2+-activated tension development at the order of PM = SL greater than AM in a dose-dependent manner. The changes of half-maximal pCa50 at 1 mmol/l VP were 0.25, 0.25, and 0.15, respectively. For Ca2+ uptake and release from the SR, VP as well as DT (0.1-3.0 mmol/l) in the uptake phase decreased caffeine-induced tension transients in a dose-dependent fashion. At 0.01-3.0 mmol/l, the drugs directly induced Ca2+ release from the SR or enhanced caffeine-induced tension transients with the exception that in PM, DT attenuated caffeine-induced tension transients. Thus, VP and DT have similar intracellular mechanisms of action in striated muscle. Both drugs induced calcium release from the SR and increase Ca2+ sensitivity of the contractile proteins, and thus could be the underlying mechanisms for potentiating twitch tension, and inducing contracture in skeletal muscle.

  17. Contracture of Slow Striated Muscle during Calcium Deprivation

    PubMed Central

    Irwin, Richard L.; Hein, Manfred M.

    1963-01-01

    When deprived of calcium the slow striated muscle fibers of the frog develop reversible contractures in either hypertonic or isotonic solutions. While calcium deprivation continues because of a flowing calcium-free solution the muscles relax slowly and completely. Restoration of calcium during contracture relaxes the muscle promptly to initial tension. When relaxed during calcium lack the return of calcium does not change tension and the muscle stays relaxed. When contractures are induced by solutions containing small amounts of calcium relaxation does not occur or requires several hours. The rate of tension development depends upon the rate at which calcium moves outward since the contractures develop slower in low concentrations of calcium and are absent or greatly slowed in a stagnant calcium-free solution. Withdrawal of calcium prevents the contractile responses to ACh, KCl, or electrical stimulation through the nerve. Muscles return to their original excitability after calcium is restored. Origin of the contractures is unrelated to nerve activity since they are maximal during transmission failure from calcium lack, occur in denervated muscles, and are not blocked by high concentrations of d-tubocurarine, procaine, or atropine. The experiments also indicate that the contractures do not originate from repetitive activity of muscle membranes. The findings are most simply explained by relating the outward movement of calcium as a link for initiating contraction in slow type striated muscle. PMID:14065284

  18. Expression of various sarcomeric tropomyosin isoforms in equine striated muscles.

    PubMed

    Dube, Syamalima; Chionuma, Henry; Matoq, Amr; Alshiekh-Nasany, Ruham; Abbott, Lynn; Poiesz, Bernard J; Dube, Dipak K

    2017-01-01

    In order to better understand the training and athletic activity of horses, we must have complete understanding of the isoform diversity of various myofibrillar protein genes like tropomyosin. Tropomyosin (TPM), a coiled-coil dimeric protein, is a component of thin filament in striated muscles. In mammals, four TPM genes (TPM1, TPM2, TPM3, and TPM4) generate a multitude of TPM isoforms via alternate splicing and/or using different promoters. Unfortunately, our knowledge of TPM isoform diversity in the horse is very limited. Hence, we undertook a comprehensive exploratory study of various TPM isoforms from horse heart and skeletal muscle. We have cloned and sequenced two sarcomeric isoforms of the TPM1 gene called TPM1α and TPM1κ, one sarcomeric isoform of the TPM2 and one of the TPM3 gene, TPM2α and TPM3α respectively. By qRT-PCR using both relative expression and copy number, we have shown that TPM1α expression compared to TPM1κ is very high in heart. On the other hand, the expression of TPM1α is higher in skeletal muscle compared to heart. Further, the expression of TPM2α and TPM3α are higher in skeletal muscle compared to heart. Using western blot analyses with CH1 monoclonal antibody we have shown the high expression levels of sarcomeric TPM proteins in cardiac and skeletal muscle. Due to the paucity of isoform specific antibodies we cannot specifically detect the expression of TPM1κ in horse striated muscle. To the best of our knowledge this is the very first report on the characterization of sarcmeric TPMs in horse striated muscle.

  19. Expression of various sarcomeric tropomyosin isoforms in equine striated muscles

    PubMed Central

    Dube, Syamalima; Chionuma, Henry; Matoq, Amr; Alshiekh-Nasany, Ruham; Abbott, Lynn; Poiesz, Bernard J.; Dube, Dipak K.

    2017-01-01

    In order to better understand the training and athletic activity of horses, we must have complete understanding of the isoform diversity of various myofibrillar protein genes like tropomyosin. Tropomyosin (TPM), a coiled-coil dimeric protein, is a component of thin filament in striated muscles. In mammals, four TPM genes (TPM1, TPM2, TPM3, and TPM4) generate a multitude of TPM isoforms via alternate splicing and/or using different promoters. Unfortunately, our knowledge of TPM isoform diversity in the horse is very limited. Hence, we undertook a comprehensive exploratory study of various TPM isoforms from horse heart and skeletal muscle. We have cloned and sequenced two sarcomeric isoforms of the TPM1 gene called TPM1α and TPM1κ, one sarcomeric isoform of the TPM2 and one of the TPM3 gene, TPM2α and TPM3α respectively. By qRT-PCR using both relative expression and copy number, we have shown that TPM1α expression compared to TPM1κ is very high in heart. On the other hand, the expression of TPM1α is higher in skeletal muscle compared to heart. Further, the expression of TPM2α and TPM3α are higher in skeletal muscle compared to heart. Using western blot analyses with CH1 monoclonal antibody we have shown the high expression levels of sarcomeric TPM proteins in cardiac and skeletal muscle. Due to the paucity of isoform specific antibodies we cannot specifically detect the expression of TPM1κ in horse striated muscle. To the best of our knowledge this is the very first report on the characterization of sarcmeric TPMs in horse striated muscle. PMID:28717602

  20. Role of titin in vertebrate striated muscle.

    PubMed Central

    Tskhovrebova, L; Trinick, J

    2002-01-01

    Titin is a giant muscle protein with a molecular weight in the megaDalton range and a contour length of more than 1 microm. Its size and location within the sarcomere structure determine its important role in the mechanism of muscle elasticity. According to the current consensus, elasticity stems directly from more than one type of spring-like behaviour of the I-band portion of the molecule. Starting from slack length, extension of the sarcomere first causes straightening of the molecule. Further extension then induces local unfolding of a unique sequence, the PEVK region, which is named due to the preponderance of these amino-acid residues. High speeds of extension and/or high forces are likely to lead to unfolding of the beta-sandwich domains from which the molecule is mainly constructed. A release of tension leads to refolding and recoiling of the polypeptide. Here, we review the literature and present new experimental material related to the role of titin in muscle elasticity. In particular, we analyse the possible influence of the arrangement and environment of titin within the sarcomere structure on its extensible behaviour. We suggest that, due to the limited conformational space, elongation and compression of the molecule within the sarcomere occur in a more ordered way or with higher viscosity and higher forces than are observed in solution studies of the isolated protein. PMID:11911777

  1. Ultrastructure and mechanical activity expressed by striated muscle in culture.

    PubMed

    Coleman, A W; Siegel, R; Coleman, J R

    1978-01-01

    Newly devised assay procedures for quantitating the mechanical capabilities of striated muscle fibers grown in cell culture have permitted the correlation of cytological features with the ability to respond mechanically to electrical and chemical stimuli during development. By developmental timing and by physiological characteristics, three distinct mechanical activities can be distinguished: : TWITCH, contracture and wave propagation (escalation). Parallel electron microscopy studies suggest that contracture and escalation require significantly greater internal membrane development than twitch. The assay procedures have revealed that fibers developed in culture from genetically dystrophic chick muscle cells display a heightened electrical threshold for a twich response, but are otherwise similar to normal fibers. Cultured chick fibers, whether of leg or breast origin, exhibit similar ultrastructural and mechanical properties; yet these are different from those of in vivo adult muscle and may represent the avian striated muscle archetype expressed in the absence of innervation. Primary or cell line cultures of rat muscle produce far fewer mechanically active fibers than do avian cell cultures. The influence of culture conditions and cell source, whether avian or mammalian, on the extent of differentiation expressed in culture is so great that our understanding of studies on cultured muscle fibers would benefit from some characterization of both morphological and contractile properties of the fibers being used.

  2. Contractile properties of esophageal striated muscle: comparison with cardiac and skeletal muscles in rats.

    PubMed

    Shiina, Takahiko; Shima, Takeshi; Masuda, Kazuaki; Hirayama, Haruko; Iwami, Momoe; Takewaki, Tadashi; Kuramoto, Hirofumi; Shimizu, Yasutake

    2010-01-01

    The external muscle layer of the mammalian esophagus consists of striated muscles. We investigated the contractile properties of esophageal striated muscle by comparison with those of skeletal and cardiac muscles. Electrical field stimulation with single pulses evoked twitch-like contractile responses in esophageal muscle, similar to those in skeletal muscle in duration and similar to those in cardiac muscle in amplitude. The contractions of esophageal muscle were not affected by an inhibitor of gap junctions. Contractile responses induced by high potassium or caffeine in esophageal muscle were analogous to those in skeletal muscle. High-frequency stimulation induced a transient summation of contractions followed by sustained contractions with amplitudes similar to those of twitch-like contractions, although a large summation was observed in skeletal muscle. The results demonstrate that esophageal muscle has properties similar but not identical to those of skeletal muscle and that some specific properties may be beneficial for esophageal peristalsis.

  3. Mechanical Stretch-Induced Activation of ROS/RNS Signaling in Striated Muscle

    PubMed Central

    Ward, Christopher W.; Prosser, Benjamin L.

    2014-01-01

    Significance: Mechanical activation of reactive oxygen species (ROS) and reactive nitrogen species (RNS) occurs in striated muscle and affects Ca2+ signaling and contractile function. ROS/RNS signaling is tightly controlled, spatially compartmentalized, and source specific. Recent Advances: Here, we review the evidence that within the contracting myocyte, the trans-membrane protein NADPH oxidase 2 (Nox2) is the primary source of ROS generated during contraction. We also review a newly characterized signaling cascade in cardiac and skeletal muscle in which the microtubule network acts as a mechanotransduction element that activates Nox2-dependent ROS generation during mechanical stretch, a pathway termed X-ROS signaling. Critical Issues: In the heart, X-ROS acts locally and affects the sarcoplasmic reticulum (SR) Ca2+ release channels (ryanodine receptors) and tunes Ca2+ signaling during physiological behavior, but excessive X-ROS can promote Ca2+-dependent arrhythmias in pathology. In skeletal muscle, X-ROS sensitizes Ca2+-permeable sarcolemmal “transient receptor potential” channels, a pathway that is critical for sustaining SR load during repetitive contractions, but when in excess, it is maladaptive in diseases such as Duchenne Musclar dystrophy. Future Directions: New advances in ROS/RNS detection as well as molecular manipulation of signaling pathways will provide critical new mechanistic insights into the details of X-ROS signaling. These efforts will undoubtedly reveal new avenues for therapeutic intervention in the numerous diseases of striated muscle in which altered mechanoactivation of ROS/RNS production has been identified. Antioxid. Redox Signal. 20, 929–936. PMID:23971496

  4. Mechanical tension and spontaneous muscle twitching precede the formation of cross-striated muscle in vivo

    PubMed Central

    Weitkunat, Manuela; Brasse, Martina; Bausch, Andreas R.

    2017-01-01

    Muscle forces are produced by repeated stereotypical actomyosin units called sarcomeres. Sarcomeres are chained into linear myofibrils spanning the entire muscle fiber. In mammalian body muscles, myofibrils are aligned laterally, resulting in their typical cross-striated morphology. Despite this detailed textbook knowledge about the adult muscle structure, it is still unclear how cross-striated myofibrils are built in vivo. Here, we investigate the morphogenesis of Drosophila abdominal muscles and establish them as an in vivo model for cross-striated muscle development. By performing live imaging, we find that long immature myofibrils lacking a periodic actomyosin pattern are built simultaneously in the entire muscle fiber and then align laterally to give mature cross-striated myofibrils. Interestingly, laser micro-lesion experiments demonstrate that mechanical tension precedes the formation of the immature myofibrils. Moreover, these immature myofibrils do generate spontaneous Ca2+-dependent contractions in vivo, which, when chemically blocked, result in cross-striation defects. Taken together, these results suggest a myofibrillogenesis model in which mechanical tension and spontaneous muscle twitching synchronize the simultaneous self-organization of different sarcomeric protein complexes to build highly regular cross-striated myofibrils spanning the length of large muscle fibers. PMID:28174246

  5. Ultrastructure of mouse striated muscle fibers following pravastatin administration.

    PubMed

    Bergman, Michael; Salman, Hertzel; Djaldetti, Meir; Alexandrova, Svetlana; Punsky, Igor; Bessler, Hanna

    2003-01-01

    To examine the effect of pravastatin administration on striated muscle ultrastructure, 10 BalbC mice were given pravastatin 40 mg/kg/day for 3 weeks. At the end of the study, blood was withdrawn for evaluation of the serum creatine phospho-kinase (CPK) level and the muscles of the hind legs, as well as the heart and liver of the animals were examined with a light and transmission electron microscope. After treatment with pravastatin the results showed a 101% increase in serum CPK level in comparison to untreated controls. Hematoxillin-eosin stained tissues of pravastatin treated mice did not show any abnormal findings. While the ultrastructure of the heart and liver of the treated animals appeared normal, the muscle fibers showed a marked alterations of the mitochondria, which were increased in size compared to those of the controls. The cristae were heavily damaged and even completely destructed, giving the mitochondria appearance of empty vacuoles. The findings are in favor of a specificity of pravastatin for striated muscles.

  6. Regulation of actin filament length in erythrocytes and striated muscle.

    PubMed

    Fowler, V M

    1996-02-01

    Actin filaments polymerize in vitro to lengths which display an exponential distribution, yet in many highly differentiated cells they can be precisely maintained at uniform lengths in elaborate supramolecular structures. Recent results obtained using two classic model systems, the erythrocyte membrane cytoskeleton and the striated muscle sarcomere, reveal surprising similarities and instructive differences in the molecules and mechanisms responsible for determining and maintaining actin filament lengths in these two systems. Tropomodulin caps the slow-growing, pointed filament ends in muscle and in erythrocytes. CapZ caps the fast-growing, barbed filament ends in striated muscle, whereas a newly discovered barbed end capping protein, adducin, may cap the barbed filament ends in erythrocytes. The mechanisms responsible for specifying the characteristic filament lengths in these systems are more elusive and may include strict control of the relative amounts of actin filament capping proteins and side-binding proteins, molecular templates (e.g. tropomyosin and nebulin) and/or verniers (e.g. tropomyosin).

  7. Elemental distribution in striated muscle and the effects of hypertonicity: Electron probe analysis of cryo sections

    PubMed Central

    Somlyo, AV; Shuman, H; Somlyo, AP

    1977-01-01

    A method of rapid freezing in supercooled Freon 22 (monochlorodifluoromethane) followed by cryoultramicrotomy is described and shown to yield ultrathin sections in which both the cellular ultrastructure and the distribution of diffusible ions across the cell membrane are preserved and intracellular compartmentalization of diffusabler ions can be quantitated. Quantitative electron probe analysis (Shuman, H., A.V. Somlyo, and A.P. Somlyo. 1976. Ultramicros. 1:317-339.) of freeze-dried ultrathin cryto sections was found to provide a valid measure of the composition of cells and cellular organelles and was used to determine the ionic composition of the in situ terminal cisternae of the sarcoplasmic reticulum (SR), the distribution of CI in skeletal muscle, and the effects of hypertonic solutions on the subcellular composition if striated muscle. There was no evidence of sequestered CI in the terminal cisternae of resting muscles, although calcium (66mmol/kg dry wt +/- 4.6 SE) was detected. The values of [C1](i) determined with small (50-100 nm) diameter probes over cytoplasm excluding organelles over nuclei or terminal cisternae were not significantly different. Mitochondria partially excluded C1, with a cytoplasmic/ mitochondrial Ci ratio of 2.4 +/- 0.88 SD. The elemental concentrations (mmol/kg dry wt +/- SD) of muscle fibers measured with 0.5-9-μm diameter electron probes in normal frog striated muscle were: P, 302 +/- 4.3; S, 189 +/- 2.9;C1, 24 +/- 1.1;K, 404 +/- 4.3, and Mg, 39 +/- 2.1. It is concluded that: (a) in normal muscle the "excess CI" measured with previous bulk chemical analyses and flux studies is not compartmentalized in the SR or in other cellular organelles, and (b) the cytoplasmic C1 in low [K](0) solutions exceeds that predicted by a passive electrochemical distribution. Hypertonic 2.2 X NaCl, 2.5 X sucrose, or 2.2 X Na isethionate produced: (a) swollen vacuoles, frequently paired, adjacent to the Z lines and containing significantly higher than

  8. 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.

  9. Mitochondria in the middle: exercise preconditioning protection of striated muscle

    PubMed Central

    Rodriguez, Dinah A.; Hord, Jeffrey M.

    2016-01-01

    Abstract Cellular and physiological adaptations to an atmosphere which became enriched in molecular oxygen spurred the development of a layered system of stress protection, including antioxidant and stress response proteins. At physiological levels reactive oxygen and nitrogen species regulate cell signalling as well as intracellular and intercellular communication. Exercise and physical activity confer a variety of stressors on skeletal muscle and the cardiovascular system: mechanical, metabolic, oxidative. Transient increases of stressors during acute bouts of exercise or exercise training stimulate enhancement of cellular stress protection against future insults of oxidative, metabolic and mechanical stressors that could induce injury or disease. This phenomenon has been termed both hormesis and exercise preconditioning (EPC). EPC stimulates transcription factors such as Nrf‐1 and heat shock factor‐1 and up‐regulates gene expression of a cadre of cytosolic (e.g. glutathione peroxidase and heat shock proteins) and mitochondrial adaptive or stress proteins (e.g. manganese superoxide dismutase, mitochondrial KATP channels and peroxisome proliferator activated receptor γ coactivator‐1 (PGC‐1)). Stress response and antioxidant enzyme inducibility with exercise lead to protection against striated muscle damage, oxidative stress and injury. EPC may indeed provide significant clinical protection against ischaemia–reperfusion injury, Type II diabetes and ageing. New molecular mechanisms of protection, such as δ‐opioid receptor regulation and mitophagy, reinforce the notion that mitochondrial adaptations (e.g. heat shock proteins, antioxidant enzymes and sirtuin‐1/PGC‐1 signalling) are central to the protective effects of exercise preconditioning. PMID:27060608

  10. Muscle on a chip: in vitro contractility assays for smooth and striated muscle.

    PubMed

    Grosberg, Anna; Nesmith, Alexander P; Goss, Josue A; Brigham, Mark D; McCain, Megan L; Parker, Kevin Kit

    2012-01-01

    To evaluate the viability of a muscle tissue, it is essential to measure the tissue's contractile performance as well as to control its structure. Accurate contractility data can aid in development of more effective and safer drugs. This can be accomplished with a robust in vitro contractility assay applicable to various types of muscle tissue. The devices developed in this work were based on the muscular thin film (MTF) technology, in which an elastic film is manufactured with a 2D engineered muscle tissue on one side. The tissue template is made by patterning extracellular matrix with microcontact printing. When muscle cells are seeded on the film, they self-organize with respect to the geometric cues in the matrix to form a tissue. Several assays based on the "MTF on a chip" technology are demonstrated. One such assay incorporates the contractility assay with striated muscle into a fluidic channel. Another assay platform incorporates the MTFs in a multi-well plate, which is compatible with automated data collection and analysis. Finally, we demonstrate the possibility of analyzing contractility of both striated and smooth muscle simultaneously on the same chip. In this work, we assembled an ensemble of contractility assays for striated and smooth muscle based on muscular thin films. Our results suggest an improvement over current methods and an alternative to isolated tissue preparations. Our technology is amenable to both primary harvests cells and cell lines, as well as both human and animal tissues. Copyright © 2012 Elsevier Inc. All rights reserved.

  11. Muscle lim protein isoform negatively regulates striated muscle actin dynamics and differentiation.

    PubMed

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

    2014-07-01

    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, whereas 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/cofilin-2-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. The findings of the present study unveil a novel player in muscle physiology and pathophysiology that is implicated in myogenesis as a negative regulator of myotube formation, as well as in differentiated striated muscles as a contributor to sarcomeric integrity. © 2014 FEBS.

  12. 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

  13. Muscle on a Chip: In Vitro Contractility Assays for Smooth and Striated Muscle

    PubMed Central

    Grosberg, Anna; Nesmith, Alexander P.; Goss, Josue A.; Brigham, Mark D.; McCain, Megan L.; Parker, Kevin Kit

    2012-01-01

    Introduction To evaluate the viability of a muscle tissue, it is essential to measure the tissue’s contractile performance as well as to control its structure. Accurate contractility data can aid in development of more effective and safer drugs. This can be accomplished with a robust in vitro contractility assay applicable to various types of muscle tissue. Methods The devices developed in this work were based on the muscular thin film (MTF) technology, in which an elastic film is manufactured with a 2D engineered muscle tissue on one side. The tissue template is made by patterning extracellular matrix with microcontact printing. When muscle cells are seeded on the film, they self-organize with respect to the geometric cues in the matrix to form a tissue. Results Several assays based on the “MTF on a chip” technology are demonstrated. One such assay incorporates the contractility assay with striated muscle into a fluidic channel. Another assay platform incorporates the MTFs in a multi-well plate, which is compatible with automated data collection and analysis. Finally, we demonstrate the possibility of analyzing contractility of both striated and smooth muscle simultaneously on the same chip. Discussion In this work, we assembled an ensemble of contractility assays for striated and smooth muscle based on muscular thin films. Our results suggest an improvement over current methods and an alternative to isolated tissue preparations. Our technology is amenable to both primary harvests cells and cell lines, as well as both human and animal tissues. PMID:22521339

  14. Emerging importance of oxidative stress in regulating striated muscle elasticity.

    PubMed

    Beckendorf, Lisa; Linke, Wolfgang A

    2015-02-01

    The contractile function of striated muscle cells is altered by oxidative/nitrosative stress, which can be observed under physiological conditions but also in diseases like heart failure or muscular dystrophy. Oxidative stress causes oxidative modifications of myofilament proteins and can impair myocyte contractility. Recent evidence also suggests an important effect of oxidative stress on muscle elasticity and passive stiffness via modifications of the giant protein titin. In this review we provide a short overview of known oxidative modifications in thin and thick filament proteins and then discuss in more detail those oxidative stress-related modifications altering titin stiffness directly or indirectly. Direct modifications of titin include reversible disulfide bonding within the cardiac-specific N2-Bus domain, which increases titin stiffness, and reversible S-glutathionylation of cryptic cysteines in immunoglobulin-like domains, which only takes place after the domains have unfolded and which reduces titin stiffness in cardiac and skeletal muscle. Indirect effects of oxidative stress on titin can occur via reversible modifications of protein kinase signalling pathways (especially the NO-cGMP-PKG axis), which alter the phosphorylation level of certain disordered titin domains and thereby modulate titin stiffness. Oxidative stress also activates proteases such as matrix-metalloproteinase-2 and (indirectly via increasing the intracellular calcium level) calpain-1, both of which cleave titin to irreversibly reduce titin-based stiffness. Although some of these mechanisms require confirmation in the in vivo setting, there is evidence that oxidative stress-related modifications of titin are relevant in the context of biomarker design and represent potential targets for therapeutic intervention in some forms of muscle and heart disease.

  15. Calcium movement of sarcoplasmic reticulum from hindlimb suspended muscle

    NASA Astrophysics Data System (ADS)

    Yoshioka, Toshitada; Shirota, Takuhiko; Tazoe, Takashi; Yamashita-Goto, Katsumasa

    1996-02-01

    The function of the sarcoplasmic reticulum (SR) was examined in the slow soleus and fast extensor digitorum longus (EDL) muscles of rats submitted to 14 days of weightlessness produced by hindlimb suspension (HS). Ca 2+ uptake, Ca 2+ release and passive Ca 2+ leakage through the SR membrane were investigated using a method of caffeine-induced contracture on the single mechanically skinned fibers. In the SR of suspended soleus muscles, the rate of Ca 2+ uptake was higher than in the control muscles. However, there was no difference between the suspended and control muscles in the rate of Ca 2+ uptake of the SR in EDL after HS. In soleus muscles, Ca 2+ movements of the SR from the suspended muscle acquired the properties that were similar to those of the control fast muscle. The study of Ca 2+ leakage showed that the velocity and amount of passive Ca 2+ leakage from SR in soleus and EDL were apparently increased after HS. The results suggested that the functional properties of the SR membrane in slow and fast muscles were changed after HS.

  16. Donnan potentials from striated muscle liquid crystals. Lattice spacing dependence.

    PubMed Central

    Aldoroty, R A; Garty, N B; April, E W

    1987-01-01

    Electrochemical potentials were measured as a function of myofilament packing density in crayfish striated muscle. The A-band striations are supramolecular smectic B1 lattice assemblies of myosin filaments and the I-band striations are nematic liquid crystals of actin filaments. Both A- and I-bands generate potentials derived from the fixed charge that is associated with structural proteins. In the reported experiments, filament packing density was varied by osmotically reducing lattice volume. The electrochemical potentials were measured from the A- and I-bands in the relaxed condition over a range of lattice volumes. From the measurements of relative cross-sectional area, unit-cell volume (obtained by low-angle x-ray diffraction) and previously determined effective linear charge densities (Aldoroty, R.A., N.B. Garty, and E.W. April, 1985, Biophys. J., 47:89-96), Donnan potentials can be predicted for any amount of compression. In the relaxed condition, the predicted Donnan potentials correspond to the measured electrochemical potentials. In the rigor condition, however, a net increase in negative charge associated with the myosin filament is observed. The predictability of the data demonstrates the applicability of Donnan equilibrium theory to the measurement of electrochemical potentials from liquid-crystalline systems. Moreover, the relationship between filament spacing and the Donnan potential is consistent with the concept that surface charge provides the necessary electrostatic force to stabilize the myofilament lattice. PMID:3567311

  17. Discontinuity of sarcoplasmic reticulum in the mid-sarcomere region in flight muscle of dragonflies.

    PubMed

    de Eguileor, M; Valvassori, R; Lanzavecchia, G

    1980-01-01

    The sarcoplasmic reticulum organization of dragonfly flight muscles is analyzed, with particular reference to the doubling existing at H-band level. This doubling could be explained as a consequence of a regular discontinuity in the sarcoplasmic reticulum covering myofibrils. In each sarcomere, two sleeves of the sarcoplasmic reticulum seem to overlap forming a telescopic system which can slide outside each other during the lengthening and shortening movements of the fiber.

  18. Characterization of the sarcoplasmic reticulum proteins in the thermogenic muscles of fish.

    PubMed

    Block, B A; O'Brien, J; Meissner, G

    1994-12-01

    Marlins, sailfish, spearfishes, and swordfish have extraocular muscles that are modified into thermogenic organs beneath the brain. The modified muscle cells, called heater cells, lack organized myofibrils and are densely packed with sarcoplasmic reticulum (SR), transverse (T) tubules, and mitochondria. Thermogenesis in the modified extraocular muscle fibers is hypothesized to be associated with increased energy turnover due to Ca2+ cycling at the SR. In this study, the proteins associated with sequestering and releasing Ca2+ from the SR (ryanodine receptor, Ca2+ ATPase, calsequestrin) of striated muscle cells were characterized in the heater SR using immunoblot and immunofluorescent techniques. Immunoblot analysis with a monoclonal antibody that recognizes both isoforms of nonmammalian RYRs indicates that the fish heater cells express only the alpha RYR isoform. The calcium dependency of [3H]ryanodine binding to the RYR isoform expressed in heater indicates functional identity with the non-mammalian alpha RYR isoform. Fluorescent labeling demonstrates that the RYR is localized in an anastomosing network throughout the heater cell cytoplasm. Measurements of oxalate supported 45Ca2+ uptake, Ca2+ ATPase activity, and [32P]phosphoenzyme formation demonstrate that the SR contains a high capacity for Ca2+ uptake via an ATP dependent enzyme. Immunoblot analysis of calsequestrin revealed a significant amount of the Ca2+ binding protein in the heater cell SR. The present study provides the first direct evidence that the heater SR system contains the proteins necessary for Ca2+ release, re-uptake and sequestration, thus supporting the hypothesis that thermogenesis in the modified muscle cells is achieved via an ATP-dependent cycling of Ca2+ between the SR and cytosolic compartments.

  19. The striated muscles in pulmonary arterial hypertension: adaptations beyond the right ventricle.

    PubMed

    Manders, Emmy; Rain, Silvia; Bogaard, Harm-Jan; Handoko, M Louis; Stienen, Ger J M; Vonk-Noordegraaf, Anton; Ottenheijm, Coen A C; de Man, Frances S

    2015-09-01

    Pulmonary arterial hypertension (PAH) is a fatal lung disease characterised by progressive remodelling of the small pulmonary vessels. The daily-life activities of patients with PAH are severely limited by exertional fatigue and dyspnoea. Typically, these symptoms have been explained by right heart failure. However, an increasing number of studies reveal that the impact of the PAH reaches further than the pulmonary circulation. Striated muscles other than the right ventricle are affected in PAH, such as the left ventricle, the diaphragm and peripheral skeletal muscles. Alterations in these striated muscles are associated with exercise intolerance and reduced quality of life. In this Back to Basics article on striated muscle function in PAH, we provide insight into the pathophysiological mechanisms causing muscle dysfunction in PAH and discuss potential new therapeutic strategies to restore muscle dysfunction.

  20. Overexpression of TEAD-1 in transgenic mouse striated muscles produces a slower skeletal muscle contractile phenotype.

    PubMed

    Tsika, Richard W; Schramm, Christine; Simmer, Gretchen; Fitzsimons, Daniel P; Moss, Richard L; Ji, Juan

    2008-12-26

    TEA domain (TEAD) transcription factors serve important functional roles during embryonic development and in striated muscle gene expression. Our previous work has implicated a role for TEAD-1 in the fast-to-slow fiber-type transition in response to mechanical overload. To investigate whether TEAD-1 is a modulator of slow muscle gene expression in vivo, we developed transgenic mice expressing hemagglutinin (HA)-tagged TEAD-1 under the control of the muscle creatine kinase promoter. We show that striated muscle-restricted HA-TEAD-1 expression induced a transition toward a slow muscle contractile protein phenotype, slower shortening velocity (Vmax), and longer contraction and relaxation times in adult fast twitch extensor digitalis longus muscle. Notably, HA-TEAD-1 overexpression resulted in an unexpected activation of GSK-3alpha/beta and decreased nuclear beta-catenin and NFATc1/c3 protein. These effects could be reversed in vivo by mechanical overload, which decreased muscle creatine kinase-driven TEAD-1 transgene expression, and in cultured satellite cells by TEAD-1-specific small interfering RNA. These novel in vivo data support a role for TEAD-1 in modulating slow muscle gene expression.

  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. 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.

  3. Nuclear tropomyosin and troponin in striated muscle: new roles in a new locale?

    PubMed

    Chase, P Bryant; Szczypinski, Mark P; Soto, Elliott P

    2013-08-01

    Tropomyosin and troponin have well known Ca(2+)-regulatory functions in the striated muscle sarcomere. In this review, we summarize experimental evidence that tropomyosin and troponin are localized, with as yet unidentified functional roles, in the striated muscle cell nucleus. We also apply bioinformatics approaches that predict localization of some tropomyosin and troponin to the nucleus, and that SUMOylation could be a covalent modification that modulates their nuclear localization and function. Further, we provide examples of cardiomyopathy mutations that alter the predicted likelihood of nuclear localization and SUMOylation of tropomyosin. These observations suggest novel mechanisms by which cardiomyopathy mutations in tropomyosin and troponin might alter not only cardiac contractility but also nuclear function.

  4. Muscle energy metabolism: structural and functional features in different types of porcine striated muscles.

    PubMed

    Huber, Korinna; Petzold, Johanna; Rehfeldt, Charlotte; Ender, Klaus; Fiedler, Ilse

    2007-01-01

    Striated muscles exhibit a wide range of metabolic activity levels. Heart and diaphragm are muscles with continuous contractile performance, which requires life-long function. In contrast, skeletal muscles like longissimus muscle can adapt metabolism from resting to different stages of exercise. The aim of this study was to compare the morphological features of these three muscles and the expression of genes that are important for energy metabolism. Therefore, histochemical studies were performed for determination of muscle fibre type composition. Oxidative and glycolytic capacity was assessed by measuring isocitrate dehydrogenase (ICDH) and lactate dehydrogenase (LDH) activities. The mRNA expression of glucose transporter 4 (GLUT 4), growth hormone receptor (GHR) and AMP-activated kinase (AMPK) alpha(1) and alpha(2) subunits was studied by semiquantitative Northern blotting. Heart, and to a slightly lesser extent diaphragm were highly oxidative muscles characterised by high expression of oxidative muscle fibres and ICDH activity. Longissimus muscle exhibited the highest percentage of glycolytic fibres and LDH activity. GLUT 4 mRNA was lowest in heart reflecting the dependency of heart muscle on fatty acids as major energy source. Higher expression of GLUT 4 in diaphragm indicated that glucose is an important energy substrate in this oxidative muscle. Highest GLUT 4 expression in longissimus should be essential for the refilling of glycogen stores after exercise. AMPK subunits, which are important stimulators of GLUT 4 protein insertion into the sarcolemma, are also highest expressed in longissimus muscle indicating the strong capacity to adapt energy metabolism to large changes in energy demand. Interestingly, AMPK alpha(1) subunit expression on protein level is strongly restricted to muscle fibres containing type I myosin in this muscle. GHR mRNA expression was also highest in longissimus muscle indicating that an enhanced effect of growth hormone, which is

  5. 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

  6. Multiparity causes uncoordinated activity of pelvic- and perineal-striated muscles and urodynamic changes in rabbits.

    PubMed

    Martínez-Gómez, Margarita; Mendoza-Martínez, Germán; Corona-Quintanilla, Dora Luz; Fajardo, Víctor; Rodríguez-Antolín, Jorge; Castelán, Francisco

    2011-12-01

    Temporal and coordinated activation of pelvic- (pubococcygeous) and perineal- (bulbospongiosus and ischiocavernosus) striated muscles occurs during micturition in female rabbits. We have hypothesized that the coordinated activation of pelvic and perineal muscles is modified during the micturition of young multiparous rabbits. Young virgin and multiparous female chinchilla rabbits were used to simultaneously record cystometrograms and electromyograms of the pubococcygeous, ischocavernosus, and bulbospongiosus muscles. Bladder function was assessed using standard urodynamic variables. The temporal coordination of pelvic- and perineal-striated muscle activity was changed in multiparous rabbits. The cystometrogram recordings were different than those obtained from virgin rabbits, as seen in alterations of the threshold volume, the residual volume, the voiding duration, and the maximum pressure. In rabbits, we find that multiparity causes uncoordinated activity of pubococcygeous, ischiocavernosus, and bulbospongiosus muscles and modifies the urodynamics.

  7. The ESCRT-II proteins are involved in shaping the sarcoplasmic reticulum in C. elegans.

    PubMed

    Lefebvre, Christophe; Largeau, Céline; Michelet, Xavier; Fourrage, Cécile; Maniere, Xavier; Matic, Ivan; Legouis, Renaud; Culetto, Emmanuel

    2016-04-01

    The sarcoplasmic reticulum is a network of tubules and cisternae localized in close association with the contractile apparatus, and regulates Ca(2+)dynamics within striated muscle cell. The sarcoplasmic reticulum maintains its shape and organization despite repeated muscle cell contractions, through mechanisms which are still under investigation. The ESCRT complexes are essential to organize membrane subdomains and modify membrane topology in multiple cellular processes. Here, we report for the first time that ESCRT-II proteins play a role in the maintenance of sarcoplasmic reticulum integrity inC. elegans ESCRT-II proteins colocalize with the sarcoplasmic reticulum marker ryanodine receptor UNC-68. The localization at the sarcoplasmic reticulum of ESCRT-II and UNC-68 are mutually dependent. Furthermore, the characterization of ESCRT-II mutants revealed a fragmentation of the sarcoplasmic reticulum network, associated with an alteration of Ca(2+)dynamics. Our data provide evidence that ESCRT-II proteins are involved in sarcoplasmic reticulum shaping.

  8. Thienylhydrazone derivative increases sarcoplasmic reticulum Ca2+ release in mammalian skeletal muscle.

    PubMed

    Zapata-Sudo, Gisele; Sudo, Roberto T; Maronas, Patricia A; Silva, Gisele L M; Moreira, Orlando R; Aguiar, Marli I S; Barreiro, Eliezer J

    2003-05-30

    3,4-Methylenedioxybenzoyl-2-thienylhydrazone (L-294) is a cardiac inotropic drug whose action is mediated by an increase in intracellular Ca(2+) concentration as a result of enhanced Ca(2+) accumulation in the sarcoplasmic reticulum. In the present study we tested whether this new thienylhydrazone derivative was effective in mammalian skeletal muscle. We investigated the effect of L-294 on the contractility of isolated skeletal muscle, on Ca(2+) uptake and release by sarcoplasmic reticulum in skinned fibers and in membrane vesicles. L-294 increased in a dose-dependent manner tension of isolated rat soleus muscle. In skinned type I fibers, L-294 induced tension and did not alter sarcoplasmic reticulum loading with Ca(2+). L-294 reduced the threshold Ca(2+) to induce Ca(2+) release and did not affect the ATP-dependent accumulation of Ca(2+) by sarcoplasmic reticulum vesicles. These results suggest that L-294 is an inotropic agent in skeletal muscle through an increase in the amount of Ca(2+) released from the sarcoplasmic reticulum.

  9. Transcriptional networks regulating the costamere, sarcomere, and other cytoskeletal structures in striated muscle.

    PubMed

    Estrella, Nelsa L; Naya, Francisco J

    2014-05-01

    Structural abnormalities in striated muscle have been observed in numerous transcription factor gain- and loss-of-function phenotypes in animal and cell culture model systems, indicating that transcription is important in regulating the cytoarchitecture. While most characterized cytoarchitectural defects are largely indistinguishable by histological and ultrastructural criteria, analysis of dysregulated gene expression in each mutant phenotype has yielded valuable information regarding specific structural gene programs that may be uniquely controlled by each of these transcription factors. Linking the formation and maintenance of each subcellular structure or subset of proteins within a cytoskeletal compartment to an overlapping but distinct transcription factor cohort may enable striated muscle to control cytoarchitectural function in an efficient and specific manner. Here we summarize the available evidence that connects transcription factors, those with established roles in striated muscle such as MEF2 and SRF, as well as other non-muscle transcription factors, to the regulation of a defined cytoskeletal structure. The notion that genes encoding proteins localized to the same subcellular compartment are coordinately transcriptionally regulated may prompt rationally designed approaches that target specific transcription factor pathways to correct structural defects in muscle disease.

  10. CAS-1, a C. elegans cyclase-associated protein, is required for sarcomeric actin assembly in striated muscle.

    PubMed

    Nomura, Kazumi; Ono, Kanako; Ono, Shoichiro

    2012-09-01

    Assembly of contractile apparatuses in striated muscle requires precisely regulated reorganization of the actin cytoskeletal proteins into sarcomeric organization. Regulation of actin filament dynamics is one of the essential processes of myofibril assembly, but the mechanism of actin regulation in striated muscle is not clearly understood. Actin depolymerizing factor (ADF)/cofilin is a key enhancer of actin filament dynamics in striated muscle in both vertebrates and nematodes. Here, we report that CAS-1, a cyclase-associated protein in Caenorhabditis elegans, promotes ADF/cofilin-dependent actin filament turnover in vitro and is required for sarcomeric actin organization in striated muscle. CAS-1 is predominantly expressed in striated muscle from embryos to adults. In vitro, CAS-1 binds to actin monomers and enhances exchange of actin-bound ATP/ADP even in the presence of UNC-60B, a muscle-specific ADF/cofilin that inhibits the nucleotide exchange. As a result, CAS-1 and UNC-60B cooperatively enhance actin filament turnover. The two proteins also cooperate to shorten actin filaments. A cas-1 mutation is homozygous lethal with defects in sarcomeric actin organization. cas-1-mutant embryos and worms have aggregates of actin in muscle cells, and UNC-60B is mislocalized to the aggregates. These results provide genetic and biochemical evidence that cyclase-associated protein is a critical regulator of sarcomeric actin organization in striated muscle.

  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. Cannabinoid receptor antagonist-induced striated muscle toxicity and ethylmalonic-adipic aciduria in beagle dogs.

    PubMed

    Tomlinson, Lindsay; Tirmenstein, Mark A; Janovitz, Evan B; Aranibar, Nelly; Ott, Karl-Heinz; Kozlosky, John C; Patrone, Laura M; Achanzar, William E; Augustine, Karen A; Brannen, Kimberly C; Carlson, Kenneth E; Charlap, Jeffrey H; Dubrow, Katherine M; Kang, Liya; Rosini, Laura T; Panzica-Kelly, Julieta M; Flint, Oliver P; Moulin, Frederic J; Megill, John R; Zhang, Haiying; Bennett, Michael J; Horvath, Joseph J

    2012-10-01

    Ibipinabant (IBI), a potent cannabinoid-1 receptor (CB1R) antagonist, previously in development for the treatment of obesity, causes skeletal and cardiac myopathy in beagle dogs. This toxicity was characterized by increases in muscle-derived enzyme activity in serum and microscopic striated muscle degeneration and accumulation of lipid droplets in myofibers. Additional changes in serum chemistry included decreases in glucose and increases in non-esterified fatty acids and cholesterol, and metabolic acidosis, consistent with disturbances in lipid and carbohydrate metabolism. No evidence of CB1R expression was detected in dog striated muscle as assessed by polymerase chain reaction, immunohistochemistry, Western blot analysis, and competitive radioligand binding. Investigative studies utilized metabonomic technology and demonstrated changes in several intermediates and metabolites of fatty acid metabolism including plasma acylcarnitines and urinary ethylmalonate, methylsuccinate, adipate, suberate, hexanoylglycine, sarcosine, dimethylglycine, isovalerylglycine, and 2-hydroxyglutarate. These results indicated that the toxic effect of IBI on striated muscle in beagle dogs is consistent with an inhibition of the mitochondrial flavin-containing enzymes including dimethyl glycine, sarcosine, isovaleryl-CoA, 2-hydroxyglutarate, and multiple acyl-CoA (short, medium, long, and very long chain) dehydrogenases. All of these enzymes converge at the level of electron transfer flavoprotein (ETF) and ETF oxidoreductase. Urinary ethylmalonate was shown to be a biomarker of IBI-induced striated muscle toxicity in dogs and could provide the ability to monitor potential IBI-induced toxic myopathy in humans. We propose that IBI-induced toxic myopathy in beagle dogs is not caused by direct antagonism of CB1R and could represent a model of ethylmalonic-adipic aciduria in humans.

  13. 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. © 2016 The Author(s).

  14. Polarization of Tryptophan Fluorescence from Single Striated Muscle Fibers

    PubMed Central

    dos Remedios, C. G.; Millikan, R. G. C.; Morales, M. F.

    1972-01-01

    Instrumentation has been developed to detect rapidly the polarization of tryptophan fluorescence from single muscle fibers in rigor, relaxation, and contraction. The polarization parameter (P⊥) obtained by exiciting the muscle tryptophans with light polarized perpendicular to the long axis of the muscle fiber had a magnitude P⊥ (relaxation) > P⊥ (contraction) > P⊥ (rigor) for the three types of muscle fibers examined (glycerinated rabbit psoas, glycerinated dorsal longitudinal flight muscle of Lethocerus americanus, and live semitendinosus of Rana pipiens). P⊥ from single psoas fibers in rigor was found to increase as the sarcomere length increased but in relaxed fibers P⊥ was independent of sarcomere length. After rigor, pyrophosphate produced little or no change in P⊥, but following an adenosine triphosphate (ATP)-containing solution, pyrophosphate produced a value of P⊥ that fell between the contraction and relaxation values. Sinusoidal or square wave oscillations of the muscle of amplitude 0.5–2.0% of the sarcomere length and frequency 1, 2, or 5 Hz were applied in rigor when the myosin cross-bridges are considered to be firmly attached to the thin filaments. No significant changes in P⊥ were observed in either rigor or relaxation. The preceding results together with our present knowledge of tryptophan distribution in the contractile proteins has led us to the conclusion that the parameter P⊥ is a probe of the contractile state of myosin which is probably sensitive to the orientation of the myosin S1 subfragment. PMID:4332133

  15. Synapse formation between clonal neuroblastoma X glioma hybrid cells and striated muscle cells.

    PubMed Central

    Nelson, P; Christian, C; Nirenberg, M

    1976-01-01

    Clonal neuroblastoma X glioma hybrid cells were shown to form synapses with cultured, striated muscle cells. The properties of the synapses between hybrid and muscle cells were similar to those of the normal, neuromuscular synapse at an early stage of development. The number of synapses formed and the efficiency of transmission across synapses were found to be regulated, apparently independently, by components in the culture medium. Under appropriate conditions synapses were found with 20% of the hybrid-muscle cell pairs examined; thus, the hybrid cells form synapses with relatively high frequency. Images PMID:1061105

  16. 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.

  17. 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.

  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. Reduced effect of caffeine on twitch contraction of oesophageal striated muscle from stroke-prone spontaneously hypertensive rats.

    PubMed

    Sekiguchi, Fumiko; Kawata, Kyoko; Shimamura, Keiichi; Sunano, Satoru

    2003-04-01

    1. There are known differences in the sensitivity to caffeine between skeletal muscle (soleus) of normotensive Wistar-Kyoto (WKY) rats and spontaneously hypertensive rats (SHR). The present study was performed in order to examine differences in the effects of caffeine on twitch contraction between visceral striated muscle using the outer layer of the oesophagus from WKY rats and stroke-prone SHR (SHRSP). 2. Caffeine, at concentrations ranging from 0.3 to 10 mmol/L, exhibited potentiating effects on twitch contraction in preparations from both WKY rats and SHRSP. The potentiating effect of caffeine was markedly less prominent in preparations from SHRSP compared with preparations from WKY rats. 3. The rate of contraction and relaxation, the time to peak tension and 80% relaxation time were not significantly altered by caffeine at concentrations lower than 3 mmol/L in preparations from either strain. 4. With 10 mmol/L caffeine, the rate of relaxation was markedly reduced and the 80% relaxation time was prolonged, with no significant changes in the rate of contraction, in preparations from WKY rats. These changes were significantly smaller in preparations from SHRSP. 5. The duration of the action potential was greater in preparations from SHRSP than in preparations from WKY rats, although the membrane potential and the amplitude of the action potential were not significantly different between preparations from WKY rats and SHRSP. 6. Caffeine, at 10 mmol/L, prolonged the duration of the action potential in preparations from both strains. The effect of caffeine was not different between preparations from WKY rats and SHRSP. 7. The results of the present study suggest that caffeine augments release of Ca2+ from the sarcoplasmic reticulum (SR) at low concentrations and attenuates Ca2+ re-uptake at 10 mmol/L. Decreased reactivity of SR to caffeine may be a cause of the lesser potentiation of twitch contraction by caffeine in preparations from SHRSP.

  20. Loss of skeletal muscle strength by ablation of the sarcoplasmic reticulum protein JP45.

    PubMed

    Delbono, Osvaldo; Xia, Jinyu; Treves, Susan; Wang, Zhong-Min; Jimenez-Moreno, Ramon; Payne, Anthony M; Messi, María Laura; Briguet, Alexandre; Schaerer, Florian; Nishi, Miyuki; Takeshima, Hiroshi; Zorzato, Francesco

    2007-12-11

    Skeletal muscle constitutes approximately 40% of the human body mass, and alterations in muscle mass and strength may result in physical disability. Therefore, the elucidation of the factors responsible for muscle force development is of paramount importance. Excitation-contraction coupling (ECC) is a process during which the skeletal muscle surface membrane is depolarized, causing a transient release of calcium from the sarcoplasmic reticulum that activates the contractile proteins. The ECC machinery is complex, and the functional role of many of its protein components remains elusive. This study demonstrates that deletion of the gene encoding the sarcoplasmic reticulum protein JP45 results in decreased muscle strength in young mice. Specifically, this loss of muscle strength in JP45 knockout mice is caused by decreased functional expression of the voltage-dependent Ca(2+) channel Ca(v)1.1, which is the molecule that couples membrane depolarization and calcium release from the sarcoplasmic reticulum. These results point to JP45 as one of the molecules involved in the development or maintenance of skeletal muscle strength.

  1. Fine structures in the light diffraction pattern of striated muscle.

    PubMed

    Leung, A F

    1984-10-01

    Single skeletal muscle fibres of frog were illuminated with a He-Ne, argon-ion or rhodamine 6G dye laser. The fine structures lying within the diffraction columns moved parallel to the fibre axis without changing their pattern when either the wavelength or the incident angle of the laser beam was varied, or when the fibre was stretched slightly. However, their pattern remained nearly constant when the fibre was submerged in hypotonic or hypertonic solution. As the illumination of about 1 mm or 0.1 mm width scanned along the length of the fibre, new structures emerged while others faded away giving rise to the notion that the diffraction columns were moving in the direction of the scan. A decrease in the illumination width caused the structures lying on the periphery of the diffraction column to disappear and the width of the remaining structures to increase. Measurements rule out the existence of large diffraction planes in these muscles. In addition, they indicate that the fine structures come from the diffraction of the whole rather than independent components of the illuminated volume. The origin of the fine structures is explained by two diffraction models.

  2. Kinetic studies of Ca2+ release from sarcoplasmic reticulum of normal and malignant hyperthermia susceptible pig muscles.

    PubMed

    Kim, D H; Sreter, F A; Ohnishi, S T; Ryan, J F; Roberts, J; Allen, P D; Meszaros, L G; Antoniu, B; Ikemoto, N

    1984-09-05

    The time-course of Ca2+ release from sarcoplasmic reticulum isolated from muscles of normal pigs and those of pigs susceptible to malignant hyperthermia were investigated using stopped-flow spectrophotometry and arsenazo III as a Ca2+ indicator. Several methods were used to trigger Ca2+ release: (a) addition of halothane (e.g., 0.2 mM); (b) an increase of extravesicular Ca2+ concentration ([Ca2+0]); (c) a combination of (a) and (b), and (d) replacement of ions (potassium gluconate with choline chloride) to produce membrane depolarization. The initial rates of Ca2+ release induced by either halothane or Ca2+ alone, or both, are at least 70% higher in malignant hyperthermic sarcoplasmic reticulum than in normal. The amount of Ca2+ released by halothane at low [Ca2+0] in malignant hyperthermic sarcoplasmic reticulum is about twice as large as in normal sarcoplasmic reticulum. Membrane depolarization led to biphasic Ca2+ release in both malignant hyperthermic and normal sarcoplasmic reticulum, the rate constant of the rapid phase of Ca2+ release induced by membrane depolarization being significantly higher in malignant hyperthermic sarcoplasmic reticulum (k = 83 s-1) than in normal (k = 37 s-1). Thus, all types of Ca2+ release investigated (a, b, c and d) have higher rates in malignant hyperthermic sarcoplasmic reticulum than normal sarcoplasmic reticulum. These results suggest that the putative Ca2+ release channels located in the sarcoplasmic reticulum are altered in malignant hyperthermic sarcoplasmic reticulum.

  3. Fiber type characterization of striated muscles related to micturition in female rabbits.

    PubMed

    López-García, Kenia; Mariscal-Tovar, Silvia; Martínez-Gómez, Margarita; Jiménez-Estrada, Ismael; Castelán, Francisco

    2014-04-01

    Pelvic and perineal striated muscles are relevant for reproduction and micturition in female mammals. Damage to these muscles is associated with pelvic organ prolapse and stress urinary incontinence. The fiber type composition of skeletal muscle influences the susceptibility for damage and/or regeneration. The aim of the present study was to determine the fiber type composition of a perineal muscle, the bulbospongiosus, and a pelvic muscle, the pubococcygeus. Both muscles were harvested from adult female rabbits (8-10 months old). NADH-TR (nicotinamide adenine dinucleotide tetrazolium reductase) histochemistry was undertaken to identify oxidative and glycolytic muscle fibers. Alkaline (pH 9.4) ATP-ase (actomyosin adenosine triphosphatase) histochemistry was used to classify type I, type IIb or type IIa/IId muscle fibers. Results showed that the content of glycolytic fibers in the bulbospongiosus muscle was higher than that of oxidative fibers. Meanwhile, the opposite was true for the pubococcygeus. In the bulbospongiosus muscle, the content of type IIb muscle fibers was higher than that of type I, but was similar to that of type IIa/IId. In contrast, the content of each fiber type was similar in the pubococcygeus muscle. The relative proportion of fibers in bulbospongiosus and pubococcygeus muscles is consistent with their function during voiding and storage phases of micturition.

  4. 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.

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

    PubMed

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

    2010-07-15

    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 mum 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' approximately 0.25-0.5 microm 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 approximately 25 nm in diameter and approximately 2.8 microm 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 degrees 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 V(max) (2.15 +/-0.26 L(0) s(-1), where L(0) 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/V(max) 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, L(R)) and at low strain rates (from 0.16 to 0.91 L(R) s(-1) ), corresponding to a range of V/V(max) from 0.073 to 0.42. During the exhalant phase of the jet the range of

  6. 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

  7. Ultrastructure of intramural ganglia in the striated muscle portions of the guinea pig oesophagus

    PubMed Central

    MORIKAWA, SHUNICHI; KOMURO, TERUMASA

    1999-01-01

    The ultrastructure of the myenteric plexus located in the striated muscle portion of the guinea pig oesophagus was examined and compared with that of the plexus associated with the smooth muscle portion of the rest of the digestive tract. The oesophageal ganglia had essentially the same architecture as those of the smooth muscle portion, such as a compact neuropil without the intervention of connective tissue and blood vessels. Some features, however, were particular to the striated muscle part of the oesophagus. It was clearly demonstrated that myelinated fibres, probably sensory terminals of vagal origin, join the myenteric ganglia. Synapses and terminal varicosities are sparsely distributed within the ganglia and fewer morphological types of axon varicosities could be distinguished compared with other regions. Glial cells are well developed in the oesophageal myenteric ganglia. These cells outnumber the ganglion cells, having a higher ratio than in the lower digestive tract, and form numerous cytoplasmic lamellar processes. The lamellar processes, located at the surface of the ganglia, considerably reduce the area of neuronal membrane which directly contacts the basal lamina. The role of these lamellar processes in the oesophageal ganglia is discussed. PMID:10473298

  8. Experimental Periodontitis in the Potentialization of the Effects of Immobilism in the Skeletal Striated Muscle.

    PubMed

    Leite, Marcela Aparecida; de Mattia, Tatiane Morgenstern; Kakihata, Camila Mayumi Martin; Bortolini, Bruna Martinazzo; de Carli Rodrigues, Pedro Henrique; Bertolini, Gladson Ricardo Flor; Brancalhão, Rose Meire Costa; Ribeiro, Lucinéia de Fátima Chasko; Nassar, Carlos Augusto; Nassar, Patrícia Oehlmeyer

    2017-08-18

    This study aims to evaluate if ligature-induced periodontitis can potentiates the deleterious effects of immobilization in the skeletal striated muscle, contributing to the development of muscle atrophy due to disuse. Forty Wistar rats were divided into four groups: (1) Control Group (CG), (2) Periodontal Disease (PDG), (3) Immobilized (IG), and (4) Immobilized with Periodontal Disease (IPDG). Periodontal disease was induced for 30 days, with ligature method, and the immobilization was performed with cast bandage for 15 days. Prior to euthanasia, nociceptive threshold and muscular grasping force were evaluated. Afterwards, the soleus muscle was dissected and processed for sarcomere counting and morphological/morphometric analysis. For data analysis, was used the one-way ANOVA and post-test Tukey (p < 0.05). The IG and IPDG presented lower muscle weight, lower muscular grip strength, and less number of sarcomeres compared to CG. The PDG showed reduction of muscle strength and nociceptive threshold after 15 days of periodontal disease and increased connective tissue compared to CG. The IPDG presented lower muscle length and nociceptive threshold. The IG presented reduction in cross-sectional area and smaller diameter, increase in the number of nuclei and a nucleus/fiber ratio, decrease in the number of capillaries and capillary/fiber ratio, with increase in connective tissue. The IPDG had increased nucleus/fiber ratio, decreased capillaries, and increased connective tissue when compared to the IG. The IPDG presented greater muscle tissue degeneration and increased inflammatory cells compared to the other groups. Ligature-induced periodontitis potentiated the deleterious effects of immobilization of the skeletal striated muscle.

  9. 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.

  10. An early post-traumatic reaction of lymph-heart striated muscle fibers in adult frog Rana temporaria during the first postoperative week: An electron microscopic and autoradiographic study.

    PubMed

    Krylova, Marina I; Bogolyubov, Dmitry S

    2015-12-01

    According to the current opinion, lymph-heart striated muscle represents a specialized type of skeletal muscles in frogs. Here, we studied muscle fibers in mechanically damaged lymph hearts during the first postoperative week using electron-microscopic autoradiography. We present evidence that both, the satellite cells and pre-existing muscle fibers bordering the site of injury, contribute directly to the lymph-heart muscle regeneration. Several muscle fibers located in the vicinity of the damaged area displayed features of nuclear and sarcoplasmic activation. We also observed ultrastructural changes indicating activation of a few satellite cells, namely decondensation of chromatin, enlargement of nuclei and nucleoli, appearance of free ribosomes and rough endoplasmic reticulum tubules in the cytoplasm. Electron-microscopic autoradiography showed that 4 h after single (3)H-thymidine administration on the seventh day after injury not only the activated satellite cells, but also some nuclei of myofibers bordering the injured zone are labeled. We showed that both, the myonuclei of fibers displaying the signs of degenerative/reparative processes in the sarcoplasm and the myonuclei of the fibers enriched with highly organized myofibrils, can re-enter into the S-phase. Our results indicate that the nuclei of lymph-heart myofibers can reactivate DNA synthesis during regenerative myogenesis, unlike the situation in regenerating frog skeletal muscle where myogenic cells do not synthesize DNA at the onset of myofibrillogenesis.

  11. 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.

  12. 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

  13. In vitro binding of dantrolene to sarcoplasmic reticulum of rabbit skeletal muscle.

    PubMed

    Dehpour, A R; Mofakham, S; Mahmoudian, M

    1982-03-15

    Dantrolene upon binding to microsomes containing sarcoplasmic reticulum of rabbit thigh muscle exhibits a fluorescence with emission at 490 nm, which shows a blue shift of 35 nm compared with its fluorescence in ethylacetate. Using fluorescence techniques, dantrolene binding to microsomes isolated from rabbit thigh muscle was investigated. From Scatchard plots of binding studies, the association constant (Kass) and the number of binding sites of dantrolene to sarcoplasmic reticulum were calculated, which was found to be 9.6 X 10(4) M-1 and 1.71 mumole/g of membrane proteins, respectively. In the presence of verapamil (1.25 X 10(-4) M), another calcium antagonist, the binding of dantrolene to microsomes was enhanced. However, at a high concentration of verapamil (3.75 X 10(-4) M), the Scatchard plot of dantrolene binding was found to be biphasic.

  14. 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.

  15. Transient kinetics of Ca2+ transport of sarcoplasmic reticulum. A comparison of cardiac and skeletal muscle.

    PubMed

    Sumida, M; Wang, T; Mandel, F; Froehlich, J P; Schwartz, A

    1978-12-25

    Current evidence supports similar functions and mechanisms for cardiac sarcoplasmic reticulum (CSR) as for skeletal sarcoplasmic reticulum (SSR). It is thought that the slower relaxation rate of cardiac muscle compared to fast skeletal muscle reflects the lower ATPase activity and calcium transport of CSR. Possible quantitative differences is phosphorylation, dephosphorylation, and calcium transport of the isolated preparations are studied using a quench-flow apparatus. The results show that both CSR and SSR bind calcium tightly in the absence of ATP, and coupling of E approximately P formation and calcium transport occurs in the transient phase of ATP hydrolysis. The rate of phosphorylation (t-1/2 - 10 ms) of sarcoplasmic reticulum (SR) preloaded with calcium is the same for cardiac and skeletal preparations. However, the rates of dissociation of extra vesicular calcium (10 s-1 versus 15 s-1), phosphorylation of calcium-free SR, and dephosphorylation of E approximately P (8 s-1 versus 12 s-1) are lower for CSR than for SSR. By computer simulation, the apparent rate constants associated with the reduced rates of phosphorylation of calcium-free SR were: 12 s-1 for CSR and 63 s-1 for SSR in the presence of high Mg2+. The difference in the rates may be partly responsible for the lower levels of ATPase and calcium transport activity with characterize cardiac muscle preparations.

  16. Oesophageal tone and sensation in the transition zone between proximal striated and distal smooth muscle oesophagus.

    PubMed

    Karamanolis, G; Stevens, W; Vos, R; Tack, J; Clave, P; Sifrim, D

    2008-04-01

    Previous studies have shown that the proximal striated muscle oesophagus is less compliant and more sensitive than the distal smooth muscle oesophagus. Conventional and high resolution manometry described a transition zone between striated and smooth muscle oesophagus. We aimed to evaluate oesophageal tone and sensitivity at the transition zone of oesophagus in healthy volunteers. In 18 subjects (seven men, mean age: 28 years) an oesophageal barostat study was performed. Tone and sensitivity were assessed using stepwise isobaric distensions with the balloon located at transition zone and at distal oesophagus in random order. To study the effect induced on transition zone by a previous distension at the distal oesophagus and vice versa, identical protocol was repeated after 7 days with inverted order. Initial distension of a region is referred to as 'naïf' distension and distension of a region following the distension of the other segment as 'primed' distension. Assessment of three oesophageal symptoms (chest pain, heartburn and 'other') was obtained at the end of every distension step. Compliance was significantly higher in the transition zone than in the distal oesophagus (1.47 +/- 0.14 vs 1.09 +/- 0.09 mL mmHg(-1), P = 0.03) after 'naif' distensions. This difference was not observed during 'primed' distensions. Higher sensitivity at transition zone level was found in 11/18 (61%) subjects compared to 6/18 (33%, P < 0.05) at smooth muscle oesophagus. Chest pain and 'other' symptom were more often induced by distention of the transition zone, whereas heartburn was equally triggered by distension of either region. The transition zone is more complaint and more sensitive than smooth muscle oesophagus.

  17. 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.

  18. The role of pelvic and perineal striated muscles in urethral function during micturition in female rabbits.

    PubMed

    Corona-Quintanilla, Dora L; Zempoalteca, René; Arteaga, Lourdes; Castelán, Francisco; Martínez-Gómez, Margarita

    2014-04-01

    To evaluate the role of pelvic and perineal striated muscles on urethral function during micturition. Pubococcygeus, or both bulbospongiosus and ischiocavernosus muscles were electrically stimulated during the voiding phase of micturition, and bladder and urethral pressure were simultaneously recorded in urethane anesthetized female rabbits. Bladder and urethral function were assessed measuring urodynamic and urethral pressure variables obtained before and during the stimulation of muscles. Two-tail paired t-tests were carried out in order to determine significant differences (P < 0.05) between groups. Electric stimulation of the pubococcygeus during voiding decreased voiding efficiency and voided volume, whereas residual volume, the duration of voiding, the interval between bladder contraction and urethral resistance increased. Simultaneously, there was an increase in maximum urethral pressure, as well as an increase in the pressure to return to baseline and in the pressure required to close the urethra. Electrical stimulation of bulbospongiosus and ischiocavernosus muscles increased voiding efficiency, voiding duration, and the maximal pressure in bladder. Meanwhile, the maximal urethral pressure, the time related to the rise of urethral pressure, and the urethral pressure required to close the urethra decreased. The stimulation of pelvic and perineal muscles have opposing roles in urethral function during micturition. Pubococcygeus muscles facilitate urethral closure, while they inhibit bladder contraction. In contrast, bulbospongiosus and ischiocavernosus muscles prevent urethral contraction while they promote bladder contraction. © 2013 Wiley Periodicals, Inc.

  19. Effects of procaine on calcium accumulation by the sarcoplasmic reticulum of mechanically disrupted rat cardiac muscle.

    PubMed Central

    Stephenson, D G; Wendt, I R

    1986-01-01

    The ability of the sarcoplasmic reticulum of skinned cardiac muscle of the rat to accumulate and release Ca2+ was studied in the presence and absence of procaine. Ca2+ accumulation was estimated from the magnitude of the caffeine- (30 mM) induced force transient in a weakly Ca2+ buffered solution. The relative area under the caffeine-induced force transient was up to 4-fold greater when 5 mM-procaine had been present during the preceding period of Ca2+ loading, than that after an equivalent period of Ca2+ loading in the absence of procaine. Procaine antagonized the caffeine-induced release of Ca2+ when present in the Ca2+ releasing solution, however, the ability of procaine to attenuate the caffeine-induced Ca2+ release diminished as the extent to which the sarcoplasmic reticulum was loaded with Ca2+ increased. In the presence of 1 mM-Mg2+ procaine also markedly attenuated the small spontaneous force oscillations (5-10% P0) associated with the cyclic release and reuptake of Ca2+ by the sarcoplasmic reticulum. When the Mg2+ concentration was reduced to 0.1 mM, procaine initially suppressed the small spontaneous oscillations in force, however, large force oscillations (40-80% P0) of lower frequency were invariably initiated after 20-60 s exposure to 5 mM-procaine. Procaine (5 mM) produced a slight shift (approximately 0.04 pCa unit) of the force-pCa relation toward lower Ca2+ concentrations. This effect is too small to influence in any substantial way the apparent effects of procaine on the sarcoplasmic reticulum. The results indicate that whilst procaine is indeed able to suppress Ca2+ release under certain circumstances, in its presence the net accumulation of Ca2+ by the sarcoplasmic reticulum can be markedly enhanced. PMID:3746672

  20. Molecular characterization and expression pattern of the porcine STARS, a striated muscle-specific expressed gene.

    PubMed

    Peng, Y B; Guan, H P; Fan, B; Zhao, S H; Xu, X W; Li, K; Zhu, M J; Yerle, M; Liu, B

    2008-10-01

    STARS (striated muscle activator of Rho signaling) promotes the nuclear localization of MRTFs and mediates SRF transcription, which provides a potential muscle-specific mechanism for linking changes in the actin cytoskeleton structure with muscle gene expression. In this study, the full-length cDNA of the porcine STARS was cloned. The open reading frame of this gene contains 1,155 bp and encodes a protein of 384 amino acids, which is 79, 73, and 77% identical with human, mouse, and rat STARS genes, respectively. RT-PCR revealed that STARS is specifically expressed in heart and skeletal muscles. STARS is also distinctly different in different muscle developmental stages. The result indicates that its expression increased gradually from 33 dpc (days postcoitum) to postnatal muscles, and peaked 28 days postnatal. The porcine STARS was mapped to SSC4p13 using the somatic cell hybrid panel and the radiation hybrid panel IMpRH (LOD score 11.98). The data show that STARS is closely linked to marker SW871. A T/G single nucleotide polymorphism in the coding sequence, detected as Bsh1236I PCR-RFLP, displays allele frequency differences in six pig breeds.

  1. Regulation of structure and function of sarcomeric actin filaments in striated muscle of the nematode Caenorhabditis elegans

    PubMed Central

    Ono, Shoichiro

    2014-01-01

    The nematode Caenorhabditis elegans has been used as a valuable system to study structure and function of striated muscle. The body wall muscle of C. elegans is obliquely striated muscle with highly organized sarcomeric assembly of actin, myosin, and other accessary proteins. Genetic and molecular biological studies in C. elegans have identified a number of genes encoding structural and regulatory components for the muscle contractile apparatuses, and many of them have counterparts in mammalian cardiac and skeletal muscles or striated muscles in other invertebrates. Applicability of genetics, cell biology, and biochemistry has made C. elegans an excellent system to study mechanisms of muscle contractility and assembly and maintenance of myofibrils. This review focuses on the regulatory mechanisms of structure and function of actin filaments in the C. elegans body wall muscle. Sarcomeric actin filaments in C. elegans muscle are associated with the troponin-tropomyosin system that regulates the actin-myosin interaction. Proteins that bind to the side and ends of actin filaments support ordered assembly of thin filaments. Furthermore, regulators of actin dynamics play important roles in initial assembly, growth, and maintenance of sarcomeres. The knowledge acquired in C. elegans can serve as bases to understand the basic mechanisms of muscle structure and function. PMID:25125169

  2. Seasonal changes in proteolytic activity of calpains in striated muscles of long-tailed ground squirrel Spermophilus undulatus.

    PubMed

    Popova, S S; Vikhlyantsev, I M; Zakharova, N M; Podlubnaya, Z A; Fesenko, E E

    2017-01-01

    Seasonal changes in proteolytic activity and content of calpains in striated muscles of the longtailed ground squirrel Spermophilus undulatus were studied by casein zymography and Western blotting analysis. The results testify to hyperactivation of calpain proteases in the skeletal muscles of awakened animals during the "winter" activity. The observed changes are discussed in the context of adaptation of skeletal muscles of long-tailed ground squirrels to hibernation.

  3. Contractile properties of the striated adductor muscle in the bay scallop Argopecten irradians at several temperatures.

    PubMed

    Olson, J M; Marsh, R L

    1993-03-01

    The isometric and isotonic contractile properties of the cross-striated adductor muscle of the bay scallop (Argopecten irradians) were measured in vitro at 10, 15 and 20 degrees C. The length at which twitch force was maximal as a function of the closed length in situ (L0/Lcl) averaged 1.38 +/- 0.01 (mean +/- S.E.M.) at 10 degrees C. This length is very close to the typical length at maximum gape during natural swimming at this temperature. Passive force was very low over the range of lengths measured here; at L0, passive force averaged approximately 0.08 N cm-2, or only 0.5% of the corresponding peak twitch force. The mean peak isometric twitch force (Ptw,max) at 10 degrees C was 21.43 +/- 0.68 N cm-2 (S.E.M.), and the ratio of peak twitch force to tetanic force (Ptw,max/P0) averaged 0.89 +/- 0.01. Temperature did not affect either twitch force (Ptw), once fatigue was taken into account, or Ptw,max/P0. In contrast, the time-related properties of twitch contractions (latent period, tL; time to peak tension, tPtw; and time from peak tension to half-relaxation, t50%R) were positively modified by temperature at all temperatures measured (Q10 > 1.8). All three properties were more temperature-sensitive over the range 10-15 degrees C than over the range 15-20 degrees C. The force-velocity relationships of the striated adductor muscle were fitted to the hyperbolic-linear (HYP-LIN) equation. The force-velocity curves of the striated adductor muscle of the scallop were strongly influenced by temperature. Maximal velocity at zero force (Vmax), and therefore maximal power output, increased significantly with temperature. The Q10 over the temperature range 10-15 degrees C (1.42) was significantly lower than that over the range 15-20 degrees C (2.41). The shape of the force-velocity relationship, assessed through comparisons of the power ratio (Wmax/VmaxP0), was not influenced by temperature.

  4. Cannabinoid CB1 Receptors Are Localized in Striated Muscle Mitochondria and Regulate Mitochondrial Respiration

    PubMed Central

    Mendizabal-Zubiaga, Juan; Melser, Su; Bénard, Giovanni; Ramos, Almudena; Reguero, Leire; Arrabal, Sergio; Elezgarai, Izaskun; Gerrikagoitia, Inmaculada; Suarez, Juan; Rodríguez De Fonseca, Fernando; Puente, Nagore; Marsicano, Giovanni; Grandes, Pedro

    2016-01-01

    The cannabinoid type 1 (CB1) receptor is widely distributed in the brain and peripheral organs where it regulates cellular functions and metabolism. In the brain, CB1 is mainly localized on presynaptic axon terminals but is also found on mitochondria (mtCB1), where it regulates cellular respiration and energy production. Likewise, CB1 is localized on muscle mitochondria, but very little is known about it. The aim of this study was to further investigate in detail the distribution and functional role of mtCB1 in three different striated muscles. Immunoelectron microscopy for CB1 was used in skeletal muscles (gastrocnemius and rectus abdominis) and myocardium from wild-type and CB1-KO mice. Functional assessments were performed in mitochondria purified from the heart of the mice and the mitochondrial oxygen consumption upon application of different acute delta-9-tetrahydrocannabinol (Δ9-THC) concentrations (100 nM or 200 nM) was monitored. About 26% of the mitochondrial profiles in gastrocnemius, 22% in the rectus abdominis and 17% in the myocardium expressed CB1. Furthermore, the proportion of mtCB1 versus total CB1 immunoparticles was about 60% in the gastrocnemius, 55% in the rectus abdominis and 78% in the myocardium. Importantly, the CB1 immunolabeling pattern disappeared in muscles of CB1-KO mice. Functionally, acute 100 nM or 200 nM THC treatment specifically decreased mitochondria coupled respiration between 12 and 15% in wild-type isolated mitochondria of myocardial muscles but no significant difference was noticed between THC treated and vehicle in mitochondria isolated from CB1-KO heart. Furthermore, gene expression of key enzymes involved in pyruvate synthesis, tricarboxylic acid (TCA) cycle and mitochondrial respiratory chain was evaluated in the striated muscle of CB1-WT and CB1-KO. CB1-KO showed an increase in the gene expression of Eno3, Pkm2, and Pdha1, suggesting an increased production of pyruvate. In contrast, no significant difference was

  5. 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

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

    PubMed

    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

    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. 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. These findings position STARS as an important regulator of skeletal muscle growth and regeneration.

  7. A theory on auto-oscillation and contraction in striated muscle.

    PubMed

    Sato, Katsuhiko; Ohtaki, Masako; Shimamoto, Yuta; Ishiwata, Shin'ichi

    2011-05-01

    It is widely accepted that muscle cells take either force-generating or relaxing state in an all-or-none fashion through the so-called excitation-contraction coupling. On the other hand, the membrane-less contractile apparatus takes the third state, i.e., the auto-oscillation (SPOC) state, at the activation level that is intermediate between full activation and relaxation. Here, to explain the dynamics of all three states of muscle, we construct a novel theoretical model based on the balance of forces not only parallel but also perpendicular to the long axis of myofibrils, taking into account the experimental fact that the spacing of myofilament lattice changes with sarcomere length and upon contraction. This theory presents a phase diagram composed of several states of the contractile apparatus and explains the dynamic behavior of SPOC, e.g., periodical changes in sarcomere length with the saw-tooth waveform. The appropriate selection of the constant of the molecular friction due to the cross-bridge formation can explain the difference in the SPOC periods observed under various activating conditions and in different muscle types, i.e., skeletal and cardiac. The theory also predicts the existence of a weak oscillation state at the boundary between SPOC and relaxation regions in the phase diagram. Thus, the present theory comprehensively explains the characteristics of auto-oscillation and contraction in the contractile system of striated muscle.

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

    PubMed

    Jayasinghe, Isuru D; Clowsley, Alexander H; Munro, Michelle; Hou, Yufeng; Crossman, David J; Soeller, Christian

    2015-01-07

    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.

  9. 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

  10. 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.

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

    PubMed Central

    Obinata, Takashi; Ono, Kanako

    2011-01-01

    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 Ca2+-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. PMID:21866271

  12. Cystic Fibrosis Transmembrane Conductance Regulator in Sarcoplasmic Reticulum of Airway Smooth Muscle. Implications for Airway Contractility

    PubMed Central

    Cook, Daniel P.; Rector, Michael V.; Bouzek, Drake C.; Michalski, Andrew S.; Gansemer, Nicholas D.; Reznikov, Leah R.; Li, Xiaopeng; Stroik, Mallory R.; Ostedgaard, Lynda S.; Abou Alaiwa, Mahmoud H.; Thompson, Michael A.; Prakash, Y. S.; Krishnan, Ramaswamy; Meyerholz, David K.; Seow, Chun Y.

    2016-01-01

    Rationale: An asthma-like airway phenotype has been described in people with cystic fibrosis (CF). Whether these findings are directly caused by loss of CF transmembrane conductance regulator (CFTR) function or secondary to chronic airway infection and/or inflammation has been difficult to determine. Objectives: Airway contractility is primarily determined by airway smooth muscle. We tested the hypothesis that CFTR is expressed in airway smooth muscle and directly affects airway smooth muscle contractility. Methods: Newborn pigs, both wild type and with CF (before the onset of airway infection and inflammation), were used in this study. High-resolution immunofluorescence was used to identify the subcellular localization of CFTR in airway smooth muscle. Airway smooth muscle function was determined with tissue myography, intracellular calcium measurements, and regulatory myosin light chain phosphorylation status. Precision-cut lung slices were used to investigate the therapeutic potential of CFTR modulation on airway reactivity. Measurements and Main Results: We found that CFTR localizes to the sarcoplasmic reticulum compartment of airway smooth muscle and regulates airway smooth muscle tone. Loss of CFTR function led to delayed calcium reuptake following cholinergic stimulation and increased myosin light chain phosphorylation. CFTR potentiation with ivacaftor decreased airway reactivity in precision-cut lung slices following cholinergic stimulation. Conclusions: Loss of CFTR alters porcine airway smooth muscle function and may contribute to the airflow obstruction phenotype observed in human CF. Airway smooth muscle CFTR may represent a therapeutic target in CF and other diseases of airway narrowing. PMID:26488271

  13. 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. Copyright © 2014 Elsevier B.V. All rights reserved.

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

    PubMed

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

    1993-11-01

    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.

  15. 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

  16. Release of calcium into the myofibrillar space in response to active shortening of striated muscle.

    PubMed

    Edman, K A P; Caputo, C

    2017-10-01

    The study was undertaken to explore whether shortening of striated muscle during activity is associated with release of bound Ca(2+) into the myofibrillar space as has previously been proposed in order to explain the depressant effect of active shortening. The experiments were carried out on single muscle fibres isolated from the anterior tibialis muscle of Rana temporaria. The fibres were loaded with the calcium sensitive indicator Fluo-3. The fibres, stimulated to produce a partially fused isometric tetanus, were subjected to a shortening ramp or, alternatively, to a stretch ramp during activity while force, fibre length, sarcomere length and the Fluo-3 signal were recorded. A shortening ramp performed during a partially fused tetanus caused an increase in the myofibrillar free calcium concentration and produced, simultaneously, a decrease in active force. The isometric force recovered gradually after the shortening ramp, while the intracellular Ca(2+) concentration stayed above the control level during the remainder of the stimulation period. A stretch ramp applied during a partially fused tetanus caused a considerably smaller change in the myofibrillar Ca(2+) concentration. The results provide evidence that the myosin cross-bridges interact with the calcium binding sites on the thin filaments during active shortening, causing sustained release of calcium and reduced contractile strength. © 2017 Scandinavian Physiological Society. Published by John Wiley & Sons Ltd.

  17. External gill motility and striated muscle presence in the embryos of anuran amphibians.

    PubMed

    Nokhbatolfoghahai, M; Downie, J R; Atherton, L

    2013-02-01

    Anuran external gills were assessed for motility and striated muscle content in 16 species from seven families. Motility of three kinds was observed. Pulsatory movements related to heart beat rhythm were common. In embryos developing to a late stage before hatching, movements of the whole embryo were frequent, with gills rearranging as a consequence. The only clearly active movement, presumably muscle driven, was 'gill flicking', a posterior movement of the entire gill into the body either on one side only, or both together, followed by a return to the normal spread-out position. Some species may actively spread their gills when hanging from the water surface film, but we did not observe this. In some species, active gill movement developed over time, but we were not able to follow all species over such a developmental sequence. The relationship between active motility and muscle content was good in most cases. Observations on late stage embryos of the aromobatid Mannophryne trinitatis are presented for the first time. In one species, we noted spread external gills being used to adhere hatchlings to a surface.

  18. 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.

  19. Expression of sarcoplasmic reticulum Ca(2+)-ATPase isoforms in marlin and swordfish muscle and heater cells.

    PubMed

    Tullis, A; Block, B A

    1996-07-01

    The superior rectus muscles of marlin, swordfish, sailfish, and spearfish are modified for generating heat rather than force. This study focuses on the sarcoplasmic reticulum calcium-adenosinetriphosphatase (SR Ca(2+)-ATPase) to gain further insight into the muscle fiber type origin of the billfish "heater cell." Direct sequencing and immunolocalization demonstrated that marlin and swordfish epaxial swimming muscles express two forms of the SR Ca(2+)-ATPase in a fiber type-specific manner; red slow-twitch skeletal and cardiac muscles express the same SERCA2 message, whereas white fast-twitch skeletal muscles express a SERCA1 message. Thus the expression pattern of the SR Ca2+ pump is similar in both billfish and tetrapod muscles. Molecular and immunological studies revealed that billfish heater tissue and superior rectus muscle express both fast and slow SR Ca2+ pump isoforms. Immunohistochemical results suggest that heater cells and most extraocular muscle fibers express the fast SR Ca2+ pump. Expression of the fast SR Ca(2+)-ATPase by heater cells has implications for heater cell origin and thermogenic control.

  20. 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

  1. 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

  2. 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.

  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. 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.

  6. 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

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

    PubMed

    Luther, Pradeep K; Squire, John M

    2014-12-03

    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.

  8. SARCOPLASMIC RETICULUM AND THE TEMPERATURE-DEPENDENT CONTRACTION OF SMOOTH MUSCLE IN CALCIUM-FREE SOLUTIONS

    PubMed Central

    Somlyo, Andrew P.; Devine, Carrick E.; Somlyo, Avril V.; North, Stanley R.

    1971-01-01

    The contractile response of turtle oviduct smooth muscle to acetylcholine after 30 min of incubation of muscles in Ca-free, 4 mM ethylene (bis) oxyethylenenitrilotetraacetic acid (EGTA) solutions at room temperature was greater than the contractile response after 30 min of incubation in the Ca-free medium at 37°C. Incubation in Ca-free solution at 37°C before stimulation with acetylcholine in Ca-free solutions at room temperature also reduced the contractile response, suggesting that activator calcium was lost from the fibers at a faster rate at higher temperatures. Electron micrographs of turtle oviduct smooth muscle revealed a sarcoplasmic reticulum (SR) occupying approximately 4% of the nucleus- and mitochondria-free cell volume. Incubation of oviduct smooth muscle with ferritin confirmed that the predominantly longitudinally oriented structures described as the SR did not communicate with the extracellular space. The SR formed fenestrations about the surface vesicles, and formed close contacts (couplings) with the surface membrane and surface vesicles in oviduct and vena caval smooth muscle; it is suggested that these are sites of electromechanical coupling. Calculation of the calcium requirements for smooth muscle contraction suggest that the amount of SR observed in the oviduct smooth muscle could supply the activator calcium for the contractions observed in Ca-free solutions. Incubation of oviduct smooth muscle in hypertonic solutions increased the electron opacity of the fibers. A new feature of some of the surface vesicles observed in oviduct, vena caval, and aortic smooth muscle was the presence of approximately 10 nm striations running approximately parallel to the openings of the vesicles to the extracellular space. Thick, thin, and intermediate filaments were observed in turtle oviduct smooth muscle, although the number of thick filaments seen in the present study appeared less than that previously found in mammalian smooth muscles. PMID:4331503

  9. 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.

  10. 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

  11. Matching of sarcoplasmic reticulum and contractile properties in rat fast- and slow-twitch muscle fibres.

    PubMed

    Trinh, Huong H; Lamb, Graham D

    2006-07-01

    1. The twitch characteristics (fast-twitch or slow-twitch) of skeletal muscle fibres are determined not only by the contractile apparatus properties of the fibre, but also by the time-course of Ca2+ release and re-uptake by the sarcoplasmic reticulum (SR). The present study examined, in individual fibres from non-transforming muscle of the rat, whether particular SR properties are matched to the contractile apparatus properties of the fibre, in particular in the case of fibres with fast-twitch contractile apparatus located in a slow-twitch muscle, namely the soleus. 2. Force was recorded in single, mechanically skinned fibres from extensor digitorum longus (EDL), gastrocnemius, peroneus longus and soleus muscles. Using repeated cycles in which the SR was emptied of all releasable Ca2+ and then reloaded, it was possible to determine the relative amount of Ca2+ present in the SR endogenously, the maximum SR capacity and the rate of Ca2+ loading. The sensitivity of the contractile apparatus to Ca2+ and Sr2+ was used to classify the fibres as fast-twitch (FT), slow-twitch (ST) or mixed (< 3% of the fibres examined) and thereby identify the likely troponin C and myosin heavy chain types present. 3. There was no significant difference in SR properties between the groups of FT fibres obtained from the four different muscles, including soleus. Despite some overlap in the SR properties of individual fibres between the FT and ST groups, the properties of the FT fibres in all four muscles studied were significantly different from those of the ST and mixed fibres. 4. In general, in FT fibres the SR had a larger capacity and the endogenous Ca2+ content was a relatively lower percentage of maximum compared with ST fibres. Importantly, in terms of their SR properties, FT fibres from soleus muscle more closely resembled FT fibres from other muscles than they did ST fibres from soleus muscle.

  12. 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.

  13. Quantitative Measurement of Ca2+ in the Sarcoplasmic Reticulum Lumen of Mammalian Skeletal Muscle

    PubMed Central

    Ziman, Andrew P.; Ward, Christopher W.; Rodney, George G.; Lederer, W. Jonathan; Bloch, Robert J.

    2010-01-01

    Skeletal muscle stores Ca2+ in the sarcoplasmic reticulum (SR) and releases it to initiate contraction, but the concentration of luminal Ca2+ in the SR ([Ca2+]SR) and the amount that is released by physiological or pharmacological stimulation has been difficult to measure. Here we present a novel, yet simple and direct, method that provides the first quantitative estimates of static content and dynamic changes in [Ca2+]SR in mammalian skeletal muscle, to our knowledge. The method uses fluo-5N loaded into the SR of single, mammalian skeletal muscle cells (murine flexor digitorum brevis myofibers) and confocal imaging to detect and calibrate the signals. Using this method, we have determined that [Ca2+]SR, free is 390 μM. 4-Chloro-m-cresol, an activator of the skeletal muscle ryanodine receptor, reduces [Ca2+]SR, free to ∼8 μM, when values are corrected for background fluorescence from cytoplasmic pools of dye. Prolonged electrical stimulation (10 s) at 50 Hz releases 88% of the SR Ca2+ content, whereas stimulation at 1 Hz (10 s) releases only 20%. Our results lay the foundation for molecular modeling of the dynamics of luminal SR Ca2+ and for future studies of the role of SR Ca2+ in healthy and diseased mammalian muscle. PMID:20959112

  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. Distribution of Myosin Attachment Times Predicted from Viscoelastic Mechanics of Striated Muscle

    PubMed Central

    Palmer, Bradley M.; Wang, Yuan; Miller, Mark S.

    2011-01-01

    We demonstrate that viscoelastic mechanics of striated muscle, measured as elastic and viscous moduli, emerge directly from the myosin crossbridge attachment time, tatt, also called time-on. The distribution of tatt was modeled using a gamma distribution with shape parameter, p, and scale parameter, β. At 5 mM MgATP, β was similar between mouse α-MyHC (16.0 ± 3.7 ms) and β-MyHC (17.9 ± 2.0 ms), and p was higher (P < 0.05) for β-MyHC (5.6 ± 0.4 no units) compared to α-MyHC (3.2 ± 0.9). At 1 mM MgATP, p approached a value of 10 in both isoforms, but β rose only in the β-MyHC (34.8 ± 5.8 ms). The estimated mean tatt (i.e., pβ product) was longer in the β-MyHC compared to α-MyHC, and became prolonged in both isoforms as MgATP was reduced as expected. The application of our viscoelastic model to these isoforms and varying MgATP conditions suggest that tatt is better modeled as a gamma distribution due to its representing multiple temporal events occurring within tatt compared to a single exponential distribution which assumes only one temporal event within tatt. PMID:22190855

  16. Differential sensitivity of C2-C12 striated muscle cells to lovastatin and pravastatin.

    PubMed

    Gadbut, A P; Caruso, A P; Galper, J B

    1995-10-01

    One of the major side-effects of the use of HMG CoA reductase inhibitors for the treatment of hypercholesterolemia is the development of myositis and, in some patients undergoing concomitant immunosuppressive treatment, the development of rhabdomyolysis. Experiments outlined in these studies demonstrate that inhibitors of HMG-CoA reductase activity which differ primary in the substitution of a methyl group for a hydroxyl group have differential effects on both cholesterol levels and cell viability in a striated muscle cell model, the mouse C2-C12 myoblast. Thus, concentrations as high as 200 microM of pravastatin had little effect on total cholesterol level while 25 microM of lovastatin decreased cellular cholesterol by over 90%. Simvastatin and lovastatin decreased viability of C2-C12 myoblasts by nearly 50% at concentrations as low as 1 and 5 microM, respectively, and decreased viability by almost 90% at 10 and 15 microM respectively. However, 300 microM of pravastatin decreased cell viability by less than 50%. The order of potency for the effects on cell viability wassimvastatin>lovastatin>pravastatin. The possible relationship between effects on cell viability and the development of myositis is discussed.

  17. 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

    2015-10-05

    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. Copyright © 2015 Elsevier B.V. All rights reserved.

  18. 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.

  19. Electron microscope tomography: further demonstration of nanocontacts between caveolae and smooth muscle sarcoplasmic reticulum

    PubMed Central

    Gherghiceanu, Mihaela; Popescu, LM

    2007-01-01

    Abstract A spatial relationship between caveolae and sarcoplasmic reticulum (SR) in smooth muscle cells (SMC) was previously reported in computer-assisted three-dimensional reconstruction from transmission electron microscope serial sections. The knowledge of the three-dimensional organization of the cortical space of SMC is essential to understand caveolae function at the cellular level. Cellular tomography using transmission electron microscopy tomography (EMT) is the only available technology to reliably chart the inside of a cell and is therefore an essential technology in the study of organellar nanospatial relationships. Using EMT we further demonstrate here that caveolae and peripheral SR in visceral SMC build constantly spatial units, presumably responsible for a vectorial control of free Ca2+ cytoplasmic concentrations in definite nanospaces. PMID:18205711

  20. Modifications of Ca2+ transport induced by glutathione in sarcoplasmic reticulum membranes of frog skeletal muscle.

    PubMed

    Belia, S; Vecchiet, J; Vecchiet, L; Fanó, G

    2000-04-01

    The Ca2+ transport across the membrane of vesicles purified from the sarcoplasmic reticulum (SR) of frog skeletal muscle is modified by raising the concentration of the reduced form of glutathione (GSH). Passive release of Ca2+ is inhibited through the direct action of GSH on ryanodine receptors while active uptake is increased by a dose-dependent stimulation of Ca2+ pumps (Ca2+ -ATPase). These effects are physiological since the concentrations of GSH utilised (0.01-10.0 mM) are compatible with the in vivo concentration of this antioxidant. They are independent of the external Ca2+ concentration and are specific for the reduced form of glutathione, since the disulphide form (GSSG) or other GSH-derivatives do not induce these effects.

  1. Calcium Uptake and Release through Sarcoplasmic Reticulum in the Inferior Oblique Muscles of Patients with Inferior Oblique Overaction

    PubMed Central

    Kim, Hee Seon; Chang, Yoon-Hee; Kim, Do Han; Park, So Ra; Han, Sueng-Han

    2006-01-01

    We characterized and compared the characteristics of Ca2+ movements through the sarcoplasmic reticulum of inferior oblique muscles in the various conditions including primary inferior oblique overaction (IOOA), secondary IOOA, and controls, so as to further understand the pathogenesis of primary IOOA. Of 15 specimens obtained through inferior oblique myectomy, six were from primary IOOA, 6 from secondary IOOA, and the remaining 3 were controls from enucleated eyes. Ryanodine binding assays were performed, and Ca2+ uptake rates, calsequestrins and SERCA levels were determined. Ryanodine bindings and sarcoplasmic reticulum Ca2+ uptake rates were significantly decreased in primary IOOA (p<0.05). Western blot analysis conducted to quantify calsequestrins and SERCA, found no significant difference between primary IOOA, secondary IOOA, and the controls. Increased intracellular Ca2+ concentration due to reduced sarcoplasmic reticulum Ca2+ uptake may play a role in primary IOOA. PMID:16642550

  2. Maximum shortening velocity of lymphatic muscle approaches that of striated muscle.

    PubMed

    Zhang, Rongzhen; Taucer, Anne I; Gashev, Anatoliy A; Muthuchamy, Mariappan; Zawieja, David C; Davis, Michael J

    2013-11-15

    Lymphatic muscle (LM) is widely considered to be a type of vascular smooth muscle, even though LM cells uniquely express contractile proteins from both smooth muscle and cardiac muscle. We tested the hypothesis that LM exhibits an unloaded maximum shortening velocity (Vmax) intermediate between that of smooth muscle and cardiac muscle. Single lymphatic vessels were dissected from the rat mesentery, mounted in a servo-controlled wire myograph, and subjected to isotonic quick release protocols during spontaneous or agonist-evoked contractions. After maximal activation, isotonic quick releases were performed at both the peak and plateau phases of contraction. Vmax was 0.48 ± 0.04 lengths (L)/s at the peak: 2.3 times higher than that of mesenteric arteries and 11.4 times higher than mesenteric veins. In cannulated, pressurized lymphatic vessels, shortening velocity was determined from the maximal rate of constriction [rate of change in internal diameter (-dD/dt)] during spontaneous contractions at optimal preload and minimal afterload; peak -dD/dt exceeded that obtained during any of the isotonic quick release protocols (2.14 ± 0.30 L/s). Peak -dD/dt declined with pressure elevation or activation using substance P. Thus, isotonic methods yielded Vmax values for LM in the mid to high end (0.48 L/s) of those the recorded for phasic smooth muscle (0.05-0.5 L/s), whereas isobaric measurements yielded values (>2.0 L/s) that overlapped the midrange of values for cardiac muscle (0.6-3.3 L/s). Our results challenge the dogma that LM is classical vascular smooth muscle, and its unusually high Vmax is consistent with the expression of cardiac muscle contractile proteins in the lymphatic vessel wall.

  3. Maximum shortening velocity of lymphatic muscle approaches that of striated muscle

    PubMed Central

    Zhang, Rongzhen; Taucer, Anne I.; Gashev, Anatoliy A.; Muthuchamy, Mariappan; Zawieja, David C.

    2013-01-01

    Lymphatic muscle (LM) is widely considered to be a type of vascular smooth muscle, even though LM cells uniquely express contractile proteins from both smooth muscle and cardiac muscle. We tested the hypothesis that LM exhibits an unloaded maximum shortening velocity (Vmax) intermediate between that of smooth muscle and cardiac muscle. Single lymphatic vessels were dissected from the rat mesentery, mounted in a servo-controlled wire myograph, and subjected to isotonic quick release protocols during spontaneous or agonist-evoked contractions. After maximal activation, isotonic quick releases were performed at both the peak and plateau phases of contraction. Vmax was 0.48 ± 0.04 lengths (L)/s at the peak: 2.3 times higher than that of mesenteric arteries and 11.4 times higher than mesenteric veins. In cannulated, pressurized lymphatic vessels, shortening velocity was determined from the maximal rate of constriction [rate of change in internal diameter (−dD/dt)] during spontaneous contractions at optimal preload and minimal afterload; peak −dD/dt exceeded that obtained during any of the isotonic quick release protocols (2.14 ± 0.30 L/s). Peak −dD/dt declined with pressure elevation or activation using substance P. Thus, isotonic methods yielded Vmax values for LM in the mid to high end (0.48 L/s) of those the recorded for phasic smooth muscle (0.05–0.5 L/s), whereas isobaric measurements yielded values (>2.0 L/s) that overlapped the midrange of values for cardiac muscle (0.6–3.3 L/s). Our results challenge the dogma that LM is classical vascular smooth muscle, and its unusually high Vmax is consistent with the expression of cardiac muscle contractile proteins in the lymphatic vessel wall. PMID:23997104

  4. [Calcium leak of sarcoplasmic reticulum induces degradation of troponin I in skeletal muscle fibers.].

    PubMed

    Li, Quan; Wang, Yun-Ying; Li, Hui; Jiao, Bo; Yu, Zhi-Bin

    2009-06-25

    The troponin I subunit (TnI) was used as a molecular marker to explore the relationship between the resting intracellular Ca(2+) concentration and myofibril degradation in muscle fibers. The isolated soleus muscle strips of rats were treated by caffeine and H2O2. Caffeine is an opener to increase the calcium release channel open probability of sarcoplasmic reticulum (SR) in contraction phase. H2O2 induces a calcium leak of SR calcium release channel in relaxation phase. The expression and degradation of TnI were detected by Western blot. The resting tension of tetanic contraction and expression of TnI were not changed, but the developed tension was lowered in isolated soleus muscle strips during 40 min of calcium-free Krebs perfusion. Low concentrations of caffeine (1 and 5 mmol/L) perfusion induced a transient increase in resting tension during fatigue period, but did not alter the extent of fatigue, recovery rate after fatigue and expression of TnI in muscle strips. High concentration of caffeine (10 mmol/L) perfusion induced a progressive increase in resting tension, a higher rate of fatigue and a decrease in recovery rate after fatigue in muscle strips. There was a detectable degradation of TnI in soleus after 10 mmol/L caffeine treatment. H2O2 perfusion facilitated a progressive increase in resting tension in a dose-dependent manner, but did not influence the fatigue rate of tetanic contraction. The recovery rate after fatigue showed a quick resumption before decline during H2O2 perfusion. Degradation of TnI occurred in 5 and 10 mmol/L H2O2-treated soleus muscles. Since resting tension is dependent on intracellular Ca(2+) concentration, the above-mentioned results suggest that SR Ca(2+) leakage in relaxation phase may induce a degradation of TnI in skeletal muscle fibers.

  5. 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.

  6. Cylindrical Spirals in Skeletal Muscles Originate From the Longitudinal Sarcoplasmic Reticulum.

    PubMed

    Xu, Jing-Wen; Liu, Fu-Chen; Li, Wei; Zhao, Yu-Ying; Zhao, Dan-Dan; Luo, Yue-Bei; Lu, Jian-Qiang; Yan, Chuan-Zhu

    2016-02-01

    Cylindrical spirals (CSs) are rare but distinct subsarcolemmal accumulations in skeletal muscle fibers. To date, CSs have been reported in only 16 patients with a variety of neuromuscular conditions. The origin and composition of CSs are unknown, although there are some morphologic similarities between CSs and tubular aggregates (TAs). To clarify the nature of CSs, we characterized the sarcoplasmic reticulum (SR) and other intracellular membrane system proteins in CSs of muscle biopsies from 2 adult Chinese siblings. Immunohistochemical studies revealed subsarcolemmal immunoreactivity for sarco/endoplasmic reticulum Ca2þ-ATPase 1 (SERCA 1) in the longitudinal SR, but no immunoreactivity for calsequestrin in the terminal cisternae or type 1 ryanodine receptor (RYR1) in the junctional SR. Muscles biopsied from 2 patients with TAs showed immunoreactivity not only for SERCA1 but also for other SR proteins, including calsequestrin and RYR1. CSs exhibited no immunoreactivity for the Golgi apparatus marker GM130, the nuclear membrane emerin, desmin, the autophagosome marker LC3, the lysosomal membrane marker LAMP2, dystrophin, or myosin. Our results suggest CSs may originate only from the longitudinal SR, whereas TAs are composed of both the junctional and longitudinal SR. Immunochemical staining with antibodies against calsequestrin and RYR1 help to distinguish these 2 pathological alterations.

  7. Cytoplasmic γ-actin and tropomodulin isoforms link to the sarcoplasmic reticulum in skeletal muscle fibers

    PubMed Central

    Gokhin, David S.

    2011-01-01

    The sarcoplasmic reticulum (SR) serves as the Ca2+ reservoir for muscle contraction. Tropomodulins (Tmods) cap filamentous actin (F-actin) pointed ends, bind tropomyosins (Tms), and regulate F-actin organization. In this paper, we use a genetic targeting approach to examine the effect of Tmod1 deletion on the organization of cytoplasmic γ-actin (γcyto-actin) in the SR of skeletal muscle. In wild-type muscle fibers, γcyto-actin and Tmod3 defined an SR microdomain that was distinct from another Z line–flanking SR microdomain containing Tmod1 and Tmod4. The γcyto-actin/Tmod3 microdomain contained an M line complex composed of small ankyrin 1.5 (sAnk1.5), γcyto-actin, Tmod3, Tm4, and Tm5NM1. Tmod1 deletion caused Tmod3 to leave its SR compartment, leading to mislocalization and destabilization of the Tmod3–γcyto-actin–sAnk1.5 complex. This was accompanied by SR morphological defects, impaired Ca2+ release, and an age-dependent increase in sarcomere misalignment. Thus, Tmod3 regulates SR-associated γcyto-actin architecture, mechanically stabilizes the SR via a novel cytoskeletal linkage to sAnk1.5, and maintains the alignment of adjacent myofibrils. PMID:21727195

  8. Intensity of light diffraction from striated muscle as a function of incident angle.

    PubMed Central

    Baskin, R J; Lieber, R L; Oba, T; Yeh, Y

    1981-01-01

    In a recently developed theory of light diffraction by single striated muscle fibers, we considered only the case of normal beam incidence. The present investigation represents both an experimental and theoretical extension of the previous work to arbitrary incident angle. Angle scan profiles over a 50 degrees range of incident angle (+25 degrees to -25 degrees) were obtained at different sarcomere lengths. Left and right first-order scan peak separations were found to be a function of sarcomere length (separation angle = 2 theta B), and good agreement was found between theory and experiment. Our theoretical analysis further showed that a myofibrillar population with a single common skew angle can yield an angle scan profile containing many peaks. Thus, it is not necessary to associate each peak with a different skew population. Finally, we have found that symmetry angle, theta s, also varies with sarcomere length, but not in a regular manner. Its value at a given sarcomere length is a function of a particular region of a given fiber and represents the average skew angle of all the myofibril populations illuminated. The intensity of a diffraction order line is considered to be principally the resultant of two interference phenomena. The first is a volume-grating phenomenon which results from the periodic A-I band structure of the fiber (with some contribution from Z bands and H zones). The second is Bragg reflection from skew planes, if the correct relation between incident angle and skew angle is met. This may result in intensity asymmetry between the left and right first order lines. PMID:6976802

  9. Calcium releasing action of quercetin on sarcoplasmic reticulum from frog skeletal muscle.

    PubMed

    Kurebayashi, N; Ogawa, Y

    1984-10-01

    The release of Ca by quercetin from the sarcoplasmic reticulum has been claimed to be a result of the well-known inhibition of Ca2+-ATPase activity, or to be due to an intrinsic property of quercetin. To get a clearer understanding of the effect of quercetin, we examined it using fragmented sarcoplasmic reticulum (FSR) from bullfrog skeletal muscle. The rapid phase of Ca release (hereafter simply referred to as "Ca release") from loaded FSR was almost completed within 5 s after addition of quercetin in the presence of ATP. It cannot be ascribed to the inhibition of Ca2+-ATPase activity on the basis of following findings. First, when Ca uptake was driven by carbamylphosphate, no or little Ca release was observed in marked contrast to a stronger reduction in the rate of Ca uptake. Secondly, procaine reverses the Ca releasing action of quercetin, whereas it show a synergistic action in the inhibition of Ca2+-ATPase activity. Thirdly, HFSR released more Ca than LFSR, while the Ca2+-ATPase activities of both fractions were inhibited to a similar extent. The Ca release by quercetin is enhanced by ATP or beta, gamma-methylene adenosine triphosphate, and decreased by procaine or a high concentration of Mg2+. In the presence of 2.5 mM caffeine, the amount of Ca2+ released by quercetin was decreased, and the dose-effect relationship was shifted to higher doses of quercetin. This indicates that quercetin and caffeine probably overlap in the site(s) of the action, but that quercetin is dissimilar from halothane in the mode of its Ca-releasing action.

  10. Microtubule structures underlying the sarcoplasmic reticulum support peripheral coupling sites to regulate smooth muscle contractility.

    PubMed

    Pritchard, Harry A T; Gonzales, Albert L; Pires, Paulo W; Drumm, Bernard T; Ko, Eun A; Sanders, Kenton M; Hennig, Grant W; Earley, Scott

    2017-09-19

    Junctional membrane complexes facilitate excitation-contraction coupling in skeletal and cardiac muscle cells by forming subcellular invaginations that maintain close (≤20 nm) proximity of ryanodine receptors (RyRs) on the sarcoplasmic reticulum (SR) with voltage-dependent Ca(2+) channels in the plasma membrane. In fully differentiated smooth muscle cells, junctional membrane complexes occur as distributed sites of peripheral coupling. We investigated the role of the cytoskeleton in maintaining peripheral coupling and associated Ca(2+) signaling networks within native smooth muscle cells of mouse and rat cerebral arteries. Using live-cell confocal and superresolution microscopy, we found that the tight interactions between the SR and the plasma membrane in these cells relied on arching microtubule structures present at the periphery of smooth muscle cells and were independent of the actin cytoskeleton. Loss of peripheral coupling associated with microtubule depolymerization altered the spatiotemporal properties of localized Ca(2+) sparks generated by the release of Ca(2+) through type 2 RyRs (RyR2s) on the SR and decreased the number of sites of colocalization between RyR2s and large-conductance Ca(2+)-activated K(+) (BK) channels. The reduced BK channel activity associated with the loss of SR-plasma membrane interactions was accompanied by increased pressure-induced constriction of cerebral resistance arteries. We conclude that microtubule structures maintain peripheral coupling in contractile smooth muscle cells, which is crucial for the regulation of contractility and cerebral vascular tone. Copyright © 2017 The Authors, some rights reserved; exclusive licensee American Association for the Advancement of Science. No claim to original U.S. Government Works.

  11. Contracture Coupling of Slow Striated Muscle in Non-Ionic Solutions and Replacement of Calcium, Sodium, and Potassium

    PubMed Central

    Irwin, Richard L.; Hein, Manfred M.

    1964-01-01

    The development of contracture related to changes of ionic environment (ionic contracture coupling) has been studied in the slowly responding fibers of frog skeletal muscle. When deprived of external ions for 30 minutes by use of solutions of sucrose, mannitol, or glucose, the slow skeletal muscle fibers, but not the fast, develop pronounced and easily reversible contractures. Partial replacement of the non-ionic substance with calcium or sodium reduces the development of the contractures but replacement by potassium does not. The concentration of calcium necessary to prevent contracture induced by a non-ionic solution is greater than that needed to maintain relaxation in ionic solutions. To suppress the non-ionic-induced contractures to the same extent as does calcium requires several fold higher concentrations of sodium. Two types of ionic contracture coupling occur in slow type striated muscle fibers: (a) a calcium deprivation type which develops maximally at full physiological concentration of external sodium, shows a flow rate dependency for the calcium-depriving fluid, and is lessened when the sodium concentration is decreased by replacement with sucrose; (b) a sodium deprivation type which occurs maximally without external sodium, is lessened by increasing the sodium concentration, and has no flow rate dependency for ion deprivation. Both types of contracture are largely prevented by the presence of sufficient calcium. There thus seem to be calcium- and sodium-linked processes at work in the ionic contracture coupling of slow striated muscle. PMID:14127603

  12. Quantitative second harmonic generation imaging and modeling of the optical clearing mechanism in striated muscle and tendon.

    PubMed

    LaComb, Ronald; Nadiarnykh, Oleg; Carey, Shawn; Campagnola, Paul J

    2008-01-01

    We have investigated the mechanisms and capabilities of optical clearing in conjunction with second harmonic generation (SHG) imaging in tendon and striated muscle. Our approach combines three-dimensional (3-D) SHG imaging of the axial attenuation and directional response with Monte Carlo simulation (based on measured bulk optical properties) of the creation intensity and propagation through the tissues. Through these experiments and simulations, we show that reduction of the primary filter following glycerol treatment dominates the axial attenuation response in both muscle and tendon. However, these disparate tissue types are shown to clear through different mechanisms of the glycerol-tissue interaction. In the acellular tendon, glycerol application reduces scattering by both index matching as well as increasing the interfibril separation. This results in an overall enhancement of the 3-D SHG intensity, where good agreement is found between experiment and simulation. Through analysis of the axial response as a function of glycerol concentration in striated muscle, we conclude that the mechanism in this tissue arises from matching of the refractive index of the cytoplasm of the muscle cells with that of the surrounding higher-index collagenous perimysium. We further show that the proportional decrease in the scattering coefficient mu(s) with increasing glycerol fraction can be well-approximated by Mie theory.

  13. Effects of imperatoxin A on local sarcoplasmic reticulum Ca(2+) release in frog skeletal muscle.

    PubMed Central

    Shtifman, A; Ward, C W; Wang, J; Valdivia, H H; Schneider, M F

    2000-01-01

    We have investigated the effects of imperatoxin A (IpTx(a)) on local calcium release events in permeabilized frog skeletal muscle fibers, using laser scanning confocal microscopy in linescan mode. IpTx(a) induced the appearance of Ca(2+) release events from the sarcoplasmic reticulum that are approximately 2 s and have a smaller amplitude (31 +/- 2%) than the "Ca(2+) sparks" normally seen in the absence of toxin. The frequency of occurrence of long-duration imperatoxin-induced Ca(2+) release events increased in proportion to IpTx(a) concentrations ranging from 10 nM to 50 nM. The mean duration of imperatoxin-induced events in muscle fibers was independent of toxin concentration and agreed closely with the channel open time in experiments on isolated frog ryanodine receptors (RyRs) reconstituted in planar lipid bilayer, where IpTx(a) induced opening of single Ca(2+) release channels to prolonged subconductance states. These results suggest involvement of a single molecule of IpTx(a) in the activation of a single Ca(2+) release channel to produce a long-duration event. Assuming the ratio of full conductance to subconductance to be the same in the fibers as in bilayer, the amplitude of a spark relative to the long event indicates involvement of at most four RyR Ca(2+) release channels in the production of short-duration Ca(2+) sparks. PMID:10920014

  14. Altered turnover of calcium regulatory proteins of the sarcoplasmic reticulum in aged skeletal muscle.

    PubMed

    Ferrington, D A; Krainev, A G; Bigelow, D J

    1998-03-06

    We have measured the in vivo protein turnover for the major calcium regulatory proteins of the sarcoplasmic reticulum from the skeletal muscle of young adult (7 months) and aged (28 months) Fischer 344 rats. From the time course of the incorporation and decay of protein-associated radioactivity after a pulse injection of [14C]leucine and correcting for leucine reutilization, in young rats, the apparent half-lives for calsequestrin, the 53-kDa glycoprotein, and ryanodine receptor are 5.4 +/- 0.4, 6.3 +/- 1.3, and 8.3 +/- 1.3 days, respectively. A half-life of 14.5 +/- 2.5 days was estimated for the Ca-ATPase isolated from young muscle. Differences in protein turnover associated with aging were determined using sequential injection of two different isotopic labels ([14C]leucine and [3H]leucine) to provide an estimate of protein synthesis and degradation within the same animal. The Ca-ATPase and ryanodine receptor isolated from aged muscle exhibits 27 +/- 5% and 25 +/- 3% slower protein turnover, respectively, relative to that from young muscle. In contrast, the 53-kDa glycoprotein exhibits a 25 +/- 5% more rapid turnover in aged SR, while calsequestrin exhibits no age-dependent alteration in turnover. Statistical analysis comparing the sensitivity of various methods for discriminating different rates of protein turnover validates the approach used in this study and demonstrates that the use of two isotopic labels provides at least a 6-fold more sensitive means to detect age-related differences in protein turnover relative to other methods.

  15. Major contribution of sarcoplasmic reticulum Ca2+ depletion during long-lasting activation of skeletal muscle

    PubMed Central

    Robin, Gaëlle

    2013-01-01

    Depolarization of skeletal muscle fibers induces sarcoplasmic reticulum (SR) Ca2+ release and contraction that progressively decline while depolarization is maintained. Voltage-dependent inactivation of SR Ca2+ release channels and SR Ca2+ depletion are the two processes proposed to explain the decline of SR Ca2+ release during long-lasting depolarizations. However, the relative contribution of these processes, especially under physiological conditions of activation, is not clearly established. Using Fura-2 and Fluo-5N to monitor cytosolic and SR Ca2+ changes, respectively, in voltage-controlled mouse muscle fibers, we show that 2-min conditioning depolarizations reduce voltage-activated cytosolic Ca2+ signals with a V1/2 of −53 mV but also induce SR Ca2+ depletion that decreased the releasable pool of Ca2+ with the same voltage sensitivity. In contrast, measurement of SR Ca2+ changes indicated that SR Ca2+ release channels were inactivated after SR had been depleted and in response to much higher depolarizations with a V1/2 of −13 mV. In response to trains of action potentials, cytosolic Ca2+ signals decayed with time, whereas SR Ca2+ changes remained stable over 1-min stimulation, demonstrating that SR Ca2+ depletion is exclusively responsible for the decline of SR Ca2+ release under physiological conditions of excitation. These results suggest that previous studies using steady-state inactivation protocols to investigate the voltage dependence of Ca2+ release inactivation in fact probed the voltage dependence of SR Ca2+ depletion, and that SR Ca2+ depletion is the only process that leads to Ca2+ release decline during continuous stimulation of skeletal muscle. PMID:23630339

  16. The Influence of Hydrogen Ion Concentration on Calcium Binding and Release by Skeletal Muscle Sarcoplasmic Reticulum

    PubMed Central

    Nakamaru, Yoshiaki; Schwartz, Arnold

    1972-01-01

    Calcium release and binding produced by alterations in pH were investigated in isolated sarcoplasmic reticulum (SR) from skeletal muscle. When the pH was abruptly increased from 6.46 to 7.82, after calcium loading for 30 sec, 80–90 nanomoles (nmole) of calcium/mg protein were released. When the pH was abruptly decreased from 7.56 to 6.46, after calcium loading for 30 sec, 25–30 nmole of calcium/mg protein were rebound. The calcium release process was shown to be a function of pH change: 57 nmole of calcium were released per 1 pH unit change per mg protein. The amount of adenosine triphosphate (ATP) bound to the SR was not altered by the pH changes. The release phenomenon was not due to alteration of ATP concentration by the increased pH. Native actomyosin was combined with SR in order to study the effectiveness of calcium release from the SR by pH change in inducing super-precipitation of actomyosin. It was found that SR, in an amount high enough to inhibit superprecipitation at pH 6.5, did not prevent the process when the pH was suddenly increased to 7.3, indicating that the affinity of SR for calcium depends specifically on pH. These data suggest the possible participation of hydrogen ion concentration in excitation-contraction coupling. PMID:5007263

  17. Ethanol enhances caffeine-induced Ca2+-release channel activation in skeletal muscle sarcoplasmic reticulum.

    PubMed

    Oba, T; Koshita, M; Yamaguchi, M

    1997-02-01

    When sarcoplasmic reticulum (SR) vesicles prepared from frog skeletal muscles were actively loaded with Ca2+, pretreatment of the SR with 2.2 mM (0.01%) ethanol for 30 s significantly potentiated 5 mM caffeine-induced release of Ca2+ from 16.7 +/- 3.7 nmol/mg protein in control without ethanol to 28.0 +/- 2.6 nmol/mg (P < 0.05, n = 5). Ethanol alone caused no release of Ca2+ from the SR. Exposure of the Ca2+-release channel, incorporated into planar lipid bilayers, to 2 mM caffeine significantly increased open probability (Po) and mean open time, but unitary conductance was not affected. Ethanol (2.2 mM) enhanced caffeine-induced Ca2+-release channel activity, with Po reaching 3.02-fold and mean open time 2.85-fold the values in the absence of ethanol. However, ethanol alone did not affect electrical parameters of single-channel current, over a concentration range of 2.2 mM (0.01%) to 217 mM (1%). The synergistic action of ethanol and caffeine on the channel activity could be attributable to enhancement of caffeine-induced release of Ca2+ from the SR vesicles in the presence of ethanol.

  18. Calsequestrin depolymerizes when calcium is depleted in the sarcoplasmic reticulum of working muscle

    PubMed Central

    Manno, Carlo; Figueroa, Lourdes C.; Gillespie, Dirk; Fitts, Robert; Kang, ChulHee; Franzini-Armstrong, Clara; Rios, Eduardo

    2017-01-01

    Calsequestrin, the only known protein with cyclical storage and supply of calcium as main role, is proposed to have other functions, which remain unproven. Voluntary movement and the heart beat require this calcium flow to be massive and fast. How does calsequestrin do it? To bind large amounts of calcium in vitro, calsequestrin must polymerize and then depolymerize to release it. Does this rule apply inside the sarcoplasmic reticulum (SR) of a working cell? We answered using fluorescently tagged calsequestrin expressed in muscles of mice. By FRAP and imaging we monitored mobility of calsequestrin as [Ca2+] in the SR--measured with a calsequestrin-fused biosensor--was lowered. We found that calsequestrin is polymerized within the SR at rest and that it depolymerized as [Ca2+] went down: fully when calcium depletion was maximal (a condition achieved with an SR calcium channel opening drug) and partially when depletion was limited (a condition imposed by fatiguing stimulation, long-lasting depolarization, or low drug concentrations). With fluorescence and electron microscopic imaging we demonstrated massive movements of calsequestrin accompanied by drastic morphological SR changes in fully depleted cells. When cells were partially depleted no remodeling was found. The present results support the proposed role of calsequestrin in termination of calcium release by conformationally inducing closure of SR channels. A channel closing switch operated by calsequestrin depolymerization will limit depletion, thereby preventing full disassembly of the polymeric calsequestrin network and catastrophic structural changes in the SR. PMID:28069951

  19. Freeze-fracture studies of nexuses between smooth muscle cells. Close relationship to sarcoplasmic reticulum

    PubMed Central

    1977-01-01

    The freeze-fracture appearance of the nexus was compared in the smooth muscle of guinea pig sphincter pupillac, portal vein, pulmonary artery, taenia coli, uretzr, and vas diferens, mouse vas deferens, chicken gizzard and anterior mesenteric artery, and toad stomach. Nexuses are particularly numerous in the guinea pig sphincter pupillae; they are usually oval and their average area is 0.15 mum2, although some as large as 0.6 mum2 were seen. Small aggregations of particles were observed which would not be recognizable as nexuses in thin section. What constitutes the minimum size of a nexus is discussed. It is estimated that the number of nexuses per cell in this preparation is of the order of tens rather than hundreds. All nexuses examined had 6-9-nm particles in the PF face, with corresponding 3-4-nm pits on the EF face forming a polygonal tending towards a hexagonal lattice. The nexuses are arranged in rows parallel to the main axis of the cell, usually alternating with longitudinal rows of plasmalemmal vesicles. Many nexuses in the guinea pig sphincter pupillae, chicken gizzard, and toad stomach show a close relationship with sarcoplasmic reticulum. The possibility that this may have some role in current flow across this specialized junction is discussed. PMID:401506

  20. Relationships between the sarcoplasmic reticulum and sarcolemmal calcium transport revealed by rapidly cooling rabbit ventricular muscle

    PubMed Central

    1986-01-01

    Rabbit right ventricular papillary muscles were cooled from 30 to approximately 1 degree C immediately after discontinuing electrical stimulation (0.5 Hz). This produced a contracture that was 30-50% of the preceding twitch magnitude and required 20-30 s to develop. The contractures were identical in cooling solutions with normal (144 mM) or low (2.0 mM) Na. They were therefore not Na-withdrawal contractures. Contracture activation was considerably slower than muscle cooling (approximately 2.5 s to cool below 2 degrees C). Cooling contractures were suppressed by caffeine treatment (10.0 mM). Rapid cooling did not cause sufficient membrane depolarization (16.5 +/- 1.2 mV after 30 s of cooling) to produce either a voltage-dependent activation of contracture or a gated entry of Ca from the extracellular space. Contractures induced by treating resting muscles with 5 X 10(-5) M strophanthidin at 30 degrees C exhibited pronounced tension noise. The Fourier spectrum of this noise revealed a periodic component (2-3 Hz) that disappeared when the muscle was cooled. Cooling contractures decayed with rest (t1/2 = 71.0 +/- 9.3 s). This decay accelerated in the presence of 10.0 mM caffeine and was prevented and to some extent reversed when extracellular Na was reduced to 2.0 mM. 20 min of rest resulted in a net decline in intracellular Ca content of 1.29 +/- 0.38 mmol/kg dry wt. I infer that cooling contractures are principally activated by Ca from the sarcoplasmic reticulum (SR). The properties of these contractures suggest that they may provide a convenient relative index of the availability of SR Ca for contraction. The rest decay of cooling contractures (and hence the decay in the availability of activating Ca) is consistent with the measured loss in analytic Ca during rest. The results suggest that contraction in heart muscle can be regulated by an interaction between sarcolemmal and SR Ca transport. PMID:3783123

  1. Activation of inositol trisphosphate-sensitive Ca2+ channels of sarcoplasmic reticulum from frog skeletal muscle.

    PubMed Central

    Suárez-Isla, B A; Alcayaga, C; Marengo, J J; Bull, R

    1991-01-01

    1. The modulation by Ca2+ of the activation by inositol 1,4,5-trisphosphate (IP3) of Ca2+ channels present in native sarcoplasmic reticulum membranes from frog skeletal muscle was studied after channel incorporation into planar phospholipid bilayers in the presence of Ca2+ or Ba2+ as current carrier species. 2. Channel activity expressed as fractional open time (Po) was low (less than or equal to 0.15) in the presence of varying free Ca2+ concentrations bathing the myoplasmic face of the channel (cis side), and did not increase significantly between 0.01 and 30 microM-Ca2+. 3. Channel activation mediated by IP3 could be elicited from free Ca2+ levels similar to those of resting skeletal muscle (about 0.1 microM) and was found to be strongly regulated by the free Ca2+ concentration present at the myoplasmic moiety of the channel. 4. Channel activation by 10 microM-IP3 depended on the Ca2+ concentration on the cis side. Po reached a maximum between pCa 7.0 and 6.0, but decreased at higher concentrations of free Ca2+. Thus, Ca2+ exerted a modulatory influence on IP3-mediated activation in a concentration range where the channel was insensitive to Ca2+. 5. The results indicate that Ca2+ ions act as modulators of IP3 efficacy to open the channel. This could arise from an interaction of Ca2+ with the channel gating mechanism or with the agonist binding site. PMID:1667801

  2. Non-proteolytic functions of calpain-3 in sarcoplasmic reticulum in skeletal muscles.

    PubMed

    Ojima, Koichi; Ono, Yasuko; Ottenheijm, Coen; Hata, Shoji; Suzuki, Hidenori; Granzier, Henk; Sorimachi, Hiroyuki

    2011-04-01

    Mutations in CAPN3/Capn3, which codes for skeletal muscle-specific calpain-3/p94 protease, are responsible for limb-girdle muscular dystrophy type 2A. Using "knock-in" (referred to as Capn3(CS/CS)) mice, in which the endogenous calpain-3 is replaced with a mutant calpain-3:C129S, which is a proteolytically inactive but structurally intact calpain-3, we demonstrated in our previous studies that loss of calpain-3 protease activity causes muscular dystrophy [Ojima, K. et al. (2010) J. Clin. Invest. 120, 2672-2683]. However, compared to Capn3-null (Capn3(-/-)) mice, Capn3(CS/CS) mice showed less severe dystrophic symptoms. This suggests that calpain-3 also has a non-proteolytic function. This study aimed to elucidate the non-proteolytic functions of calpain-3 through comparison of Capn3(CS/CS) mice with Capn3(-/-) mice. We found that calpain-3 is a component of the sarcoplasmic reticulum (SR), and that calpain-3 interacts with, but does not proteolyze, typical SR components such as ryanodine receptor and calsequestrin. Furthermore, Capn3(CS/CS) mice showed that the nonenzymatic role of calpain-3 is required for proper Ca(2+) efflux from the SR to cytosol during muscle contraction. These results indicate that calpain-3 functions as a nonenzymatic element for the Ca(2+) efflux machinery in the SR, rather than as a protease. Thus, defects in the nonenzymatic function of calpain-3 must also be involved in the pathogenesis of limb-girdle muscular dystrophy type 2A.

  3. Dynamic measurement of the calcium buffering properties of the sarcoplasmic reticulum in mouse skeletal muscle

    PubMed Central

    Manno, Carlo; Sztretye, Monika; Figueroa, Lourdes; Allen, Paul D; Ríos, Eduardo

    2013-01-01

    The buffering power, B, of the sarcoplasmic reticulum (SR), ratio of the changes in total and free [Ca2+], was determined in fast-twitch mouse muscle cells subjected to depleting membrane depolarization. Changes in total SR [Ca2+] were measured integrating Ca2+ release flux, determined with a cytosolic [Ca2+] monitor. Free [Ca2+]SR was measured using the cameleon D4cpv-Casq1. In 34 wild-type (WT) cells average B during the depolarization (ON phase) was 157 (SEM 26), implying that of 157 ions released, 156 were bound inside the SR. B was significantly greater when BAPTA, which increases release flux, was present in the cytosol. B was greater early in the pulse – when flux was greatest – than at its end, and greater in the ON than in the OFF. In 29 Casq1-null cells, B was 40 (3.6). The difference suggests that 75% of the releasable calcium is normally bound to calsequestrin. In the nulls the difference in B between ON and OFF was less than in the WT but still significant. This difference and the associated decay in B during the ON were not artifacts of a slow SR monitor, as they were also found in the WT when [Ca2+]SR was tracked with the fast dye fluo-5N. The calcium buffering power, binding capacity and non-linear binding properties of the SR measured here could be accounted for by calsequestrin at the concentration present in mammalian muscle, provided that its properties were substantially different from those found in solution. Its affinity should be higher, or KD lower than the conventionally accepted 1 mm; its cooperativity (n in a Hill fit) should be higher and the stoichiometry of binding should be at the higher end of the values derived in solution. The reduction in B during release might reflect changes in calsequestrin conformation upon calcium loss. PMID:23148320

  4. Intramembrane charge movement and sarcoplasmic calcium release in enzymatically isolated mammalian skeletal muscle fibres.

    PubMed Central

    Szentesi, P; Jacquemond, V; Kovács, L; Csernoch, L

    1997-01-01

    1. Single muscle fibres were dissociated enzymatically from the extensor digitorum longus and communis muscles of rats and guinea-pigs. The fibres were mounted into a double Vaseline gap experimental chamber and the events in excitation-contraction coupling were studied under voltage clamp conditions. 2. The voltage dependence of intramembrane charge movement followed a two-state Boltzmann distribution with maximal available charge of 26.1 +/- 1.5 and 26.1 +/- 1.3 nC microF-1, mid-point voltage of -35.1 +/- 5.0 and -42.2 +/- 1.2 mV and steepness of 16.7 +/- 2.2 and 17.0 +/- 1.9 mV (means +/- S.E.M., n = 7 and 4) in rats and guinea-pigs, respectively. 3. Intracellular calcium concentration ([Ca2+]i) was monitored using the calcium-sensitive dyes antipyrylazo III, fura-2 and mag-fura-5. Resting [Ca2+]i was similar in rats and guinea-pigs with 125 +/- 18 and 115 +/- 8 nM (n = 10 and 9), respectively, while the maximal increase for a 100 ms depolarization to 0 mV was larger in rats (6.3 +/- 1.0 microM; n = 7), than in guinea-pigs (2.8 +/- 0.3; n = 4). 4. The rate of calcium release (Rrel) from the sarcoplasmic reticulum (SR) displayed an early peak followed by a fast and a slow decline to a quasi maintained steady level. After normalizing Rrel to the estimated SR calcium content (1.2 +/- 0.1 and 0.9 +/- 0.1 mM in rats and guinea-pigs, respectively) and correcting for depletion of calcium in the SR the peak and steady levels at 0 mV, respectively, were found to be 2.50 +/- 0.08 and 0.81 +/- 0.06% ms-1 in rats and 2.43 +/- 0.25 and 0.88 +/- 0.01% ms-1 in guinea-pigs. The voltage dependence was essentially the same in both species, but different from that in amphibians. 5. These experiments show that enzymatic isolation yields functionally intact mammalian skeletal muscle fibres for Vaseline gap experiments. The data also suggest a close connection in the regulation of the different kinetic components of SR calcium release in mammalian skeletal muscle. PMID:9423180

  5. 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.

  6. Striated muscle activator of Rho signaling is required for myotube survival but does not influence basal protein synthesis or degradation.

    PubMed

    Wallace, Marita A; Russell, Aaron P

    2013-08-15

    Skeletal muscle mass is regulated by sensing and transmitting extracellular mechanical stress signals to intracellular signaling pathways controlling protein synthesis and degradation. Striated muscle activator of Rho signaling (STARS) is a muscle-specific actin-binding protein that is sensitive to extracellular stress signals. STARS stimulates actin polymerization and influences serum response factor (SRF) and peroxisome proliferator-activated receptor-γ coactivator (PGC)-1α transcription of genes involved in muscle growth, structure, and contraction. The role of STARS in skeletal muscle cells is not well understood. This study investigated whether STARS influenced C2C12 myotube growth by regulating protein synthesis and degradation. The influence of STARS on Pgc-1α, Srf, and Errα mRNA levels, as well as several of their downstream targets involved in muscle cell growth, contraction, and metabolism, was also investigated. STARS overexpression increased actin polymerization, with no effect on protein synthesis, protein degradation, or Akt phosphorylation. STARS overexpression increased Pgc-1α, Srf, Ckmt2, Cpt-1β, and Mhc1 mRNA. STARS knockdown reduced actin polymerization and increased cell death and dead cell protease activity. It also increased markers of inflammation (Casp1, Il-1β, and Mcp-1), regeneration (Socs3 and Myh8), and fast myosin isoforms (Mhc2a and Mhc2x). We show for the first time in muscle cells that STARS overexpression increases actin polymerization and shifts the muscle cell to a more oxidative phenotype. The suppression of STARS causes cell death and increases markers of necrosis, inflammation, and regeneration. As STARS levels are suppressed in clinical models associated with increased necrosis and inflammation, such as aging and limb immobilization, rescuing STARS maybe a future therapeutic strategy to maintain skeletal muscle function and attenuate contraction-induced muscle damage.

  7. Ca2+ release from the sarcoplasmic reticulum compared in amphibian and mammalian skeletal muscle

    PubMed Central

    1996-01-01

    Puzzled by recent reports of differences in specific ligand binding to muscle Ca2+ channels, we quantitatively compared the flux of Ca2+ release from the sarcoplasmic reticulum (SR) in skeletal muscle fibers of an amphibian (frog) and a mammal (rat), voltage clamped in a double Vaseline gap chamber. The determinations of release flux were carried out by the "removal" method and by measuring the rate of Ca2+ binding to dyes in large excess over other Ca2+ buffers. To have a more meaningful comparison, the effects of stretching the fibers, of rapid changes in temperature, and of changes in the Ca2+ content of the SR were studied in both species. In both frogs and rats, the release flux had an early peak followed by fast relaxation to a lower sustained release. The peak and steady values of release flux, Rp and Rs, were influenced little by stretching. Rp in frogs was 31 mM/s (SEM = 4, n = 24) and in rats 7 +/- 2 mM/s (n = 12). Rs was 9 +/- 1 and 3 +/- 0.7 mM/s in frogs and rats, respectively. Transverse (T) tubule area, estimated from capacitance measurements and normalized to fiber volume, was greater in rats (0.61 +/- 0.04 microns-1) than in frogs (0.48 +/- 0.04 micron-1), as expected from the greater density of T tubuli. Total Ca in the SR was estimated as 3.4 +/- 0.6 and 1.9 +/- 0.3 mmol/liter myoplasmic water in frogs and rats. With the above figures, the steady release flux per unit area of T tubule was found to be fourfold greater in the frog, and the steady permeability of the junctional SR was about threefold greater. The ratio Rp/Rs was approximately 2 in rats at all voltages, whereas it was greater and steeply voltage dependent in frogs, going through a maximum of 6 at -40 mV, then decaying to approximately 3.5 at high voltage. Both Rp and Rs depended strongly on the temperature, but their ratio, and its voltage dependence, did not. Assuming that the peak of Ca2+ release is contributed by release channels not in contact with voltage sensors, or not under

  8. Small, Membrane-bound, Alternatively Spliced Forms of Ankyrin 1 Associated with the Sarcoplasmic Reticulum of Mammalian Skeletal Muscle

    PubMed Central

    Zhou, Daixing; Birkenmeier, Connie S.; Williams, McRae W.; Sharp, John J.; Barker, Jane E.; Bloch, Robert J.

    1997-01-01

    We have recently found that the erythroid ankyrin gene, Ank1, expresses isoforms in mouse skeletal muscle, several of which share COOH-terminal sequence with previously known Ank1 isoforms but have a novel, highly hydrophobic 72–amino acid segment at their NH2 termini. Here, through the use of domainspecific peptide antibodies, we report the presence of the small ankyrins in rat and rabbit skeletal muscle and demonstrate their selective association with the sarcoplasmic reticulum. In frozen sections of rat skeletal muscle, antibodies to the spectrin-binding domain (anti-p65) react only with a 210-kD Ank1 and label the sarcolemma and nuclei, while antibodies to the COOH terminus of the small ankyrin (anti-p6) react with peptides of 20 to 26 kD on immunoblots and decorate the myoplasm in a reticular pattern. Mice homozygous for the normoblastosis mutation (gene symbol nb) are deficient in the 210-kD ankyrin but contain normal levels of the small ankyrins in the myoplasm. In nb/nb skeletal muscle, anti-p65 label is absent from the sarcolemma, whereas anti-p6 label shows the same distribution as in control skeletal muscle. In normal skeletal muscle of the rat, anti-p6 decorates Z lines, as defined by antidesmin distribution, and is also present at M lines where it surrounds the thick myosin filaments. Immunoblots of the proteins isolated with rabbit sarcoplasmic reticulum indicate that the small ankyrins are highly enriched in this fraction. When expressed in transfected HEK 293 cells, the small ankyrins are distributed in a reticular pattern resembling the ER if the NH2-terminal hydrophobic domain is present, but they are uniformly distributed in the cytosol if this domain is absent. These results suggest that the small ankyrins are integral membrane proteins of the sarcoplasmic reticulum. We propose that, unlike the 210-kD form of Ank1, previously localized to the sarcolemma and believed to be a part of the supporting cytoskeleton, the small Ank1 isoforms may

  9. Acute vascular endothelial growth factor expression during hypertrophy is muscle phenotype specific and localizes as a striated pattern within fibres.

    PubMed

    Parvaresh, Kevin C; Huber, Ashley M; Brochin, Robert L; Bacon, Phoebe L; McCall, Gary E; Huey, Kimberly A; Hyatt, Jon-Philippe K

    2010-11-01

    Skeletal muscle hypertrophy requires the co-ordinated expression of locally acting growth factors that promote myofibre growth and concurrent adaptive changes in the microvasculature. These studies tested the hypothesis that vascular endothelial growth factor (VEGF) and heparin-binding epidermal growth factor (HB-EGF) expression are upregulated during the early stages of compensatory muscle growth induced by chronic functional overload (FO). Bilateral FO of the plantaris and soleus muscles was induced for 3 or 7 days in the hindlimbs of adult female Sprague-Dawley rats (n = 5 per group) and compared with control (non-FO) rats. Relative muscle mass (in mg (kg body weight)(-1)) increased by 18 and 24% after 3 days and by 20 and 33% after 7 days in the plantaris and soleus muscles, respectively. No differences in HB-EGF mRNA or protein were observed in either muscle of FO rats relative to control muscles. The VEGF mRNA was similar in the soleus muscles of FO and control rats, whereas a significant elevation occurred at 3 and 7 days of FO in the plantaris muscle. However, VEGF protein expression after 3 days of FO exhibited a differential response; expression in the soleus muscle decreased 1.6-fold, whereas that in the plantaris muscle increased 1.8-fold compared with the control muscle. After 7 days of FO, VEGF protein remained elevated within the plantaris muscle, but returned to basal levels in the soleus. Robust basal HB-EGF and VEGF protein expression was consistently seen in control muscles. In all groups, immunohistochemistry for VEGF protein displayed a distinct striated expression pattern within myofibres, with considerably less labelling in extracellular spaces. Constitutive expression of HB-EGF and VEGF in control myofibres is consistent with housekeeping roles for these growth factors in skeletal muscle tissue. However, the specific patterns of VEGF expression in these muscles during FO may reflect the chronic changes in neural recruitment between muscles

  10. ZASP Interacts with the Mechanosensing Protein Ankrd2 and p53 in the Signalling Network of Striated Muscle

    PubMed Central

    Martinelli, Valentina C.; Kyle, W. Buck; Kojic, Snezana; Vitulo, Nicola; Li, Zhaohui; Belgrano, Anna; Maiuri, Paolo; Banks, Lawrence; Vatta, Matteo; Valle, Giorgio; Faulkner, Georgine

    2014-01-01

    ZASP is a cytoskeletal PDZ-LIM protein predominantly expressed in striated muscle. It forms multiprotein complexes and plays a pivotal role in the structural integrity of sarcomeres. Mutations in the ZASP protein are associated with myofibrillar myopathy, left ventricular non-compaction and dilated cardiomyopathy. The ablation of its murine homologue Cypher results in neonatal lethality. ZASP has several alternatively spliced isoforms, in this paper we clarify the nomenclature of its human isoforms as well as their dynamics and expression pattern in striated muscle. Interaction is demonstrated between ZASP and two new binding partners both of which have roles in signalling, regulation of gene expression and muscle differentiation; the mechanosensing protein Ankrd2 and the tumour suppressor protein p53. These proteins and ZASP form a triple complex that appears to facilitate poly-SUMOylation of p53. We also show the importance of two of its functional domains, the ZM-motif and the PDZ domain. The PDZ domain can bind directly to both Ankrd2 and p53 indicating that there is no competition between it and p53 for the same binding site on Ankrd2. However there is competition for this binding site between p53 and a region of the ZASP protein lacking the PDZ domain, but containing the ZM-motif. ZASP is negative regulator of p53 in transactivation experiments with the p53-responsive promoters, MDM2 and BAX. Mutations in the ZASP ZM-motif induce modification in protein turnover. In fact, two mutants, A165V and A171T, were not able to bind Ankrd2 and bound only poorly to alpha-actinin2. This is important since the A165V mutation is responsible for zaspopathy, a well characterized autosomal dominant distal myopathy. Although the mechanism by which this mutant causes disease is still unknown, this is the first indication of how a ZASP disease associated mutant protein differs from that of the wild type ZASP protein. PMID:24647531

  11. Ryanodine modification of cardiac muscle responses to potassium-free solutions. Evidence for inhibition of sarcoplasmic reticulum calcium release

    PubMed Central

    1983-01-01

    To test whether ryanodine blocks the release of calcium from the sarcoplasmic reticulum in cardiac muscle, we examined its effects on the aftercontractions and transient depolarizations or transient inward currents developed by guinea pig papillary muscles and voltage-clamped calf cardiac Purkinje fibers in potassium-free solutions. Ryanodine (0.1-1.0 microM) abolished or prevented aftercontractions and transient depolarizations by the papillary muscles without affecting any of the other sequelae of potassium removal. In the presence of 4.7 mM potassium and at a stimulation rate of 1 Hz, ryanodine had only a small variable effect on papillary muscle force development and action potential characteristics. In calf Purkinje fibers, ryanodine (1 nM-1 microM) completely blocked the aftercontractions and transient inward currents without altering the steady state current-voltage relationship. Ryanodine also abolished the twitch in potassium-free solutions, but it enhanced the tonic force during depolarizing voltage- clamp steps. This latter effect was dependent on the combination of ryanodine and potassium-free solutions. The slow inward current was not blocked by 1 microM ryanodine, but ryanodine did appear to abolish an outward current that remained in the presence of 0.5 mM 4- aminopyridine. Our observations are consistent with the hypothesis that ryanodine, by inhibiting the release of calcium from the sarcoplasmic reticulum, prevents the oscillations in intracellular calcium that activate the transient inward currents and aftercontractions associated with calcium overload states. PMID:6631403

  12. Effects of preslaughter stress levels on the post-mortem sarcoplasmic proteomic profile of gilthead seabream muscle.

    PubMed

    Silva, Tomé S; Cordeiro, Odete D; Matos, Elisabete D; Wulff, Tune; Dias, Jorge P; Jessen, Flemming; Rodrigues, Pedro M

    2012-09-19

    Fish welfare is an important concern in aquaculture, not only due to the ethical implications but also for productivity and quality-related reasons. The purpose of this study was to track soluble proteome expression in post-mortem gilthead seabream muscle and to observe how preslaughter stress affects these post-mortem processes. For the experiment, two groups of gilthead seabream (n = 5) were subjected to distinct levels of preslaughter stress, with three muscle samples being taken from each fish. Proteins were extracted from the muscle samples, fractionated, and separated by 2DE. Protein identification was performed by MALDI-TOF-TOF MS. Analysis of the results indicates changes on several cellular pathways, with some of these changes being attributable to oxidative and proteolytic activity on sarcoplasmic proteins, together with leaking of myofibrillar proteins. These processes appear to have been hastened by preslaughter stress, confirming that it induces clear post-mortem changes in the muscle proteome of gilthead seabream.

  13. High intracellular [Ca2+] alters sarcoplasmic reticulum function in skinned skeletal muscle fibres of the rat

    PubMed Central

    Lamb, G D; Cellini, M A

    1999-01-01

    The effect on sarcoplasmic reticulum (SR) function of exposure to high intracellular [Ca2+] was studied in mechanically skinned fibres from the extensor digitorum longus muscle of the rat, using caffeine to assay the SR Ca2+ content. A 15 s exposure to 50 μm Ca2+ irreversibly reduced the ability of the SR to load/retain Ca2+ and completely abolished depolarization-induced Ca2+ release, whereas a 90 s exposure to 10 μm Ca2+ had no detectable effect on either function. The reduction in net SR Ca2+ uptake: (a) was near-maximal with treatment at 50 μm Ca2+, (b) was unrelated to voltage-sensor function, and (c) persisted unchanged for > 20 min. The reduction was primarily due to a threefold increase in leakage of Ca2+ out of the SR. This increased leakage was not substantially blocked by the presence of 10 mM Mg2+ or 2 μm Ruthenium Red. The adverse effect on SR function of exposure to high [Ca2+] could also be observed by the reduction in the ability of the SR to maintain a low [Ca2+] within the skinned fibre in the face of elevated [Ca2+] in the bathing solution. When bathed in a solution with ≈1.5 μm Ca2+ (0.75 mM CaEGTA-EGTA), skinned fibres produced only low force responses for many minutes, but after high [Ca2+] treatment (15 s exposure to 50 μm Ca2+) they showed large, steady or oscillatory force responses. These findings indicate that, in addition to uncoupling the Ca2+ release channels from the voltage sensors, exposure of skinned fibres to high [Ca2+] causes a persistent increase in resting Ca2+ efflux from the SR. Such efflux in an intact fibre would alter the distribution of Ca2+ between the SR, the cytoplasm and the extracellular solution. These results may be relevant to the basis of low-frequency fatigue and possibly other conditions in muscle. PMID:10457093

  14. Calcium release by noradrenaline from central sarcoplasmic reticulum in rabbit main pulmonary artery smooth muscle.

    PubMed Central

    Kowarski, D; Shuman, H; Somlyo, A P; Somlyo, A V

    1985-01-01

    The subcellular composition of relaxed and noradrenaline-contracted rabbit main pulmonary artery smooth muscle cells was measured by electron probe X-ray microanalysis of cryosections of rapidly frozen tissue. Some of the preparations were made permeable with saponin and exposed to a known free Ca ion concentration, rapidly frozen, freeze-substituted, and also analysed by electron probe X-ray microanalysis. 98% of intracellular K could be replaced by Rb. This was done to remove the K peak that partially overlaps the Ca peak in the X-ray spectra. The final [Rb]i plus residual [K]i was not significantly different from the [K]i of normal tissue. The [Ca]i in Rb-containing tissue was not significantly different from the [Ca]i in normal, K-containing tissue. Non-mitochondrial micro-regions containing high [Ca] (up to 33 mmol/kg dry wt.) were found at sites 200 nm or more away from the plasma membrane. These micro-regions also contained high [P]. We consider the identification of these regions containing high [Ca] as sarcoplasmic reticulum (s.r.), validated by: (a) conventional electron micrographs that show no other structures in main pulmonary artery smooth muscle in sufficient quantity and location to account for the frequency of these regions, (b) the previous localization of strontium, a functional calcium analogue, in the central s.r. in these smooth muscles (Somlyo & Somlyo, 1971 a), (c) the present demonstration that the central s.r. in this tissue can accumulate large amounts of calcium oxalate. The proportion of regions containing high [Ca] (greater than 12.0 mmol/kg dry wt.) was significantly higher in relaxed (35 of 330 measurements) than in the contracted (14 of 337) tissues (P less than 0.005), or 26 of 34 vs. 6 of 31 high [Ca] measurements in regions identified as s.r. through their high phosphorus content (P less than 0.006). This difference is thought to represent Ca release from the central s.r. There was no significant difference (P greater than 0

  15. Myostatin, a profibrotic factor and the main inhibitor of striated muscle mass, is present in the penile and vascular smooth muscle.

    PubMed

    Kovanecz, I; Masouminia, M; Gelfand, R; Vernet, D; Rajfer, J; Gonzalez-Cadavid, N F

    2017-09-01

    Myostatin is present in striated myofibers but, except for myometrial cells, has not been reported within smooth muscle cells (SMC). We investigated in the rat whether myostatin is present in SMC within the penis and the vascular wall and, if so, whether it is transcriptionally expressed and associated with the loss of corporal SMC occurring in certain forms of erectile dysfunction (ED). Myostatin protein was detected by immunohistochemistry/fluorescence and western blots in the perineal striated muscles, and also in the SMC of the penile corpora, arteries and veins, and aorta. Myostatin was found in corporal SMC cultures, and its transcriptional expression (and its receptor) was shown there by DNA microarrays. Myostatin protein was measured by western blots in the penile shaft of rats subjected to bilateral cavernosal nerve resection (BCNR), that were left untreated, or treated (45 days) with muscle-derived stem cells (MDSC), or concurrent daily low-dose sildenafil. Myostatin was not increased by BCNR (compared with sham operated animals), but over expressed after treatment with MDSC. This was reduced by concurrent sildenafil. The presence of myostatin in corporal and vascular SMC, and its overexpression in the corpora by MDSC therapy, may have relevance for the stem cell treatment of corporal fibrosis and ED.

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

    USDA-ARS?s Scientific Manuscript database

    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...

  17. Effect of gadolinium on the ryanodine receptor/sarcoplasmic reticulum calcium release channel of skeletal muscle.

    PubMed

    Sárközi, Sándor; Szegedi, Csaba; Lukács, Balázs; Ronjat, Michel; Jóna, István

    2005-01-01

    The effect of gadolinium ions on the sarcoplasmic reticulum (SR) calcium release channel/ryanodine receptor (RyR1) was studied using heavy SR (HSR) vesicles and RyR1 isolated from rabbit fast twitch muscle. In the [(3)H]ryanodine binding assay, 5 microM Gd(3+) increased the K(d) of the [(3)H]ryanodine binding of the vesicles from 33.8 nM to 45.6 nM while B(max), referring to the binding capacity, was not affected significantly. In the presence of 18 nM[(3)H]ryanodine and 100 microM free Ca(2+), Gd(3+) inhibited the binding of the radiolabeled ryanodine with an apparent K(d) value of 14.7 microM and a Hill coefficient of 3.17. In (45)Ca(2+) experiments the time constant of (45)Ca(2+) efflux from HSR vesicles increased from 90.9 (+/- 11.1) ms to 187.7 (+/- 24.9) ms in the presence of 20 microM gadolinium. In single channel experiments gadolinium inhibited the channel activity from both the cytoplasmic (cis) (IC(50) = 5.65 +/- 0.33 microM, n(Hill) = 4.71) and the luminal (trans) side (IC(50) = 5.47 +/- 0.24 microM, n(Hill) = 4.31). The degree of inhibition on the cis side didn't show calcium dependency in the 100 microM to 1 mM Ca(2+) concentration range which indicates no competition with calcium on its regulatory binding sites. When Gd(3+) was applied at the trans side, EGTA was present at the cis side to prevent the binding of Gd(+3) to the cytoplasmic calcium binding regulatory sites of the RyR1 if Gd(3+) accidentally passed through the channel. The inhibition of the channel did not show any voltage dependence, which would be the case if Gd(3+) exerted its effect after getting to the cis side. Our results suggest the presence of inhibitory binding sites for Gd(3+) on both sides of the RyR1 with similar Hill coefficients and IC(50) values.

  18. Effects of procaine and caffeine on calcium release from the sarcoplasmic reticulum in frog skeletal muscle.

    PubMed Central

    Klein, M G; Simon, B J; Schneider, M F

    1992-01-01

    1. Resting myoplasmic free [Ca2+] and [Ca2+] transients (delta [Ca2+]) were measured in single voltage-clamped frog skeletal muscle fibres in the presence and absence of procaine, caffeine or procaine plus caffeine using Fura-2 fluorescence and antipyrylazo III (Ap III) absorbance signals. The rate of release (Rrel) of calcium from the sarcoplasmic reticulum (SR) was calculated from the calcium transients and corrected for the relatively small decline due to depletion of calcium from the SR. 2. Procaine (1 mM) reversibly suppressed delta [Ca2+] and the corresponding Rrel by about 40% for 60-100 ms depolarizing steps to -40 to +20 mV. Procaine had little effect on either the waveform or voltage dependence of the Rrel records. 3. [Ca2+] transients calculated from Fura-2 fluorescence changes in the presence or absence of procaine had similar time courses and amplitudes as those calculated from the Ap III absorbance changes suggesting that 1 mM-procaine did not interfere with the ability of Ap III or Fura-2 to monitor delta [Ca2+]. 4. Although 1 mM-procaine depressed Rrel it had no effect on intramembrane charge movements (IQ) calculated from membrane currents recorded simultaneously with delta [Ca2+]. 5. Procaine (1 mM) reversibly inhibited the potentiating effect of 0.5 mM-caffeine on delta [Ca2+]. The amplitude and waveform of the Rrel records were similar in control fibres and in the presence of 1 mM-procaine plus 0.5 mM-caffeine. 6. In the presence of 0.5 mM-caffeine delta [Ca2+] after 10-20 ms voltage steps exhibited an increase in the time to peak and a slower decay time course compared with caffeine-free controls, suggestive of significant calcium-induced calcium release in the presence of caffeine. These effects of caffeine were completely and reversibly blocked by 1 mM-procaine. 7. In the absence of caffeine, 1 mM-procaine caused a small decrease in time to peak of delta [Ca2+] after 10-30 ms duration voltage steps compared to the bracketing control and wash

  19. The sarcoplasmic reticulum and sarcolemma together form a passive Ca2+ trap in colonic smooth muscle.

    PubMed

    Bradley, Karen N; Craig, John W; Muir, Thomas C; McCarron, John G

    2004-07-01

    In smooth muscle, active Ca(2+) uptake into regions of sarcoplasmic reticulum (SR) which are closely apposed to the sarcolemma has been proposed to substantially limit the increase in the cytoplasmic Ca(2+) concentration ([Ca(2+)](c)) following Ca(2+) influx, i.e. the 'superficial buffer barrier hypothesis'. The present study has re-examined this proposal. The results suggest that the SR close to the sarcolemma acts as a passive barrier to Ca(2+) influx limiting [Ca(2+)](c) changes; for this, SR Ca(2+) pump activity is not required. In single voltage-clamped colonic myocytes, sustained opening of the ryanodine receptor (RyR) (and depletion of the SR) using ryanodine increased the amplitude of depolarisation-evoked Ca(2+) transients and accelerated the rate of [Ca(2+)](c) decline following depolarisation. These results could be explained by a reduction in the Ca(2+) buffer power of the cytosol taking place when RyR are opened (i.e. the SR is 'leaky'). Indeed, determination of the Ca(2+) buffer power confirmed it was reduced by approximately 40%. Inhibition of the SR Ca(2+) pump (with thapsigargin) also depleted the SR of Ca(2+) but did not reduce the Ca(2+) buffer power or increase depolarisation-evoked Ca(2+) transients and slowed (rather than accelerated) Ca(2+) removal. However, thapsigargin prevented the ryanodine-induced increase in [Ca(2+)](c) decline following depolarisation. Together, these results suggest that when the SR was rendered 'leaky' (a) more of the Ca(2+) entering the cell reached the bulk cytoplasm and (b) Ca(2+) was removed more quickly at the end of cell activation. Under physiological circumstances in the absence of blocking drugs, it is proposed that the SR limits the [Ca(2+)](c) increase following influx without the need for active Ca(2+) uptake. The SR and sarcolemma may form a passive physical barrier to Ca(2+) influx, a Ca(2+) trap, which limits the [Ca(2+)](c) rise occurring during depolarisation by about 50% and from which the ion only

  20. 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

  1. Correction of multiple striated muscles in murine Pompe disease through adeno-associated virus-mediated gene therapy.

    PubMed

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

    2008-08-01

    Glycogen storage disease type II (Pompe disease; MIM 232300) stems from the deficiency of acid alpha-glucosidase (GAA; acid maltase; EC 3.2.1.20), which primarily involves cardiac and skeletal muscles. An adeno-associated virus 2/8 (AAV2/8) vector containing the muscle creatine kinase (MCK) (CK1) reduced glycogen content by approximately 50% in the heart and quadriceps in GAA-knockout (GAA-KO) mice; furthermore, an AAV2/8 vector containing the hybrid alpha-myosin heavy chain enhancer-/MCK enhancer-promoter (MHCK7) cassette reduced glycogen content by >95% in heart and >75% in the diaphragm and quadriceps. Transduction with an AAV2/8 vector was higher in the quadriceps than in the gastrocnemius. An AAV2/9 vector containing the MHCK7 cassette corrected GAA deficiency in the distal hindlimb, and glycogen accumulations were substantially cleared by human GAA (hGAA) expression therein; however, the analogous AAV2/7 vector achieved much lower efficacy. Administration of the MHCK7-containing vectors significantly increased striated muscle function as assessed by increased Rotarod times at 18 weeks after injection, whereas the CK1-containing vector did not increase Rotarod performance. Importantly, type IIb myofibers in the extensor digitalis longus (EDL) 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.

  2. Fiber type composition of pubococcygeus and bulbospongiosus striated muscles is modified by multiparity in the rabbit.

    PubMed

    López-García, Kenia; Mariscal-Tovar, Silvia; Serrano-Meneses, Martín Alejandro; Castelán, Francisco; Martínez-Gómez, Margarita; Jiménez-Estrada, Ismael

    2017-08-01

    We analyzed the effect of multiparity on the fiber type composition of two skeletal muscles involved in the maintenance of the micturition process, the pelvic pubococcygeus (Pc) and perineal bulbospongiosus (Bs) muscles in nulliparous and multiparous rabbits (Oryctolagus cuniculus). We used the basic ATPase and NADH-TR techniques to identify and characterize slow, intermediate, and fast fiber types and glycolitic and oxidative fibers in muscles, respectively. Pc muscles of multiparous rabbits present relatively high percentages of slow and intermediate fibers but a low percentage of fast fibers (P < 0.05) as compared to Pc muscles from nulliparous rabbits, while percentages of glycolytic and oxidative fibers were similar (P > 0.05). Bs muscles of multiparous rabbits had a higher proportion of intermediate and glycolytic fibers (P < 0.05) than muscles of nulliparous. Both, Pc and Bs muscles of nulliparous and multiparous rabbits contain slow fibers with similar large cross sectional area, but fast fibers in multiparous muscles showed small cross sectional area than in nulliparous. Multiparity modified the fiber type composition of Pc and Bs muscles in female rabbits. We propose that the contractile force and the physiological role of both muscles during micturition are affected because of the observed changes in the relative composition of muscle fiber types. © 2016 Wiley Periodicals, Inc.

  3. Cannabinoid signalling inhibits sarcoplasmic Ca(2+) release and regulates excitation-contraction coupling in mammalian skeletal muscle.

    PubMed

    Oláh, Tamás; Bodnár, Dóra; Tóth, Adrienn; Vincze, János; Fodor, János; Reischl, Barbara; Kovács, Adrienn; Ruzsnavszky, Olga; Dienes, Beatrix; Szentesi, Péter; Friedrich, Oliver; Csernoch, László

    2016-12-15

    Marijuana was found to cause muscle weakness, although the exact regulatory role of its receptors (CB1 cannabinoid receptor; CB1R) in the excitation-contraction coupling (ECC) of mammalian skeletal muscle remains unknown. We found that CB1R activation or its knockout did not affect muscle force directly, whereas its activation decreased the Ca(2+) -sensitivity of the contractile apparatus and made the muscle fibres more prone to fatigue. We demonstrate that CB1Rs are not connected to the inositol 1,4,5-trisphosphate pathway either in myotubes or in adult muscle fibres. By contrast, CB1Rs constitutively inhibit sarcoplasmic Ca(2+) release and sarcoplasmic reticulum Ca(2+) ATPase during ECC in a Gi/o protein-mediated way in adult skeletal muscle fibres but not in myotubes. These results help with our understanding of the physiological effects and pathological consequences of CB1R activation in skeletal muscle and may be useful in the development of new cannabinoid drugs. Marijuana was found to cause muscle weakness, although it is unknown whether it affects the muscles directly or modulates only the motor control of the central nervous system. Although the presence of CB1 cannabinoid receptors (CB1R), which are responsible for the psychoactive effects of the drug in the brain, have recently been demonstrated in skeletal muscle, it is unclear how CB1R-mediated signalling affects the contraction and Ca²⁺ homeostasis of mammalian skeletal muscle. In the present study, we demonstrate that in vitro CB1R activation increased muscle fatigability and decreased the Ca(2+) -sensitivity of the contractile apparatus, whereas it did not alter the amplitude of single twitch contractions. In myotubes, CB1R agonists neither evoked, nor influenced inositol 1,4,5-trisphosphate (IP3 )-mediated Ca(2+) transients, nor did they alter excitation-contraction coupling. By contrast, in isolated muscle fibres of wild-type mice, although CB1R agonists did not evoke IP3 -mediated Ca(2

  4. (-)-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. © 2014 FEBS.

  5. (−)-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

  6. [Striated lichen].

    PubMed

    Méndez-Santillán, E

    1991-02-01

    Striated lichen is a rare disease seen primarily in pediatric ages. Of more of less quick appearance, generally persisting for a year, and later spontaneously involuting. Its etiology and pathogenia are unkown. It is a disease essentially clinically diagnosed and should be differentiated from other entities of linear distribution.

  7. Expression of distinct classes of titin isoforms in striated and smooth muscles by alternative splicing, and their conserved interaction with filamins.

    PubMed

    Labeit, Siegfried; Lahmers, Sunshine; Burkart, Christoph; Fong, Chi; McNabb, Mark; Witt, Stephanie; Witt, Christian; Labeit, Dietmar; Granzier, Henk

    2006-09-29

    While the role of titin as a sarcomeric protein is well established, its potential functional role(s) in smooth muscles and non-muscle tissues are controversial. We used a titin exon array to search for which part(s) of the human titin transcriptional unit encompassing 363 exons is(are) expressed in non-striated muscle tissues. Expression profiling of adult smooth muscle tissues (aorta, bladder, carotid, stomach) identified alternatively spliced titin isoforms, encompassing 80 to about 100 exons. These exons code for parts of the titin Z-disk, I-band and A-band regions, allowing the truncated smooth muscle titin isoform to link Z-disks/dense bodies together with thick filaments. Consistent with the array data, Western blot studies detected the expression of approximately 1 MDa smooth muscle titin in adult smooth muscles, reacting with selected Z-disc, I-band, and A-band titin antibodies. Immunofluorescence with these antibodies located smooth muscle titin in the cytoplasm of cultured human aortic smooth muscle cells and in the tunica media of intact adult bovine aorta. Real time PCR studies suggested that smooth muscle titins are expressed from a promoter located 35 kb or more upstream of the transcription initiation site used for striated muscle titin, driving expression of a bi-cistronic mRNA, coding 5' for the anonymous gene FL39502, followed 3' by titin, respectively. Our work showed that smooth muscle and striated muscle titins share in their conserved amino-terminal regions binding sites for alpha-actinin and filamins: Yeast two-hybrid screens using Z2-Zis1 titin baits identified prey clones coding for alpha-actinin-1 and filamin-A from smooth muscle, and alpha-actinin-2/3, filamin-C, and nebulin from skeletal muscle cDNA libraries, respectively. This suggests that the titin Z2-Zis1 domain can link filamins and alpha-actinin together in the periphery of the Z-line/dense bodies in a fashion that is conserved in smooth and striated muscles.

  8. Direct x-ray observation of a single hexagonal myofilament lattice in native myofibrils of striated muscle.

    PubMed Central

    Iwamoto, Hiroyuki; Nishikawa, Yukihiro; Wakayama, Jun'ichi; Fujisawa, Tetsuro

    2002-01-01

    A striated muscle fiber consists of thousands of myofibrils with crystalline hexagonal myofilament lattices. Because the lattices are randomly oriented, the fiber gives rise to an equatorial x-ray diffraction pattern, which is essentially a rotary-averaged "powder diffraction," carrying only information about the distance between the lattice planes. We were able to record an x-ray diffraction pattern from a single myofilament lattice, very likely originating from a single myofibril from the flight muscle of a bumblebee, by orienting the incident x-ray microbeam along the myofibrillar axis (end-on diffraction). The pattern consisted of a number of hexagonally symmetrical diffraction spots whose originating lattice planes were readily identified. This also held true for some of the weak higher order reflections. The spot-like appearance of reflections implies that the lattice order is extremely well maintained for a distance of millimeters, covering up to a thousand of approximately 2.5-microm-long sarcomeres connected in series. The results open the possibility of applying the x-ray microdiffraction technique to study many other micrometer-sized assemblies of functional biomolecules in the cell. PMID:12124287

  9. 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.

  10. 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

  11. On the Rate of Synthesis of Individual Proteins within and between Different Striated Muscles of the Rat.

    PubMed

    Hesketh, Stuart; Srisawat, Kanchana; Sutherland, Hazel; Jarvis, Jonathan; Burniston, Jatin

    2016-03-15

    striated muscles.

  12. On the Rate of Synthesis of Individual Proteins within and between Different Striated Muscles of the Rat

    PubMed Central

    Hesketh, Stuart; Srisawat, Kanchana; Sutherland, Hazel; Jarvis, Jonathan; Burniston, Jatin

    2016-01-01

    The turnover of muscle protein is responsive to different (patho)-physiological conditions but little is known about the rate of synthesis at the level of individual proteins or whether this varies between different muscles. We investigated the synthesis rate of eight proteins (actin, albumin, ATP synthase alpha, beta enolase, creatine kinase, myosin essential light chain, myosin regulatory light chain and tropomyosin) in the extensor digitorum longus, diaphragm, heart and soleus of male Wistar rats (352 ± 30 g body weight). Animals were assigned to four groups (n = 3, in each), including a control and groups that received deuterium oxide (2H2O) for 4 days, 7 days or 14 days. Deuterium labelling was initiated by an intraperitoneal injection of 10 μL/g body weight of 99.9% 2H2O-saline, and was maintained by administration of 5% (v/v) 2H2O in drinking water provided ad libitum. Homogenates of the isolated muscles were analysed by 2-dimensional gel electrophoresis and matrix-assisted laser desorption ionisation time of flight mass spectrometry. Proteins were identified against the SwissProt database using peptide mass fingerprinting. For each of the eight proteins investigated, the molar percent enrichment (MPE) of 2H and rate constant (k) of protein synthesis was calculated from the mass isotopomer distribution of peptides based on the amino acid sequence and predicted number of exchangeable C–H bonds. The average MPE (2.14% ± 0.2%) was as expected and was consistent across muscles harvested at different times (i.e., steady state enrichment was achieved). The synthesis rate of individual proteins differed markedly within each muscle and the rank-order of synthesis rates differed among the muscles studied. After 14 days the fraction of albumin synthesised (23% ± 5%) was significantly (p < 0.05) greater than for other muscle proteins. These data represent the first attempt to study the synthesis rates of individual proteins across a number of different striated

  13. Sarcoplasmic reticulum calcium release in frog skeletal muscle fibres estimated from Arsenazo III calcium transients.

    PubMed Central

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

    1983-01-01

    Single twitch fibres, dissected from frog muscle, were injected with the metallochromic dye Arsenazo III. Changes in dye-related absorbance measured at 650 or 660 nm were used to estimate the time course of myoplasmic free [Ca2+] following either action potential stimulation or voltage-clamp depolarization (temperature, 15-17 degrees C). The amplitude of the Ca2+ transient decreased when fibres were stretched to sarcomere spacings approaching 4 microns. The effect appeared to be less marked in H2O Ringer than in D2O Ringer, where a reduction of about 40% was observed in going from 3.0 microns to 3.7-3.9 microns. In fibres heavily injected with dye (1.5-2.2 mM-dye) at least 0.1 mM-Ca2+ was complexed with Arsenazo III following a single action potential, implying that at least 0.1 mM-Ca2+ was released from the sarcoplasmic reticulum (s.r.) into the myoplasm. Computer simulations were carried out to estimate the flux of Ca2+ between the s.r. and myoplasm (in fibres containing no more that 0.8 mM-dye). The amounts and time courses of Ca2+ bound to the Ca2+-regulatory sites on troponin and to the Ca2+, Mg2+ sites on parvalbumin were estimated from the free [Ca2+] wave form and the law of mass action. In the computations the total myoplasmic [Ca2+] was taken as the total amount of Ca2+ existing either as free ion or as ion complexed with dye, troponin or parvalbumin. The time derivative of total myoplasmic [Ca2+] was used as an estimate of net Ca2+ flux (release minus uptake) from the s.r. into myoplasm. Rate constants for formation of cation: receptor complex were taken from published values. For the Ca2+-regulatory sites on troponin, three sets of rate constants, corresponding to two values of dissociation constant (0.2 and 2 microM) were used. Each set of three simulations was carried out both with and without parvalbumin. The simulations show that following action potential stimulation, 0.2-0.3 mM-Ca2+ enters the myoplasm from the s.r. The wave form of s.r. Ca2

  14. Effect of sarcoplasmic reticulum calcium depletion on intramembranous charge movement in frog cut muscle fibers

    PubMed Central

    1995-01-01

    Cut muscle fibers from Rana temporaria (sarcomere length, 3.3-3.5 microns; temperature, 13-16 degrees C) were mounted in a double Vaseline-gap chamber and equilibrated for at least an hour with an internal solution that contained 20 mM EGTA and phenol red and an external solution that contained predominantly TEA-gluconate; both solutions were nominally Ca-free. The increase in total myoplasmic concentration of Ca (delta[CaT]) produced by sarcoplasmic reticulum (SR) Ca release was estimated from the change in pH produced when the released Ca was complexed by EGTA (Pape, P.C., D.-S. Jong, and W.K. Chandler. 1995. Journal of General Physiology. 106:259-336). The resting value of SR Ca content, [CaSR]R (expressed as myoplasmic concentration), was taken to be equal to the value of delta[CaT] obtained during a step depolarization (usually to -50 to -40 mV) that was sufficiently long (200-750 ms) to release all of the readily releasable Ca from the SR. In ten fibers, the first depolarization gave [CaSR]R = 839-1,698 microM. Progressively smaller values were obtained with subsequent depolarizations until, after 30-40 depolarizations, the value of [CaSR]R had usually been reduced to < 10 microM. Measurements of intramembranous charge movement, Icm, showed that, as the value of [CaSR]R decreased, ON-OFF charge equality held and the amount of charge moved remained constant. ON Icm showed brief initial I beta components and prominent I gamma "humps", even after the value of [CaSR]R was < 10 microM. Although the amplitude of the hump component decreased during depletion, its duration increased in a manner that preserved the constancy of ON charge. In the depleted state, charge movement was steeply voltage dependent, with a mean value of 7.2 mV for the Boltzmann factor k. These and other results are not consistent with the idea that there is one type of charge, Q beta, and that I gamma is a movement of Q beta caused by SR Ca release, as proposed by Pizarro, Csernoch, Uribe

  15. A Novel Striated Muscle-Specific Myosin-Blocking Drug for the Study of Neuromuscular Physiology

    PubMed Central

    Heredia, Dante J.; Schubert, Douglas; Maligireddy, Siddhardha; Hennig, Grant W.; Gould, Thomas W.

    2016-01-01

    The failure to transmit neural action potentials (APs) into muscle APs is referred to as neuromuscular transmission failure (NTF). Although synaptic dysfunction occurs in a variety of neuromuscular diseases and impaired neurotransmission contributes to muscle fatigue, direct evaluation of neurotransmission by measurement of successfully transduced muscle APs is difficult due to the subsequent movements produced by muscle. Moreover, the voltage-gated sodium channel inhibitor used to study neurotransmitter release at the adult neuromuscular junction is ineffective in embryonic tissue, making it nearly impossible to precisely measure any aspect of neurotransmission in embryonic lethal mouse mutants. In this study we utilized 3-(N-butylethanimidoyl)-4-hydroxy-2H-chromen-2-one (BHC), previously identified in a small-molecule screen of skeletal muscle myosin inhibitors, to suppress movements without affecting membrane currents. In contrast to previously characterized drugs from this screen such as N-benzyl-p-toluene sulphonamide (BTS), which inhibit skeletal muscle myosin ATPase activity but also block neurotransmission, BHC selectively blocked nerve-evoked muscle contraction without affecting neurotransmitter release. This feature allowed a detailed characterization of neurotransmission in both embryonic and adult mice. In the presence of BHC, neural APs produced by tonic stimulation of the phrenic nerve at rates up to 20 Hz were successfully transmitted into muscle APs. At higher rates of phrenic nerve stimulation, NTF was observed. NTF was intermittent and characterized by successful muscle APs following failed ones, with the percentage of successfully transmitted muscle APs diminishing over time. Nerve stimulation rates that failed to produce NTF in the presence of BHC similarly failed to produce a loss of peak muscle fiber shortening, which was examined using a novel optical method of muscle fatigue, or a loss of peak cytosolic calcium transient intensity, examined

  16. A Novel Striated Muscle-Specific Myosin-Blocking Drug for the Study of Neuromuscular Physiology.

    PubMed

    Heredia, Dante J; Schubert, Douglas; Maligireddy, Siddhardha; Hennig, Grant W; Gould, Thomas W

    2016-01-01

    The failure to transmit neural action potentials (APs) into muscle APs is referred to as neuromuscular transmission failure (NTF). Although synaptic dysfunction occurs in a variety of neuromuscular diseases and impaired neurotransmission contributes to muscle fatigue, direct evaluation of neurotransmission by measurement of successfully transduced muscle APs is difficult due to the subsequent movements produced by muscle. Moreover, the voltage-gated sodium channel inhibitor used to study neurotransmitter release at the adult neuromuscular junction is ineffective in embryonic tissue, making it nearly impossible to precisely measure any aspect of neurotransmission in embryonic lethal mouse mutants. In this study we utilized 3-(N-butylethanimidoyl)-4-hydroxy-2H-chromen-2-one (BHC), previously identified in a small-molecule screen of skeletal muscle myosin inhibitors, to suppress movements without affecting membrane currents. In contrast to previously characterized drugs from this screen such as N-benzyl-p-toluene sulphonamide (BTS), which inhibit skeletal muscle myosin ATPase activity but also block neurotransmission, BHC selectively blocked nerve-evoked muscle contraction without affecting neurotransmitter release. This feature allowed a detailed characterization of neurotransmission in both embryonic and adult mice. In the presence of BHC, neural APs produced by tonic stimulation of the phrenic nerve at rates up to 20 Hz were successfully transmitted into muscle APs. At higher rates of phrenic nerve stimulation, NTF was observed. NTF was intermittent and characterized by successful muscle APs following failed ones, with the percentage of successfully transmitted muscle APs diminishing over time. Nerve stimulation rates that failed to produce NTF in the presence of BHC similarly failed to produce a loss of peak muscle fiber shortening, which was examined using a novel optical method of muscle fatigue, or a loss of peak cytosolic calcium transient intensity, examined

  17. High endocytotic activity occurs periodically in the endplate region of denervated mouse striated muscle fibers.

    PubMed

    Lawoko, G; Tågerud, S

    1995-08-01

    High endocytotic activity after denervation of skeletal muscle occurs in a proportion of muscle fibers (both slow and fast fiber types) in the endplate region. The present study was performed in order to examine if a periodicity in the endocytotic activity could explain why the process is not observed in all fibers at a given time. Three markers, horseradish peroxidase (HRP), rhodamine B isothiocyanate-labeled dextran, and fluorescein isothiocyanate-labeled dextran were used to demonstrate endocytotic activity of muscle fibers of the denervated mouse hemidiaphragm in vivo. Acetylcholine esterase staining was used in conjunction with HRP uptake to determine the proportion of denervated muscle fibers with endocytotic activity in the endplate region at any one time. The results show that 25-50% of the muscle fibers display high endocytotic activity in the endplate region at a given time 10 days after denervation. The existence of a periodicity in this endocytotic activity is suggested by results obtained using two different endocytotic markers administered at time intervals of 0-7 days. We conclude that loss of contact with the innervating motorneuron induces a high endocytotic activity which occurs periodically in the perisynaptic region of skeletal muscle fibers.

  18. Triclosan impairs excitation–contraction coupling and Ca2+ dynamics in striated muscle

    PubMed Central

    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.

    2012-01-01

    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 Ca2+ transients followed by complete failure of ECC, independent of Ca2+ store depletion or block of RyR1 channels. TCS also completely blocks excitation-coupled Ca2+ entry. Voltage clamp experiments showed that TCS partially inhibits L-type Ca2+ currents of cardiac and skeletal muscle, and [3H]PN200 binding to skeletal membranes is noncompetitively inhibited by TCS in the same concentration range that enhances [3H]ryanodine binding. TCS potently impairs orthograde and retrograde signaling between L-type Ca2+ and RyR channels in skeletal muscle, and L-type Ca2+ 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. PMID:22891308

  19. 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.

  20. 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.

  1. 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.

  2. 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

  3. 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

  4. Novel diabetes mellitus treatment: mature canine insulin production by canine striated muscle through gene therapy.

    PubMed

    Niessen, S J M; Fernandez-Fuente, M; Mahmoud, A; Campbell, S C; Aldibbiat, A; Huggins, C; Brown, A E; Holder, A; Piercy, R J; Catchpole, B; Shaw, J A M; Church, D B

    2012-07-01

    Muscle-targeted gene therapy using insulin genes has the potential to provide an inexpensive, low maintenance alternative or adjunctive treatment method for canine diabetes mellitus. A canine skeletal muscle cell line was established through primary culture, as well as through transdifferentiation of canine fibroblasts after infection with a myo-differentiation gene containing adenovirus vector. A novel mutant furin-cleavable canine preproinsulin gene insert (cppI4) was designed and created through de novo gene synthesis. Various cell lines, including the generated canine muscle cell line, were transfected with nonviral plasmids containing cppI4. Insulin and desmin immunostaining were used to prove insulin production by muscle cells and specific canine insulin ELISA to prove mature insulin secretion into the medium. The canine myoblast cultures proved positive on desmin immunostaining. All cells tolerated transfection with cppI4-containing plasmid, and double immunostaining for insulin and desmin proved present in the canine cells. Canine insulin ELISA assessment of medium of cppI4-transfected murine myoblasts and canine myoblast and fibroblast mixture proved presence of mature fully processed canine insulin, 24 and 48 h after transfection. The present study provides proof of principle that canine muscle cells can be induced to produce and secrete canine insulin on transfection with nonviral plasmid DNA containing a novel mutant canine preproinsulin gene that produces furin-cleavable canine preproinsulin. This technology could be developed to provide an alternative canine diabetes mellitus treatment option or to provide a constant source for background insulin, as well as C-peptide, alongside current treatment options.

  5. Impact of a nickel-reduced stainless steel implant on striated muscle microcirculation: a comparative in vivo study.

    PubMed

    Kraft, C N; Burian, B; Perlick, L; Wimmer, M A; Wallny, T; Schmitt, O; Diedrich, O

    2001-12-05

    The impairment of skeletal muscle microcirculation by a biomaterial may have profound consequences. With moderately good physical and corrosion characteristics, implant-quality stainless steel is particularly popular in orthopedic surgery. However, due to the presence of a considerable amount of nickel in the alloy, concern has been voiced in respect to local tissue responses. More recently a stainless steel alloy with a significant reduction of nickel has become commercially available. We, therefore, studied in vivo nutritive perfusion and leukocytic response of striated muscle to this nickel-reduced alloy, and compared these results with those of the materials conventional stainless steel and titanium. Using the hamster dorsal skinfold chamber preparation and intravital microscopy, we could demonstrate that reduction of the nickel quantity in a stainless steel implant has a positive effect on local microvascular parameters. Although the implantation of a conventional stainless steel sample led to a distinct and persistent activation of leukocytes combined with disruption of the microvascular endothelial integrity, marked leukocyte extravasation, and considerable venular dilation, animals with a nickel-reduced stainless steel implant showed only a moderate increase of these parameters, with a clear tendency of recuperation. Titanium implants merely caused a transient increase of leukocyte-endothelial cell interaction within the first 120 min, and no significant change in macromolecular leakage, leukocyte extravasation, or venular diameter. Pending biomechanical and corrosion testing, nickel-reduced stainless steel may be a viable alternative to conventional implant-quality stainless steel for biomedical applications. Concerning tolerance by the local vascular system, titanium currently remains unsurpassed.

  6. Striated muscle activator of Rho signalling (STARS) is reduced in ageing human skeletal muscle and targeted by miR-628-5p.

    PubMed

    Russell, A P; Wallace, M A; Kalanon, M; Zacharewicz, E; Della Gatta, P A; Garnham, A; Lamon, S

    2017-06-01

    The striated muscle activator of Rho signalling (STARS) is a muscle-specific actin-binding protein. The STARS signalling pathway is activated by resistance exercise and is anticipated to play a role in signal mechanotransduction. Animal studies have reported a negative regulation of STARS signalling with age, but such regulation has not been investigated in humans. Ten young (18-30 years) and 10 older (60-75 years) subjects completed an acute bout of resistance exercise. Gene and protein expression of members of the STARS signalling pathway and miRNA expression of a subset of miRNAs, predicted or known to target members of STARS signalling pathway, were measured in muscle biopsies collected pre-exercise and 2 h post-exercise. For the first time, we report a significant downregulation of the STARS protein in older subjects. However, there was no effect of age on the magnitude of STARS activation in response to an acute bout of exercise. Finally, we established that miR-628-5p, a miRNA regulated by age and exercise, binds to the STARS 3'UTR to directly downregulate its transcription. This study describes for the first time the resistance exercise-induced regulation of STARS signalling in skeletal muscle from older humans and identifies a new miRNA involved in the transcriptional control of STARS. © 2016 The Authors. Acta Physiologica published by John Wiley & Sons Ltd on behalf of Scandinavian Physiological Society.

  7. High-speed video imaging and digital analysis of microscopic features in contracting striated muscle cells

    NASA Astrophysics Data System (ADS)

    Roos, Kenneth P.; Taylor, Stuart R.

    1993-02-01

    The rapid motion of microscopic features such as the cross striations of single contracting muscle cells are difficult to capture with conventional optical microscopes, video systems, and image processing approaches. An integrated digital video imaging microscope system specifically designed to capture images from single contracting muscle cells at speeds of up to 240 Hz and to analyze images to extract features critical for the understanding of muscle contraction is described. This system consists of a brightfield microscope with immersion optics coupled to a high-speed charge-coupled device (CCD) video camera, super-VHS (S- VHS) and optical media disk video recording (OMDR) systems, and a semiautomated digital image analysis system. Components are modified to optimize spatial and temporal resolution to permit the evaluation of submicrometer features in real physiological time. This approach permits the critical evaluation of the magnitude, time course, and uniformity of contractile function throughout the volume of a single living cell with higher temporal and spatial resolutions than previously possible.

  8. Thyroid hormone regulates Ca(2+)-ATPase mRNA levels of sarcoplasmic reticulum during neonatal development of fast skeletal muscle.

    PubMed

    van der Linden, G C; Simonides, W S; van Hardeveld, C

    1992-12-01

    In gastrocnemius muscle from newborn rats the mRNA for the fast sarcoplasmic reticulum (SR) Ca(2+)-ATPase isoform (SERCA1) comprised over 90% of total SR Ca(2+)-ATPase mRNA content and increased 5-fold between day 5 and 20 after birth, whereas in hypothyroid muscle the SERCA1 message level remained constant. Triiodothyronine (T3) treatment of 2-day-old euthyroid rats induced a precocious stimulation of SERCA1 mRNA levels, indicating that T3 is the determining factor in the stimulation of SERCA1 message levels and that this stimulation underlies the previously reported effect of the thyroid status on the neonatal development of SR Ca(2+)-ATPase activity. The low mRNA level for the slow SR Ca(2+)-ATPase isoform (SERCA2) was constant in both euthyroid and hypothyroid muscle development. Nevertheless, T3 treatment of hypothyroid neonates induced a transient stimulation of SERCA2 message levels, indicating that SERCA2 is responsive to higher levels of T3.

  9. Sarcoplasmic reticulum Ca2+ depletion in adult skeletal muscle fibres measured with the biosensor D1ER.

    PubMed

    Jiménez-Moreno, Ramón; Wang, Zhong-Ming; Messi, María Laura; Delbono, Osvaldo

    2010-04-01

    The endoplasmic/sarcoplasmic reticulum (ER/SR) plays a crucial role in cytoplasmic signalling in a variety of cells. It is particularly relevant to skeletal muscle fibres, where this organelle constitutes the main Ca2+ store for essential functions, such as contraction. In this work, we expressed the cameleon biosensor D1ER by in vivo electroporation in the mouse flexor digitorum brevis (FDB) muscle to directly assess SR Ca2+ depletion in response to electrical and pharmacological stimulation. The main conclusions are: (1) D1ER is expressed in the SR of FDB fibres according to both di-8-(amino naphthyl ethenyl pyridinium) staining experiments and reductions in the Förster resonance energy transfer signal consequent to SR Ca2+ release; (2) the amplitude of D1ER citrine/cyan fluorescent protein (CFP) ratio evoked by either 4-chloro-m-cresol (4-CmC) or electrical stimulation is directly proportional to the basal citrine/CFP ratio, which indicates that SR Ca2+ modulates ryanodine-receptor-isoform-1-mediated SR Ca2+ release in the intact muscle fibre; (3) SR Ca2+ release, measured as D1ER citrine/CFP signal, is voltage-dependent and follows a Boltzmann function; and (4) average SR Ca2+ depletion is 20% in response to 4-CmC and 6.4% in response to prolonged sarcolemmal depolarization. These results indicate that significantly depleting SR Ca2+ content under physiological conditions is difficult.

  10. Specific protein-protein interactions of calsequestrin with junctional sarcoplasmic reticulum of skeletal muscle

    SciTech Connect

    Damiani, E.; Margreth, A. )

    1990-11-15

    Minor protein components of triads and of sarcoplasmic reticulum (SR) terminal cisternae (TC), i.e. 47 and 37 kDa peptides and 31-30 kDa and 26-25 kDa peptide doublets, were identified from their ability to bind {sup 125}I calsequestrin (CS) in the presence of EGTA. The CS-binding peptides are specifically associated with the junctional membrane of TC, since they could not be detected in junctional transverse tubules and in longitudinal SR fragments. The 31-30 kDa peptide doublet, exclusively, did not bind CS in the presence of Ca{sup 2+}. Thus, different types of protein-protein interactions appear to be involved in selective binding of CS to junctional TC.

  11. 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.

  12. 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.

  13. Ionic currents underlying fast action potentials in the obliquely striated muscle cells of the octopus arm.

    PubMed

    Rokni, Dan; Hochner, Binyamin

    2002-12-01

    The octopus arm provides a unique model for neuromuscular systems of flexible appendages. We previously reported the electrical compactness of the arm muscle cells and their rich excitable properties ranging from fast oscillations to overshooting action potentials. Here we characterize the voltage-activated ionic currents in the muscle cell membrane. We found three depolarization-activated ionic currents: 1) a high-voltage-activated L-type Ca(2+) current, which began activating at approximately -35 mV, was eliminated when Ca(2+) was substituted by Mg(2+), was blocked by nifedipine, and showed Ca(2+)-dependent inactivation. This current had very rapid activation kinetics (peaked within milliseconds) and slow inactivation kinetics (tau in the order of 50 ms). 2) A delayed rectifier K(+) current that was totally blocked by 10 mM TEA and partially blocked by 10 mM 4-aminopyridine (4AP). This current exhibited relatively slow activation kinetics (tau in the order of 15 ms) and inactivated only partially with a time constant of ~150 ms. And 3) a transient A-type K(+) current that was totally blocked by 10 mM 4AP and was partially blocked by 10 mM TEA. This current exhibited very fast activation kinetics (peaked within milliseconds) and inactivated with a time constant in the order of 60 ms. Inactivation of the A-type current was almost complete at -40 mV. No voltage-dependent Na(+) current was found in these cells. The octopus arm muscle cells generate fast (~3 ms) overshooting spikes in physiological conditions that are carried by a slowly inactivating L-type Ca(2+) current.

  14. 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.

  15. 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.

  16. 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.

  17. Mechanosensitive channels in striated muscle and the cardiovascular system: not quite a stretch anymore.

    PubMed

    Stiber, Jonathan A; Seth, Malini; Rosenberg, Paul B

    2009-08-01

    Stretch-activated or mechanosensitive channels transduce mechanical forces into ion fluxes across the cell membrane. These channels have been implicated in several aspects of cardiovascular physiology including regulation of blood pressure, vasoreactivity, and cardiac arrhythmias, as well as the adverse remodeling associated with cardiac hypertrophy and heart failure. This review discusses mechanosensitive channels in skeletal muscle and the cardiovascular system and their role in disease pathogenesis. We describe the regulation of gating of mechanosensitive channels including direct mechanisms and indirect activation by signaling pathways, as well as the influence on activation of these channels by the underlying cytoskeleton and scaffolding proteins. We then focus on the role of transient receptor potential channels, several of which have been implicated as mechanosensitive channels, in the pathogenesis of adverse cardiac remodeling and as potential therapeutic targets in the treatment of heart failure.

  18. In Vitro and In Vivo Single Myosin Step-Sizes in Striated Muscle a

    PubMed Central

    Burghardt, Thomas P.; Sun, Xiaojing; Wang, Yihua; Ajtai, Katalin

    2016-01-01

    Myosin in muscle transduces ATP free energy into the mechanical work of moving actin. It has a motor domain transducer containing ATP and actin binding sites, and, mechanical elements coupling motor impulse to the myosin filament backbone providing transduction/mechanical-coupling. The mechanical coupler is a lever-arm stabilized by bound essential and regulatory light chains. The lever-arm rotates cyclically to impel bound filamentous actin. Linear actin displacement due to lever-arm rotation is the myosin step-size. A high-throughput quantum dot labeled actin in vitro motility assay (Qdot assay) measures motor step-size in the context of an ensemble of actomyosin interactions. The ensemble context imposes a constant velocity constraint for myosins interacting with one actin filament. In a cardiac myosin producing multiple step-sizes, a “second characterization” is step-frequency that adjusts longer step-size to lower frequency maintaining a linear actin velocity identical to that from a shorter step-size and higher frequency actomyosin cycle. The step-frequency characteristic involves and integrates myosin enzyme kinetics, mechanical strain, and other ensemble affected characteristics. The high-throughput Qdot assay suits a new paradigm calling for wide surveillance of the vast number of disease or aging relevant myosin isoforms that contrasts with the alternative model calling for exhaustive research on a tiny subset myosin forms. The zebrafish embryo assay (Z assay) performs single myosin step-size and step-frequency assaying in vivo combining single myosin mechanical and whole muscle physiological characterizations in one model organism. The Qdot and Z assays cover “bottom-up” and “top-down” assaying of myosin characteristics. PMID:26728749

  19. Human recombinant erythropoietin protects the striated muscle microcirculation of the dorsal skinfold from postischemic injury in mice.

    PubMed

    Contaldo, Claudio; Meier, Christoph; Elsherbiny, Ahmed; Harder, Yves; Trentz, Otmar; Menger, Michael D; Wanner, Guido A

    2007-07-01

    Erythropoietin (EPO) has been proposed as a novel cytoprotectant in ischemia-reperfusion (I/R) injury of the brain, heart, and kidney. However, whether EPO exerts its protection by prevention of postischemic microcirculatory deterioration is unknown. We have investigated the effect of EPO on I/R-induced microcirculatory dysfunctions. We used the mouse dorsal skinfold chamber preparation to study nutritive microcirculation and leukocyte-endothelial cell interaction in striated muscle of the dorsal skinfold by in vivo fluorescence microscopy before 3 h of ischemia and during 5 days of reperfusion. Animals were pretreated with EPO (5,000 U/kg body wt) 1 or 24 h before ischemia. Vehicle-treated I/R-injured animals served as controls. Additional animals underwent sham operation only or were pretreated with EPO but not subjected to I/R. I/R significantly (P < 0.05) reduced functional capillary density, increased microvascular permeability, and enhanced venular leukocyte-endothelial cell interaction during early reperfusion. These findings were associated with pronounced (P < 0.05) arteriolar constriction and diminution of blood flow during late reperfusion. Pretreatment with EPO induced EPO receptor and endothelial nitric oxide synthase expression at 6 h of reperfusion (P < 0.05). In parallel, EPO significantly (P < 0.05) reduced capillary perfusion failure and microvascular hyperpermeability during early reperfusion and arteriolar constriction and flow during late reperfusion. EPO pretreatment substantially (P < 0.05) diminished I/R-induced leukocytic inflammation by reducing the number of rolling and firmly adhering leukocytes in postcapillary venules. EPO applied 1 h before ischemia induced angiogenic budding and sprouting at 1 and 3 days of reperfusion and formation of new capillary networks at 5 days of reperfusion. Thus our study demonstrates for the first time that EPO effectively attenuates I/R injury by preserving nutritive perfusion, reducing leukocytic

  20. Structure and interactions of the carboxyl terminus of striated muscle alpha-tropomyosin: it is important to be flexible.

    PubMed Central

    Greenfield, Norma J; Palm, Thomas; Hitchcock-DeGregori, Sarah E

    2002-01-01

    Tropomyosin (TM) binds to and regulates the actin filament. We used circular dichroism and heteronuclear NMR to investigate the secondary structure and interactions of the C terminus of striated muscle alpha-TM, a major functional determinant, using a model peptide, TM9a(251-284). The (1)H(alpha) and (13)C(alpha) chemical shift displacements show that residues 252 to 277 are alpha-helical but residues 278 to 284 are nonhelical and mobile. The (1)H(N) and (13)C' displacements suggest that residues 257 to 269 form a coiled coil. Formation of an "overlap" binary complex with a 33-residue N-terminal chimeric peptide containing residues 1 to 14 of alpha-TM perturbs the (1)H(N) and (15)N resonances of residues 274 to 284. Addition of a fragment of troponin T, TnT(70-170), to the binary complex perturbs most of the (1)H(N)-(15)N cross-peaks. In addition, there are many new cross-peaks, showing that the binding is asymmetric. Q263, in a proposed troponin T binding site, shows two sets of side-chain (15)N-(1)H cross-peaks, indicating conformational flexibility. The conformational equilibrium of the side chain changes upon formation of the binary and ternary complexes. Replacing Q263 with leucine greatly increases the stability of TM9a(251-284) and reduces its ability to form the binary and ternary complexes, showing that conformational flexibility is crucial for the binding functions of the C terminus. PMID:12414708

  1. Skeletal Muscle Myofibrillar and Sarcoplasmic Protein Synthesis Rates Are Affected Differently by Altitude-Induced Hypoxia in Native Lowlanders

    PubMed Central

    Holm, Lars; Haslund, Mads Lyhne; Robach, Paul; van Hall, Gerrit; Calbet, Jose A. L.; Saltin, Bengt; Lundby, Carsten

    2010-01-01

    As a consequence to hypobaric hypoxic exposure skeletal muscle atrophy is often reported. The underlying mechanism has been suggested to involve a decrease in protein synthesis in order to conserve O2. With the aim to challenge this hypothesis, we applied a primed, constant infusion of 1-13C-leucine in nine healthy male subjects at sea level and subsequently at high-altitude (4559 m) after 7–9 days of acclimatization. Physical activity levels and food and energy intake were controlled prior to the two experimental conditions with the aim to standardize these confounding factors. Blood samples and expired breath samples were collected hourly during the 4 hour trial and vastus lateralis muscle biopsies obtained at 1 and 4 hours after tracer priming in the overnight fasted state. Myofibrillar protein synthesis rate was doubled; 0.041±0.018 at sea-level to 0.080±0.018%⋅hr−1 (p<0.05) when acclimatized to high altitude. The sarcoplasmic protein synthesis rate was in contrast unaffected by altitude exposure; 0.052±0.019 at sea-level to 0.059±0.010%⋅hr−1 (p>0.05). Trends to increments in whole body protein kinetics were seen: Degradation rate elevated from 2.51±0.21 at sea level to 2.73±0.13 µmol⋅kg−1⋅min−1 (p = 0.05) at high altitude and synthesis rate similar; 2.24±0.20 at sea level and 2.43±0.13 µmol⋅kg−1⋅min−1 (p>0.05) at altitude. We conclude that whole body amino acid flux is increased due to an elevated protein turnover rate. Resting skeletal muscle myocontractile protein synthesis rate was concomitantly elevated by high-altitude induced hypoxia, whereas the sarcoplasmic protein synthesis rate was unaffected by hypoxia. These changed responses may lead to divergent adaptation over the course of prolonged exposure. PMID:21187972

  2. Release and recycling of calcium by the sarcoplasmic reticulum in guinea-pig portal vein smooth muscle.

    PubMed Central

    Bond, M; Kitazawa, T; Somlyo, A P; Somlyo, A V

    1984-01-01

    The amplitude of interrupted contractions evoked by noradrenaline or caffeine in Ca2+-free, high-K+ solutions containing EGTA or La3+ was determined in small (40-60 micron thick) bundles of guinea-pig portal anterior mesenteric vein. Interrupted contractions were produced by removing the stimulating agent as soon as the amplitude of the tension record reached its peak. The distribution of intracellular Ca2+ was determined, with electron probe X-ray microanalysis, in cryosections of preparations frozen in the relaxed state and at the peak of noradrenaline-induced contractions. Interrupted contractions of maximal or near-maximal amplitudes could be evoked every 2 min for up to 15 min in the virtual absence of extracellular Ca2+. If noradrenaline was allowed to remain in the solution throughout the period of spontaneous relaxation, a subsequent contraction could no longer be evoked in the absence of extracellular Ca2+. Interrupted contractions, similar to those evoked by noradrenaline, could also be stimulated by caffeine. The amplitude of reproducible interrupted contractions in Ca2+-free, high-K+ solution was graded with noradrenaline concentration. The ability of these smooth muscles to contract repeatedly and maximally in Ca2+-free solutions indicates the recycling of Ca2+ released from an intracellular store. The occurrence of these contractions in high-K+ (depolarizing) solutions supports the conclusion (Devine, Somlyo & Somlyo, 1972) that the release of intracellular Ca2+ is one of the mechanisms of pharmacomechanical coupling. The number of subplasmalemmal regions in which high Ca concentrations (greater than 10 mmol/kg dry wt.) were detected, with approximately 75 nm diameter electron probes, was reduced in muscles frozen at the peak of contraction, from 4.7/cell periphery in the relaxed to 1.4/cell periphery in the contracted preparations. In freeze-substituted smooth muscles, in which the membranes of the junctional sarcoplasmic reticulum could be

  3. Expression of eight distinct MHC isoforms in bovine striated muscles: evidence for MHC-2B presence only in extraocular muscles.

    PubMed

    Toniolo, L; Maccatrozzo, L; Patruno, M; Caliaro, F; Mascarello, F; Reggiani, C

    2005-11-01

    This study aimed to analyse the expression of myosin heavy chain (MHC) isoforms in bovine muscles, with particular attention to the MHC-2B gene. Diaphragm, longissimus dorsi, masseter, several laryngeal muscles and two extraocular muscles (rectus lateralis and retractor bulbi) were sampled in adult male Bos taurus (age 18-24 months, mass 400-500 kg) and analysed by RT-PCR, gel electrophoresis and immunohistochemistry. Transcripts and proteins corresponding to eight MHC isoforms were identified: MHC-alpha and MHC-beta/slow (or MHC-1), two developmental isoforms (MHC-embryonic and MHC-neonatal), three adult fast isoforms (MHC-2A, MHC-2X and MHC-2B) and the extraocular isoform MHC-Eo. All eight MHC isoforms were found to be co-expressed in extrinsic eye muscles, retractor bulbi and rectus lateralis, four (beta/slow, 2A, 2X, neonatal) in laryngeal muscles, three (beta/slow, 2A and 2X) in trunk and limb muscles and two (beta/slow and alpha) in masseter. The expression of MHC-2B and MHC-Eo was restricted to extraocular muscles. Developmental MHC isoforms (neonatal and embryonic) were only found in specialized muscles in the larynx and in the eye. MHC-alpha was only found in extraocular and masseter muscle. Single fibres dissected from masseter, diaphragm and longissimus were classified into five groups (expressing, respectively, beta/slow, alpha, slow and 2A, 2A and 2X) on the basis of MHC isoform electrophoretical separation, and their contractile properties [maximum shortening velocity (v(0)) and isometric tension (P(0))] were determined. v(0) increased progressively from slow to fast 2A and fast 2X, whereas hybrid 1-2A fibres and fibres containing MHC-alpha were intermediate between slow and fast 2A.

  4. 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.

  5. A novel isoform of delta-sarcoglycan is localized at the sarcoplasmic reticulum of mouse skeletal muscle

    PubMed Central

    Estrada, Francisco J.; Mornet, Dominique; Rosas-Vargas, Haydeé; Angulo, Alexandra; Hernández, Manuel; Becker, Viola; Rendón, Alvaro; Ramos-Kuri, Manuel; Coral-Vázquez, Ramón M.

    2006-01-01

    The sarcoglycan-sarcospan complex (α-, β-, χ-, δ-, ε-, and ζ-SG-SSPN), a component of the dystrophin-associated glycoprotein complex (DAGC), is located at the sarcolemma of muscle fibers where it contributes to maintain cell integrity during contraction-relaxation cycles; χ-and δ-SG are also expressed in the sarcoplasmic reticulum (SR). In this study, we report the identification of a novel isoform of murine δ-SG produced by alternative splicing that we named δ-SG3. This isoform is present at transcript level in several tissues, with its highest expression in skeletal and cardiac muscle. The δ-SG3 protein lacks the last 122 amino acids at the C-terminal, which are replaced by 10 new amino acids (EGFLNMQLAG). Interestingly, double immunofluorescence analysis for δ-SG3 and the dihydropyridine receptor (DHPR) shows a close localization of these two proteins. We propose the subcellular distribution of this novel δ-SG3 isoform at the SR and its involvement in intracellular calcium concentration regulation. PMID:16403451

  6. Peeled mammalian skeletal muscle fibers. Possible stimulation of Ca2+ release via a transverse tubule-sarcoplasmic reticulum mechanism

    PubMed Central

    1985-01-01

    Single muscle fibers from rabbit soleus and adductor magnus and from semitendinosus muscles were peeled to remove the sarcolemma and then stimulated to release Ca2+ by (a) caffeine application or (b) ionic depolarization accomplished via substitution of choline chloride for potassium propionate at constant [K+] X [Cl-] in the bathing solution. Each stimulus, ionic or caffeine, elicited an isometric tension transient that appeared to be due to Ca2+ released from the sarcoplasmic reticulum (SR). The peak magnitude of the ionic (Cl- - induced) tension transient increased with increasing Cl- concentration. The application of ouabain to fibers after peeling had no effect on either type of tension transient. However, soaking the fibers in a ouabain solution before peeling blocked the Cl- -induced but not the caffeine-induced tension transient, which suggests that ouabain's site of action is extracellular, perhaps inside transverse tubules (TTs). Treating the peeled fibers with saponin, which should disrupt TTs to a greater extent than SR membrane, greatly reduced or eliminated the Cl- - induced tension transient without significantly altering the caffeine- induced tension transient. These results suggest that the Cl- -induced tension transient is elicited via stimulation of sealed, polarized TTs rather than via ionic depolarization of the SR. PMID:4056734

  7. Sensory and autonomic neurons project both to the smooth retractor penis and to the striated bulbospongiosus muscles. Neurochemical features of the sympathetic subset.

    PubMed

    Botti, Maddalena; Gazza, Ferdinando; Ragionieri, Luisa; Minelli, Luisa Bo; Panu, Rino

    2012-08-01

    Aim of the present study was to verify, by means of double retrograde neuronal tracers technique, the hypothesis that a subpopulation of sensory and autonomic neurons send collateral axons to both smooth and striated genital muscles. We also wanted to define the neurochemical content of the eventually retrogradelly double labeled (RDL) neurons in the sympathetic trunk ganglia (STG). We used six intact pigs and we injected the tracer Diamidino Yellow (DY) in the smooth left retractor penis muscle (RPM) and the tracer Fast Blue (FB) in the striated left bulbospongiosus muscle (BSM). Rare (2 ± 0.6) RDL neurons were found in the ipsilateral S2 spinal ganglion (SG), 220 ± 42 in the ipsilateral STGs, from L3 to S3, 19 ± 15 in the contralateral S1-S2 ones and 22 ± 5 in the bilateral caudal mesenteric ganglia (CMG). The RDL neurons of the STG were IR for TH (85 ± 13%), DβH (69 ± 17%), NPY (69 ± 23%), nNOS (60 ± 11%), LENK (54 ± 19%), VIP (53±26%), SOM (40 ± 8%), CGRP (34 ± 12%), SP (31 ± 16%), and VAChT (28 ± 3%). Our research highlights the presence of sensory and sympathetic neurons with qualitatively different neurochemical content sending axons both to the smooth RPM and to the striated BSM of the pig. These RDL neurons are likely to project to the smooth vasal musculature to create the ideal physiological conditions in which these muscles can optimize the erectile function. Copyright © 2012 Wiley Periodicals, Inc.

  8. Role of Ca(2+) in the rapid cooling-induced Ca(2+) release from sarcoplasmic reticulum in ferret cardiac muscles.

    PubMed

    Tanaka, Etsuko; Konishi, Masato; Kurihara, Satoshi

    2012-05-01

    Rapid lowering of the solution temperature (rapid cooling, RC) from 24 to 3°C within 3 s releases considerable amounts of Ca(2+) from the sarcoplasmic reticulum (SR) in mammalian cardiac muscles. In this study, we investigated the intracellular mechanism of RC-induced Ca(2+) release, especially the role of Ca(2+), in ferret ventricular muscle. Saponin-treated skinned trabeculae were placed in a glass capillary, and the amount of Ca(2+) released from the SR by RC and caffeine (50 mM) was measured with fluo-3. It was estimated that in the presence of ATP about 45% of the Ca(2+) content in the SR was released by RC. The amount of SR Ca(2+) released by RC was unchanged by the replacement of ATP by AMP-PCP (a non-hydrolysable ATP analogue and agonist for the ryanodine receptor but not for the Ca(2+) pump of SR), suggesting that the suppression of the Ca(2+) pump of SR at low temperature might not be a major mechanism in RC-induced Ca(2+) release. The free Ca(2+) concentration of the solution used for triggering RC-induced Ca(2+) release was estimated to be only about 20 nM with fluo-3 or aequorin. When this solution was applied to the preparation at 3°C, only a small amount of Ca(2+) was released from SR presumably by the Ca(2+)-induced Ca(2+) release (CICR) mechanism. Thus, in mammalian cardiac muscles, RC releases a part of the (<50%) stored Ca(2+) contained in the SR, and the mechanism of RC-induced Ca(2+) release may differ from that of CICR, which is thought to play a role in frog skeletal muscle fibres that express ryanodine receptors of different types.

  9. Proteomic study of muscle sarcoplasmic proteins using AUT-PAGE/SDS-PAGE as two-dimensional gel electrophoresis.

    PubMed

    Picariello, Gianluca; De Martino, Alessandra; Mamone, Gianfranco; Ferranti, Pasquale; Addeo, Francesco; Faccia, Michele; Spagnamusso, Salvatore; Di Luccia, Aldo

    2006-03-20

    In the present study, an alternative procedure for two-dimensional (2D) electrophoretic analysis in proteomic investigation of the most represented basic muscle water-soluble proteins is suggested. Our method consists of Acetic acid-Urea-Triton polyacrylamide gel (AUT-PAGE) analysis in the first dimension and standard sodium dodecyl sulphate polyacrylamide gel (SDS-PAGE) in the second dimension. Although standard two-dimensional Immobilized pH Gradient-Sodium Dodecyl-Sulphate (2D IPG-SDS) gel electrophoresis has been successfully used to study these proteins, most of the water-soluble proteins are spread on the alkaline part of the 2D map and are poorly focused. Furthermore, the similarity in their molecular weights impairs resolution of the classical approach. The addition of Triton X-100, a non-ionic detergent, into the gel induces a differential electrophoretic mobility of proteins as a result of the formation of mixed micelles between the detergent and the hydrophobic moieties of polypeptides, separating basic proteins with a criterion similar to reversed phase chromatography based on their hydrophobicity. The acid pH induces positive net charges, increasing with the isoelectric point of proteins, thus allowing enhanced resolution in the separation. By using 2D AUT-PAGE/SDS electrophoresis approach to separate water-soluble proteins from fresh pork and from dry-cured products, we could spread proteins over a greater area, achieving a greater resolution than that obtained by IPG in the pH range 3-10 and 6-11. Sarcoplasmic proteins undergoing proteolysis during the ripening of products were identified by Matrix Assisted Laser Desorption/Ionization-Time of Flight (MALDI-ToF) mass spectrometry peptide mass fingerprinting in a easier and more effective way. Two-dimensional AUT-PAGE/SDS electrophoresis has allowed to simplify separation of sarcoplasmic protein mixtures making this technique suitable in the defining of quality of dry-cured pork products by immediate

  10. Inhibition of sarcoplasmic Ca2+-ATPase increases caffeine- and halothane-induced contractures in muscle bundles of malignant hyperthermia susceptible and healthy individuals

    PubMed Central

    Schuster, Frank; Müller, Rainer; Hartung, Edmund; Roewer, Norbert; Anetseder, Martin

    2005-01-01

    Background Malignant hyperthermia (MH) is triggered by halogenated anaesthetics and depolarising muscle relaxants, leading to an uncontrolled hypermetabolic state of skeletal muscle. An uncontrolled sarcoplasmic Ca2+ release is mediated via the ryanodine receptor. A compensatory mechanism of increased sarcoplasmic Ca2+-ATPase activity was described in pigs and in transfected cell lines. We hypothesized that inhibition of Ca2+ reuptake via the sarcoplasmic Ca2+-ATPase (SERCA) enhances halothane- and caffeine-induced muscle contractures in MH susceptible more than in non-susceptible skeletal muscle. Methods With informed consent, surplus muscle bundles of 7 MHS (susceptible), 7 MHE (equivocal) and 16 MHN (non-susceptible) classified patients were mounted to an isometric force transducer, electrically stimulated, preloaded and equilibrated. Following 15 min incubation with cyclopiazonic acid (CPA) 25 μM, the European MH standard in-vitro-contracture test protocol with caffeine (0.5; 1; 1.5; 2; 3; 4 mM) and halothane (0.11; 0.22; 0.44; 0.66 mM) was performed. Data as median and quartiles; Friedman- and Wilcoxon-test for differences with and without CPA; p < 0.05. Results Initial length, weight, maximum twitch height, predrug resting tension and predrug twitch height of muscle bundles did not differ between groups. CPA increased halothane- and caffeine-induced contractures significantly. This increase was more pronounced in MHS and MHE than in MHN muscle bundles. Conclusion Inhibition of the SERCA activity by CPA enhances halothane- and caffeine-induced contractures especially in MHS and MHE skeletal muscle and may help for the diagnostic assignment of MH susceptibility. The status of SERCA activity may play a significant but so far unknown role in the genesis of malignant hyperthermia. PMID:15946384

  11. Potentiation of sarcoplasmic reticulum Ca2+ release by 2,3-butanedione monoxime in crustacean muscle.

    PubMed

    Györke, S; Dettbarn, C; Palade, P

    1993-06-01

    The effect of the chemical phosphatase 2,3-butanedione monoxime (BDM) on various aspects of excitation/contraction coupling in crustacean muscle was investigated. Despite having a depressant effect on vertebrate skeletal and cardiac muscle, BDM was a potentiator of contraction in crustacean muscle. At concentrations of 1-3 mM BDM caused an increase of potassium contractures in bundles of fibers isolated from crayfish muscle. At higher concentrations BDM caused oscillatory contractions by itself. In single voltage-clamped cut muscle fibers loaded with rhod-2, BDM (0.5-2 mM) potentiated the magnitude and duration of intracellular Ca2+ transients elicited by depolarization. At the same time BDM did not affect the rate of Ca2+ removal from the myoplasm under conditions where Ca2+ release was blocked by tetracaine. Nor did BDM increase Ca2+ entry; in fact it caused a decrease in the amplitude of the inward Ca2+ current (ICa). In microsomes isolated from lobster muscle, BDM also potentiated Ca2+ release induced by caffeine and at higher concentrations (above 3 mM) induced release by itself. At the same time it had little effect on Ca2+ uptake. These results indicate that BDM potentiates Ca2+ release in crustacean muscle possibly by dephosphorylation of the Ca(2+)-release channel.

  12. Primary structure of myosin from the striated adductor muscle of the Atlantic scallop, Pecten maximus, and expression of the regulatory domain.

    PubMed

    Janes, D P; Patel, H; Chantler, P D

    2000-01-01

    We have determined the complete cDNA and deduced amino acid sequences of the heavy chain, regulatory light chain and essential light chain which constitute the molecular structure of myosin from the striated adductor muscle of the scallop, Pecten maximus. The deduced amino acid sequences of P. maximus regulatory light chain, essential light chain and heavy chain comprise 156, 156 and 1940 amino acids, respectively. These myosin peptide sequences, obtained from the most common of the eastern Atlantic scallops, are compared with those from three other molluscan myosins: the striated adductor muscles of Argopecten irradians and Placopecten magellanicus, and myosin from the siphon retractor muscle of the squid, Loligo pealei. The Pecten heavy chain sequence resembles those of the other two scallop sequences to a much greater extent as compared with the squid sequence, amino acid identities being 97.5% (A. irradians), 95.6% (P. magellanicus) and 73.6% (L. pealei), respectively. Myosin heavy chain residues that are known to be important for regulation are conserved in Pecten maximus. Using these Pecten sequences, we have overexpressed the regulatory light chain, and a combination of essential light chain and myosin heavy chain fragment, separately, in E. coli BL21 (DE3) prior to recombination, thereby producing Pecten regulatory domains without recourse to proteolytic digestion. The expressed regulatory domain was shown to undergo a calcium-dependent increase (approximately 7%) in intrinsic tryptophan fluorescence with a mid-point at a pCa of 6.6.

  13. Ca2+-releasing effect of cerivastatin on the sarcoplasmic reticulum of mouse and rat skeletal muscle fibers.

    PubMed

    Inoue, Ryotaku; Tanabe, Mitsuo; Kono, Keita; Maruyama, Kei; Ikemoto, Takaaki; Endo, Makoto

    2003-11-01

    We analyzed the effect of HMG-CoA reductase inhibitors on Ca(2+) release from the sarcoplasmic reticulum (SR) using chemically skinned skeletal muscle fibers from the mouse and the rat. Cerivastatin (>20 microM) released Ca(2+) from the SR, while pravastatin showed only a little effect. The rates of Ca(2+) release were increased by cerivastatin at all Ca(2+) concentrations tested. Cerivastatin-induced Ca(2+) release in the presence of Ca(2+) was affected by adenosine monophosphate, Mg(2+), and procaine in essentially the same way as for caffeine-induced Ca(2+) release. The Ca(2+)-uptake capacity of the SR was reduced after co-treatment with ryanodine and cerivastatin at pCa 6.0 to a much greater extent than with ryanodine alone. Thus, cerivastatin-induced Ca(2+) release in the presence of Ca(2+) must be a result of the activation of the Ca(2+)-induced Ca(2+) release (CICR) mechanism of the ryanodine receptor. However, even when CICR was maximally inhibited by Mg(2+) and procaine, or in the practical absence of Ca(2+) (pCa >8), cerivastatin still caused Ca(2+) release. These results indicate that cerivastatin causes Ca(2+) release also by activating some other mechanism(s) in addition to the activation of CICR. Either or both of these effects might be related to its adverse effect, rhabdomyolysis.

  14. Phosphorylation of anchoring protein by calmodulin protein kinase associated to the sarcoplasmic reticulum of rabbit fast-twitch muscle.

    PubMed

    Damiani, E; Sacchetto, R; Margreth, A

    2000-12-09

    Regulatory phosphorylation of phospholamban and of SR Ca(2+)-ATPase SERCA2a isoform by endogenous CaM-K II in slow-twitch skeletal and cardiac sarcoplasmic reticulum (SR) is well documented, but much less is known of the exact functional role of CaM K II in fast-twitch muscle SR. Recently, it was shown that RNA splicing of brain-specific alpha CaM K II, gives rise to a truncated protein (alpha KAP), consisting mainly of the association domain, serving to anchor CaM K II to SR membrane in rat skeletal muscle [Bayer, K.-U., et al. (1998) EMBO J. 19, 5598-5605]. In the present study, we searched for the presence of alpha KAP in sucrose-density purified SR membrane fractions from representative fast-twitch and slow-twitch limb muscles, both of the rabbit and the rat, using immunoblot techniques and antibody directed against the association domain of alpha CaM K II. Putative alpha KAP was immunodetected as a 23-kDa electrophoretic component on SDS-PAGE of the isolated SR from fast-twitch but not from slow-twitch muscle, and was further identified as a specific substrate of endogenous CaM K II, in the rabbit. Immunodetected, (32)P-labeled, non-calmodulin binding protein, behaved as a single 23-kDa protein species under several electrophoretic conditions. The 23-kDa protein, with defined properties, was isolated as a complex with 60-kDa delta CaM K II isoform, by sucrose-density sedimentation analysis. Moreover, we show here that putative alphaKAP, in spite of its inability to bind CaM in ligand blot overlay, co-eluted with delta CaM K II from CaM-affinity columns. That raises the question of whether CaM K II-mediated phosphorylation of alpha KAP and triadin together might be involved in a molecular signaling pathway important for SR Ca(2+)-release in fast-twitch muscle SR.

  15. 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.

  16. 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.

  17. Effect of taurine on sarcoplasmic reticulum function and force in skinned fast-twitch skeletal muscle fibres of the rat.

    PubMed

    Bakker, Anthony J; Berg, Helen M

    2002-01-01

    We examined the effect of taurine on depolarisation-induced force responses and sarcoplasmic reticulum (SR) function in mechanically skinned skeletal muscle fibres from the extensor digitorum longus (EDL) of the rat. Taurine (20 mM) produced a small but significant (P < 0.01) decrease in the sensitivity of the contractile apparatus to Ca(2+) (increase in the [Ca(2+)] corresponding to 50 % of maximum force of about 7 %; n = 10) and in maximum force (92.0 +/- 1.0 % of controls) in the skinned fibres. Taurine had no statistically significant effect on the slope of the force-pCa curve. Depolarisation-induced force responses in the skinned fibres were markedly increased in peak value by 20 mM taurine, to 120.8 +/- 5.3 % of control measurements (P = 0.0006, n = 27). Taurine (20 mM) significantly increased the SR Ca(2+) accumulation in the skinned fibres by 34.6 +/- 9.3 % compared to control conditions (measured by comparing the integral of caffeine contractures in fibres previously loaded with Ca(2+) in the absence or presence of taurine; P = 0.0014, n = 10). Taurine (20 mM) also increased both the peak and rate of rise of caffeine-induced force responses in the fibres by 29.2 +/- 9.7 % (P = 0.0298, n = 6) and 27.6 +/- 8.9 % (P = 0.037), respectively, compared with controls. This study shows that taurine is a modulator of contractile function in mammalian skeletal muscle. Taurine may increase the size of depolarisation-induced force responses by augmenting SR Ca(2+) accumulation and release.

  18. 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

  19. Effect of taurine on sarcoplasmic reticulum function and force in skinned fast-twitch skeletal muscle fibres of the rat

    PubMed Central

    Bakker, Anthony J; Berg, Helen M

    2002-01-01

    We examined the effect of taurine on depolarisation-induced force responses and sarcoplasmic reticulum (SR) function in mechanically skinned skeletal muscle fibres from the extensor digitorum longus (EDL) of the rat. Taurine (20 mm) produced a small but significant (P < 0.01) decrease in the sensitivity of the contractile apparatus to Ca2+ (increase in the [Ca2+] corresponding to 50 % of maximum force of about 7 %; n = 10) and in maximum force (92.0 ± 1.0 % of controls) in the skinned fibres. Taurine had no statistically significant effect on the slope of the force-pCa curve. Depolarisation-induced force responses in the skinned fibres were markedly increased in peak value by 20 mm taurine, to 120.8 ± 5.3 % of control measurements (P = 0.0006, n = 27). Taurine (20 mm) significantly increased the SR Ca2+ accumulation in the skinned fibres by 34.6 ± 9.3 % compared to control conditions (measured by comparing the integral of caffeine contractures in fibres previously loaded with Ca2+ in the absence or presence of taurine; P = 0.0014, n = 10). Taurine (20 mm) also increased both the peak and rate of rise of caffeine-induced force responses in the fibres by 29.2 ± 9.7 % (P = 0.0298, n = 6) and 27.6 ± 8.9 % (P = 0.037), respectively, compared with controls. This study shows that taurine is a modulator of contractile function in mammalian skeletal muscle. Taurine may increase the size of depolarisation-induced force responses by augmenting SR Ca2+ accumulation and release. PMID:11773327

  20. Histological distinction of mechanical and thermal defects produced by nanosecond laser pulses in striated muscle at 1064 nm

    NASA Astrophysics Data System (ADS)

    Gratzl, Thomas; Dohr, Gottfried; Schmidt-Kloiber, Heinz; Reichel, Erich

    1991-06-01

    For the therapeutic application of laser light it is necessary to minimize defects in the non-irradiated tissue. These defects depend on the primary mechanism of interaction which is determined by the duration of laser action. In the case of continuous wave laserlight a tissue layer surrounding the irradiated volume is thermally affected. On using laser pulsed with a certain energy this layer becomes smaller with decreasing pulse duration. With the pulses of a Q-switched laser tissue cutting will be obtained by the laser-induced breakdown (LIB). Thereby shockwaves are emitted which stress the tissue mechanically. Even in this case thermal lesions can be found. To be able to distinguish between thermal and mechanical effects by histological examination, experiments were performed with ns- and microsecond(s) -laserpulses under the same conditions. A Nd:YAG-laser at 1064 nm was used either Q-switched (pulse duration: 8 ns) or flashlamp-pulsed (100 microsecond(s) ) with a pulse repetition rate of 10 Hz. The beam was focused through air below the tissue surface (focal length in air: 80 mm). The beam geometry in the focal region was identical for both cases. The position of the focal plane relative to the surface was exactly controlled, as it influences extension and kind of the defect. To produce evaluable defects in the microsecond(s) experiments 200 laserpulses with an energy of 340 mJ per pulse had to be applied. The unfixed striated muscle samples of Sprague Dawley rats were immediately dissected prior to laser exposure. For the microsecond(s) experiments the defect region could be divided into 4 zones surrounding a crater, which was found at a focal plane position 2 mm below the surface. Zone 1 shows vacuoles and intensive staining. In zone 2 the myofibrils were displaced and torn apart. Zone 3 represents a sharply bordered intensively stained region. In zone 4 muscle cells are contracted. The zones are all of thermal origin, which could be derived from experiments

  1. Effect of 23-day muscle disuse on sarcoplasmic reticulum Ca2+ properties and contractility in human type I and type II skeletal muscle fibers.

    PubMed

    Lamboley, C R; Wyckelsma, V L; Perry, B D; McKenna, M J; Lamb, G D

    2016-08-01

    Inactivity negatively impacts on skeletal muscle function mainly through muscle atrophy. However, recent evidence suggests that the quality of individual muscle fibers is also altered. This study examined the effects of 23 days of unilateral lower limb suspension (ULLS) on specific force and sarcoplasmic reticulum (SR) Ca(2+) content in individual skinned muscle fibers. Muscle biopsies of the vastus lateralis were taken from six young healthy adults prior to and following ULLS. After disuse, the endogenous SR Ca(2+) content was ∼8% lower in type I fibers and maximal SR Ca(2+) capacity was lower in both type I and type II fibers (-11 and -5%, respectively). The specific force, measured in single skinned fibers from three subjects, decreased significantly after ULLS in type II fibers (-23%) but not in type I fibers (-9%). Western blot analyses showed no significant change in the amounts of myosin heavy chain (MHC) I and MHC IIa following the disuse, whereas the amounts of sarco(endo)plasmic reticulum Ca(2+)-ATPase 1 (SERCA1) and calsequestrin increased by ∼120 and ∼20%, respectively, and the amount of troponin I decreased by ∼21%. These findings suggest that the decline in force and power occurring with muscle disuse is likely to be exacerbated in part by reductions in maximum specific force in type II fibers, and in the amount of releasable SR Ca(2+) in both fiber types, the latter not being attributable to a reduced calsequestrin level. Furthermore, the ∼3-wk disuse in human elicits change in SR properties, in particular a more than twofold upregulation in SERCA1 density, before any fiber-type shift.

  2. 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

  3. COORDINATED DEVELOPMENT OF THE SARCOPLASMIC RETICULUM AND T SYSTEM DURING POSTNATAL DIFFERENTIATION OF RAT SKELETAL MUSCLE

    PubMed Central

    Schiaffino, S.; Margreth, A.

    1969-01-01

    An electron microscope study has been carried out on rat psoas muscle, during the early postnatal stages of development. Among the several subcellular components, the sarcotubular system undergoes the most striking modifications during this period. In muscle fibers of the newborn rat, junctional contacts between the T system and the SR are sparse and are, mostly, longitudinally or obliquely oriented. The T tubules do not penetrate deeply into the muscle cell, as indicated by the predominantly peripheral location of the triads and the persistence, at these stages of development, of a highly branched subsarcolemmal system of tubules. Diadic associations of junctional SR elements with the plasma membrane are also occasionally observed. The early SR elaborations incompletely delineate the myofibrils, at both the A- and I-band level. Longitudinal sections show irregularly oriented SR tubules, running continuously over successive sarcomeres. Flattened junctional cisterns filled with granular material are sparse and laterally interconnected, at circumscribed sites, with the SR tubules. Between 1 and 2 wk postpartum, transversal triadic contacts are extensively established, at the A-I band level, and the SR network differentiates into two portions in register with the A and I band, respectively. At 10–15 days after birth, the SR provides a transversely continuous double sheet around the myofibrils at the I-band level, whereas it forms a single discontinuous layer at the A-band level. The relationship that these morphological modifications of the sarcotubular system may bear to previously described biochemical and physiological changes of rat muscle fibers after birth is discussed. PMID:5814005

  4. Striated perineal muscles: location of autonomic, sensory, and somatic neurons projecting to the male pig bulbospongiosus muscle.

    PubMed

    Botti, Maddalena; Ragionieri, Luisa; Gazza, Ferdinando; Acone, Franca; Bo Minelli, Luisa; Panu, Rino

    2009-11-01

    The location, number, and size of the neurons innervating the bulbospongiosus muscle (BSM) were studied in male pigs, by means of Fast Blue (FB) retrograde transport. After injection of FB into the left BSM, labeled neurons were found bilaterally in the L2-S4 sympathetic trunk ganglia (STGs), in the caudal mesenteric ganglia (CMGs), in the microganglia of the pelvic plexus (PGs), in a dorsolateral area with respect to the central canal of S1-S3 segments of the spinal cord (SC) and in the S1-S4 ipsilateral and S2-S3 contralateral spinal ganglia (SGs). The mean number of labeled FB cells was 3,122 +/- 1,968 in STGs, 979 +/- 667 in CMGs, 108 +/- 104 in PGs, 89 +/- 39 in SC and 77 +/- 23 in SGs. The area of the multipolar neurons was 852 +/- 22 microm(2) in the STGs, 878 +/- 23 microm(2) in the CMGs and 922 +/- 31 microm(2) in the PGs. The multipolar SC neurons had an area of 1,057 +/- 38 microm(2), while pseudounipolar SG cells had dimensions of 2,281 +/- 129 microm(2). Our research enables us to highlight two peculiarities regarding the innervation of the boar BSM: the very high number of labeled autonomic neurons and the particular localization of the motor somatic nucleus.

  5. 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

  6. Complete primary structure of a scallop striated muscle myosin heavy chain. Sequence comparison with other heavy chains reveals regions that might be critical for regulation.

    PubMed

    Nyitray, L; Goodwin, E B; Szent-Györgyi, A G

    1991-10-05

    We have determined the primary structure of the myosin heavy chain (MHC) of the striated adductor muscle of the scallop Aequipecten irradians by cloning and sequencing its cDNA. It is the first heavy chain sequence obtained in a directly Ca(2+)-regulated myosin. The 1938-amino acid sequence has an overall structure similar to other MHCs. The subfragment-1 region of the scallop MHC has a 59-62% sequence identity with sarcomeric and a 52-53% identity with nonsarcomeric (smooth and metazoan nonmuscle) MHCs. The heavy chain component of the regulatory domain (Kwon, H., Goodwin, E. B., Nyitray, L., Berliner, E., O'Neall-Hennessey, E., Melandri, F. D., and Szent-Györgyi, A. G. (1990) Proc. Natl. Acad. Sci. U.S.A. 87, 4771-4775) starts at either Leu-755 or Val-760. Ca(2+)-sensitive Trp residues (Wells, C., Warriner, K. E., and Bagshaw, C. R. (1985) Biochem. J. 231, 31-38) are located near the C-terminal end of this segment (residues 818-827). More detailed sequence comparison with other MHCs reveals that the 50-kDa domain and the N-terminal two-thirds of the 20-kDa domain differ substantially between sarcomeric and nonsarcomeric myosins. In contrast, in the light chain binding region of the regulatory domain (residues 784-844) the scallop sequence shows greater homology with regulated myosins (smooth muscle, nonmuscle, and invertebrate striated muscles) than with unregulated ones (vertebrate skeletal and heart muscles). The N-terminal 25-kDa domain also contains several residues which are preserved only in regulated myosins. These results indicate that certain heavy chain sites might be critical for regulation. The rod has features typical of sarcomeric myosins. It is 52-60% and 30-33% homologous with sarcomeric and nonsarcomeric MHCs, respectively. A Ser-rich tailpiece (residues 1918-1938) is apparently nonhelical.

  7. 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.

  8. Decrease in sarcoplasmic reticulum calcium content, not myofilament function, contributes to muscle twitch force decline in isolated cardiac trabeculae

    PubMed Central

    Milani-Nejad, Nima; Brunello, Lucia; Gyorke, Sándor; Janssen, Paul M.L.

    2014-01-01

    We set out to determine the factors responsible for twitch force decline in isolated intact rat cardiac trabeculae. The contractile force of trabeculae declined over extended periods of isometric twitch contractions. The force-frequency relationship within the frequency range of 4–8 Hz, at 37 °C, became more positive and the frequency optimum shifted to higher rates with this decline in baseline twitch tensions. The post-rest potentiation (37 °C), a phenomenon highly dependent on calcium handling mechanisms, became more pronounced with decrease in twitch tensions. We show that the main abnormality during muscle run-down was not due to a deficit in the myofilaments; maximal tension achieved using a K+ contracture protocol was either unaffected or only slightly decreased. Conversely, the sarcoplasmic reticulum (SR) calcium content, as assessed by rapid cooling contractures (from 27 °C to 0 °C), decreased, and had a close association with the declining twitch tensions (R2 ~ 0.76). SR Ca2+-ATPase, relative to Na+/Ca2+ exchanger activity, was not altered as there was no significant change in paired rapid cooling contracture ratios. Furthermore, confocal microscopy detected no abnormalities in the overall structure of the cardiomyocytes and t-tubules in the cardiac trabeculae (~23 °C). Overall, the data indicates that the primary mechanism responsible for force run-down in multi-cellular cardiac preparations is a decline in the SR calcium content and not the maximal tension generation capability of the myofilaments. PMID:25056841

  9. Molecular Characterization of Striated Muscle-Specific Gab1 Isoform as a Critical Signal Transducer for Neuregulin-1/ErbB Signaling in Cardiomyocytes

    PubMed Central

    Yasui, Taku; Masaki, Takeshi; Arita, Yoh; Ishibashi, Tomohiko; Inagaki, Tadakatsu; Okazawa, Makoto; Oka, Toru; Shioyama, Wataru; Yamauchi-Takihara, Keiko; Komuro, Issei; Sakata, Yasushi; Nakaoka, Yoshikazu

    2016-01-01

    Grb2-associated binder (Gab) docking proteins regulate signals downstream of a variety of growth factors and receptor tyrosine kinases. Neuregulin-1 (NRG-1), a member of epidermal growth factor family, plays a critical role for cardiomyocyte proliferation and prevention of heart failure via ErbB receptors. We previously reported that Gab1 and Gab2 in the myocardium are essential for maintenance of myocardial function in the postnatal heart via transmission of NRG-1/ErbB-signaling through analysis of Gab1/Gab2 cardiomyocyte-specific double knockout mice. In that study, we also found that there is an unknown high-molecular weight (high-MW) Gab1 isoform (120 kDa) expressed exclusively in the heart, in addition to the ubiquitously expressed low-MW (100 kDa) Gab1. However, the high-MW Gab1 has been molecularly ill-defined to date. Here, we identified the high-MW Gab1 as a striated muscle-specific isoform. The high-MW Gab1 has an extra exon encoding 27 amino acid residues between the already-known 3rd and 4th exons of the ubiquitously expressed low-MW Gab1. Expression analysis by RT-PCR and immunostaining with the antibody specific for the high-MW Gab1 demonstrate that the high-MW Gab1 isoform is exclusively expressed in striated muscle including heart and skeletal muscle. The ratio of high-MW Gab1/ total Gab1 mRNAs increased along with heart development. The high-MW Gab1 isoform in heart underwent tyrosine-phosphorylation exclusively after intravenous administration of NRG-1, among several growth factors. Adenovirus-mediated overexpression of the high-MW Gab1 induces more sustained activation of AKT after stimulation with NRG-1 in cardiomyocytes compared with that of β-galactosidase. On the contrary, siRNA-mediated knockdown of the high-MW Gab1 significantly attenuated AKT activation after stimulation with NRG-1 in cardiomyocytes. Taken together, these findings suggest that the striated muscle-specific high-MW isoform of Gab1 has a crucial role for NRG-1/ErbB signaling

  10. Role of sarcoplasmic reticulum Ca2+ content in Ca2+ entry of bovine airway smooth muscle cells.

    PubMed

    Bazán-Perkins, Blanca; Flores-Soto, Edgar; Barajas-López, Carlos; Montaño, Luis M

    2003-10-01

    Depletion of intracellular Ca(2+) stores induces the opening of an unknown Ca(2+ )entry pathway to the cell. We measured the intracellular free-Ca(2+) concentration ([Ca(2+)]i) at different sarcoplasmic reticulum (SR) Ca(2+) content in fura-2-loaded smooth muscle cells isolated from bovine tracheas. The absence of Ca(2+) in the extracellular medium generated a time-dependent decrement in [Ca(2+)]i which was proportional to the reduction in the SR-Ca(2+) content. This SR-Ca(2+) level was indirectly determined by measuring the amount of Ca(2+) released by caffeine. Ca(2+) restoration at different times after Ca(2+)-free incubation (2, 4, 6 and 10 min) induced an increment of [Ca(2+)]i. This increase in [Ca(2+)]i was considered as Ca(2+) entry to the cell. The rate of this entry was slow (~0.3 nM/s) when SR-Ca(2+) content was higher than 50% (2 and 4 min in Ca(2+)-free medium), and significantly ( p<0.01) accelerated (>1.0 nM/s) when SR-Ca(2+) content was lower than 50% (6 and 10 min in Ca(2+)-free medium). Thapsigargin significantly induced a higher rate of this Ca(2+) entry ( p<0.01). Variations in Ca(2+) influx after SR-Ca(2+) depletion were estimated more directly by a Mn(2+) quench approach. Ca(2+) restoration to the medium 4 min after Ca(2+) removal did not modify the Mn(2+) influx. However, when Ca(2+) was added after 10 min in Ca(2+)-free medium, an increment of Mn(2+) influx was observed, corroborating an increase in Ca(2+) entry. The fast Ca(2+) influx was Ni(2+) sensitive but was not affected by other known capacitative Ca(2+) entry blockers such as La(3+), Mg(2+), SKF 96365 and 2-APB. It was also not affected by the blockage of L-type Ca2(+) channels with methoxyverapamil or by the sustained K(+)-induced depolarisation. The slow Ca(2+) influx was only sensitive to SKF 96365. In conclusion, our results indicate that in bovine airway smooth muscle cells Ca(2+) influx after SR-Ca(2+) depletion has two rates: A) The slow Ca(2+) influx, which occurred in cells

  11. Sarcoplasmic reticulum lumenal Ca2+ has access to cytosolic activation and inactivation sites of skeletal muscle Ca2+ release channel.

    PubMed Central

    Tripathy, A; Meissner, G

    1996-01-01

    The effects of sarcoplasmic reticulum lumenal (trans) Ca2+ on cytosolic (cis) ATP-activated rabbit skeletal muscle Ca2+ release channels (ryanodine receptors) were examined using the planar lipid bilayer method. Single channels were recorded in symmetric 0.25 M KCl media with K+ as the major current carrier. With nanomolar [Ca2+] in both bilayer chambers, the addition of 2 mM cytosolic ATP greatly increased the number of short channel openings. As lumenal [Ca2+] was increased from < 0.1 microM to approximately 250 microM, increasing channel activities and events with long open time constants were seen at negative holding potentials. Channel activity remained low at positive holding potentials. Further increase in lumenal [Ca2+] to 1, 5, and 10 mM resulted in a decrease in channel activities at negative holding potentials and increased activities at positive holding potentials. A voltage-dependent activation by 50 microM lumenal Ca2+ was also observed when the channel was minimally activated by < 1 microM cytosolic Ca2+ in the absence of ATP. With microM cytosolic Ca2+ in the presence or absence of 2 mM ATP, single-channel activities showed no or only a weak voltage dependence. Other divalent cations (Mg2+, Ba2+) could not replace lumenal Ca2+. On the contrary, cytosolic ATP-activated channel activities were decreased as lumenal Ca2+ fluxes were reduced by the addition of 1-5 mM BaCl2 or MgCl2 to the lumenal side, which contained 50 microM Ca2+. An increase in [KCl] from 0.25 M to 1 M also reduced single-channel activities. Addition of the "fast" Ca2+ buffer 1,2-bis(2-aminophenoxy)ethanetetraacetic acid (BAPTA) to the cls chamber increased cytosolic ATP-, lumenal Ca(2+)-activated channel activities to a nearly maximum level. These results suggested that lumenal Ca2+ flowing through the skeletal muscle Ca2+ release channel may regulate channel activity by having access to cytosolic Ca2+ activation and Ca2+ inactivation sites that are located in "BAPTA

  12. Three-dimensional structure of the M-region (bare zone) of vertebrate striated muscle myosin filaments by single-particle analysis.

    PubMed

    Al-Khayat, Hind A; Kensler, Robert W; Morris, Edward P; Squire, John M

    2010-11-12

    The rods of anti-parallel myosin molecules overlap at the centre of bipolar myosin filaments to produce an M-region (bare zone) that is free of myosin heads. Beyond the M-region edges, myosin molecules aggregate in a parallel fashion to yield the bridge regions of the myosin filaments. Adjacent myosin filaments in striated muscle A-bands are cross-linked by the M-band. Vertebrate striated muscle myosin filaments have a 3-fold rotational symmetry around their long axes. In addition, at the centre of the M-region, there are three 2-fold axes perpendicular to the filament long axis, giving the whole filament dihedral 32-point group symmetry. Here we describe the three-dimensional structure obtained by a single-particle analysis of the M-region of myosin filaments from goldfish skeletal muscle under relaxing conditions and as viewed in negative stain. This is the first single-particle reconstruction of isolated M-regions. The resulting three-dimensional reconstruction reveals details to about 55 Å resolution of the density distribution in the five main nonmyosin densities in the M-band (M6', M4', M1, M4 and M6) and in the myosin head crowns (P1, P2 and P3) at the M-region edges. The outermost crowns in the reconstruction were identified specifically by their close similarity to the corresponding crown levels in our previously published bridge region reconstructions. The packing of myosin molecules into the M-region structure is discussed, and some unidentified densities are highlighted. Copyright © 2010 Elsevier Ltd. All rights reserved.

  13. 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

  14. 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.

  15. 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.

  16. 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.

  17. 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

  18. Bromo-eudistomin D, a novel inducer of calcium release from fragmented sarcoplasmic reticulum that causes contractions of skinned muscle fibers.

    PubMed

    Nakamura, Y; Kobayashi, J; Gilmore, J; Mascal, M; Rinehart, K L; Nakamura, H; Ohizumi, Y

    1986-03-25

    Bromo-eudistomin D induced a contraction of the chemically skinned fibers from skeletal muscle at concentrations of 10 microM or more. This contractile response to bromo-eudistomin D was completely blocked by 10 mM procaine. The extravascular Ca2+ concentrations of the heavy fractions of the fragmented sarcoplasmic reticulum (HSR) were measured directly by a Ca2+ electrode to examine the effect of bromo-eudistomin D on the sarcoplasmic reticulum. After the HSR was loaded with Ca2+ by the ATP-dependent Ca2+ pump, the addition of 10 microM bromo-eudistomin D caused Ca2+ release that was followed by spontaneous Ca2+ reuptake. In the presence of 2 microM ruthenium red or 4 mM MgCl2, no Ca2+ release was induced by 20 microM bromo-eudistomin D. The rate of 45Ca2+ efflux from HSR, which had been passively preloaded with 45Ca2+, was accelerated 7 times by 10 microM bromo-eudistomin D. The concentration of bromo-eudistomin D for half-maximum effect on the apparent efflux rate was 1.5 microM, while that of caffeine was 0.6 mM. The bromo-eudistomin D-evoked efflux of 45Ca2+ was abolished by 2 microM ruthenium red or 0.5 mM MgCl2. Bromo-eudistomin D was found to be 400 times more potent than caffeine in its Ca2+-releasing action but was similar in its action in other respects. These results indicate that bromo-eudistomin D may induce Ca2+ release from the sarcoplasmic reticulum through physiologically relevant Ca2+ channels.

  19. A Novel Conserved Isoform of the Ubiquitin Ligase UFD2a/UBE4B Is Expressed Exclusively in Mature Striated Muscle Cells

    PubMed Central

    Mammen, Andrew L.; Mahoney, James A.; St. Germain, Amanda; Badders, Nisha; Taylor, J. Paul; Rosen, Antony; Spinette, Sarah

    2011-01-01

    Yeast Ufd2p was the first identified E4 multiubiquitin chain assembly factor. Its vertebrate homologues later referred to as UFD2a, UBE4B or E4B were also shown to have E3 ubiquitin ligase activity. UFD2a function in the brain has been well established in vivo, and in vitro studies have shown that its activity is essential for proper condensation and segregation of chromosomes during mitosis. Here we show that 2 alternative splice forms of UFD2a, UFD2a-7 and -7/7a, are expressed sequentially during myoblast differentiation of C2C12 cell cultures and during cardiotoxin-induced regeneration of skeletal muscle in mice. UFD2a-7 contains an alternate exon 7, and UFD2a-7/7a, the larger of the 2 isoforms, contains an additional novel exon 7a. Analysis of protein or mRNA expression in mice and zebrafish revealed that a similar pattern of isoform switching occurs during developmental myogenesis of cardiac and skeletal muscle. In vertebrates (humans, rodents, zebrafish), UFD2a-7/7a is expressed only in mature striated muscle. This unique tissue specificity is further validated by the conserved presence of 2 muscle-specific splicing regulatory motifs located in the 3′ introns of exons 7 and 7a. UFD2a interacts with VCP/p97, an AAA-type ATPase implicated in processes whose functions appear to be regulated, in part, through their interaction with one or more of 15 previously identified cofactors. UFD2a-7/7a did not interact with VCP/p97 in yeast 2-hybrid experiments, which may allow the ATPase to bind cofactors that facilitate its muscle-specific functions. We conclude that the regulated expression of these UFD2a isoforms most likely imparts divergent functions that are important for myogenisis. PMID:22174917

  20. 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.

  1. Effects of Diaspirin Crosslinked Hemoglobin (DCLHb) on microcirculation and local tissue pO2 of striated skin muscle following resuscitation from hemorrhagic shock.

    PubMed

    Hungerer, Sven; Nolte, Dirk; Botzlar, Andreas; Messmer, Konrad

    2006-01-01

    The hemoglobin based oxygen carrier (HBOC) Diaspirin Crosslinked Hemoglobin (DCLHb) has been developed to substitute not only the blood volume, but also to restore the oxygen-carrying properties of blood during hemorrhagic shock. However, it has been suggested that HBOCs may enhance the formation of free oxygen radicals through the release of free iron ions via the Haber-Weiss reaction. The aim of this study was to investigate the effects of DCLHb on the microcirculation, leukocyte-endothelial cell interaction and local tissue oxygenation in striated skin muscle of Syrian golden hamsters during and after resuscitation from hemorrhagic shock. In particular we focused on the local tissue oxygenation after resuscitation with DCLHb (hemoglobin content 10 g%) compared to resuscitation using autologous blood diluted to a hemoglobin content of 10 g%. Hemorrhagic shock was induced for 45 minutes by bleeding the animals at a rate of 33 ml/kg BW maintaining a mean arterial pressure of 30 +/- 5 mmHg. Animals were resuscitated either with 33 ml/kg BW 6% Dextran-60.000 or with 10 g% DCLHb. The control group received shed blood diluted with Ringers to a hemoglobin content of 10 g%. Intravital microscopy was used for investigation of the microcirculatory parameters and a multiwire platinum surface electrode for measurement of local tissue pO2 in striated skin muscle in the dorsal skinfold chamber of Syrian golden hamsters. Resuscitation from hemorrhagic shock with 10 g% AUB revealed significant increase of leukocytes rolling in postcapillary venules at 30 to 120 minutes after resuscitation compared to baseline values. DCLHb turned out to reduce the number of firmly adherent leukocytes after resuscitation compared to 10 g% AUB. Microvascular permeability as an indicator for functional endothelial integrity revealed no significant differences between the groups. DCLHb and 10 g% AUB led to a significant increase in local tissue oxygenation after resuscitation from hemorrhagic shock

  2. The Influence of Sarcoplasmic Reticulum Ca2+ Concentration on Ca2+ Sparks and Spontaneous Transient Outward Currents in Single Smooth Muscle Cells

    PubMed Central

    ZhuGe, Ronghua; Tuft, Richard A.; Fogarty, Kevin E.; Bellve, Karl; Fay, Fredric S.; Walsh, John V.

    1999-01-01

    Localized, transient elevations in cytosolic Ca2+, known as Ca2+ sparks, caused by Ca2+ release from sarcoplasmic reticulum, are thought to trigger the opening of large conductance Ca2+-activated potassium channels in the plasma membrane resulting in spontaneous transient outward currents (STOCs) in smooth muscle cells. But the precise relationships between Ca2+ concentration within the sarcoplasmic reticulum and a Ca2+ spark and that between a Ca2+ spark and a STOC are not well defined or fully understood. To address these problems, we have employed two approaches using single patch-clamped smooth muscle cells freshly dissociated from toad stomach: a high speed, wide-field imaging system to simultaneously record Ca2+ sparks and STOCs, and a method to simultaneously measure free global Ca2+ concentration in the sarcoplasmic reticulum ([Ca2+]SR) and in the cytosol ([Ca2+]CYTO) along with STOCs. At a holding potential of 0 mV, cells displayed Ca2+ sparks and STOCs. Ca2+ sparks were associated with STOCs; the onset of the sparks coincided with the upstroke of STOCs, and both had approximately the same decay time. The mean increase in [Ca2+]CYTO at the time and location of the spark peak was ∼100 nM above a resting concentration of ∼100 nM. The frequency and amplitude of spontaneous Ca2+ sparks recorded at −80 mV were unchanged for a period of 10 min after removal of extracellular Ca2+ (nominally Ca2+-free solution with 50 μM EGTA), indicating that Ca2+ influx is not necessary for Ca2+sparks. A brief pulse of caffeine (20 mM) elicited a rapid decrease in [Ca2+]SR in association with a surge in [Ca2+]CYTO and a fusion of STOCs, followed by a fast restoration of [Ca2+]CYTO and a gradual recovery of [Ca2+]SR and STOCs. The return of global [Ca2+]CYTO to rest was an order of magnitude faster than the refilling of the sarcoplasmic reticulum with Ca2+. After the global [Ca2+]CYTO was fully restored, recovery of STOC frequency and amplitude were correlated with the

  3. [Electron microscopic studies of striated muscles of the Japanese quail (Coturnix coturnix japonica) obtained from eggs centrifuged during incubation].

    PubMed

    Belák, M; Kocisová, J; Marcaník, J; Boda, K; Skarda, R

    1981-01-01

    Reported in this paper are results obtained from investigations into the effect of hypergravitation on embryogenesis of striped muscles in Japanese quail. Specimens of striped muscles were excised from the birds after hatching and used in the study. The eggs had been centrifuged on zero day as well as on the first and 15th days of incubation. The muscle fibrils were found to proliferate in all groups, that is the control group, the group with centrifugation on zero day, and that with centrifugation on the first and 15th days. The process thus is characterised as physiological, known to occur prenatally and postnatally in developing individuals.

  4. 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.

  5. Beta 1D integrin displaces the beta 1A isoform in striated muscles: localization at junctional structures and signaling potential in nonmuscle cells.

    PubMed

    Belkin, A M; Zhidkova, N I; Balzac, F; Altruda, F; Tomatis, D; Maier, A; Tarone, G; Koteliansky, V E; Burridge, K

    1996-01-01

    The cytoplasmic domains of integrins provide attachment of these extracellular matrix receptors to the cytoskeleton and play a critical role in integrin-mediated signal transduction. In this report we describe the identification, expression, localization, and initial functional characterization of a novel form of beta 1 integrin, termed beta 1D. This isoform contains a unique alternatively spliced cytoplasmic domain of 50 amino acids, with the last 24 amino acids encoded by an additional exon. Of these 24 amino acids, 11 are conserved when compared to the beta 1A isoform, but 13 are unique (Zhidkova, N. I., A. M. Belkin, and R. Mayne. 1995. Biochem. Biophys. Res. Commun. 214:279-285; van der Flier, A., I. Kuikman, C. Baudoin, R, van der Neuf, and A. Sonnenberg. 1995. FEBS Lett. 369:340-344). Using an anti-peptide antibody against the beta 1D integrin subunit, we demonstrated that the beta 1D isoform is synthesized only in skeletal and cardiac muscles, while very low amounts of beta 1A were detected by immunoblot in striated muscles. Whereas beta 1A could not be detected in adult skeletal muscle fibers and cardiomyocytes by immunofluorescence, beta 1D was localized to the sarcolemma of both cell types. In skeletal muscle, beta 1D was concentrated in costameres, myotendinous, and neuromuscular junctions. In cardiac muscle this beta 1 isoform was found in costamers and intercalated discs. beta 1D was associated with alpha 7A and alpha 7B in adult skeletal muscle. In cardiomyocytes of adult heart, alpha 7B was the major partner for the beta 1D isoform. beta 1D could not be detected in proliferating C2C12 myoblasts, but it appeared immediately after myoblast fusion and its amount continued to rise during myotube growth and maturation. In contrast, expression of the beta 1A isoform was downregulated during myodifferentiation in culture and it was completely displaced by beta 1D in mature differentiated myotubes. We also analyzed some functional properties of the beta 1D

  6. ATP Consumption by Sarcoplasmic Reticulum Ca2+ Pumps Accounts for 40-50% of Resting Metabolic Rate in Mouse Fast and Slow Twitch Skeletal Muscle

    PubMed Central

    Smith, Ian Curtis; Bombardier, Eric; Vigna, Chris; Tupling, A. Russell

    2013-01-01

    The main purpose of this study was to directly quantify the relative contribution of Ca2+ cycling to resting metabolic rate in mouse fast (extensor digitorum longus, EDL) and slow (soleus) twitch skeletal muscle. Resting oxygen consumption of isolated muscles (VO2, µL/g wet weight/s) measured polarographically at 30°C was ~20% higher (P<0.05) in soleus (0.326 ± 0.022) than in EDL (0.261 ± 0.020). In order to quantify the specific contribution of Ca2+ cycling to resting metabolic rate, the concentration of MgCl2 in the bath was increased to 10 mM to block Ca2+ release through the ryanodine receptor, thus eliminating a major source of Ca2+ leak from the sarcoplasmic reticulum (SR), and thereby indirectly inhibiting the activity of the sarco(endo) plasmic reticulum Ca2+-ATPases (SERCAs). The relative (%) reduction in muscle VO2 in response to 10 mM MgCl2 was similar between soleus (48.0±3.7) and EDL (42.4±3.2). Using a different approach, we attempted to directly inhibit SERCA ATPase activity in stretched EDL and soleus muscles (1.42x optimum length) using the specific SERCA inhibitor cyclopiazonic acid (CPA, up to 160 µM), but were unsuccessful in removing the energetic cost of Ca2+ cycling in resting isolated muscles. The results of the MgCl2 experiments indicate that ATP consumption by SERCAs is responsible for 40–50% of resting metabolic rate in both mouse fast- and slow-twitch muscles at 30°C, or 12–15% of whole body resting VO2. Thus, SERCA pumps in skeletal muscle could represent an important control point for energy balance regulation and a potential target for metabolic alterations to oppose obesity. PMID:23840903

  7. ATP consumption by sarcoplasmic reticulum Ca²⁺ pumps accounts for 40-50% of resting metabolic rate in mouse fast and slow twitch skeletal muscle.

    PubMed

    Smith, Ian Curtis; Bombardier, Eric; Vigna, Chris; Tupling, A Russell

    2013-01-01

    The main purpose of this study was to directly quantify the relative contribution of Ca²⁺ cycling to resting metabolic rate in mouse fast (extensor digitorum longus, EDL) and slow (soleus) twitch skeletal muscle. Resting oxygen consumption of isolated muscles (VO₂, µL/g wet weight/s) measured polarographically at 30°C was ~20% higher (P<0.05) in soleus (0.326 ± 0.022) than in EDL (0.261 ± 0.020). In order to quantify the specific contribution of Ca²⁺ cycling to resting metabolic rate, the concentration of MgCl₂ in the bath was increased to 10 mM to block Ca²⁺ release through the ryanodine receptor, thus eliminating a major source of Ca²⁺ leak from the sarcoplasmic reticulum (SR), and thereby indirectly inhibiting the activity of the sarco(endo) plasmic reticulum Ca²⁺-ATPases (SERCAs). The relative (%) reduction in muscle VO₂ in response to 10 mM MgCl₂ was similar between soleus (48.0±3.7) and EDL (42.4±3.2). Using a different approach, we attempted to directly inhibit SERCA ATPase activity in stretched EDL and soleus muscles (1.42x optimum length) using the specific SERCA inhibitor cyclopiazonic acid (CPA, up to 160 µM), but were unsuccessful in removing the energetic cost of Ca²⁺ cycling in resting isolated muscles. The results of the MgCl₂ experiments indicate that ATP consumption by SERCAs is responsible for 40-50% of resting metabolic rate in both mouse fast- and slow-twitch muscles at 30°C, or 12-15% of whole body resting VO₂. Thus, SERCA pumps in skeletal muscle could represent an important control point for energy balance regulation and a potential target for metabolic alterations to oppose obesity.

  8. The expression of the neonatal sarcoplasmic reticulum Ca2+ pump (SERCA1b) hints to a role in muscle growth and development.

    PubMed

    Zádor, Erno; Vangheluwe, Peter; Wuytack, Frank

    2007-04-01

    The neonatal isoform of sarcoplasmic/endoplasmic reticulum Ca2+ ATPase 1 (SERCA1b) is a Ca2+ pump with a well-known developmentally regulated transcript level but an undefined protein expression and function. Specific antibodies were generated to show that SERCA1b is exclusively expressed in myoblasts and myotubes of cultured and regenerating muscle. However, the SERCA1b protein was not detectable in normal adult fast and slow muscles. Studies of the in vitro differentiating myogenic cell lines C2C12 and sol8 showed that SERCA1b is the main SERCA1 protein isoform induced during differentiation and that it is found in the myotubes. Remarkably in BC3H1 cells, which show incomplete differentiation and are reluctant to form myotubes, express the SERCA1b mRNA but not the corresponding protein. SERCA1b protein was also absent from stretched or denervated adult soleus, in spite of the fact that its mRNA level was upregulated. SERCA1b accounts for nearly the total of SERCA1 expression in the diaphragm of newborn mice, which suggests that the insufficient function and development of the diaphragm in the SERCA1 null mutant mice may be due to the lack of SERCA1b. Our studies point to an important regulation of SERCA1b expression at the protein level and hints to a role in the growth of the developing muscle.

  9. [Striated and delayed nephrography].

    PubMed

    Marlois, O; Padovani, J; Faure, F; Devred, P; Grangier, M L; Panuel, M

    1985-10-01

    About a case of striated and delayed nephrogram seen on a diabetic child, authors come back to the different etiologies. Among them, the tubular precipitation of Tamm-Horsfall protein seems to be given like on the right possibilities. Whatever is its etiology, the mechanism of striated appearance is always the same, being founded on the radiated disposal of the collecting ducts and on a tubular stasis beeing with iodine concentration.

  10. The increase in non-cross-bridge forces after stretch of activated striated muscle is related to titin isoforms.

    PubMed

    Cornachione, Anabelle S; Leite, Felipe; Bagni, Maria Angela; Rassier, Dilson E

    2016-01-01

    Skeletal muscles present a non-cross-bridge increase in sarcomere stiffness and tension on Ca(2+) activation, referred to as static stiffness and static tension, respectively. It has been hypothesized that this increase in tension is caused by Ca(2+)-dependent changes in the properties of titin molecules. To verify this hypothesis, we investigated the static tension in muscles containing different titin isoforms. Permeabilized myofibrils were isolated from the psoas, soleus, and heart ventricle from the rabbit, and tested in pCa 9.0 and pCa 4.5, before and after extraction of troponin C, thin filaments, and treatment with the actomyosin inhibitor blebbistatin. The myofibrils were tested with stretches of different amplitudes in sarcomere lengths varying between 1.93 and 3.37 μm for the psoas, 2.68 and 4.21 μm for the soleus, and 1.51 and 2.86 μm for the ventricle. Using gel electrophoresis, we confirmed that the three muscles tested have different titin isoforms. The static tension was present in psoas and soleus myofibrils, but not in ventricle myofibrils, and higher in psoas myofibrils than in soleus myofibrils. These results suggest that the increase in the static tension is directly associated with Ca(2+)-dependent change in titin properties and not associated with changes in titin-actin interactions.

  11. Expression of cardiac alpha-actin spares extraocular muscles in skeletal muscle alpha-actin diseases--quantification of striated alpha-actins by MRM-mass spectrometry.

    PubMed

    Ravenscroft, Gianina; Colley, Stephen M J; Walker, Kendall R; Clement, Sophie; Bringans, Scott; Lipscombe, Richard; Fabian, Victoria A; Laing, Nigel G; Nowak, Kristen J

    2008-12-01

    As with many skeletal muscle diseases, the extraocular muscles (EOMs) are spared in skeletal muscle alpha-actin diseases, with no ophthalmoplegia even in severely affected patients. We hypothesised that the extraocular muscles sparing in these patients was due to significant expression of cardiac alpha-actin, the alpha-actin isoform expressed in heart and foetal skeletal muscle. We have shown by immunochemistry, Western blotting and a novel MRM-mass spectrometry technique, comparable levels of cardiac alpha-actin in the extraocular muscles of human, pig and sheep to those in the heart. The sparing of extraocular muscles in skeletal muscle alpha-actin disease is thus probably due to greater levels of cardiac alpha-actin, than the negligible amounts in skeletal muscles, diluting out the effects of the mutant skeletal muscle alpha-actin.

  12. Evolutionarily conserved sequences of striated muscle myosin heavy chain isoforms. Epitope mapping by cDNA expression.

    PubMed

    Miller, J B; Teal, S B; Stockdale, F E

    1989-08-05

    A cDNA expression strategy was used to localize amino acid sequences which were specific for fast, as opposed to slow, isoforms of the chicken skeletal muscle myosin heavy chain (MHC) and which were conserved in vertebrate evolution. Five monoclonal antibodies (mAbs), termed F18, F27, F30, F47, and F59, were prepared that reacted with all of the known chicken fast MHC isoforms but did not react with any of the known chicken slow nor with smooth muscle MHC isoforms. The epitopes recognized by mAbs F18, F30, F47, and F59 were on the globular head fragment of the MHC, whereas the epitope recognized by mAb F27 was on the helical tail or rod fragment. Reactivity of all five mAbs also was confined to fast MHCs in the rat, with the exception of mAb F59, which also reacted with the beta-cardiac MHC, the single slow MHC isoform common to both the rat heart and skeletal muscle. None of the five epitopes was expressed on amphioxus, nematode, or Dictyostelium MHC. The F27 and F59 epitopes were found on shark, electric ray, goldfish, newt, frog, turtle, chicken, quail, rabbit, and rat MHCs. The epitopes recognized by these mAbs were conserved, therefore, to varying degrees through vertebrate evolution and differed in sequence from homologous regions of a number of invertebrate MHCs and myosin-like proteins. The sequence of those epitopes on the head were mapped using a two-part cDNA expression strategy. First, Bal31 exonuclease digestion was used to rapidly generate fragments of a chicken embryonic fast MHC cDNA that were progressively deleted from the 3' end. These cDNA fragments were expressed as beta-galactosidase/MHC fusion proteins using the pUR290 vector; the fusion proteins were tested by immunoblotting for reactivity with the mAbs; and the approximate locations of the epitopes were determined from the sizes of the cDNA fragments that encoded a particular epitope. The epitopes were then precisely mapped by expression of overlapping cDNA fragments of known sequence that

  13. Action of lovastatin, simvastatin, and pravastatin on sterol synthesis and their antiproliferative effect in cultured myoblasts from human striated muscle.

    PubMed

    van Vliet, A K; Nègre-Aminou, P; van Thiel, G C; Bolhuis, P A; Cohen, L H

    1996-11-08

    Lovastatin, simvastatin, and pravastatin are fairly strong inhibitors of sterol synthesis in human myoblasts in culture. Lovastatin and simvastatin have IC50 values of 19 +/- 6 nM and 4.0 +/- 2.3 nM, respectively. Pravastatin is a weaker inhibitor of sterol synthesis (IC50 value of 110 +/- 38 nM). Through inhibition of mevalonate production, these compounds have a distinct inhibiting effect on cell proliferation. Because proliferation of myoblasts is important in the repair of damaged skeletal muscle, experiments were performed to investigate the effect of lovastatin, simvastatin, and pravastatin on cell proliferation and cell viability. The more potent inhibitors of sterol synthesis, lovastatin, and simvastatin, were able to inhibit the proliferation of these cells during 3 days of incubation with drug concentrations of 1 microM for lovastatin and 0.1 microM or 1 microM for simvastatin. DNA synthesis was decreased by more than 80% in the presence of 1 microM of lovastatin or simvastatin. In contrast, under these conditions, pravastatin had no influence on cell proliferation or DNA synthesis, which is probably related to the lack of inhibition of sterol synthesis by pravastatin on extended incubation. The three 3-hydroxy-3-methylglutaryl coenzyme A reductase inhibitors did not disturb cell viability because mitochondrial dehydrogenase activity and ATP content remained proportional to the number of cells in the culture at any concentration used.

  14. The effect of pH on the calcium dependence of calcium accumulation in dog cardiac muscle sarcoplasmic reticulum.

    PubMed

    Grassi de Gende, A O

    1988-12-01

    Net Ca2+ accumulation in vesicles of dog cardiac sarcoplasmic reticulum (CSR) was evaluated at three different pHs: 6.0, 6.8 and 7.6. The Ca2+ sequestration by CSR depends on Ca2+ concentration and on pH values. The curves that show the relationship between Ca2+ accumulated by CSR and external Ca2+ concentrations were shifted with pH changes, both in the absence and in the presence of potassium oxalate. Considering the curve at pH 6.8 as reference, a lower Ca concentration was needed to obtain the half-maximal value in Ca sequestration under pH 7.6 (0.04 +/- 0.006 and 0.79 +/- 0.09 microM at pH 7.6 and 6.8, respectively). Opposite results were obtained under pH 6.0 (13.66 +/- 1.29 microM). Net calcium release during active accumulation of Ca2+ and Ca2+ efflux from passively 45Ca2+ loaded CSR microsomes were significantly higher at alkaline pH than at acidic pH. The results suggest that in CSR alkaline pH would promote the increase in the rates of both, Ca2+ release and active Ca2+ accumulation, while opposite effects would be expected under acidic pH. Therefore, pH changes may regulate both, the Ca2+ level upon which the SR Ca2+ pump works (permeability effect) and the sequestration rate of the Ca2+ pump (variation in the affinity for calcium).

  15. Nanospan, an alternatively spliced isoform of sarcospan, localizes to the sarcoplasmic reticulum in skeletal muscle and is absent in limb girdle muscular dystrophy 2F.

    PubMed

    Peter, Angela K; Miller, Gaynor; Capote, Joana; DiFranco, Marino; Solares-Pérez, Alhondra; Wang, Emily L; Heighway, Jim; Coral-Vázquez, Ramón M; Vergara, Julio; Crosbie-Watson, Rachelle H

    2017-06-06

    Sarcospan (SSPN) is a transmembrane protein that interacts with the sarcoglycans (SGs) to form a tight subcomplex within the dystrophin-glycoprotein complex that spans the sarcolemma and interacts with laminin in the extracellular matrix. Overexpression of SSPN ameliorates Duchenne muscular dystrophy in murine models. Standard cloning approaches were used to identify nanospan, and nanospan-specific polyclonal antibodies were generated and validated. Biochemical isolation of skeletal muscle membranes and two-photon laser scanning microscopy were used to analyze nanospan localization in muscle from multiple murine models. Duchenne muscular dystrophy biopsies were analyzed by immunoblot analysis of protein lysates as well as indirect immunofluorescence analysis of muscle cryosections. Nanospan is an alternatively spliced isoform of sarcospan. While SSPN has four transmembrane domains and is a core component of the sarcolemmal dystrophin-glycoprotein complex, nanospan is a type II transmembrane protein that does not associate with the dystrophin-glycoprotein complex. We demonstrate that nanospan is enriched in the sarcoplasmic reticulum (SR) fractions and is not present in the T-tubules. SR fractions contain membranes from three distinct structural regions: a region flanking the T-tubules (triadic SR), a SR region across the Z-line (ZSR), and a longitudinal SR region across the M-line (LSR). Analysis of isolated murine muscles reveals that nanospan is mostly associated with the ZSR and triadic SR, and only minimally with the LSR. Furthermore, nanospan is absent from the SR of δ-SG-null (Sgcd(-/-)) skeletal muscle, a murine model for limb girdle muscular dystrophy 2F. Analysis of skeletal muscle biopsies from Duchenne muscular dystrophy patients reveals that nanospan is preferentially expressed in type I (slow) fibers in both control and Duchenne samples. Furthermore, nanospan is significantly reduced in Duchenne biopsies. Alternative splicing of proteins from the SG

  16. Nanospan, an alternatively spliced isoform of sarcospan, localizes to the sarcoplasmic reticulum in skeletal muscle and is absent in limb girdle muscular dystrophy 2F.

    PubMed

    Peter, Angela K; Miller, Gaynor; Capote, Joana; DiFranco, Marino; Solares-Pérez, Alhondra; Wang, Emily L; Heighway, Jim; Coral-Vázquez, Ramón M; Vergara, Julio; Crosbie-Watson, Rachelle H

    2017-01-01

    Sarcospan (SSPN) is a transmembrane protein that interacts with the sarcoglycans (SGs) to form a tight subcomplex within the dystrophin-glycoprotein complex that spans the sarcolemma and interacts with laminin in the extracellular matrix. Overexpression of SSPN ameliorates Duchenne muscular dystrophy in murine models. Standard cloning approaches were used to identify nanospan, and nanospan-specific polyclonal antibodies were generated and validated. Biochemical isolation of skeletal muscle membranes and two-photon laser scanning microscopy were used to analyze nanospan localization in muscle from multiple murine models. Duchenne muscular dystrophy biopsies were analyzed by immunoblot analysis of protein lysates as well as indirect immunofluorescence analysis of muscle cryosections. Nanospan is an alternatively spliced isoform of sarcospan. While SSPN has four transmembrane domains and is a core component of the sarcolemmal dystrophin-glycoprotein complex, nanospan is a type II transmembrane protein that does not associate with the dystrophin-glycoprotein complex. We demonstrate that nanospan is enriched in the sarcoplasmic reticulum (SR) fractions and is not present in the T-tubules. SR fractions contain membranes from three distinct structural regions: a region flanking the T-tubules (triadic SR), a SR region across the Z-line (ZSR), and a longitudinal SR region across the M-line (LSR). Analysis of isolated murine muscles reveals that nanospan is mostly associated with the ZSR and triadic SR, and only minimally with the LSR. Furthermore, nanospan is absent from the SR of δ-SG-null (Sgcd(-/-)) skeletal muscle, a murine model for limb girdle muscular dystrophy 2F. Analysis of skeletal muscle biopsies from Duchenne muscular dystrophy patients reveals that nanospan is preferentially expressed in type I (slow) fibers in both control and Duchenne samples. Furthermore, nanospan is significantly reduced in Duchenne biopsies. Alternative splicing of proteins from the SG

  17. The use of the indicator fluo-5N to measure sarcoplasmic reticulum calcium in single muscle fibres of the cane toad

    PubMed Central

    Kabbara, Akram A; Allen, David G

    2001-01-01

    Single fibres from the lumbrical muscles of the cane toad (Bufo marinus) were incubated in fluo-5N AM for 2 h at 35 °C in order to load the indicator into the sarcoplasmic reticulum. Fluo-5N is a low-affinity calcium indicator (KCa 90 μm). Successful sarcoplasmic reticulum (SR) loading was indicated by a fluorescence signal that declined during contraction. Confocal microscopy showed that the dye loaded principally in lines perpendicular to the long axis of the fibre that repeated each sarcomere. This is consistent with much of the dye residing in the SR. To establish the site of loading, fibres were exposed to 30 mm caffeine in the presence of 20 μm 2,5-di(tert-butyl)1,4-hydroquinone (TBQ, an SR pump inhibitor) which should release most Ca2+ from the SR; this procedure reduced the fluorescence to 46 ± 4 % of the control value. To determine how much indicator was in the myoplasm, fibres were exposed to 100 μg ml−1 saponin which permeabilizes the surface membrane; saponin treatment reduced the fluorescence to 51 ± 2 % of the control value. During maximally activated tetani (100 Hz stimulation rate, 22 °C) the component of signal from the SR declined by 33 ± 4 %. During relaxation the SR signal recovered in two phases with time constants of 0.38 ± 0.14 s and 10.1 ± 1.7 s. Partially activated tetani (30 Hz stimulation rate) showed a smaller SR signal. Application of the SR Ca2+ pump inhibitor TBQ slowed the rate of recovery of the SR signal. Muscle fatigue was produced by repeated short tetani until tension was reduced to 50 %. The SR signal during the periods between tetani declined steadily and the SR Ca2+ signal was eventually reduced to 71 ± 8 % of the control signal. This signal recovered in two phases when the muscle was rested. An initial phase had a time constant of 1.7 ± 0.2 s so that by 20 s of recovery the SR Ca2+ signal was 86 ± 7 % of control; the second phase was slower and by 5 min the SR Ca2+ signal was back to control values (98 ± 5

  18. The use of the indicator fluo-5N to measure sarcoplasmic reticulum calcium in single muscle fibres of the cane toad.

    PubMed

    Kabbara, A A; Allen, D G

    2001-07-01

    1. Single fibres from the lumbrical muscles of the cane toad (Bufo marinus) were incubated in fluo-5N AM for 2 h at 35 degrees C in order to load the indicator into the sarcoplasmic reticulum. Fluo-5N is a low-affinity calcium indicator (K(Ca) 90 microM). Successful sarcoplasmic reticulum (SR) loading was indicated by a fluorescence signal that declined during contraction. 2. Confocal microscopy showed that the dye loaded principally in lines perpendicular to the long axis of the fibre that repeated each sarcomere. This is consistent with much of the dye residing in the SR. 3. To establish the site of loading, fibres were exposed to 30 mM caffeine in the presence of 20 microM 2,5-di(tert-butyl)1,4-hydroquinone (TBQ, an SR pump inhibitor) which should release most Ca(2+) from the SR; this procedure reduced the fluorescence to 46 +/- 4 % of the control value. To determine how much indicator was in the myoplasm, fibres were exposed to 100 microg ml(-1) saponin which permeabilizes the surface membrane; saponin treatment reduced the fluorescence to 51 +/- 2 % of the control value. 4. During maximally activated tetani (100 Hz stimulation rate, 22 degrees C) the component of signal from the SR declined by 33 +/- 4 %. During relaxation the SR signal recovered in two phases with time constants of 0.38 +/- 0.14 s and 10.1 +/- 1.7 s. Partially activated tetani (30 Hz stimulation rate) showed a smaller SR signal. Application of the SR Ca(2+) pump inhibitor TBQ slowed the rate of recovery of the SR signal. 5. Muscle fatigue was produced by repeated short tetani until tension was reduced to 50 %. The SR signal during the periods between tetani declined steadily and the SR Ca(2+) signal was eventually reduced to 71 +/- 8 % of the control signal. This signal recovered in two phases when the muscle was rested. An initial phase had a time constant of 1.7 +/- 0.2 s so that by 20 s of recovery the SR Ca(2+) signal was 86 +/- 7 % of control; the second phase was slower and by 5 min the

  19. Contractile properties and sarcoplasmic reticulum calcium content in type I and type II skeletal muscle fibres in active aged humans

    PubMed Central

    Lamboley, C R; Wyckelsma, V L; Dutka, T L; McKenna, M J; Murphy, R M; Lamb, G D

    2015-01-01

    This study examined the contractile properties and sarcoplasmic reticulum (SR) Ca2+ content in mechanically skinned vastus lateralis muscle fibres of Old (70 ± 4 years) and Young (22 ± 3 years) humans to investigate whether changes in muscle fibre properties contribute to muscle weakness in old age. In type II fibres of Old subjects, specific force was reduced by ∼17% and Ca2+ sensitivity was also reduced (pCa50 decreased ∼0.05 pCa units) relative to that in Young. S-Glutathionylation of fast troponin I (TnIf) markedly increased Ca2+ sensitivity in type II fibres, but the increase was significantly smaller in Old versus Young (+0.136 and +0.164 pCa unit increases, respectively). Endogenous and maximal SR Ca2+ content were significantly smaller in both type I and type II fibres in Old subjects. In fibres of Young, the SR could be nearly fully depleted of Ca2+ by a combined caffeine and low Mg2+ stimulus, whereas in fibres of Old the amount of non-releasable Ca2+ was significantly increased (by > 12% of endogenous Ca2+ content). Western blotting showed an increased proportion of type I fibres in Old subjects, and increased amounts of calsequestrin-2 and calsequestrin-like protein. The findings suggest that muscle weakness in old age is probably attributable in part to (i) an increased proportion of type I fibres, (ii) a reduction in both maximum specific force and Ca2+ sensitivity in type II fibres, and also a decreased ability of S-glutathionylation of TnIf to counter the fatiguing effects of metabolites on Ca2+ sensitivity, and (iii) a reduction in the amount of releasable SR Ca2+ in both fibre types. Key points Muscle weakness in old age is due in large part to an overall loss of skeletal muscle tissue, but it remains uncertain how much also stems from alterations in the properties of the individual muscle fibres. This study examined the contractile properties and amount of stored intracellular calcium in single muscle fibres of Old (70

  20. Identification of 30 kDa protein for Ca(2+) releasing action of myotoxin a with a mechanism common to DIDS in skeletal muscle sarcoplasmic reticulum.

    PubMed

    Hirata, Y; Nakahata, N; Ohkura, M; Ohizumi, Y

    1999-08-12

    The molecular mechanism of Ca(2+) release by myotoxin a (MTYX), a polypeptide toxin isolated from the venom of prairie rattlesnakes (Crotalus viridis viridis), was investigated in the heavy fraction of sarcoplasmic reticulum (HSR) of rabbit skeletal muscles. [(125)I]MYTX bound to four HSR proteins (106, 74, 53 and 30 kDa) on polyvinylidene difluoride (PVDF) membrane. DIDS, 4, 4'-diisothiocyanatostilbene-2,2'-disulfonic acid, bound predominantly to 30 kDa protein on the PVDF membrane, the molecular weight of which was similar to one of the MYTX binding proteins. The maximum (45)Ca(2+) release induced by caffeine (30 mM) was further increased in the presence of MYTX (10 microM) or DIDS (30 microM), whereas that induced by DIDS (30 microM) was not affected by MYTX (10 microM). MYTX inhibited [(3)H]DIDS binding to HSR in a concentration-dependent manner. Furthermore, [(125)I]MYTX binding to 30 kDa protein was inhibited by DIDS in a concentration-dependent manner. These results suggest that MYTX and DIDS release Ca(2+) from HSR in a common mechanism. The 30 kDa protein may be a target protein for the Ca(2+) releasing action of MYTX and DIDS.

  1. The volume of the T-system and its association with the sarcoplasmic reticulum in slow muscle fibres of the frog

    PubMed Central

    Flitney, F. W.

    1971-01-01

    1. A study has been made of the T-system and sarcoplasmic reticulum (SR) in slow muscle fibres of the frog, Rana temporaria. 2. The size of the T-system was measured by an autoradiographic method, using tritium-labelled albumin as a marker. Its volume, expressed as a fraction of that of the fibre, was found to be 1·8 × 10-3, as compared with a figure of 3·9 × 10-3 for the T-system in a twitch fibre. 3. The spatial distribution of the T-tubules, and their association with the SR, was studied with the electron microscope, employing ferritin and the enzyme peroxidase as markers. The observations show (a) the tubules form a three dimensional, rather than transverse, network and (b) the area of triadic (and diadic) contact with the SR is 5-10 × smaller than in a twitch fibre. 4. The possibility that the T-system and SR of the slow fibre participate in linking membrane excitation with contraction is discussed in the light of these findings. ImagesFig. 1Fig. 2Plate 2Figs. 1 and 2Plate 4 PMID:5571928

  2. 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.

  3. Interaction between endoplasmic/sarcoplasmic reticulum stress (ER/SR stress), mitochondrial signaling and Ca2+ regulation in airway smooth muscle (ASM)1

    PubMed Central

    Delmotte, Philippe; Sieck, Gary C.

    2015-01-01

    Airway inflammation is a key aspect of diseases such as asthma. Several inflammatory cytokines (e.g., TNFα and IL-13) increase cytosolic Ca2+ ([Ca2+]cyt) responses to agonist stimulation and Ca2+ 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 [Ca2+]cyt induced by agonists leads to a transient increase in mitochondrial Ca2+ ([Ca2+]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 [Ca2+]mito is blunted despite enhanced [Ca2+]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 Ca2+ buffering, mitochondrial fragmentation, and increased cell proliferation. PMID:25506723

  4. The Ca(2+)-ATPase of the sarcoplasmic reticulum in skeletal and cardiac muscle. An overview from the very beginning to more recent prospects.

    PubMed

    Hasselbach, W

    1998-09-16

    The discovery of the ATP-driven calcium pump in the sarcoplasmic reticulum membranes reaches back to the postwar (World War II) years and would not be possible without the generous support by the American scientific community. It was this community that in pre- and postwar years gave shelter to many European scientists, which in return stimulated scientific development in the United States. These pre- and postwar relations helped to establish the calcium pump as a physiologically relevant mechanism in all kinds of cells. The pump and its counterpart, the calcium release channel, proved to be controlled by various intrinsic mechanisms. Rising hydrogen concentrations as occurring in ischemic muscles switch off pump activity and counteract allosterically caffeine-induced calcium release (CICR). Rising phosphate or the presence of other calcium-precipitating anions, on the other hand, prevents pump inhibition by intraluminal calcium precipitation, which, simultaneously, can increase the quantity of releasable calcium. The inactivation of CICR by removing medium chloride must be considered as a hint of additional mechanisms by which calcium-dependent activity regulation can be modified.

  5. Sarcoplasmic reticulum calcium release compared in slow-twitch and fast-twitch fibres of mouse muscle.

    PubMed

    Baylor, S M; Hollingworth, S

    2003-08-15

    Experiments were carried out to compare the amplitude and time course of Ca2+ release from the sarcoplasmic reticulum (SR) in intact slow-twitch and fast-twitch mouse fibres. Individual fibres within small bundles were injected with furaptra, a low-affinity, rapidly responding Ca2+ indicator. In response to a single action potential at 16 degrees C, the peak amplitude and half-duration of the change in myoplasmic free [Ca2+] (Delta[Ca2+]) differed significantly between fibre types (slow-twitch: peak amplitude, 9.4 +/- 1.0 microM (mean +/- S.E.M.); half-duration, 7.7 +/- 0.6 ms; fast-twitch: peak amplitude 18.5 +/- 0.5 microM; half-duration, 4.9 +/- 0.3 ms). SR Ca2+ release was estimated from Delta[Ca2+] with a computational model that calculated Ca2+ binding to the major myoplasmic Ca2+ buffers (troponin, ATP and parvalbumin); buffer concentrations and reaction rate constants were adjusted to reflect fibre-type differences. In response to an action potential, the total concentration of released Ca2+ (Delta[CaT]) and the peak rate of Ca2+ release ((d/dt)Delta[CaT]) differed about 3-fold between the fibre types (slow-twitch: Delta[CaT], 127 +/- 7 microM; (d/dt)Delta[CaT], 70 +/- 6 microM ms-1; fast-twitch: Delta[CaT], 346 +/- 6 microM; (d/dt)Delta[CaT], 212 +/- 4 microM ms-1). In contrast, the half-duration of (d/dt)Delta[CaT] was very similar in the two fibre types (slow-twitch, 1.8 +/- 0.1 ms; fast-twitch, 1.6 +/- 0.0 ms). When fibres were stimulated with a 5-shock train at 67 Hz, the peaks of (d/dt)Delta[CaT] in response to the second and subsequent shocks were much smaller than that due to the first shock; the later peaks, expressed as a fraction of the amplitude of the first peak, were similar in the two fibre types (slow-twitch, 0.2-0.3; fast-twitch, 0.1-0.3). The results support the conclusion that individual SR Ca2+ release units function similarly in slow-twitch and fast-twitch mammalian fibres.

  6. Identification of a 97-kDa mastoparan-binding protein involving in Ca(2+) release from skeletal muscle sarcoplasmic reticulum.

    PubMed

    Hirata, Y; Nakahata, N; Ohizumi, Y

    2000-06-01

    Mastoparan (MP) and radiolabeled [Tyr(3)]MP caused a transient Ca(2+) release from the heavy fraction of sarcoplasmic reticulum, which was inhibited by ryanodine. MP enhanced [(3)H]ryanodine binding in a concentration-dependent manner with an EC(50) value of approximately 0.3 microM. The (45)Ca(2+) release was accelerated by MP, [Tyr(3)]MP, or caffeine in a concentration-dependent manner. The EC(50) values for MP, [Tyr(3)]MP, and caffeine were approximately 2. 0 microM, 7.7 microM, and 1.8 mM, respectively. MP, like caffeine, shifted the stimulatory limb of a bell-shaped curve of Ca(2+) dependence to the left. (45)Ca(2+) release induced by caffeine was completely inhibited by typical blockers of Ca(2+)-induced Ca(2+) release, such as Mg(2+), ruthenium red, or procaine. However, (45)Ca(2+) release induced by MP was completely inhibited by Mg(2+), but it was only partially inhibited by ruthenium red or procaine. The rate of (45)Ca(2+) release induced by MP was further increased in the presence of caffeine, showing that the MP binding site is different from that of caffeine on Ca(2+) release channels. We succeeded in the synthesis of (125)I-[Tyr(3)]MP with a high specific activity. (125)I-[Tyr(3)]MP bound specifically to heavy fraction of sarcoplasmic reticulum with a K(d) value of 4.0 microM and a B(max) value of 3.0 nmol/mg. Furthermore, (125)I-[Tyr(3)]MP specifically cross-linked to the 97-kDa protein without direct binding to ryanodine receptor. The protein was not triadin or Ca(2+)-pump, because antitriadin antibody and anti-Ca(2+)-pump antibody did not immunoprecipitate the protein. These results suggest that the 97-kDa MP-binding protein may have an important role in the excitation-contraction coupling of skeletal muscle.

  7. Acute effects of taurine on sarcoplasmic reticulum Ca2+ accumulation and contractility in human type I and type II skeletal muscle fibers.

    PubMed

    Dutka, T L; Lamboley, C R; Murphy, R M; Lamb, G D

    2014-10-01

    Taurine occurs in high concentrations in muscle and is implicated in numerous physiological processes, yet its effects on many aspects of contractility remain unclear. Using mechanically skinned segments of human vastus lateralis muscle fibers, we characterized the effects of taurine on sarcoplasmic reticulum (SR) Ca2+ accumulation and contractile apparatus properties in type I and type II fibers. Prolonged myoplasmic exposure (>10 min) to taurine substantially increased the rate of accumulation of Ca2+ by the SR in both fiber types, with no change in the maximum amount accumulated; no such effect was found with carnosine. SR Ca2+ accumulation was similar with 10 or 20 mM taurine, but was significantly slower at 5 mM taurine. Cytoplasmic taurine (20 mM) had no detectable effects on the responsiveness of the Ca2+ release channels in either fiber type. Taurine caused a small increase in Ca2+ sensitivity of the contractile apparatus in type I fibers, but type II fibers were unaffected; maximum Ca(2+)-activated force was unchanged in both cases. The effects of taurine on SR Ca2+ accumulation (1) only became apparent after prolonged cytoplasmic exposure, and (2) persisted for some minutes after complete removal of taurine from the cytoplasm, consistent with the hypothesis that the effects were due to an action of taurine from inside the SR. In summary, taurine potentiates the rate of SR Ca2+ uptake in both type I and type II human fibers, possibly via an action from within the SR lumen, with the degree of potentiation being significantly reduced at low physiological taurine levels. Copyright © 2014 the American Physiological Society.

  8. cGMP reduces the sarcoplasmic reticulum Ca2+ loading in airway smooth muscle cells: a putative mechanism in the regulation of Ca2+ by cGMP.

    PubMed

    Bazán-Perkins, Blanca

    2012-03-01

    Ca(2+) and cGMP have opposite roles in many physiological processes likely due to a complex negative feedback regulation between them. Examples of opposite functions induced by Ca(2+) and cGMP are smooth muscle contraction and relaxation, respectively. A main Ca(2+) storage involved in contraction is sarcoplasmic reticulum (SR); nevertheless, the role of cGMP in the regulation of SR-Ca(2+) has not been completely understood. To evaluate this role, intracellular Ca(2+) concentration ([Ca(2+)]i) was determinated by a ratiometric method in isolated myocytes from bovine trachea incubated with Fura-2/AM. The release of Ca(2+) from SR induced by caffeine was transient, whereas caffeine withdrawal was followed by a [Ca(2+)]i undershoot. Caffeine-induced Ca(2+) transient peak and [Ca(2+)]i undershoot after caffeine were reproducible in the same cell. Dibutyryl cGMP (db-cGMP) blocked the [Ca(2+)]i undershoot and reduced the subsequent caffeine peak (SR-Ca(2+) loading). Both, the opening of SR channels with ryanodine (10 μM) and the blockade of SR-Ca(2+) ATPase with cyclopiazonic acid inhibited the [Ca(2+)]i undershoot as well as the SR-Ca(2+) loading. The addition of db-cGMP to ryanodine (10 μM) incubated cells partially restored the SR-Ca(2+) loading. Cyclic GMP enhanced [Ca(2+)]i undershoot induced by the blockade of ryanodine channels with 50 μM ryanodine. In conclusion, the reduction of SR-Ca(2+) content in airway smooth muscle induced by cGMP can be explained by the combination of SR-Ca(2+) loading and the simultaneous release of SR-Ca(2+). The reduction of SR-Ca(2+) content induced by cGMP might be a putative mechanism limiting releasable Ca(2+) in response to a particular stimulus.

  9. Biphasic contractions induced by milrinone at low temperature in ferret ventricular muscle: role of the sarcoplasmic reticulum and transmembrane calcium influx.

    PubMed

    Malecot, C O; Bers, D M; Katzung, B G

    1986-08-01

    The effects of milrinone were studied in ferret papillary muscle stimulated at various rates and temperatures from 23 degrees to 36 degrees C. In voltage-clamp experiments, 50 micrograms/ml (0.237 mM) milrinone induced a 2.1-fold increase in calcium current at 28 degrees or 36 degrees C. At 50 micrograms/ml, milrinone transiently increased contractility in all muscles at 28 degrees C, but its steady-state effect was either increased (+50%) or decreased (-24.7%) steady-state twitch amplitude. A negative inotropic effect always occurred below 27 degrees C. Milrinone decreased the total twitch duration and split the twitch into two components (P1 and P2) in the absence of any evidence of aberrant conduction. Increasing milrinone concentration from 50 to 300 micrograms/ml decreased P1 and increased P2. Ryanodine (100 mM) or caffeine (10 mM) suppressed P1. Contractions elicited after 30 seconds of rest were also biphasic in the presence of milrinone, but not in its absence. P2 of post-rest contraction was increased by caffeine or calcium (10 mM) and decreased by cobalt (2 mM) when drugs were applied at the beginning of the rest. Ryanodine and caffeine also suppressed P1 of post-rest contraction. The evidence suggests that P1 may be caused by Ca release from the sarcoplasmic reticulum and P2 by increased Ca influx during the action potential via the calcium channel. It is also suggested that P2 may be present under control conditions, but to a lesser extent, and masked by a large P1.

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

    PubMed

    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; Meissner, Gerhard

    2011-05-01

    In vitro, calmodulin (CaM) and S100A1 activate the skeletal muscle ryanodine receptor ion channel (RyR1) at submicromolar Ca(2+) concentrations, whereas at micromolar Ca(2+) 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 (Ryr1(D/D)) were viable and appeared normal. However, single RyR1 channel recordings from Ryr1(D/D) mice exhibited impaired activation by CaM and S100A1 and impaired CaCaM inhibition. Isolated flexor digitorum brevis muscle fibers from Ryr1(D/D) mice had depressed Ca(2+) transients when stimulated by a single action potential. However, during repetitive stimulation, the mutant fibers demonstrated greater relative summation of the Ca(2+) transients. Consistently, in vivo stimulation of tibialis anterior muscles in Ryr1(D/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, Ryr1(D/D) fibers exhibited slowed inactivation of sarcoplasmic reticulum Ca(2+) release flux, consistent with increased summation of the Ca(2+) transient and contractile force. Peak Ca(2+) release flux was suppressed at all voltages in voltage-clamped Ryr1(D/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 Ca(2+) release, providing new insights into CaM and S100A1 regulation of skeletal muscle excitation-contraction coupling.

  11. Effects of beta-escin and saponin on the transverse-tubular system and sarcoplasmic reticulum membranes of rat and toad skeletal muscle.

    PubMed

    Launikonis, B S; Stephenson, D G

    1999-05-01

    Mechanically skinned skeletal muscle fibres from rat and toad were exposed to the permeabilizing agents beta-escin and saponin. The effects of these agents on the sealed transverse tubular system (t-system) and sarcoplasmic reticulum (SR) were examined by looking at changes in the magnitude of the force responses to t-system depolarization, the time course of the fluorescence of fura-2 trapped in the sealed t-system, and changes in the magnitude of caffeine-induced contractures following SR loading with Ca2+ under defined conditions. In the presence of 2 microg ml-1 beta-escin and saponin, the response to t-system depolarization was not completely abolished, decreasing to a plateau, and a large proportion of fura-2 remained in the sealed t-system. At 10 microg ml-1, both agents abolished the ability of both rat and toad preparations to respond to t-system depolarization after 3 min of exposure, but a significant amount of fura-2 remained in sealed t-tubules even after exposure to 100 microg ml-1 beta-escin and saponin for 10 min. beta-Escin took longer than saponin to reduce the t-system depolarizations and fura-2 content of the sealed t-system to a similar level. The ability of the SR to load Ca2+ was reduced to a lower level after treatment with beta-escin than saponin. This direct effect on the SR occurred at much lower concentrations for rat (2 microg ml-1 beta-escin and 10 microg ml-1 saponin) than toad (10 microg ml-1 beta-escin and 150 microg ml-1 saponin). The reverse order in sensitivities to beta-escin and saponin of t-system and SR membranes indicates that the mechanisms of action of beta-escin and saponin are different in the two types of membrane. In conclusion, this study shows that: (1) beta-escin has a milder action on the surface membrane than saponin; (2) beta-escin is a more potent modifier of SR function; (3) simple permeabilization of membranes is not sufficient to explain the effects of beta-escin and saponin on muscle membranes; and (4) the t

  12. 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.

  13. Modulation of the Frequency of Spontaneous Sarcoplasmic Reticulum Ca2+ Release Events (Ca2+ Sparks) by Myoplasmic [Mg2+] in Frog Skeletal Muscle

    PubMed Central

    Lacampagne, Alain; Klein, Michael G.; Schneider, Martin F.

    1998-01-01

    The modulation by internal free [Mg2+] of spontaneous calcium release events (Ca2+ “sparks”) from the sarcoplasmic reticulum (SR) was studied in depolarized notched frog skeletal muscle fibers using a laser scanning confocal microscope in line-scan mode (x vs. t). Over the range of [Mg2+] from 0.13 to 1.86 mM, decreasing the [Mg2+] induced an increase in the frequency of calcium release events in proportion to [Mg2+]−1.6. The change of event frequency was not due to changes in [Mg-ATP] or [ATP]. Analysis of individual SR calcium release event properties showed that the variation in event frequency induced by the change of [Mg2+] was not accompanied by any changes in the spatiotemporal spread (i.e., spatial half width or temporal half duration) of Ca2+ sparks. The increase in event frequency also had no effect on the distribution of event amplitudes. Finally, the rise time of calcium sparks was independent of the [Mg2+], indicating that the open time of the SR channel or channels underlying spontaneous calcium release events was not altered by [Mg2+] over the range tested. These results suggest that in resting skeletal fibers, [Mg2+] modulates the SR calcium release channel opening frequency by modifying the average closed time of the channel without altering the open time. A kinetic reaction scheme consistent with our results and those of bilayer and SR vesicle experiments indicates that physiological levels of resting Mg2+ may inhibit channel opening by occupying the site for calcium activation of the SR calcium release channel. PMID:9450940

  14. Skeletal muscle

    USDA-ARS?s Scientific Manuscript database

    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...

  15. Ultrastructural study of muscles fibers in tick Hyalomma (Hyalomma) anatolicum anatolicum (Ixodoidea: Ixodidae).

    PubMed

    Bughdadi, Faisal A

    2010-09-01

    In the present study, ticks were obtained from a colony maintained at 28 degrees C and 75% relative humidity in at the Department of Biology, University College Umm Al-Qura University, Saudi Arabia and the Transmission Electron Microscope technique (TEM) was used to describes the ultrastructure and description of muscle of the of ixodid tick Hyalomma (Hyalomma) anatolicum anatolicum. The results showed that muscles of the unfed ticks Hyalomma (Hyalomma) anatolicum anatolicum in longitudinal sections are spindle-shaped to cylindrical muscle fibers. In the unfed nymph Hyalomma (Hyalomma) anatolicum anatolicum skeletal and visceral muscles are distinguished according to structure, function and position. These muscles include the capitulum, dorsoventral and leg oblique muscles. All muscle fibers are ensheathed (covered by sheath) in a sarcolemma. Their muscle fibers have striated pattern of successive sarcomeres whose thick myosin filaments are surrounded by orbitals of up to 12 thin actin filaments. The cytoplasm of the epidermal cell appears largely devoted with complicated microtubules present in parallel with long axis of adjacent muscle fibers. The cell membrane invaginates into tubular system extending deeply into the sarcoplasm and closely associated to cisternae of sarcoplasmic reticulum. The tubular system and sarcoplasmic reticulum forming two-membered (dyads) are considered to be the main route of calcium ions whose movement are synchronized with the motor impulse to control muscles contraction. In the sarcoplasm two types of muscle fibers are recognized according to thickness and density and mitochondrial size, distribution and population. Both skeletal and visceral muscles are invaginated by tracheoles and innervated by nerve axons containing synaptic vesicles. The actin and myosin filaments are slightly interrupted and the tubular system sarcoplasmic reticulum is well demonstrated.

  16. The SH3 and cysteine-rich domain 3 (Stac3) gene is important to growth, fiber composition, and calcium release from the sarcoplasmic reticulum in postnatal skeletal muscle.

    PubMed

    Cong, Xiaofei; Doering, Jonathan; Mazala, Davi A G; Chin, Eva R; Grange, Robert W; Jiang, Honglin

    2016-01-01

    The SH3 and cysteine-rich domain 3 (Stac3) gene is specifically expressed in the skeletal muscle. Stac3 knockout mice die perinatally. In this study, we determined the potential role of Stac3 in postnatal skeletal muscle growth, fiber composition, and contraction by generating conditional Stac3 knockout mice. We disrupted the Stac3 gene in 4-week-old male mice using the Flp-FRT and tamoxifen-inducible Cre-loxP systems. RT-qPCR and western blotting analyses of the limb muscles of target mice indicated that nearly all Stac3 mRNA and more than 70 % of STAC3 protein were deleted 4 weeks after tamoxifen injection. Postnatal Stac3 deletion inhibited body and limb muscle mass gains. Histological staining and gene expression analyses revealed that postnatal Stac3 deletion decreased the size of myofibers and increased the percentage of myofibers containing centralized nuclei, with no effect on the total myofiber number. Grip strength and grip time tests indicated that postnatal Stac3 deletion decreased limb muscle strength in mice. Muscle contractile tests revealed that postnatal Stac3 deletion reduced electrostimulation-induced but not the ryanodine receptor agonist caffeine-induced maximal force output in the limb muscles. Calcium imaging analysis of single flexor digitorum brevis myofibers indicated that postnatal Stac3 deletion reduced electrostimulation- but not caffeine-induced calcium release from the sarcoplasmic reticulum. This study demonstrates that STAC3 is important to myofiber hypertrophy, myofiber-type composition, contraction, and excitation-induced calcium release from the sarcoplasmic reticulum in the postnatal skeletal muscle.

  17. Calcium buffering properties of sarcoplasmic reticulum and calcium-induced Ca2+ release during the quasi-steady level of release in twitch fibers from frog skeletal muscle

    PubMed Central

    Fénelon, Karine; Lamboley, Cédric R.H.; Carrier, Nicole

    2012-01-01

    Experiments were performed to characterize the properties of the intrinsic Ca2+ buffers in the sarcoplasmic reticulum (SR) of cut fibers from frog twitch muscle. The concentrations of total and free calcium ions within the SR ([CaT]SR and [Ca2+]SR) were measured, respectively, with the EGTA/phenol red method and tetramethylmurexide (a low affinity Ca2+ indicator). Results indicate SR Ca2+ buffering was consistent with a single cooperative-binding component or a combination of a cooperative-binding component and a linear binding component accounting for 20% or less of the bound Ca2+. Under the assumption of a single cooperative-binding component, the most likely resting values of [Ca2+]SR and [CaT]SR are 0.67 and 17.1 mM, respectively, and the dissociation constant, Hill coefficient, and concentration of the Ca-binding sites are 0.78 mM, 3.0, and 44 mM, respectively. This information can be used to calculate a variable proportional to the Ca2+ permeability of the SR, namely d[CaT]SR/dt ÷ [Ca2+]SR (denoted release permeability), in experiments in which only [CaT]SR or [Ca2+]SR is measured. In response to a voltage-clamp step to −20 mV at 15°C, the release permeability reaches an early peak followed by a rapid decline to a quasi-steady level that lasts ∼50 ms, followed by a slower decline during which the release permeability decreases by at least threefold. During the quasi-steady level of release, the release amplitude is 3.3-fold greater than expected from voltage activation alone, a result consistent with the recruitment by Ca-induced Ca2+ release of 2.3 SR Ca2+ release channels neighboring each channel activated by its associated voltage sensor. Release permeability at −60 mV increases as [CaT]SR decreases from its resting physiological level to ∼0.1 of this level. This result argues against a release termination mechanism proposed in mammalian muscle fibers in which a luminal sensor of [Ca2+]SR inhibits release when [CaT]SR declines to a low level

  18. Role of calsequestrin evaluated from changes in free and total calcium concentrations in the sarcoplasmic reticulum of frog cut skeletal muscle fibres

    PubMed Central

    Pape, Paul C; Fénelon, Karine; Lamboley, Cédric R H; Stachura, Dorothy

    2007-01-01

    Calsequestrin is a large-capacity Ca-binding protein located in the terminal cisternae of sarcoplasmic reticulum (SR) suggesting a role as a buffer of the concentration of free Ca in the SR ([Ca2+]SR) serving to maintain the driving force for SR Ca2+ release. Essentially all of the functional studies on calsequestrin to date have been carried out on purified calsequestrin or on disrupted muscle preparations such as terminal cisternae vesicles. To obtain information about calsequestrin's properties during physiological SR Ca2+ release, experiments were carried out on frog cut skeletal muscle fibres using two optical methods. One – the EGTA–phenol red method – monitored the content of total Ca in the SR ([CaT]SR) and the other used the low affinity Ca indicator tetramethylmurexide (TMX) to monitor the concentration of free Ca in the SR. Both methods relied on a large concentration of the Ca buffer EGTA (20 mm), in the latter case to greatly reduce the increase in myoplasmic [Ca2+] caused by SR Ca2+ release thereby almost eliminating the myoplasmic component of the TMX signal. By releasing almost all of the SR Ca, these optical signals provided information about [CaT]SR versus [Ca2+]SR as [Ca2+]SR varied from its resting level ([Ca2+]SR,R) to near zero. Since almost all of the Ca in the SR is bound to calsequestrin, this information closely resembles the binding curve of the Ca–calsequestrin reaction. Calcium binding to calsequestrin was found to be cooperative (estimated Hill coefficient = 2.95) and to have a very high capacity (at the start of Ca2+ release, 23 times more Ca was estimated to initiate from calsequestrin as opposed to the pool of free Ca in the SR). The latter result contrasts with an earlier report that only ∼25% of released Ca2+ comes from calsequestrin and ∼75% comes from the free pool. The value of [Ca2+]SR,R was close to the KD for calsequestrin, which has a value near 1 mm in in vitro studies. Other evidence indicates that [Ca2+]SR

  19. Calcium buffering properties of sarcoplasmic reticulum and calcium-induced Ca(2+) release during the quasi-steady level of release in twitch fibers from frog skeletal muscle.

    PubMed

    Fénelon, Karine; Lamboley, Cédric R H; Carrier, Nicole; Pape, Paul C

    2012-10-01

    Experiments were performed to characterize the properties of the intrinsic Ca(2+) buffers in the sarcoplasmic reticulum (SR) of cut fibers from frog twitch muscle. The concentrations of total and free calcium ions within the SR ([Ca(T)](SR) and [Ca(2+)](SR)) were measured, respectively, with the EGTA/phenol red method and tetramethylmurexide (a low affinity Ca(2+) indicator). Results indicate SR Ca(2+) buffering was consistent with a single cooperative-binding component or a combination of a cooperative-binding component and a linear binding component accounting for 20% or less of the bound Ca(2+). Under the assumption of a single cooperative-binding component, the most likely resting values of [Ca(2+)](SR) and [Ca(T)](SR) are 0.67 and 17.1 mM, respectively, and the dissociation constant, Hill coefficient, and concentration of the Ca-binding sites are 0.78 mM, 3.0, and 44 mM, respectively. This information can be used to calculate a variable proportional to the Ca(2+) permeability of the SR, namely d[Ca(T)](SR)/dt ÷ [Ca(2+)](SR) (denoted release permeability), in experiments in which only [Ca(T)](SR) or [Ca(2+)](SR) is measured. In response to a voltage-clamp step to -20 mV at 15°C, the release permeability reaches an early peak followed by a rapid decline to a quasi-steady level that lasts ~50 ms, followed by a slower decline during which the release permeability decreases by at least threefold. During the quasi-steady level of release, the release amplitude is 3.3-fold greater than expected from voltage activation alone, a result consistent with the recruitment by Ca-induced Ca(2+) release of 2.3 SR Ca(2+) release channels neighboring each channel activated by its associated voltage sensor. Release permeability at -60 mV increases as [Ca(T)](SR) decreases from its resting physiological level to ~0.1 of this level. This result argues against a release termination mechanism proposed in mammalian muscle fibers in which a luminal sensor of [Ca(2+)](SR) inhibits

  20. Role of malignant hyperthermia domain in the regulation of Ca2+ release channel (ryanodine receptor) of skeletal muscle sarcoplasmic reticulum.

    PubMed

    Zorzato, F; Menegazzi, P; Treves, S; Ronjat, M

    1996-09-13

    A fusion protein encompassing Gly341 of the skeletal muscle ryanodine receptor was used to raise monoclonal antibodies; epitope mapping demonstrates that monoclonal antibody 419 (mAb419) reacts with a sequence a few residues upstream from Gly341. The mAb419 was then used to probe ryanodine receptor (RYR) functions. Our results show that upon incubation of triads vesicles with mAb419 the Ca2+-induced Ca2+ release rate at pCa 8 was increased. Equilibrium evaluation of [3H]ryanodine binding at different [Ca2+] indicates that mAb419 shifted the half-maximal [Ca2+] for stimulation of ryanodine binding to lower value (0.1 versus 1.2 microM). Such functional effects may be due to a direct action of the Ab on the Ca2+ binding domain of the RYR or to the perturbation by the Ab of the intramolecular interaction between the immunopositive region and regulatory domain of the RYR. The latter hypothesis was tested directly using the optical biosensor BIAcore (Pharmacia Biotech Inc.): we show that the immunopositive RYR polypeptide is able to interact with the native RYR complex. Ligand overlays with immunopositive digoxigenin-RYR fusion protein indicate that such an interaction might occur with a calmodulin binding domain (defined by residues 3010-3225) and with a polypeptide defined by residues 799-1172. In conclusion our results suggest that the stimulation by the mAb419 of the RYR channel activity is due to the perturbation of an intramolecular interaction between the immunopositive polypeptide and a Ca2+ regulatory site probably corresponding to a calmodulin binding domain.

  1. Effects of free oxygen radicals on Ca2+ release mechanisms in the sarcoplasmic reticulum of scallop (Pecten jacobaeus) adductor muscle.

    PubMed

    Burlando, B; Viarengo, A; Pertica, M; Ponzano, E; Orunesu, M

    1997-08-01

    In vitro oxyradical effects on SR Ca2+ regulation were studied by using a SR-containing cell-free preparation from scallop (Pecten jacobaeus) adductor muscle. Ca2+ variations were fluorimetrically detected after incubation with Fluo-3 in the presence of ATP. Exposure to Fe3+/ascorbate produced dose-dependent Ca2+ release from SR vesicles, eventually leading to massive Ca2+ loss. Exposure to hypoxanthine/xanthine oxidase also caused Ca2+ release but at a much slower rate. Pre-incubations with catalase or with the hydroxyl radical scavenger KMBA led to a significant decrease in the Fe3+/ascorbate-induced Ca2+ release rate and to a delay of massive Ca2+ loss. Pre-incubations with GSH or DTT strongly reduced the Ca2+ release caused by Fe3+/ascorbate and, moreover, they prevented massive Ca2+ loss from SR vesicles. Addition of GSH or DTT after Fe3+/ascorbate promptly reduced the Ca2+ release rate and delayed massive Ca2+ release. Pre-incubation with the SR Ca2+ channel blocker ruthenium red strongly reduced the Ca2+ release caused by Fe3+/ascorbate, and also prevented massive Ca2+ loss. In the presence of ruthenium red, Fe3+/ascorbate treatments followed by Ca2+ addition revealed that Ca2+ uptake inhibition was slower than Ca2+ release. Taken together, data showed that free radicals and, in particular, hydroxyl radicals, affected the scallop SR Ca2+ regulation. This mainly occurred through Ca2+ channel opening, most likely triggered by sulfhydryl oxidation, which eventually led to massive Ca2+ release from SR vesicles. The demonstration of a specific effect of oxyradicals on SR Ca2+ channels is in line with their possible involvement in cell signaling.

  2. Ca2+ homeostasis and fast-type sarcoplasmic reticulum Ca(2+)-ATPase expression in L6 muscle cells. Role of thyroid hormone.

    PubMed

    Muller, A; van Hardeveld, C; Simonides, W S; van Rijn, J

    1992-05-01

    The effect of thyroid hormone (L-tri-iodothyronine; T3) on the cytosolic free Ca2+ concentration ([Ca2+]i) in L6 myotubes was studied at rest and during activation to explore the possible mediating role of [Ca2+]i in the T3-induced net synthesis of fast-type sarcoplasmic reticulum (SR) Ca(2+)-ATPase. The mean [Ca2+]i at rest was approx. 115 nM in myoblasts, control myotubes and T3-treated myotubes. Therefore it is unlikely that the T3-induced elevation of Ca(2+)-ATPase levels is mediated by [Ca2+]i changes. To investigate the influence of the 4-fold higher Ca(2+)-ATPase levels in T3-treated myotubes (compared with controls) on [Ca2+]i, interventions with caffeine (10 mM) and a high extracellular K+ concentration ([K+]o) (30 mM) were applied which initially mobilize Ca2+ predominantly from the SR. The results showed a lower (caffeine) or not significantly different (high [K+]o) increase in [Ca2+]i in T3-treated myotubes compared with controls. No rise in [Ca2+]i was found in myoblasts with caffeine or high [K+]o. The role of [Ca2+]i in the regulation of Ca(2+)-ATPase levels was investigated by varying [Ca2+]i through exposure of cells to different concentrations of extracellular Ca2+ (0.2-1.8 mM) and ionomycin (0.1-0.25 microM). At subnormal [Ca2+]i (55 nM) the T3-induced net synthesis of Ca(2+)-ATPase was virtually abolished, and at supranormal [Ca2+]i (195 nM) it was greatly depressed. Intermediate stimulation of net Ca(2+)-ATPase synthesis was found at [Ca2+]i of 95 and 165 nM, with an optimum at approx. 125 nM. Similar but less pronounced effects were found for the basal Ca(2+)-ATPase levels. In contracting primary rat myotubes, Ca(2+)-ATPase levels were significantly lower than in tetrodotoxin-arrested myotubes. The same results were obtained in the presence of T3. Since the mean [Ca2+]i in contracting cells is higher than in resting cells, these data agree with those obtained in the L6 cells with ionomycin. A major conclusion of this study is the existence of

  3. 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.

  4. 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 Central

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

    2011-01-01

    Abstract 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. PMID:21486805

  5. Your Muscles

    MedlinePlus

    ... of the heart because it controls the heartbeat. Skeletal Muscle Now, let's talk about the kind of muscle ... soccer ball into the goal. These are your skeletal muscles — sometimes called striated (say: STRY-ay-tud) muscle ...

  6. Novel targets for treating heart and muscle disease: stabilizing ryanodine receptors and preventing intracellular calcium leak.

    PubMed

    Lehnart, Stephan E

    2007-04-01

    Ryanodine receptors (RyRs) function as intracellular Ca(2+) release channels on the endoplasmic and sarcoplasmic reticulum membranes. In striated muscles, Ca(2+) release through RyRs controls muscle excitation-contraction coupling. RyR channel function is regulated by a cytoplasmic scaffold domain that forms a macromolecular signaling complex including calstabin (formerly known as FK506-binding protein), calmodulin, phosphodiesterase, kinase and phosphatase proteins. An increasing number of genetic and acquired diseases has been associated with intracellular Ca(2+) leak. In heart failure, for instance, the RyR complex becomes altered, resulting in chronic channel dysfunction and chronic sarcoplasmic reticulum Ca(2+) leak. Recently, the efficacy of novel Ca(2+) release channel-stabilizing drugs has been demonstrated in cardiac and skeletal muscle disease models.

  7. In vivo aging of rat skeletal muscle sarcoplasmic reticulum Ca-ATPase. Chemical analysis and quantitative simulation by exposure to low levels of peroxyl radicals.

    PubMed

    Viner, R I; Ferrington, D A; Aced, G I; Miller-Schlyer, M; Bigelow, D J; Schöneich, C

    1997-10-23

    Sarcoplasmic reticulum (SR) Ca-ATPase of young adult (5 months) and aged (28 months) Fischer 344 male rat skeletal muscle was analyzed for posttranslational modifications as a result of biological aging and their potential functional consequences. The significant differences in the amino acid composition were a 6.8% lower content of sulfhydryl groups and a ca. 4% lower content of Arg residues of the Ca-ATPase from old as compared to young rats. Based on a total of 24 Cys residues the difference in protein thiols corresponds to a loss of 1.5 mol Cys/mol Ca-ATPase as a result of in vivo aging. The loss of Cys residues was not accompanied by a loss of enzyme activity though the 'aged' Ca-ATPase was more sensitive to heat inactivation, aggregation, and tryptic digestion. A comparison of the total sulfhydryl content of all SR proteins present revealed a 13% lower amount for SR vesicles isolated from aged rats. Compared to the alterations of Cys and Arg, there was only a slight and probably physiologically insignificant increase of protein carbonyls with aging, i.e. from 0.32 to 0.46 mol carbonyl groups per mol of Ca-ATPase. When SR vesicles from young rats were exposed to AAPH-derived peroxyl radicals, there was a loss of ca. 1.38 x 10(-4) M total SR sulfhydryl groups per 4 mg SR protein/ml (corresponding to ca. 25%) and a loss of 9.6 x 10(-5) M Ca-ATPase sulfhydryl groups (corresponding to ca. 31%) per 1.6 x 10(-5) M initiating peroxyl radicals, indicating that the stoichiometry of sulfhydryl oxidation was > or = 6 oxidized thiols per initiating AAPH-derived peroxyl radical. Besides Cys, the exposure to AAPH-derived radicals caused a slight loss of Ca-ATPase Arg, Met, and Ser residues. Most importantly, the SR Ca-ATPase exposed to this low concentration of peroxyl radicals displayed physical and functional properties quantitatively comparable to those of SR Ca-ATPase isolated from aged rats, i.e. no immediate loss of activity, increased susceptibility to heat

  8. Changes in sarcoplasmic metabolite concentrations and pH associated with the catch contraction and relaxation of the anterior byssus retractor muscle of Mytilus edulis measured by phosphorus-31 nuclear magnetic resonance.

    PubMed

    Ishii, N; Mitsumori, F; Takahashi, K

    1991-06-01

    The sarcoplasmic concentrations of phosphorus metabolites and pH (pHin) were measured in the anterior byssus retractor muscle (ABRM) of Mytilus edulis by 31P nuclear magnetic resonance spectroscopy. During an active contraction induced by 10(-3) acetylcholine, the concentration of arginine phosphate ([Arg-P]in) decreased from the resting value of 7.47 +/- 0.26 (mean +/- SE, n = 8) to 6.67 +/- 0.29 (n = 6) mumol g-1, and that of inorganic phosphate (Pi) consistently increased from 0.84 +/- 0.06 (n = 7) to 1.61 +/- 0.12 (n = 5) mumol g-1. In the 'catch' state following the active contraction, these concentrations were close to their resting levels, indicating that the catch is an inactive state. 5-hydroxytryptamine caused a rapid relaxation of the catch, which was associated with a slight decrease in [Arg-P]in and an increase in pHin by ca 0.2 units. The sarcoplasmic concentration of ATP (mean, 1.6 mumol g-1) did not change throughout the contraction-relaxation cycle.

  9. Spectroscopic and ITC study of the conformational change upon Ca{sup 2+}-binding in TnC C-lobe and TnI peptide complex from Akazara scallop striated muscle

    SciTech Connect

    Yumoto, Fumiaki; Tanaka, Hiroyuki; Nagata, Koji; Miyauchi, Yumiko; Miyakawa, Takuya; Ojima, Takao; Tanokura, Masaru

    2008-04-25

    Akazara scallop (Chlamys nipponensis akazara) troponin C (TnC) of striated adductor muscle binds only one Ca{sup 2+} ion at the C-terminal EF-hand motif (Site IV), but it works as the Ca{sup 2+}-dependent regulator in adductor muscle contraction. In addition, the scallop troponin (Tn) has been thought to regulate muscle contraction via activating mechanisms that involve the region spanning from the TnC C-lobe (C-lobe) binding site to the inhibitory region of the TnI, and no alternative binding of the TnI C-terminal region to TnC because of no similarity between second TnC-binding regions of vertebrate and the scallop TnIs. To clarify the Ca{sup 2+}-regulatory mechanism of muscle contraction by scallop Tn, we have analyzed the Ca{sup 2+}-binding properties of the complex of TnC C-lobe and TnI peptide, and their interaction using isothermal titration microcalorimetry, nuclear magnetic resonance, circular dichroism, and gel filtration chromatography. The results showed that single Ca{sup 2+}-binding to the Site IV leads to a structural transition not only in Site IV but also Site III through the structural network in the C-lobe of scallop TnC. We therefore assumed that the effect of Ca{sup 2+}-binding must lead to a change in the interaction mode between the C-lobe of TnC and the TnI peptide. The change should be the first event of the transmission of Ca{sup 2+} signal to TnI in Tn ternary complex.

  10. Ryanodine receptors in smooth muscle.

    PubMed

    Guerrero-Hernández, Agustín; Gómez-Viquez, Leticia; Guerrero-Serna, Guadalupe; Rueda, Angélica

    2002-07-01

    The sarcoplasmic reticulum (SR) of smooth muscle is endowed with two different types of Ca2+ release channels, i.e. inositol 1,4,5-trisphosphate receptors (IP3Rs) and ryanodine receptors (RyRs). In general, both release channels mobilize Ca2+ from the same internal store in smooth muscle. While the importance of IP3Rs in agonist-induced contraction is well established, the role of RyRs in excitation-contraction coupling of smooth muscle is not clear. The participation of smooth muscle RyRs in the amplification of Ca2+ transients induced by either opening of Ca2+-permeable channels or IP3-triggered Ca2+ release has been studied. The efficacy of both processes to activate RyRs by calcium-induced calcium release (CICR) is highly variable and not widely present in smooth muscle. Although RyRs in smooth muscle generate Ca2+ sparks that are similar to those observed in striated muscles, the contribution of these local Ca2+ events to depolarization-induced global rise in [Ca2+]i is rather limited. Recent data suggest that RyRs are involved in regulating the luminal [Ca2+] of SR and also in smooth muscle relaxation. This review summarizes studies that were carried out mainly in muscle strips or in freshly isolated myocytes, and that were aimed to determine the physiological role of RyRs in smooth muscle.

  11. The microscopic and ultramicroscopic changes in the skeletal muscles, caused by heavy metal salts

    PubMed Central

    Tymoshenko, Alexey; Tkach, Gennadii; Sikora, Vitalii; Bumeister, Valentina; Shpetnyi, Ihor; Lyndin, Mykola; Maksymova, Olena; Maslenko, Anna

    2016-01-01

    Purpose The article is devoted to study the structural changes in the skeletal muscles caused by heavy metal salts. Materials and methods The study was conducted on 72 mature male rats. The experimental groups were given to drink water with combinations of heavy metal salts for one, two and three months. This type of water is typical for the water basins in the northern districts of the Sumy region. The study of morphological changes in the striated muscles was concluded using light and scanning electron microscopy. Results The data analysis revealed that a prolonged duration of negative factor could intensify sclerotic and edematous processes. The structure of muscle fibers was destroyed, nuclei were deformed and placed irregularly, and many petechial hemorrhages occurred. Besides, cross-striation was irregular, I and A bands were deformed and destroyed, H band was hardly visualized. The inner mitochondrial membrane and cristae become deformed. The symplastic nuclei were placed irregularly within sarcoplasm. Besides, they were swollen. Against swollen and enlarged symplastic nuclei, pyknotic nuclei were also found. The structures of sarcoplasmic reticulum were mainly dilated with deformed and ruptured areas. Conclusion Our study approves that high concentrations of heavy metal salts have a destructive influence on the skeletal striated muscles. PMID:28386464

  12. The evolutionary origin of bilaterian smooth and striated myocytes

    PubMed Central

    Brunet, Thibaut; Fischer, Antje HL; Steinmetz, Patrick RH; Lauri, Antonella; Bertucci, Paola; Arendt, Detlev

    2016-01-01

    The dichotomy between smooth and striated myocytes is fundamental for bilaterian musculature, but its evolutionary origin is unsolved. In particular, interrelationships of visceral smooth muscles remain unclear. Absent in fly and nematode, they have not yet been characterized molecularly outside vertebrates. Here, we characterize expression profile, ultrastructure, contractility and innervation of the musculature in the marine annelid Platynereis dumerilii and identify smooth muscles around the midgut, hindgut and heart that resemble their vertebrate counterparts in molecular fingerprint, contraction speed and nervous control. Our data suggest that both visceral smooth and somatic striated myocytes were present in the protostome-deuterostome ancestor and that smooth myocytes later co-opted the striated contractile module repeatedly – for example, in vertebrate heart evolution. During these smooth-to-striated myocyte conversions, the core regulatory complex of transcription factors conveying myocyte identity remained unchanged, reflecting a general principle in cell type evolution. DOI: http://dx.doi.org/10.7554/eLife.19607.001 PMID:27906129

  13. Transport of the alpha subunit of the voltage gated L-type calcium channel through the sarcoplasmic reticulum occurs prior to localization to triads and requires the beta subunit but not Stac3 in skeletal muscles.

    PubMed

    Linsley, Jeremy W; Hsu, I-Uen; Wang, Wenjia; Kuwada, John Y

    2017-09-01

    Contraction of skeletal muscle is initiated by excitation-contraction (EC) coupling during which membrane voltage is transduced to intracellular Ca(2+) release. EC coupling requires L-type voltage gated Ca2+ channels (the dihydropyridine receptor or DHPR) located at triads, which are junctions between the transverse (T) tubule and sarcoplasmic reticulum (SR) membranes, that sense membrane depolarization in the T tubule membrane. Reduced EC coupling is associated with ageing, and disruptions of EC coupling result in congenital myopathies for which there are few therapies. The precise localization of DHPRs to triads is critical for EC coupling, yet trafficking of the DHPR to triads is not well understood. Using dynamic imaging of zebrafish muscle fibers, we find that DHPR is transported along the longitudinal SR in a microtubule-independent mechanism. Furthermore, transport of DHPR in the SR membrane is differentially affected in null mutants of Stac3 or DHPRβ, two essential components of EC coupling. These findings reveal previously unappreciated features of DHPR motility within the SR prior to assembly at triads. © 2017 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

  14. Muscle weakness in Ryr1I4895T/WT knock-in mice as a result of reduced ryanodine receptor Ca2+ ion permeation and release from the sarcoplasmic reticulum

    PubMed Central

    Loy, Ryan E.; Orynbayev, Murat; Xu, Le; Andronache, Zoita; Apostol, Simona; Zvaritch, Elena; MacLennan, David H.; Meissner, Gerhard; Melzer, Werner

    2011-01-01

    The type 1 isoform of the ryanodine receptor (RYR1) is the Ca2+ release channel of the sarcoplasmic reticulum (SR) that is activated during skeletal muscle excitation–contraction (EC) coupling. Mutations in the RYR1 gene cause several rare inherited skeletal muscle disorders, including malignant hyperthermia and central core disease (CCD). The human RYR1I4898T mutation is one of the most common CCD mutations. To elucidate the mechanism by which RYR1 function is altered by this mutation, we characterized in vivo muscle strength, EC coupling, SR Ca2+ content, and RYR1 Ca2+ release channel function using adult heterozygous Ryr1I4895T/+ knock-in mice (IT/+). Compared with age-matched wild-type (WT) mice, IT/+ mice exhibited significantly reduced upper body and grip strength. In spite of normal total SR Ca2+ content, both electrically evoked and 4-chloro-m-cresol–induced Ca2+ release were significantly reduced and slowed in single intact flexor digitorum brevis fibers isolated from 4–6-mo-old IT/+ mice. The sensitivity of the SR Ca2+ release mechanism to activation was not enhanced in fibers of IT/+ mice. Single-channel measurements of purified recombinant channels incorporated in planar lipid bilayers revealed that Ca2+ permeation was abolished for homotetrameric IT channels and significantly reduced for heterotetrameric WT:IT channels. Collectively, these findings indicate that in vivo muscle weakness observed in IT/+ knock-in mice arises from a reduction in the magnitude and rate of RYR1 Ca2+ release during EC coupling that results from the mutation producing a dominant-negative suppression of RYR1 channel Ca2+ ion permeation. PMID:21149547

  15. Muscle weakness in Ryr1I4895T/WT knock-in mice as a result of reduced ryanodine receptor Ca2+ ion permeation and release from the sarcoplasmic reticulum.

    PubMed

    Loy, Ryan E; Orynbayev, Murat; Xu, Le; Andronache, Zoita; Apostol, Simona; Zvaritch, Elena; MacLennan, David H; Meissner, Gerhard; Melzer, Werner; Dirksen, Robert T

    2011-01-01

    The type 1 isoform of the ryanodine receptor (RYR1) is the Ca(2+) release channel of the sarcoplasmic reticulum (SR) that is activated during skeletal muscle excitation-contraction (EC) coupling. Mutations in the RYR1 gene cause several rare inherited skeletal muscle disorders, including malignant hyperthermia and central core disease (CCD). The human RYR1(I4898T) mutation is one of the most common CCD mutations. To elucidate the mechanism by which RYR1 function is altered by this mutation, we characterized in vivo muscle strength, EC coupling, SR Ca(2+) content, and RYR1 Ca(2+) release channel function using adult heterozygous Ryr1(I4895T/+) knock-in mice (IT/+). Compared with age-matched wild-type (WT) mice, IT/+ mice exhibited significantly reduced upper body and grip strength. In spite of normal total SR Ca(2+) content, both electrically evoked and 4-chloro-m-cresol-induced Ca(2+) release were significantly reduced and slowed in single intact flexor digitorum brevis fibers isolated from 4-6-mo-old IT/+ mice. The sensitivity of the SR Ca(2+) release mechanism to activation was not enhanced in fibers of IT/+ mice. Single-channel measurements of purified recombinant channels incorporated in planar lipid bilayers revealed that Ca(2+) permeation was abolished for homotetrameric IT channels and significantly reduced for heterotetrameric WT:IT channels. Collectively, these findings indicate that in vivo muscle weakness observed in IT/+ knock-in mice arises from a reduction in the magnitude and rate of RYR1 Ca(2+) release during EC coupling that results from the mutation producing a dominant-negative suppression of RYR1 channel Ca(2+) ion permeation.

  16. Oxidation in HiOx-packaged pork Longissimus muscle predisposes myofibrillar and sarcoplasmic proteins to N-nitrosamine formation in nitrite-curing solution.

    PubMed

    Yang, Hua; Meng, Peipei; Xiong, Youling L; Ma, Lizhen; Wang, Changlu; Zhu, Yingchun

    2013-11-01

    The effect of meat protein in situ oxidation on the formation of N-nitrosodiethylamine (NDEA) was investigated. Fresh minced pork was untreated (Con) or treated with 700mg/kg α-tocopherol (Toc) or 300mg/kg tea polyphenols (PPE), packaged in a HiOx atmosphere (78.8% O2, 18.8% CO2, 2.4% N2), then stored at 2±1°C for up to 10days. Crude myofibrillar (MP) or sarcoplasmic (SP) protein (20mg/mL) extracted from stored meat was reacted with 43μM sodium nitrite at 80°C for 1h. Lipid oxidation was totally inhibited in PPE pork but increased in Con and Toc samples after 10days. There was significant protein oxidation (losses of sulfhydryls, formation of protein carbonyls) in both MP and SP in all samples during storage. However, the Con group suffered more extensive protein oxidation than Toc and PPE and produced more NDEA (P<0.05), indicating that protein oxidation promoted nitrosation.

  17. Attachment of rattlesnake venom myotoxin a to sarcoplasmic reticulum: peroxidase conjugated method.

    PubMed Central

    Tu, A. T.; Morita, M.

    1983-01-01

    Myotoxin a is a muscle-damaging toxin isolated from the venom of Crotalus atrox (western diamondback rattlesnake) and is composed of 42 amino acid residues. Earlier electron microscopic observation indicated that the toxin causes extensive swelling of the sarcoplasmic reticulum followed by disorganization of the sarcomers. In the present paper we describe the evidence for the attachment of peroxidase-conjugated myotoxin a to the membrane of sarcoplasmic reticulum of human muscle. It is thus suggested that the attachment of the toxin to the sarcoplasmic reticulum and the subsequent swelling are the first steps in myonecrosis induced by myotoxin a. Images Fig. 3 Fig. 4 Fig. 5 PMID:6661395

  18. The Time Course of the Loss and Recovery of Contracture Ability in Frog Striated Muscle Following Exposure to Ca-Free Solutions

    PubMed Central

    Milligan, J. V.

    1965-01-01

    Using area under the contracture curve to quantitate contractures, the diffusion coefficient of calcium ions within the frog toe muscle during washout in a calcium-free solution and subsequent recovery after reintroduction of calcium to the bathing solution was calculated to be about 2 x 10-6 cm2/sec. The diffusion coefficient measured during washout was found to be independent of temperature or initial calcium ion concentration. During recovery it was found to decrease if the temperature was lowered. This was likely due to the repolarization occurring after the depolarizing effect of the calcium-free solution. The relation between contracture area and [Ca]o was found to be useful over a wider range than that between maximum tension and [Ca]o. The normalized contracture areas were larger at lower calcium concentrations if the contractures were produced with cold potassium solutions or if NO3 replaced Cl in the bathing solutions. Decreasing the potassium concentration of the contracture solution to 50 mM from 115 mM did not change the relation between [Ca]o and the normalized area. If the K concentration of the bathing solution was increased, the areas were decreased at lower concentrations of Ca. PMID:14324991

  19. UNC-89 (obscurin) binds to MEL-26, a BTB-domain protein, and affects the function of MEI-1 (katanin) in striated muscle of Caenorhabditis elegans.

    PubMed

    Wilson, Kristy J; Qadota, Hiroshi; Mains, Paul E; Benian, Guy M

    2012-07-01

    The ubiquitin proteasome system is involved in degradation of old or damaged sarcomeric proteins. Most E3 ubiquitin ligases are associated with cullins, which function as scaffolds for assembly of the protein degradation machinery. Cullin 3 uses an adaptor to link to substrates; in Caenorhabditis elegans, one of these adaptors is the BTB-domain protein MEL-26 (maternal effect lethal). Here we show that MEL-26 interacts with the giant sarcomeric protein UNC-89 (obscurin). MEL-26 and UNC-89 partially colocalize at sarcomeric M-lines. Loss of function or gain of function of mel-26 results in disorganization of myosin thick filaments similar to that found in unc-89 mutants. It had been reported that in early C. elegans embryos, a target of the CUL-3/MEL-26 ubiquitylation complex is the microtubule-severing enzyme katanin (MEI-1). Loss of function or gain of function of mei-1 also results in disorganization of thick filaments similar to unc-89 mutants. Genetic data indicate that at least some of the mel-26 loss-of-function phenotype in muscle can be attributed to increased microtubule-severing activity of MEI-1. The level of MEI-1 protein is reduced in an unc-89 mutant, suggesting that the normal role of UNC-89 is to inhibit the CUL-3/MEL-26 complex toward MEI-1.

  20. UNC-89 (obscurin) binds to MEL-26, a BTB-domain protein, and affects the function of MEI-1 (katanin) in striated muscle of Caenorhabditis elegans

    PubMed Central

    Wilson, Kristy J.; Qadota, Hiroshi; Mains, Paul E.; Benian, Guy M.

    2012-01-01

    The ubiquitin proteasome system is involved in degradation of old or damaged sarcomeric proteins. Most E3 ubiquitin ligases are associated with cullins, which function as scaffolds for assembly of the protein degradation machinery. Cullin 3 uses an adaptor to link to substrates; in Caenorhabditis elegans, one of these adaptors is the BTB-domain protein MEL-26 (maternal effect lethal). Here we show that MEL-26 interacts with the giant sarcomeric protein UNC-89 (obscurin). MEL-26 and UNC-89 partially colocalize at sarcomeric M-lines. Loss of function or gain of function of mel-26 results in disorganization of myosin thick filaments similar to that found in unc-89 mutants. It had been reported that in early C. elegans embryos, a target of the CUL-3/MEL-26 ubiquitylation complex is the microtubule-severing enzyme katanin (MEI-1). Loss of function or gain of function of mei-1 also results in disorganization of thick filaments similar to unc-89 mutants. Genetic data indicate that at least some of the mel-26 loss-of-function phenotype in muscle can be attributed to increased microtubule-severing activity of MEI-1. The level of MEI-1 protein is reduced in an unc-89 mutant, suggesting that the normal role of UNC-89 is to inhibit the CUL-3/MEL-26 complex toward MEI-1. PMID:22621901

  1. Single-channel properties of the sarcoplasmic reticulum calcium-release channel in slow- and fast-twitch muscles of Rhesus monkeys.

    PubMed

    Bastide, B; Mounier, Y

    1998-08-01

    RyR1 is the main isoform of ryanodine receptor expressed in fast- and slow-twitch mammalian skeletal muscles although differences in Ca2+-release kinetics and properties have been reported. Single-channel measurements reveal that a large proportion (82%) of Ca2+-release channels measured in slow-twitch muscle preparations have properties similar to those of the Ca2+-release channels of fast-twitch preparations, i.e. the same conductance, an identical sensitivity to caffeine and a bell-shaped Ca2+ activation curve for pCa (-log10[Ca2+]) 7 to 3. A low proportion (18%) of Ca2+-release channels observed in preparations from slow-twitch muscles were characterized by a very high activity level. These channels were not inhibited at a millimolar concentration of Ca2+. Our data suggest that the different properties of Ca2+ release in slow- and fast-twitch muscles might not be related to intrinsic properties of the Ca2+-release channels of each type of muscle but rather to the co-expression of two isoforms of ryanodine receptor and the lower amount of Ca2+-release channels expressed in slow- than in fast-twitch muscles.

  2. Effects of Partial Sarcoplasmic Reticulum Calcium Depletion on Calcium Release in Frog Cut Muscle Fibers Equilibrated with 20 mM EGTA

    PubMed Central

    Pape, Paul C.; Jong, De-Shien; Knox Chandler, W.

    1998-01-01

    Resting sarcoplasmic reticulum (SR) Ca content ([CaSR]R) was varied in cut fibers equilibrated with an internal solution that contained 20 mM EGTA and 0–1.76 mM Ca. SR Ca release and [CaSR]R were measured with the EGTA–phenol red method (Pape et al. 1995. J. Gen. Physiol. 106:259–336). After an action potential, the fractional amount of Ca released from the SR increased from 0.17 to 0.50 when [CaSR]R was reduced from 1,200 to 140 μM. This increase was associated with a prolongation of release (final time constant, from 1–2 to 10–15 ms) and of the action potential (by 1–2 ms). Similar changes in release were observed with brief stimulations to −20 mV in voltage-clamped fibers, in which charge movement (Qcm) could be measured. The peak values of Qcm and the fractional rate of SR Ca release, as well as their ON time courses, were little affected by reducing [CaSR]R from 1,200 to 140 μM. After repolarization, however, the OFF time courses of Qcm and the rate of SR Ca release were slowed by factors of 1.5–1.7 and 6.5, respectively. These and other results suggest that, after action potential stimulation of fibers in normal physiological condition, the increase in myoplasmic free [Ca] that accompanies SR Ca release exerts three negative feedback effects that tend to reduce additional release: (a) the action potential is shortened by current through Ca-activated potassium channels in the surface and/or tubular membranes; (b) the OFF kinetics of Qcm is accelerated; and (c) Ca inactivation of Ca release is increased. Some of these effects of Ca on an SR Ca channel or its voltage sensor appear to be regulated by the value of [Ca] within 22 nm of the mouth of the channel. PMID:9725889

  3. Reduction of calcium inactivation of sarcoplasmic reticulum calcium release by fura-2 in voltage-clamped cut twitch fibers from frog muscle

    PubMed Central

    1993-01-01

    Cut fibers from Rana temporaria and Rana pipiens (striation spacing, 3.9-4.2 microns) were mounted in a double Vaseline-gap chamber and studied at 14 degrees C. The Ca indicator purpurate-3,3' diacetic acid (PDAA) was introduced into the end pools and allowed to diffuse into the optical recording site. When the concentration at the site exceeded 2 mM, step depolarizations to 10 mV were applied and the [Ca] transient measured with PDAA was used to estimate Ca release from the sarcoplasmic reticulum (SR) (Baylor, S. M., W. K. Chandler, and M. W. Marshall. 1983. Journal of Physiology. 344:625-666). With depolarization, the rate of SR Ca release increased to an early peak and then rapidly decreased several-fold to a quasi-steady level. The total amount of Ca released from the SR at the time of peak rate of release appeared to be independent of SR Ca content, consistent with the idea that a single activated channel might pass, on average, a fixed number of ions, independent of the magnitude of the single channel flux. A possible explanation of this property is given in terms of locally induced Ca inactivation of Ca release. The solution in the end pools was then changed to one with PDAA plus fura-2. SR Ca release was estimated from the [Ca] transient, as before, and from the delta [Cafura-2] signal. On average, 2-3 mM fura-2 increased the quasi-steady level of the rate of SR Ca release by factors of 6.6 and 3.8, respectively, in three fibers from Rana temporaria and three fibers from Rana pipiens. The peak rate of release was increased in five of the six fibers but to a lesser extent than the quasi-steady level. In all fibers, the amplitude of the free [Ca] transient was markedly reduced. These increases in the rate of SR Ca release are consistent with the idea that Ca inactivation of Ca release develops during a step depolarization to 10 mV and that 2-3 mM fura-2 is able to reduce this inactivation by complexing Ca and thereby reducing free [Ca]. Once the concentration

  4. The effect of Mg2+ on cardiac muscle function: Is CaATP the substrate for priming myofibril cross-bridge formation and Ca2+ reuptake by the sarcoplasmic reticulum?

    PubMed Central

    Smith, G A; Vandenberg, J I; Freestone, N S; Dixon, H B

    2001-01-01

    of the kinetics of the cardiac sarcoplasmic reticulum ATPase to those of the myofibril, in particular the positive co-operativity of both Mg2+ inhibition and Ca2+ activation, leads to the conclusion that this ATPase also has an initiation step that utilizes CaATP. The first-order activation by sub-millimolar [Mg2+], similar to that of the myofibril, may be explained by Mg2+ involvement in the phosphate-release step of the ATPase. The inhibition of both the myofibril and sarcoplasmic reticulum Ca2+ transporting ATPases by Mg2+ offers an explanation for the specific requirement for phosphocreatine (PCr) for full activity of both enzymes in situ and its effect on their apparent affinities for ATP. This explanation is based on the slow diffusion of Mg2+ within the myofibril and on the contrast of PCr with both ATP and phosphoenolpyruvate, in that PCr does not bind Mg2+ under physiological conditions, whereas both the other two bind it more tightly than the products of their hydrolysis do. The switch to supply of energy by diffusion of MgATP into the myofibril when depletion of PCr raises [ATP]/[PCr] greatly, e.g. during anoxia, results in a local [Mg2+] increase, which inhibits the ATPase. It is possible that mechanisms similar to those described above occur in skeletal muscle but the Ca2+ co-operativity involved would be masked by the presence of two Ca2+ binding sites on each troponin. PMID:11237858

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

    PubMed Central

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

    2014-01-01

    Calcium cycling is integral to muscle performance during the rapid muscle contraction and relaxation of high-intensity exercise. Ca2+ 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 Ca2+ 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 Ca2+ 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 Ca2+-ATPase activity (+28%) and Ca2+ uptake (+21%) than in CON were evident across both times and days (P < 0.05), but performance was similar. In T1D, resting Ca2+-ATPase activity correlated with work performed until exhaustion (r = 0.7, P < 0.01). Ca2+-ATPase activity, but not Ca2+ uptake, was lower (−24%, P < 0.05) among the women across both times and days. Intense exercise did not alter Ca2+-ATPase activity in T1D or CON. However, sex differences were evident: Ca2+-ATPase was reduced with exercise among men but increased among women across both days (time × sex interaction, P < 0.05). Sprint training reduced Ca2+-ATPase (−8%, P < 0.05), but not Ca2+ uptake, in T1D and CON. In summary, skeletal muscle Ca2+ resequestration capacity was increased in T1D, but performance was not greater than CON. Sprint training reduced Ca2+-ATPase in T1D and CON. Sex differences in Ca2+-ATPase activity were evident and may be linked with fibre type proportion differences. PMID:24297852

  6. Sequencing of genes involved in the movement of calcium across human skeletal muscle sarcoplasmic reticulum: continuing the search for genes associated with malignant hyperthermia.

    PubMed

    Bjorksten, A R; Gillies, R L; Hockey, B M; Du Sart, D

    2016-11-01

    The genetic basis of malignant hyperthermia (MH) is not fully characterised and likely involves more than just the currently classified mutations in the gene encoding the skeletal muscle ryanodine receptor (RYR1) and the gene encoding the α1 subunit of the dihydropyridine receptor (CACNA1S). In this paper we sequence other genes involved in calcium trafficking within skeletal muscle in patients with positive in vitro contracture tests, searching for alternative genes associated with MH. We identified four rare variants in four different genes (CACNB1, CASQ1, SERCA1 and CASQ2) encoding proteins involved in calcium handling in skeletal muscle in a cohort of 30 Australian MH susceptible probands in whom prior complete sequencing of RYR1 and CACNA1S had yielded no rare variants. These four variants have very low minor allele frequencies and while it is tempting to speculate that they have a role in MH, they remain at present variants of unknown significance. Nevertheless they provide the basis for a new set of functional studies, which may indeed identify novel players in MH.

  7. Sarcoplasmic MxA expression: A valuable marker of dermatomyositis.

    PubMed

    Uruha, Akinori; Nishikawa, Atsuko; Tsuburaya, Rie S; Hamanaka, Kohei; Kuwana, Masataka; Watanabe, Yurika; Suzuki, Shigeaki; Suzuki, Norihiro; Nishino, Ichizo

    2017-01-31

    To evaluate the diagnostic value of myxovirus resistance A (MxA) expression in the cytoplasm of myofibers in the diagnosis of dermatomyositis (DM). We assessed the sensitivity and specificity of the sarcoplasmic expression of MxA in muscles with DM by immunohistochemistry in consecutive cases of DM (n = 34) and other idiopathic inflammatory myopathies (n = 120: 8 with polymyositis, 16 with anti-tRNA-synthetase antibody-associated myositis, 46 with immune-mediated necrotizing myopathy, and 50 with inclusion body myositis) and compared them with conventional pathologic hallmarks of DM, including perifascicular atrophy (PFA) and membrane attack complex (MAC) deposition on endomysial capillaries. The sensitivity and specificity of sarcoplasmic MxA expression were 71% and 98%, respectively. While the specificity was almost comparable to that of PFA and capillary MAC deposition, the sensitivity was higher, with PFA showing 47% sensitivity and 98% specificity and capillary MAC deposition showing 35% sensitivity and 93% specificity. Of note, in patients with DM with typical skin rash but no PFA, 44% of the samples showed sarcoplasmic MxA expression, which was higher than the 17% sensitivity of capillary MAC deposition in the population. Sarcoplasmic MxA expression detected by immunohistochemistry is a more sensitive marker of DM than the conventional hallmarks, indicating its practical utility in the diagnosis of DM. It may well be included in the routine immunohistochemistry panel for myositis. This study provides Class II evidence that immunohistochemistry-detected sarcoplasmic MxA expression accurately identifies patients with dermatomyositis. © 2016 American Academy of Neurology.

  8. 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 ...

  9. Oxygen-coupled redox regulation of the skeletal muscle ryanodine receptor-Ca2+ release channel by NADPH oxidase 4

    PubMed Central

    Sun, Qi-An; Hess, Douglas T.; Nogueira, Leonardo; Yong, Sandro; Bowles, Dawn E.; Eu, Jerry; Laurita, Kenneth R.; Meissner, Gerhard; Stamler, Jonathan S.

    2011-01-01

    Physiological sensing of O2 tension (partial O2 pressure, pO2) plays an important role in some mammalian cellular systems, but striated muscle generally is not considered to be among them. Here we describe a molecular mechanism in skeletal muscle that acutely couples changes in pO2 to altered calcium release through the ryanodine receptor–Ca2+-release channel (RyR1). Reactive oxygen species are generated in proportion to pO2 by NADPH oxidase 4 (Nox4) in the sarcoplasmic reticulum, and the consequent oxidation of a small set of RyR1 cysteine thiols results in increased RyR1 activity and Ca2+ release in isolated sarcoplasmic reticulum and in cultured myofibers and enhanced contractility of intact muscle. Thus, Nox4 is an O2 sensor in skeletal muscle, and O2-coupled hydrogen peroxide production by Nox4 governs the redox state of regulatory RyR1 thiols and thereby governs muscle performance. These findings reveal a molecular mechanism for O2-based signaling by an NADPH oxidase and demonstrate a physiological role for oxidative modification of RyR1. PMID:21896730

  10. Oxygen-coupled redox regulation of the skeletal muscle ryanodine receptor-Ca2+ release channel by NADPH oxidase 4.

    PubMed

    Sun, Qi-An; Hess, Douglas T; Nogueira, Leonardo; Yong, Sandro; Bowles, Dawn E; Eu, Jerry; Laurita, Kenneth R; Meissner, Gerhard; Stamler, Jonathan S

    2011-09-20

    Physiological sensing of O(2) tension (partial O(2) pressure, pO(2)) plays an important role in some mammalian cellular systems, but striated muscle generally is not considered to be among them. Here we describe a molecular mechanism in skeletal muscle that acutely couples changes in pO(2) to altered calcium release through the ryanodine receptor-Ca(2+)-release channel (RyR1). Reactive oxygen species are generated in proportion to pO(2) by NADPH oxidase 4 (Nox4) in the sarcoplasmic reticulum, and the consequent oxidation of a small set of RyR1 cysteine thiols results in increased RyR1 activity and Ca(2+) release in isolated sarcoplasmic reticulum and in cultured myofibers and enhanced contractility of intact muscle. Thus, Nox4 is an O(2) sensor in skeletal muscle, and O(2)-coupled hydrogen peroxide production by Nox4 governs the redox state of regulatory RyR1 thiols and thereby governs muscle performance. These findings reveal a molecular mechanism for O(2)-based signaling by an NADPH oxidase and demonstrate a physiological role for oxidative modification of RyR1.

  11. Role of sarcoplasmic reticulum in digitalis-induced electrical and mechanical oscillations in the heart.

    PubMed

    Chan, T L; Chau, T C; Bose, D

    1987-05-01

    The mechanism of cardiac oscillatory activity induced by digitalis was studied in the canine ventricular muscle. We determined the role of sarcoplasmic reticulum in the phenomenon of oscillatory afterpotential and mechanical aftercontractions. Additionally we wished to study the interaction between changes in stimulus interval, which are known to affect sarcoplasmic reticulum function and these oscillatory phenomena. Aftercontraction was induced by ouabagenin but it required previous stimulation of the heart. The production of aftercontraction depended on short stimulus interval and bore no relation with the size of the releasable pool of Ca in the sarcoplasmic reticulum as indicated by the size of the driven contraction. Aftercontraction was often seen without an accompanying action potential. When a depolarization was present during aftercontraction, its temporal relation with the contraction often did not show the usual delay, suggesting that electrical activity may not necessarily initiate contraction. This was supported by the finding that Mn, a Ca channel blocker, blocked normal contractions more than aftercontraction. However, inhibition of Ca release from the sarcoplasmic reticulum by ryanodine or of Ca reuptake by caffeine were effective in blocking aftercontraction and when present, oscillatory afterpotential. Abolition of aftercontraction was not due to slowing of relaxation. These studies confirm the role of sarcoplasmic reticulum in causing aftercontraction and oscillatory afterpotential during ouabagenin toxicity and also suggest that there is a relationship between the 'repriming' ability of the sarcoplasmic reticulum and the oscillatory phenomena.

  12. 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.

  13. Obscurin Is a Ligand for Small Ankyrin 1 in Skeletal Muscle

    PubMed Central

    Kontrogianni-Konstantopoulos, Aikaterini; Jones, Ellene M.; van Rossum, Damian B.; Bloch, Robert J.

    2003-01-01

    The factors that organize the internal membranes of cells are still poorly understood. We have been addressing this question using striated muscle cells, which have regular arrays of membranes that associate with the contractile apparatus in stereotypic patterns. Here we examine links between contractile structures and the sarcoplasmic reticulum (SR) established by small ankyrin 1 (sAnk1), a ∼17.5-kDa integral protein of network SR. We used yeast two-hybrid to identify obscurin, a giant Rho-GEF protein, as the major cytoplasmic ligand for sAnk1. The binding of obscurin to the cytoplasmic sequence of sAnk1 is mediated by a sequence of obscurin that is C-terminal to its last Ig-like domain. Binding was confirmed in two in vitro assays. In one, GST-obscurin, bound to glutathione-matrix, specifically adsorbed native sAnk1 from muscle homogenates. In the second, MBP-obscurin bound recombinant GST-sAnk1 in nitrocellulose blots. Kinetic studies using surface plasmon resonance yielded a KD = 130 nM. On subcellular fractionation, obscurin was concentrated in the myofibrillar fraction, consistent with its identification as sarcomeric protein. Nevertheless, obscurin, like sAnk1, concentrated around Z-disks and M-lines of striated muscle. Our findings suggest that obscurin binds sAnk1, and are the first to document a specific and direct interaction between proteins of the sarcomere and the SR. PMID:12631729

  14. Effect of taurine depletion on excitation-contraction coupling and Cl- conductance of rat skeletal muscle.

    PubMed

    De Luca, A; Pierno, S; Camerino, D C

    1996-01-25

    The pharmacological action of taurine on skeletal muscle is to stabilize sarcolemma by increasing macroscopic conductance to Cl- (GCl), whereas a proposed physiological role for the amino acid is to modulate excitation-contraction coupling mechanism via Ca2+ availability. To get insight in the physiological role of taurine in skeletal muscle, the effects of its depletion were evaluated on voltage threshold for mechanical activation and GCl with the two intracellular microelectrode method in 'point' voltage clamp mode and current clamp mode, respectively. The experiments were performed on extensor digitorum longus muscle fibers from rats depleted of taurine by a chronic 4 week treatment with guanidinoethane sulfonate, a known inhibitor of taurine transporter. The treatment significantly modified the mechanical threshold of striated fibers; i.e. at each pulse duration they needed significantly less depolarization to contract and the fitted rheobase voltage was more negative by 10 mV with respect to untreated muscle fibers. In parallel, the treatment with guanidinoethane sulfonate produced a significant 40% lowering of GCl. In vitro application of 60 mM of taurine to such depleted muscles almost completely restored the mechanical threshold and increased GCl even above the value of untreated control. However, in vitro application of 60 mM of either taurine or guanidinoethane sulfonate to untreated control muscles did not cause any change of the mechanical threshold but increased GCl by 40% and 21%, respectively. Furthermore, 100 microM of the S-(-) enantiomer of 2-(p-chlorophenoxy)propionic acid almost fully blocked GCl but did not produce any change in the mechanical threshold of normal muscle fibers. The present results show that the large amount of intracellular taurine plays a role in the excitation-contraction coupling mechanism of striated muscle fibers. This action is independent from any effect involving muscle Cl- channels, but it is likely mediated by the

  15. Nitric oxide synthase in cardiac sarcoplasmic reticulum.

    PubMed

    Xu, K Y; Huso, D L; Dawson, T M; Bredt, D S; Becker, L C

    1999-01-19

    NO. is a free radical that modulates heart function and metabolism. We report that a neuronal-type NO synthase (NOS) is located on cardiac sarcoplasmic reticulum (SR) membrane vesicles and that endogenous NO. produced by SR-associated NOS inhibits SR Ca2+ uptake. Ca2+-dependent biochemical conversion of L-arginine to L-citrulline was observed from isolated rabbit cardiac SR vesicles in the presence of NOS substrates and cofactors. Endogenous NO. was generated from the vesicles and detected by electron paramagnetic resonance spin-trapping measurements. Immunoelectron microscopy demonstrated labeling of cardiac SR vesicles by using anti-neuronal NOS (nNOS), but not anti-endothelial NOS (eNOS) or anti-inducible NOS (iNOS) antibodies, whereas skeletal muscle SR vesicles had no nNOS immunoreactivity. The nNOS immunoreactivity also displayed a pattern consistent with SR localization in confocal micrographs of sections of human myocardium. Western blotting demonstrated that cardiac SR NOS is larger than brain NOS (160 vs. 155 kDa). No immunodetection was observed in cardiac SR vesicles from nNOS knockout mice or with an anti-nNOS mu antibody, suggesting the possibility of a new nNOS-type isoform. 45Ca uptake by cardiac SR vesicles, catalyzed by Ca2+-ATPase, was inhibited by NO. produced endogenously from cardiac SR NOS, and 7-nitroindazole, a selective nNOS inhibitor, completely prevented this inhibition. These results suggest that a cardiac muscle nNOS isoform is located on SR of cardiac myocytes, where it may respond to intracellular Ca2+ concentration and modulate SR Ca2+ ion active transport in the heart.

  16. Nitric oxide synthase in cardiac sarcoplasmic reticulum

    PubMed Central

    Xu, Kai Y.; Huso, David L.; Dawson, Ted M.; Bredt, David S.; Becker, Lewis C.

    1999-01-01

    NO⋅ is a free radical that modulates heart function and metabolism. We report that a neuronal-type NO synthase (NOS) is located on cardiac sarcoplasmic reticulum (SR) membrane vesicles and that endogenous NO⋅ produced by SR-associated NOS inhibits SR Ca2+ uptake. Ca2+-dependent biochemical conversion of l-arginine to l-citrulline was observed from isolated rabbit cardiac SR vesicles in the presence of NOS substrates and cofactors. Endogenous NO⋅ was generated from the vesicles and detected by electron paramagnetic resonance spin-trapping measurements. Immunoelectron microscopy demonstrated labeling of cardiac SR vesicles by using anti-neuronal NOS (nNOS), but not anti-endothelial NOS (eNOS) or anti-inducible NOS (iNOS) antibodies, whereas skeletal muscle SR vesicles had no nNOS immunoreactivity. The nNOS immunoreactivity also displayed a pattern consistent with SR localization in confocal micrographs of sections of human myocardium. Western blotting demonstrated that cardiac SR NOS is larger than brain NOS (160 vs. 155 kDa). No immunodetection was observed in cardiac SR vesicles from nNOS knockout mice or with an anti-nNOSμ antibody, suggesting the possibility of a new nNOS-type isoform. 45Ca uptake by cardiac SR vesicles, catalyzed by Ca2+-ATPase, was inhibited by NO⋅ produced endogenously from cardiac SR NOS, and 7-nitroindazole, a selective nNOS inhibitor, completely prevented this inhibition. These results suggest that a cardiac muscle nNOS isoform is located on SR of cardiac myocytes, where it may respond to intracellular Ca2+ concentration and modulate SR Ca2+ ion active transport in the heart. PMID:9892689

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

    USDA-ARS?s Scientific Manuscript database

    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...

  18. 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.

  19. Supravital morphology of small branches of lateral striate arteries as observed with Nomarski optics.

    PubMed

    Gouveia, C J

    1996-01-01

    Lateral striate arteries were dissected from the fixed brains of 6 patients of increasing age. Small branches of arteries were observed--unprocessed and unstained--by Nomarski optics. Among the findings there was fibrous intimal proliferation, replacement of medial muscle by collagen, tortuosity, twisting or coiling. The severity of changes seemed to progress with aging. The advantages of the used methodology that aims at avoiding artifacts of processing are discussed briefly.

  20. Abnormal response to calmodulin in vitro of dystrophic chicken muscle membrane Ca2+-ATPase activity.

    PubMed

    Galindo, J; Hudecki, M S; Davis, F B; Davis, P J; Thacore, H R; Pollina, C M; Blas, S D; Schoenl, M

    1988-09-20

    A skeletal muscle membrane fraction enriched in sarcoplasmic reticulum (SR) contained Ca2+-ATPase activity which was stimulated in vitro in normal chickens (line 412) by 6 nM purified bovine calmodulin (33% increase over control, P less than 0.001). In contrast, striated muscle from chickens (line 413) affected with an inherited form of muscular dystrophy, but otherwise genetically similar to line 412, contained SR-enriched Ca2+-ATPase activity which was resistant to stimulation in vitro by calmodulin. Basal levels of Ca2+-ATPase activity (no added calmodulin) were comparable in muscles of unaffected and affected animals, and the Ca2+ optima of the enzymes in normal and dystrophic muscle were identical. Purified SR vesicles, obtained by calcium phosphate loading and sucrose density gradient centrifugation, showed the same resistance of dystrophic Ca2+-ATPase to exogenous calmodulin as the SR-enriched muscle membrane fraction. Dystrophic muscle had increased Ca2+ content compared to that of normal animals (P less than 0.04) and has been previously shown to contain increased levels of immuno- and bioactive calmodulin and of calmodulin mRNA. The calmodulin resistance of the Ca2+-ATPase in dystrophic muscle reflects a defect in regulation of cell Ca2+ metabolism associated with elevated cellular Ca2+ and calmodulin concentrations.

  1. 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

  2. A significant role of sarcoplasmic reticulum in cardiac contraction of a basal vertebrate, the river lamprey (Lampetra fluviatilis).

    PubMed

    Vornanen, M; Haverinen, J

    2013-02-01

    Cardiac contraction and relaxation are mediated by rapidly changing calcium concentration around the myofibrils. In comparison with endotherms, ectothermic hearts are more strongly dependent on extracellular calcium for contraction suggesting this trait might represent the primitive vertebrate mode of cardiac activation. This study tests the hypothesis that intracellular calcium stores play a minor role in cardiac excitation-contraction coupling of a basal vertebrate, the lamprey (Lampetra fluviatilis). Contribution of sarcoplasmic reticulum to cardiac calcium management was examined by measuring the ryanodine sensitivity of contraction, determining the number of cardiac ryanodine receptors and their calcium sensitivity, assessing the size of sarcoplasmic reticulum calcium stores in enzymatically isolated cardiac myocytes and qualitative electron microscopic observations of sarcoplasmic reticulum in cardiac muscle. An extensive network of nonjunctional sarcoplasmic reticulum around myofibrils and numerous junctional sarcoplasmic reticulum couplings at the periphery of the myocytes was present in atrial and ventricular muscle of the lamprey heart. High numbers of ryanodine receptors were present in atrial (0.16 ± 0.04 nmol mg(-1) prot) and ventricular membranes (0.27 ± 0.03) (P < 0.01), and 10 μm ryanodine inhibited large part of contraction force in atrial (37.8 ± 5.7%) and ventricular (49.2 ± 6.5%) muscle. Sarcoplasmic reticulum is well developed in the lamprey heart and plays a significant role in cardiac calcium management. This suggests that in the common ancestor of vertebrates, cardiac excitation-contraction coupling could have been fairly strongly dependent on sarcoplasmic reticulum calcium stores. Functionally, this trait might be associated with high cardiac output and active predatory lifestyle of the lamprey. © 2012 The Authors Acta Physiologica © 2012 Scandinavian Physiological Society.

  3. Normalization of cell responses in cat striate cortex

    NASA Technical Reports Server (NTRS)

    Heeger, D. J.

    1992-01-01

    Simple cells in the striate cortex have been depicted as half-wave-rectified linear operators. Complex cells have been depicted as energy mechanisms, constructed from the squared sum of the outputs of quadrature pairs of linear operators. However, the linear/energy model falls short of a complete explanation of striate cell responses. In this paper, a modified version of the linear/energy model is presented in which striate cells mutually inhibit one another, effectively normalizing their responses with respect to stimulus contrast. This paper reviews experimental measurements of striate cell responses, and shows that the new model explains a significantly larger body of physiological data.

  4. [Molecular architecture of the sarcoplasmic reticulum and its role in the ECC].

    PubMed

    Rigoard, P; Buffenoir, K; Wager, M; Bauche, S; Giot, J-P; Lapierre, F

    2009-03-01

    The sarcoplasmic reticulum (SR) plays a fundamental role in excitation-contraction coupling, which propagates the electric signal conversion along the muscle fiber's plasmic membrane to a mechanical event manifested as a muscle contraction. It plays a crucial role in calcium homeostasis and intracellular calcium storage control (storage, liberation and uptake) necessary for fiber muscle contraction and then relaxation. These functions take place at the triad, made up of individualized SR subdomains where the protein-specific organization provides efficient and fast coupling. Ryanodine receptors (RyR) and dihydropyridine receptors (DHPR) mainly act in calcium exchanges in the SR. This particular structural and molecular architecture must be correlated to its functional specificity.

  5. 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

  6. [Calcium transport in sarcoplasmic reticulum in the presence of AR-L 115 BS].

    PubMed

    Hasselbach, W

    1981-01-01

    2-[(2-Methoxy-4-methylsulfinyl)phenyl]-1H-imidazo[4,5-b]pyridine (AR-L 115 BS) is a substance with positive inotropic activity which does not influence the activity of the sarcoplasmic calcium pump. It can, therefore, be expected that AR-L 115 BS does not interfere with the distribution and movement of calcium in the resting and active muscle.

  7. 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.

  8. 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.

  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. 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.

  11. Regulation of Physiological and Metabolic Function of Muscle by Female Sex Steroids

    PubMed Central

    Spangenburg, Espen E.; Geiger, Paige C.; Leinwand, Leslie A.; Lowe, Dawn A.

    2012-01-01

    The ability of female sex steroids to regulate tissue function has long been appreciated, however their role in the regulation of striated muscle function has received considerably less attention. The purpose of this symposium review is to document recent evidence indicating the role female sex steroids have in defining the functional characteristics of striated muscle. The presentations provide substantial evidence indicating that estrogens are critical to the physiological and metabolic regulation of striated muscle, thus when considering women’s health issues striated muscle must included as an important target tissue along with other classically thought of estrogen sensitive tissues. PMID:22525764

  12. Paspalum striate mosaic virus: an Australian mastrevirus from Paspalum dilatatum.

    PubMed

    Geering, Andrew D W; Thomas, John E; Holton, Timothy; Hadfield, James; Varsani, Arvind

    2012-01-01

    Three monocot-infecting mastreviruses from Australia, all found primarily in pasture and naturalised grasses, have been characterised at the molecular level. Here, we present the full genome sequence of a fourth, Paspalum striate mosaic virus (PSMV), isolated from Paspalum dilatatum from south-east Queensland. The genome was 2816 nt long and had an organisation typical of other monocot-infecting mastreviruses. Its nearest relative is Bromus cartharticus striate mosaic virus (BCSMV), with which it shares an overall genome identity of 75%. Phylogenetic analysis of the complete genome and each of the putative viral proteins places PSMV in a group with the other three Australian striate mosaic viruses. PSMV, BCSMV and Digitaria didactyla striate mosaic virus all contain a similar, small recombinant sequence in the small intergenic region.

  13. Modification of the functional capacity of sarcoplasmic reticulum membranes in patients suffering from chronic fatigue syndrome.

    PubMed

    Fulle, Stefania; Belia, Silvia; Vecchiet, Jacopo; Morabito, Caterina; Vecchiet, Leonardo; Fanò, Giorgio

    2003-08-01

    In chronic fatigue syndrome, several reported alterations may be related to specific oxidative modifications in muscle. Since sarcoplasmic reticulum membranes are the basic structures involved in excitation-contraction coupling and the thiol groups of Ca(2+) channels of SR terminal cisternae are specific targets for reactive oxygen species, it is possible that excitation-contraction coupling is involved in this pathology. We investigated the possibility that abnormalities in this compartment are involved in the pathogenesis of chronic fatigue syndrome and consequently responsible for characteristic fatigue. The data presented here support this hypothesis and indicate that the sarcolemmal conduction system and some aspects of Ca(2+) transport are negatively influenced in chronic fatigue syndrome. In fact, both deregulation of pump activities (Na(+)/K(+) and Ca(2+)-ATPase) and alteration in the opening status of ryanodine channels may result from increased membrane fluidity involving sarcoplasmic reticulum membranes.

  14. Ultrastructure of sarcoplasmic reticulum in atrial myocardium of patients with mitral valvular disease.

    PubMed Central

    Thiedermann, K. U.; Ferrans, V. J.

    1976-01-01

    Alterations observed in the sarcoplasmic reticulum of muscle cells in left and right atrial myocardium from 10 patients with mitral valvular disease consisted of: a) proliferation of rough-surfaced endoplasmic reticulum, which formed large cisterns in perinuclear areas of hypertrophied cells and was considered indicative of increased protein synthesis; b) proliferation of free sarcoplasmic reticulum, a change that occurred in degenerated cells and appeared to be related to loss of contractile elements; c) two types of aggregates of tubules of free SR--one type was associated wtih abnormal Z-band material and was found only in cells showing loss of myofibrils and proliferation of free SR, whereas the other was not associated with either of these changes and occurred in less severely altered cells; and d) proliferation and enlargement of cisterns of extended junctional sarcoplasmic reticulum, which formed two distinct types of complexes: the first of these consisted of large, convoluted (Type A) cisterns that were wide (550 to 650 A in thickness) and did not have a central dense lamina; the second was composed of stacks of concentric or parallel (Type B) cisterns that were narrower (220 to 300 A in thickness), had a central dense lamina, and were separated from one another by layers of glycogen granules. The formation of these complexes of cisterns was regarded as an extreme form of overdevelopment of extended junctional sarcoplasmic reticulum in atrial muscle cells. Images Figure 21 Figures 22-25 Figures 1-3 Figures 26-29 Figure 4 Figure 5 Figure 6 Figures 30 and 31 Figure 7 Figure 8 Figure 9 Figure 10 Figures 32-36 Figure 11 Figure 12 Figures 37-39 Figure 15 Figure 16 Figure 17 Figure 13 Figure 14 Figures 40 and 41 Figure 18 Figures 19 and 20 PMID:1275054

  15. Characterization of sarcoplasmic calcium binding protein (SCP) variants from freshwater crayfish Procambarus clarkii.

    PubMed

    White, Alexandra J; Northcutt, Michael J; Rohrback, Suzanne E; Carpenter, Robert O; Niehaus-Sauter, Margaret M; Gao, Yongping; Wheatly, Michele G; Gillen, Christopher M

    2011-09-01

    Sarcoplasmic calcium binding protein (SCP) is an invertebrate EF-hand calcium buffering protein that has been proposed to fulfill a similar function in muscle relaxation as vertebrate parvalbumin. We have identified three SCP variants in the freshwater crayfish Procambarus clarkii. The variants (pcSCP1a, pcSCP1b, and pcSCP1c) differ across a 37 amino acid region that lies mainly between the second and third EF-hand calcium binding domains. We evaluated tissue distribution and response of the variants to cold exposure, a stress known to affect expression of parvalbumin. Expression patterns of the variants were not different and therefore do not provide a functional rationale for the polymorphism of pcSCP1. Compared to hepatopancreas, expression of pcSCP1 variants was 100,000-fold greater in axial abdominal muscle and 10-fold greater in cardiac muscle. Expression was 10-100 greater in fast-twitch deep flexor and extensor muscles compared to slow-twitch superficial flexor and extensors. In axial muscle, no significant changes of pcSCP1, calmodulin (CaM), or sarcoplasmic/endoplasmic reticulum Ca-ATPase (SERCA) expression were measured after one week of 4°C exposure. In contrast, large decreases of pcSCP1 were measured in cardiac muscle, with no changes in CaM or SERCA. Knockdown of pcSCP1 by dsRNA led to reduced muscle activity and decreased expression of SERCA. In summary, the pattern of pcSCP1 tissue expression is similar to parvalbumin, supporting a role in muscle contraction. However, the response of pcSCP1 to cold exposure differs from parvalbumin, suggesting possible functional divergence between the two proteins.

  16. Evaluation of Vascular Delivery Methodologies to Enhance rAAV6-mediated Gene Transfer to Canine Striated Musculature

    PubMed Central

    Gregorevic, Paul; Schultz, Brian R; Allen, James M; Halldorson, Jeffrey B; Blankinship, Michael J; Meznarich, Norman A; Kuhr, Christian S; Doremus, Caitlin; Finn, Eric; Liggitt, Denny; Chamberlain, Jeffrey S

    2009-01-01

    A growing body of research supports the development of recombinant adeno-associated viral (rAAV) vectors for delivery of gene expression cassettes to striated musculature as a method of treating severe neuromuscular conditions. However, it is unclear whether delivery protocols that achieve extensive gene transfer in mice can be adapted to produce similarly extensive gene transfer in larger mammals and ultimately patients. Consequently, we sought to investigate methodological modifications that would facilitate rAAV-mediated gene transfer to the striated musculature of canines. A simple procedure incorporating acute (i) occlusion of limb blood flow, (ii) exsanguination via compression bandage, and (iii) vector “dwell” time of <20 minutes, markedly enhanced the transduction of limb muscles, compared with a simple bolus limb infusion of vector. A complementary method whereby vector was infused into the jugular vein led to efficient transduction of cardiomyocytes and to a lesser degree the diaphragm. Together these methods can be used to achieve transgene expression in heart, diaphragm, and limb muscles of juvenile dogs using rAAV6 vectors. These results establish that rAAV-mediated gene delivery is a viable approach to achieving systemic transduction of striated musculature in mammals approaching the dimensions of newborn humans. PMID:19471246

  17. Effect of Sarcoplasmic Reticulum (SR) Calcium Content on SR Calcium Release Elicited by Small Voltage-Clamp Depolarizations in Frog Cut Skeletal Muscle Fibers Equilibrated with 20 mM EGTA

    PubMed Central

    Pape, Paul C.; Carrier, Nicole

    1998-01-01

    Cut muscle fibers from Rana temporaria (sarcomere length, 3.5–3.9 μm; 14–16°C) were mounted in a double Vaseline-gap chamber and equilibrated with an external solution that contained tetraethyl ammonium– gluconate and an internal solution that contained Cs as the principal cation, 20 mM EGTA, and 0 Ca. Fibers were stimulated with a voltage-clamp pulse protocol that consisted of pulses to −70, −65, −60, −45, and −20 mV, each separated by 400-ms periods at −90 mV. The change in total Ca that entered into the myoplasm (Δ[CaT]) and the Ca content of the SR ([CaSR]) were estimated with the EGTA/phenol red method (Pape, P.C., D.-S. Jong, and W.K. Chandler. 1995. J. Gen. Physiol. 106:259–336). Fibers were stimulated with the pulse protocol, usually every 5 min, so that the resting value of [CaSR] decreased from its initial value of 1,700–2,300 μM to values near or below 100 μM after 18–30 stimulations. Three main findings for the voltage pulses to −70, −65, and −60 mV are: (a) the depletion-corrected rate of Ca release (release permeability) showed little change when [CaSR] decreased from its highest level (>1,700 μM) to ∼1,000 μM; (b) as [CaSR] decreased below 1,000 μM, the release permeability increased to a maximum level when [CaSR] was near 300 μM that was on average about sevenfold larger than the values observed for [CaSR] > 1,000 μM; and (c) as [CaSR] decreased from ∼300 μM to <100 μM, the release permeability decreased, reaching half its maximum value when [CaSR] was ∼110 μM on average. It was concluded that finding b was likely due to a decrease in Ca inactivation, while finding c was likely due to a decrease in Ca-induced Ca release. PMID:9689025

  18. Function and regulation of sarcoplasmic reticulum Ca2+-ATPase: advances during the past decade and prospects for the coming decade.

    PubMed

    Arai, M

    2000-01-01

    In cardiac muscle, the contraction-relaxation cycle is tightly controlled by the regulated release and uptake of intracellular Ca2+ between sarcoplasmic reticulum and cytoplasm. A major protein controlling Ca2+ cycling is Ca2+-ATPase (SERCA2a) located in the sarcoplasmic reticulum membrane. The function of SERCA2a protein is regulated by the phosphorylatable protein, phospholamban. Phosphorylation of phospholamban releases its inhibitory effect on SERCA2a through direct molecular interaction. Recently, mice whose SERCA2a function is increased (overexpression of the gene) or lost (knock out) were developed. These mice demonstrated that SERCA2a pump levels are a major determinant of cardiac muscle contractility and relaxation. These studies open the prospect that the overexpression of SERCA2a can correct cardiac dysfunction seen in heart failure. Advances in knowledge concerning the function and gene regulation of SERCA2a are discussed in this review.

  19. 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…

  20. 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…

  1. 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.

  2. Doxorubicin cardiomyopathy is associated with a decrease in calcium release channel of the sarcoplasmic reticulum in a chronic rabbit model.

    PubMed Central

    Dodd, D A; Atkinson, J B; Olson, R D; Buck, S; Cusack, B J; Fleischer, S; Boucek, R J

    1993-01-01

    Doxorubicin is a highly effective cancer chemotherapeutic agent that produces a dose-dependent cardiomyopathy that limits its clinical usefulness. Clinical and animal studies of morphological changes during the early stages of doxorubicin-induced cardiomyopathy have suggested that the sarcoplasmic reticulum, the intracellular membrane system responsible for myoplasmic calcium regulation in adult mammalian heart, may be the early target of doxorubicin. To detect changes in the calcium pump protein or the calcium release channel (ryanodine receptor) of the sarcoplasmic reticulum during chronic doxorubicin treatment, rabbits were treated with intravenous doxorubicin (1 mg/kg) twice weekly for 12 to 18 doses. Pair-fed controls received intravenous normal saline. The severity of cardiomyopathy was scored by light and electron microscopy of left ventricular papillary muscles. Developed tension was measured in isolated atrial strips. In subcellular fractions from heart, [3H]ryanodine binding was decreased in doxorubicin-treated rabbits (0.33 +/- 0.03 pmol/mg) compared with control rabbits (0.66 +/- 0.02 pmol/mg; P < 0.0001). The magnitude of the decrease in [3H]ryanodine binding correlated with both the severity of the cardiomyopathy graded by pathology score (light and electron microscopy) and the decrease in developed tension in isolated atrial strips. Bmax for [3H]ryanodine binding and the amount of immunoreactive ryanodine receptor by Western blot analysis using sequence-specific antibody were both decreased, consistent with a decrease in the amount of calcium release channel of sarcoplasmic reticulum in doxorubicin-treated rabbits. In contrast, there was no decrease in the amount or the activity of the calcium pump protein of the sarcoplasmic reticulum in doxorubicin-treated rabbits. Doxorubicin treatment did not decrease [3H]ryanodine binding or the amount of immunoreactive calcium release channel of sarcoplasmic reticulum in skeletal muscle. Since the sarcoplasmic

  3. Flavour formation from hydrolysis of pork sarcoplasmic protein extract by a unique LAB culture isolated from Harbin dry sausage.

    PubMed

    Chen, Qian; Liu, Qian; Sun, Qinxiu; Kong, Baohua; Xiong, Youling

    2015-02-01

    The lactic acid bacteria Pediococcus pentosaceus, Lactobacillus brevis, Lactobacillus curvatus, and Lactobacillus fermentum isolated from Harbin dry sausage were assessed for their protein hydrolysis and flavour development in pork muscle sarcoplasmic protein extracts. Gel electrophoresis indicated that sarcoplasmic proteins were degraded by all of the strains, especially by P. pentosaceus and L. curvatus. Trichloroacetic acid-soluble peptides increased in all of the samples (P < 0.05), especially samples inoculated with P. pentosaceus. Samples inoculated with P. pentosaceus and L. curvatus had higher free amino acid contents than did the other two strains(P < 0.05), and glutamic acid and alanine appeared to be the predominant free amino acids. The volatile compound analysis indicated that the highest aldehydes, alcohols and acid contents were found in the sample with P. pentosaceus followed by L. curvatus. The results revealed that P. pentosaceus could be appropriate for use as a meat starter culture.

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

    PubMed

    Chen, L Y; Chen, X; Tian, X L; Yu, X H

    2000-10-01

    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 micromol l(-1) Tet reduced the initial rate of calcium uptake to 59% of control 6 min after reaction. Tet un-competitively inhibited SR Ca(2+), Mg(2+)-ATPase activity, causing the stoichiometric ratio of SR Ca(2+)/ATP to decrease to 1.43 from 2.0 of control. Inhibitory rates on SR Ca(2+),Mg(2+)-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 Ca(2+),Mg(2+)-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 [Ca(2+)](i) and myocardial contractility.

  5. 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

  6. 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

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

    PubMed

    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.

  8. 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.

  9. 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.

  10. The Fine-Scale Functional Correlation of Striate Cortex in Sighted and Blind People

    PubMed Central

    Butt, Omar H.; Benson, Noah C.; Datta, Ritobrato

    2013-01-01

    To what extent are spontaneous neural signals within striate cortex organized by vision? We examined the fine-scale pattern of striate cortex correlations within and between hemispheres in rest-state BOLD fMRI data from sighted and blind people. In the sighted, we find that corticocortico correlation is well modeled as a Gaussian point-spread function across millimeters of striate cortical surface, rather than degrees of visual angle. Blindness produces a subtle change in the pattern of fine-scale striate correlations between hemispheres. Across participants blind before the age of 18, the degree of pattern alteration covaries with the strength of long-range correlation between left striate cortex and Broca's area. This suggests that early blindness exchanges local, vision-driven pattern synchrony of the striate cortices for long-range functional correlations potentially related to cross-modal representation. PMID:24107953

  11. Modulation of Sarcoplasmic Reticulum Calcium Pump (SERCA) Function by Membrane Cholesterol during Unloading

    NASA Astrophysics Data System (ADS)

    Clarke, M. S. F.; Hammond, D. K.; Feeback, D. L.

    2002-01-01

    We have recently demonstrated by in situ immuno-localization that cholesterol is predominantly located in the sarcoplasmic reticulum (SR), rather than in the sarcolemmal/T-tubule (SL-TT) membranes of both human and rat skeletal muscle (Clarke et al., 2000, JAP). In addition, we have demonstrated that mechanical unloading of skeletal muscle in a rat hindlimb suspension model significantly increases membrane cholesterol content and that this increase is also localized to SR rather than SL-TT membranes in such atrophied muscle. Utilizing a novel fluorescent calcium staining technique in perfusion fixed soleus muscle we observed a significant positive correlation between membrane cholesterol content and free intramyofiber calcium levels during unloading. To determine if a correlation between increased SR membrane cholesterol content and increased free intramyofiber calcium levels during unloading is due to a membrane cholesterol-mediated alteration in SR calcium pump function, we also describe the effects of modulating the cholesterol content of purified SR membrane preparations on SR-Ca2+ ATPase activity and ryanodine channel activity. As an increase in free intra-cellular calcium levels have previously demonstrated to induce catabolism in a wide range of biological systems, we suggest that altered SR calcium pump function may be the underlying basis for the initiation of unloading induced muscle atrophy.

  12. Smooth muscle alpha-actinin interaction with smitin.

    PubMed

    Chi, Richard J; Olenych, Scott G; Kim, Kyoungtae; Keller, Thomas C S

    2005-07-01

    Actin-myosin II filament-based contractile structures in striated muscle, smooth muscle, and nonmuscle cells also contain the actin filament-crosslinking protein alpha-actinin. In striated muscle sarcomeres, interactions between the myosin-binding protein titin and alpha-actinin in the Z-line provide an important structural linkage. We previously discovered a titin-like protein, smitin, associated with the contractile apparatus of smooth muscle cells. Purified native smooth muscle alpha-actinin binds with nanomolar affinity to smitin in smitin-myosin coassemblies in vitro. Smooth muscle alpha-actinin also interacts with striated muscle titin. In contrast to striated muscle alpha-actinin interaction with titin and smitin, which is significantly enhanced by PIP2, smooth muscle alpha-actinin interacts with smitin and titin equally well in the presence and absence of PIP2. Using expressed regions of smooth muscle alpha-actinin, we have demonstrated smitin-binding sites in the smooth muscle alpha-actinin R2-R3 spectrin-like repeat rod domain and a C-terminal domain formed by cryptic EF-hand structures. These smitin-binding sites are highly homologous to the titin-binding sites of striated muscle alpha-actinin. Our results suggest that direct interaction between alpha-actinin and titin or titin-like proteins is a common feature of actin-myosin II contractile structures in striated muscle and smooth muscle cells and that the molecular bases for alpha-actinin interaction with these proteins are similar, although regulation of these interactions may differ according to tissue.

  13. Deoxyglucose Analysis of Retinotopic Organization in Primate Striate Cortex

    NASA Astrophysics Data System (ADS)

    Tootell, Roger B. H.; Silverman, Martin S.; Switkes, Eugene; de Valois, Russell L.

    1982-11-01

    We have anatomically analyzed retinotopic organization using the 14C-labeled 2-deoxy-D-glucose method. The method has several advantages over conventional electrophysiological mapping techniques. In the striate cortex, the anatomical substrate for retinotopic organization is suprisingly well ordered, and there seems to be a systematic relationship between ocular dominance strips and cortical magnification. The 2-deoxyglucose maps in this area appear to be largely uninfluenced by known differences in long-term metabolic activity. This method should prove useful in analyzing retinotopic organization in various visual areas of the brain and in different species.

  14. 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

  15. Prolonged exercise potentiates sarcoplasmic reticulum Ca2+ uptake in rat diaphragm.

    PubMed

    Stavrianeas, Stasinos; Spangenburg, Espen; Batts, Tim; Williams, Jay H; Klug, Gary A

    2003-03-01

    The effects of a single bout of prolonged treadmill exercise [mean=81 (13) min] on sarcoplasmic reticulum (SR) Ca(2+) release, uptake and ATPase activity were determined in the costal region of rat diaphragm (D) and red gastrocnemius (RG). Glycogen depletion measurements made immediately following exercise suggested that treadmill running substantially recruited the fibers throughout both muscles. SR Ca(2+) ATPase activity, measured in isolated SR vesicles, decreased in the RG by 33% but remained unchanged in D in response to the exercise bout. This effect in RG was matched by a 37% decline in Ca(2+) uptake and a 28% depression in Ca(2+) release when measured in muscle homogenates. Conversely, Ca(2+) uptake increased between 157% and 263% in the D in the absence of any change in Ca(2+) release. These data show that the attenuation of SR function that has been consistently observed in limb muscle over the last several decades is absent in diaphragm despite the fact that its fibers appear to experience sufficient activity to deplete their glycogen. In fact, the large increase in Ca(2+) uptake in D shows that prolonged activity actually potentiates the ability of SR vesicles to sequester Ca(2+) in the absence of any increase in energy cost. Thus, it appears necessary to re-evaluate the role of exercise in regulating Ca(2+) sequestration by the SR as different muscles may respond in ways that are dictated by their function.

  16. Inositol (1,4,5)-trisphosphate activates a calcium channel in isolated sarcoplasmic reticulum membranes.

    PubMed Central

    Suárez-Isla, B. A.; Irribarra, V.; Oberhauser, A.; Larralde, L.; Bull, R.; Hidalgo, C.; Jaimovich, E.

    1988-01-01

    Sarcoplasmic reticulum membrane vesicles isolated from frog skeletal muscle display high conductance calcium channels when fused into phospholipid bilayers. The channels are selective for calcium and barium over Tris. The fractional open time was voltage-independent (-40 to +25 mV), but was steeply dependent on the free cis [Ca2+] (P0 = 0.02 at 10 microM cis Ca2+ and 0.77 at 150 microM Ca2+; estimated Hill coefficient: 1.6). Addition of ATP (1 mM; cis) further increased P0 from 0.77 to 0.94. Calcium activation was reversed by addition of EGTA to the cis compartment. Magnesium (2 mM) increased the frequency of rapid closures and 8 mM magnesium decreased the current amplitude from 3.4 to 1.2 pA at 0 mV, suggesting a reversible fast blockade. Addition of increasing concentrations of inositol (1, 4, 5)-triphosphate (cis), increased P0 from 0.10 +/- 0.01 (mean +/- SEM) in the control to 0.85 +/- 0.02 at 50 microM in an approximately sigmoidal fashion, with an apparent half-maximal activation at 15 microM inositol (1, 4, 5)-trisphosphate in the presence of 40 microM cis Ca2+. Lower concentrations of this agonist were required to produce a significant increase in P0 when 10 microM or less cis Ca2+ were used. The channel was blocked by the addition to the cis compartment of either 0.5 mM lanthanum, 0.5 microM ruthenium red, or 200 nM ryanodine, all known inhibitors of Ca2+ release from sarcoplasmic reticulum vesicles. These results demonstrate the presence of calcium channels in the sarcoplasmic reticulum from frog skeletal muscle with a pharmacological profile consistent with a role in excitation contraction coupling and with the hypothesis that inositol ( 1,4,5)-trisphosphate is a physiological agonist in this process. PMID:2852037

  17. 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

  18. The basal apparatus. Mass isolation from the molluscan ciliated gill epithelium and a preliminary characterization of striated rootlets

    PubMed Central

    1975-01-01

    The basal apparatus, consisting of an array of interconnected basal bodies bearing bifurcating striated rootlets encompassing a nucleus, has been isolated from hypertonically deciliated columnar gill epithelial cells of the bay scallop Aequipecten irradians through gentle lysis with Triton X-100. The rootlets, 8-10 mum in length, were not easily preserved with conventional electron microscope fixatives, suggesting that the extent of their contribution to cellular architecture has been somewhat underestimated, even though Englemann described many of the structural details of the basal apparatus in 1880. The striated rootlets were soluble at high but not at low pH, in 2 M solutions of sodium azide and potassium thiocyanate but not sodium or potassium chloride, in 1% deoxycholate but not digitonin, and in the denaturing solvents 6 M guanidine-HC1, 8 M urea, and 1% sodium dodecylsulfate at 100 degrees C. The protein found consistently when rootlets were solubilized migrated on SDS-polyacrylamide gels as a closely spaced doublet with apparent molecular weights of 230,000 and 250,000 daltons. This unique protein, distinct from tropocollagen or various muscle components, has been named ankyrin because of the rootlet's anchor-like function in the cell. PMID:1090630

  19. Thyroid hormones differentially affect sarcoplasmic reticulum function in rat atria and ventricles.

    PubMed

    Kaasik, A; Minajeva, A; Paju, K; Eimre, M; Seppet, E K

    1997-11-01

    The present study was undertaken to compare the effects of hypothyroidism and hyperthyroidism on sarcoplasmic reticulum (SR) Ca(2+)-pump activity, together with assessment of the functional role of SR in providing activator Ca2+ under these altered thyroid states. In response to a shift from hypothyroid to hyperthyroid state, a 10 fold and 2 fold increase in SR Ca(2+)-pump activity in atria and ventricles, respectively, were observed. This was associated with the 8-9 fold increases in atrial contractility (+dT/dt) and relaxation (-dT/dt), but only with a 3-4 fold increase in their ventricular counterparts. Also, the recirculation fraction of activator Ca2+ (RFA) increased to a far greater extent in atria (4 fold) than in papillary muscles, and the relative increment in inhibition of developed tension by ryanodine became 3 times larger in atria than in papillary muscles. A positive force-frequency relationship (FFR) was observed in hypothyroid atria, whereas the hyperthyroid atria, hypothyroid and hyperthyroid papillary muscles showed a negative FFR. These results suggest the greater role of transsarcolemmal (SL) Ca2+ and smaller role of SR Ca2+ in activating contraction in hypothyroid atria compared to other preparations. Thyroid hormones decrease the contribution of SL and increase that of SR in providing activator Ca2+ to the greater extent in atria than in ventricles. This effect of thyroid hormones is based on larger stimulation of SR Ca(2+)-pump in atria compared to ventricles.

  20. Striated clast pavements: Products of deforming subglacial sediment?

    NASA Astrophysics Data System (ADS)

    Clark, Peter U.

    1991-05-01

    Studies of modern glaciers have recently drawn attention to the importance of subglacial sediment deformation to glacier dynamics and processes. Inferences regarding the probable shear strength of this sediment imply that large clasts may in some cases sink to underlying rigid sediment, where abrasion by overlying deforming sediment could occur. This scenario provides a formative mechanism for striated clast pavements commonly described from the base of fine-grained massive diamictons associated with the late Pleistocene Laurentide ice sheet. Such a mechanism indicates that, at the time of formation of clast pavements, overlying diamictons associated with pavements had a low yield strength (≤0.5 kPa) and were deforming mechanically like a debris flow. Clast pavements may therefore be an important criterion for recognition of sediments deposited by subglacial deformation transport.

  1. Weakly nonlinear ion waves in striated electron temperatures

    NASA Astrophysics Data System (ADS)

    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.

  2. Model simulation of the SPOC wave in a bundle of striated myofibrils.

    PubMed

    Nakagome, Koutaro; Sato, Katsuhiko; Shintani, Seine A; Ishiwata, Shin'ichi

    2016-01-01

    SPOC (spontaneous oscillatory contraction) is a phenomenon observed in striated muscle under intermediate activation conditions. Recently, we constructed a theoretical model of SPOC for a sarcomere, a unit sarcomere model, which explains the behavior of SPOC at each sarcomere level. We also constructed a single myofibril model, which visco-elastically connects the unit model in series, and explains the behaviors of SPOC at the myofibril level. In the present study, to understand the SPOC properties in a bundle of myofibrils, we extended the single myofibril model to a two-dimensional (2D) model and a three-dimensional (3D) model, in which myofibrils were elastically connected side-by-side through cross-linkers between the Z-lines and M-lines. These 2D and 3D myofibril models could reproduce various patterns of SPOC waves experimentally observed in a 2D sheet and a 3D bundle of myofibrils only by choosing different values of elastic constants of the cross-linkers and the external spring. The results of these 2D and 3D myofibril models provide insight into the SPOC properties of the higher-ordered assembly of myofibrils.

  3. Model simulation of the SPOC wave in a bundle of striated myofibrils

    PubMed Central

    Nakagome, Koutaro; Sato, Katsuhiko; Shintani, Seine A.; Ishiwata, Shin’ichi

    2016-01-01

    SPOC (spontaneous oscillatory contraction) is a phenomenon observed in striated muscle under intermediate activation conditions. Recently, we constructed a theoretical model of SPOC for a sarcomere, a unit sarcomere model, which explains the behavior of SPOC at each sarcomere level. We also constructed a single myofibril model, which visco-elastically connects the unit model in series, and explains the behaviors of SPOC at the myofibril level. In the present study, to understand the SPOC properties in a bundle of myofibrils, we extended the single myofibril model to a two-dimensional (2D) model and a three-dimensional (3D) model, in which myofibrils were elastically connected side-by-side through cross-linkers between the Z-lines and M-lines. These 2D and 3D myofibril models could reproduce various patterns of SPOC waves experimentally observed in a 2D sheet and a 3D bundle of myofibrils only by choosing different values of elastic constants of the cross-linkers and the external spring. The results of these 2D and 3D myofibril models provide insight into the SPOC properties of the higher-ordered assembly of myofibrils. PMID:27924277

  4. Affi-gel blue treatment simplifies the protein composition of sarcoplasmic reticulum vesicles.

    PubMed

    Papp, S; Dux, L; Martonosi, A

    1986-04-01

    Sarcoplasmic reticulum vesicles isolated by conventional techniques usually contain, in addition to the recognized sarcoplasmic reticulum components, several other proteins (phosphorylase, myosin, glyceraldehyde-3-phosphate dehydrogenase, etc.) in variable amounts; these proteins complicate the interpretation of chemical modification data. Incubation of sarcoplasmic reticulum vesicles with Affi-Gel blue particles for 1-4 h at 2 degrees C, followed by sedimentation of the Affi-Gel in a clinical centrifuge, simplifies the protein composition by selective adsorption of the accessory proteins, and improves the consistency of the preparations. The Affi-Gel blue treatment is recommended as part of the standard procedure for the isolation of sarcoplasmic reticulum vesicles.

  5. Calcium uptake and ATPase activity of sarcoplasmic reticulum vesicles isolated from control and selenium deficient lambs.

    PubMed

    Tripp, M J; Whanger, P D; Schmitz, J A

    1993-06-01

    The calcium uptake and ATPase activity were studied using fragmented sarcoplasmic reticulum (FSR) vesicles from normal and selenium (vitamin E)--deficient lambs. The latter group was suffering from white muscle disease (WMD). The calcium uptake of FSR vesicles from muscle of WMD lambs was reduced 10-fold as compared to those from normal lambs. An inverse relationship was found with the calcium uptake ability of the FSR vesicles and the severity of WMD. ATPase activity was nonsignificantly lower in vesicles from WMD lambs. The most active FSR vesicles from both normal and WMD lambs banded at 27% when purified on linear sucrose density gradients. The number of protein bands appearing in acrylamide gels of the purified vesicles appeared to be directly proportional to the severity of WMD. The 75Se cosedimented with the calcium uptake and ATPase activity when FSR vesicles from a lamb injected with 75Se-selenite were subjected to linear sucrose density gradient centrifugation, suggesting that selenium is incorporated into these vesicles. Injection of selenium into WMD lambs resulted in significantly greater calcium uptake activity in vesicles 18 and 38 days later as compared with untreated WMD lambs. Injection of selenium in WMD lambs resulted in a marked decrease in plasma CPK activity and a significant increase of glutathione peroxidase activity in the blood.

  6. Striated spectral activity in Jovian and Saturnian radio emission

    NASA Technical Reports Server (NTRS)

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

    1988-01-01

    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.

  7. 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

  8. 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.

  9. [Striate receptive fields mapped with single and bipartite stimuli].

    PubMed

    Lazareva, N A; Shevelev, I A; Saltykov, K A; Novikova, R V; Tikhomirov, A S; Sharaev, G A; Tsutskiridze, D Iu; Eĭdeland, P V

    2008-01-01

    In 22 acute experiments with anesthetized and immobilized adult cats, 364 maps of receptive fields (RF) of 47 striate neurons were obtained by means of single local stimuli flashed at different parts of the visual field, or with additional asynchronous activation of the RF excitatory center with oscillating bar of the optimal orientation. Under bipartite stimulation, considerable and significant decrease in the square and weight of the central excitatory RF zone was revealed in more then 75% of the studied cells. Additional excitatory zones appeared in 54% of cases, or the square and weight of the excitatory zones substantially increased, and inhibitory zones developed in 90% of cases. These effects were correlated with the degree of increase in the background firing during transition from the mode of mapping with single stimulation to that with bipartite stimulation. The mechanism and possible functional role of cooperative excitatory and inhibitory intracortical interactions in organization of receptive fields and detection of features of a visual image are discussed.

  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. Copyright © 2016 Elsevier Ltd. All rights reserved.

  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. Procaine effects on single sarcoplasmic reticulum Ca2+ release channels.

    PubMed Central

    Zahradníková, A; Palade, P

    1993-01-01

    The effects of the Ca(2+)-induced Ca2+ release blocker procaine on individual sarcoplasmic reticulum Ca2+ release channels have been examined in planar lipid bilayers. Procaine did not reduce the single channel conductance nor appreciably shorten the mean open times of the channel; rather, it increased the longest closed time. These results indicated that procaine interacted selectively with a closed state of the channel rather than with an open state. Gating of the sarcoplasmic reticulum Ca2+ release channel was described by a modified scheme of Ashley and Williams (1990. J. Gen. Physiol. 95:981-1005), including an additional long-lived closed state. Computer simulations determined that procaine was more likely to interact with this long-lived Ca(2+)-bound closed state of the channel rather than with other states of the channel. Simulations with the same model were also able to reproduce a prominent Ca(2+)-sensitive transition between "random" and "bursting" forms of gating of the channel, variations of which may account for "gearshift" behavior reported in studies with this and other single channels. PMID:8388270

  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. 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.

  15. Characteristics of sarcoplasmic proteins and their interaction with surimi and kamaboko gel.

    PubMed

    Jafarpour, A; Gorczyca, E M

    2009-01-01

    This study examined the effect of adding common carp sarcoplasmic proteins (Sp- P) on the gel characteristics of threadfin bream surimi and kamaboko while maintaining constant moisture and myofibrillar levels. Based on the temperature sweep test, which is involved in heating of surimi gel from 10 to 80 degrees C to monitor the viscoelastic properties, at temperature range of 40 to 50 degrees C, the decrease level (depth of valley) in storage modulus (G') thermograph was in proportion to the concentration of added Sp- P. Storage modulus (G') showed greater elasticity after adding Sp- P compared with the control without Sp- P. Furthermore, the breaking force and distance and consequently gel strength of the resultant kamaboko were improved significantly (P > 0.05). Thus, added Sp- P did not interfere with myofibrillar proteins during sol-gel transition phase but associated with textural quality enhancement of resultant kamaboko; however, addition of Sp- P from the dark muscle of the carp decreased the whiteness of the resultant surimi. Furthermore, according to the SEM micrographs, the gel strength could not be associated with either the number of polygonal structures/mm(2) or the area of the polygonal structures in the kamaboko gel microstructure.

  16. Transport mechanism of the sarcoplasmic reticulum Ca2+ -ATPase pump.

    PubMed

    Møller, Jesper V; Nissen, Poul; Sørensen, Thomas L-M; le Maire, Marc

    2005-08-01

    The sarcoplasmic reticulum Ca(2+)-ATPase (SERCA1a) belongs to the group of P-type ATPases, which actively transport inorganic cations across membranes at the expense of ATP hydrolysis. Three-dimensional structures of several transport intermediates of SERCA1a, stabilized by structural analogues of ATP and phosphoryl groups, are now available at atomic resolution. This has enabled the transport cycle of the protein to be described, including the coupling of Ca(2+) occlusion and phosphorylation by ATP, and of proton counter-transport and dephosphorylation. From these structures, Ca(2+)-ATPase gradually emerges as a molecular mechanical device in which some of the transmembrane segments perform Ca(2+) transport by piston-like movements and by the transmission of reciprocating movements that affect the chemical reactivity of the cytosolic globular domains.

  17. Prevention of endotoxin-induced sarcoplasmic reticulum calcium leak improves mitochondrial and myocardial dysfunction.

    PubMed

    Hassoun, Sidi Mohammed; Marechal, Xavier; Montaigne, David; Bouazza, Youcef; Decoster, Brigitte; Lancel, Steve; Neviere, Remi

    2008-09-01

    Growing evidence suggests that mitochondria function is impaired in sepsis. Here, we tested the hypothesis that lipopolysaccharide would induce mitochondrial Ca2+ overload and oxygen utilization abnormalities as consequences of sarcoplasmic reticulum Ca2+ handling derangements that are typically observed in sepsis. As lipopolysaccharide-induced sarcoplasmic reticulum dysfunction was mainly characterized by reduced sarcoplasmic reticulum Ca2+ uptake and Ca2+ leak, we tested whether dantrolene, a sarco(endo)plasmic reticulum calcium ATPase leak inhibitor, would prevent mitochondrial and cardiac contractile dysfunction. Randomized controlled trial. Experimental laboratory. Male Sprague Dawley rats. Sepsis was induced by injection of endotoxin lipopolysaccharide (10 mg/kg/intravenously). Assessment of contractile function and Ca2+ handling was performed 4 hr after lipopolysaccharide. The relative contribution of the different Ca2+ transporters to relaxation in intact cardiomyocytes was studied during successive electrically evoked twitches and caffeine stimulation. Sarcoplasmic reticulum vesicles and mitochondria from ventricles of rats treated or not with lipopolysaccharide were prepared to evaluate Ca2+ uptake-release and oxygen fluxes, respectively. Effects of dantrolene (10 mg/kg) treatment in rats were evaluated in sarcoplasmic reticulum vesicles, mitochondria, and isolated hearts. Lipopolysaccharide challenge elicited cardiac contractile dysfunction that was accompanied by severe derangements in sarcoplasmic reticulum function, i.e., reduced Ca2+ uptake and increased sarcoplasmic reticulum Ca2+ leak. Functional sarcoplasmic reticulum changes were associated with modification in the status of phospholamban phosphorylation whereas SERCA was unchanged. Rises in mitochondrial Ca2+ content observed in lipopolysaccharide-treated rats coincided with derangements in mitochondrial oxygen efficacy, i.e., reduced respiratory control ratio. Administration of dantrolene in

  18. Development of binocular vision in the kitten's striate cortex.

    PubMed

    Freeman, R D; Ohzawa, I

    1992-12-01

    Studies of the development and plasticity of the visual pathway are well documented, but a basic question remains open: what is the physiological status of the system prior to extensive visual experience? Somewhat conflicting answers have been put forward, and in a major area, binocular vision, reports have ranged from severe immaturity to well-developed maturity. This is an important question to resolve since binocular cells in the visual cortex are thought to be the neural substrate for stereoscopic depth perception. We have addressed this question by recording from single cells in the striate cortex of kittens at postnatal ages 2, 3, and 4 weeks and from adults for comparison. Gratings with sinusoidal luminance distribution are presented to left, right, or both eyes. For each cell, we determine optimal values for orientation and spatial frequency. Relative phase (retinal disparity) is then varied in a dichoptic sequence so that binocular interaction may be studied. Results are as follows. In the normal adult, we have shown in previous work that most binocular interaction in the visual cortex can be accounted for on the basis of linear summation. Results from 3 and 4 week postnatal kittens are closely similar to those from adults. All types of binocular interaction found in adults are present in kittens. This includes phase-specific and non-phase-specific suppression or facilitation. Furthermore, monocular and binocular tuning characteristics are comparable in kittens and adults. The clear changes that occur with age are optimal spatial frequencies and peak responses. In addition, at 2 weeks, there is a substantially higher proportion of monocular cells compared to other ages and correspondingly, lower relative numbers of cells that exhibit phase-specific or suppressive binocular interactions. From increases in optimal spatial frequency and interpupillary distance with age, we calculated predicted changes in binocular disparity thresholds (stereo acuity) with age

  19. Sarcoplasmic reticulum calcium mobilization in right ventricular pressure-overload hypertrophy in the ferret: relationships to diastolic dysfunction and a negative treppe.

    PubMed

    Gwathmey, J K; Morgan, J P

    1993-03-01

    In a model of right-ventricular pressure-overload hypertrophy (POH) in the ferret, action potential duration (to 90% repolarization) was found to be significantly longer (228 +/- 11 vs 314 +/- 12 ms) with no change in amplitude (85 +/- 3 vs 85 +/- 2 mV) or resting membrane potential (-79 +/- 1.5 vs -79 +/- 1 mV) for control and POH, respectively. Peak sarcoplasmic reticulum Ca2+ release (expressed as the logarithm of the fractional luminescence, -4.2 +/- 0.1 vs -4.4 +/- 0.3) and resting calcium concentrations (-5.5 +/- 0.1 vs -5.7 +/- 0.1) were not different between the two groups (control vs POH respectively). Muscles from control and POH animals demonstrated a positive force/interval relationship in the presence of physiological extracellular [Ca2+]. However, unlike muscles from control animals, muscles from animals with POH subjected to increasing frequencies of contraction in the presence of increased extracellular [Ca2+] demonstrated further impairment of diastolic relaxation and a negative treppe. Exposure of muscles from POH animals to isoproterenol returned the slowed Ca2+ uptake by the sarcoplasmic reticulum as detected with aequorin to control values, although the relaxation phase of the isometric twitch remained prolonged compared to non-hypertrophied muscles. Exposure to milrinone also abbreviated the time course of the intracellular Ca2+ transient, but did not return it to that seen in normal myocardium. The exposure of non-hypertrophied isolated muscles to caffeine resulted in similar prolongation of the isometric twitch duration to that seen in hypertrophied myocardium. Results of these experiments suggest that impaired muscle relaxation in POH reflects changes at the level of the myofilaments. Thus, although slowed intracellular calcium mobilization contributes to diastolic relaxation abnormalities, it can not be the sole factor responsible for the slowed relaxation as has been suggested.

  20. Paradoxical buffering of calcium by calsequestrin demonstrated for the calcium store of skeletal muscle

    PubMed Central

    Royer, Leandro; Sztretye, Monika; Manno, Carlo; Pouvreau, Sandrine; Zhou, Jingsong; Knollmann, Bjorn C.; Protasi, Feliciano; Allen, Paul D.

    2010-01-01

    Contractile activation in striated muscles requires a Ca2+ reservoir of large capacity inside the sarcoplasmic reticulum (SR), presumably the protein calsequestrin. The buffering power of calsequestrin in vitro has a paradoxical dependence on [Ca2+] that should be valuable for function. Here, we demonstrate that this dependence is present in living cells. Ca2+ signals elicited by membrane depolarization under voltage clamp were compared in single skeletal fibers of wild-type (WT) and double (d) Casq-null mice, which lack both calsequestrin isoforms. In nulls, Ca2+ release started normally, but the store depleted much more rapidly than in the WT. This deficit was reflected in the evolution of SR evacuability, E, which is directly proportional to SR Ca2+ permeability and inversely to its Ca2+ buffering power, B. In WT mice E starts low and increases progressively as the SR is depleted. In dCasq-nulls, E started high and decreased upon Ca2+ depletion. An elevated E in nulls is consistent with the decrease in B expected upon deletion of calsequestrin. The different value and time course of E in cells without calsequestrin indicate that the normal evolution of E reflects loss of B upon SR Ca2+ depletion. Decrement of B upon SR depletion was supported further. When SR calcium was reduced by exposure to low extracellular [Ca2+], release kinetics in the WT became similar to that in the dCasq-null. E became much higher, similar to that of null cells. These results indicate that calsequestrin not only stores Ca2+, but also varies its affinity in ways that progressively increase the ability of the store to deliver Ca2+ as it becomes depleted, a novel feedback mechanism of potentially valuable functional implications. The study revealed a surprisingly modest loss of Ca2+ storage capacity in null cells, which may reflect concurrent changes, rather than detract from the physiological importance of calsequestrin. PMID:20713548

  1. Membrane asymmetry and enhanced ultrastructural detail of sarcoplasmic reticulum revealed with use of tannic acid

    PubMed Central

    1978-01-01

    Fixation of purified sarcoplasmic reticulum (SR) membrane vesicles, using glutaraldehyde supplemented with 1% tannic acid, reveals newly visualized ultrastructure in thin sections. The trilaminar appearance of the membrane is highly asymmetric; the outer electron-opaque layer is appreciably wider (70 A) than the inner layer (20 A). The asymmetry is not referable to lack of penetration of the tannic acid since: (a) SR vesicles made permeable with 1 mM EDTA, pH 8.5, show similar asymmetry; (b) treatment of SR with trypsin results in progressive loss in protein content and decrease in the thickness of the outer layer, until in the limit the trilayer has a symmetric appearance; (c) within the same muscle section, the SR membrane appears highly asymmetric whereas the sarcolemma has a more symmetric appearance; (d) reconstituted SR vesicles have a symmetric appearance with equally broad inner and outer layers (approximately 70 A); the symmetric structure is confirmed by freeze-fracture and negative staining electron microscopy. Heavy and light SR vesicles obtained by isopycnic density sedimentation of purified SR have the same asymmetric appearance of the membrane and seem to differ mainly in that the heavy vesicles contain internal contents consisting largely of Ca++-binding protein. The asymmetry of the SR membrane is referable mainly to the unidirectional alignment of the Ca++ pump protein, the major component (90% of the protein) of the membrane. The asymmetry of the SR membrane can be visualized now for the first time in situ in thin sections of muscle. PMID:83321

  2. ELECTRICAL AND MECHANICAL CHARACTERISTICS OF A VERY FAST LOBSTER MUSCLE

    PubMed Central

    Mendelson, Martin

    1969-01-01

    The remotor muscle of the second antenna of the American lobster is functionally divided into two parts. One part produces slow, powerful contractions and is used for postural control. The other part produces very brief twitches, can follow frequencies over 100/sec without fusion and is probably used for sound production. This great speed is due, in part, to synchronous arrival of nerve impulses at multiple terminals, a very brief membrane electrical response and electrical continuity throughout large volumes of sarcoplasm. Calculations indicate that the very extensive sarcoplasmic reticulum is probably responsible for the rapid decline of tension in this muscle. PMID:5792339

  3. Voltage-sensitive dyes reveal a modular organization in monkey striate cortex

    NASA Astrophysics Data System (ADS)

    Blasdel, Gary G.; Salama, Guy

    1986-06-01

    Voltage-sensitive dyes allow neuronal activity to be studied by non-invasive optical techniques. They provide an attractive means of investigating striate cortex, where important response properties are organized in two dimensions. In the present study, patterns of ocular dominance and orientation selectivity were obtained repeatedly from the same patch of cortex using the dye merocyanine oxazolone, together with current image-processing techniques. The patterns observed agree with most established features of monkey striate cortex and suggest a new unit of cortical organization; one that is modular in structure and which appears to link the organization of orientation selectivity with that of ocular dominance.

  4. The reaction of N-(1-pyrene)maleimide with sarcoplasmic reticulum.

    PubMed Central

    Papp, S; Kracke, G; Joshi, N; Martonosi, A

    1986-01-01

    The excimer fluorescence of the adduct of N-(1-pyrene)maleimide (PMI) with the Ca2+-ATPase was proposed as a probe of ATPase-ATPase interactions in sarcoplasmic reticulum (Lüdi and Hasselbach, Eur. J. Biochem., 1983, 130:5-8). We tested this proposition by analyzing the spectral properties and stoichiometry of the adducts of pyrenemaleimide with sarcoplasmic reticulum and with dithiothreitol and by comparing the effects of various detergents on the excimer fluorescence of the two adducts, with their influence on the sedimentation characteristics, ATPase activity, and light scattering of the pyrenemaleimide-labeled sarcoplasmic reticulum. These studies indicate that pyrenemaleimide reacts nearly randomly with several SH groups on the Ca2+-ATPase, and suggest that the observed excimer fluorescence of pyrenemaleimide-labeled sarcoplasmic reticulum may reflect intramolecular phenomena rather than ATPase-ATPase interactions. Further work is required to establish the relative contribution of intra- and intermolecular mechanisms to the excimer fluorescence. PMID:2937461

  5. Altered sarcoplasmic reticulum calcium cycling—targets for heart failure therapy

    PubMed Central

    Kho, Changwon; Lee, Ahyoung; Hajjar, Roger J.

    2013-01-01

    Cardiac myocyte function is dependent on the synchronized movements of Ca2+ into and out of the cell, as well as between the cytosol and sarcoplasmic reticulum. These movements determine cardiac rhythm and regulate excitation–contraction coupling. Ca2+ cycling is mediated by a number of critical Ca2+-handling proteins and transporters, such as L-type Ca2+ channels (LTCCs) and sodium/calcium exchangers in the sarcolemma, and sarcoplasmic/endoplasmic reticulum calcium ATPase 2a (SERCA2a), ryanodine receptors, and cardiac phospholamban in the sarcoplasmic reticulum. The entry of Ca2+ into the cytosol through LTCCs activates the release of Ca2+ from the sarcoplasmic reticulum through ryanodine receptor channels and initiates myocyte contraction, whereas SERCA2a and cardiac phospholamban have a key role in sarcoplasmic reticulum Ca2+ sequesteration and myocyte relaxation. Excitation–contraction coupling is regulated by phosphorylation of Ca2+-handling proteins. Abnormalities in sarcoplasmic reticulum Ca2+ cycling are hallmarks of heart failure and contribute to the pathophysiology and progression of this disease. Correcting impaired intracellular Ca2+ cycling is a promising new approach for the treatment of heart failure. Novel therapeutic strategies that enhance myocyte Ca2+ homeostasis could prevent and reverse adverse cardiac remodeling and improve clinical outcomes in patients with heart failure. PMID:23090087

  6. 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.

  7. Role of Golgi derived clathrin coated vesicles in the transport of calsequestrin to the sarcoplasmic reticulum of developing myotubes

    SciTech Connect

    Thomas, K.M.

    1988-01-01

    The sarcoplasmic reticulum (SR) is the major intracellular calcium sequestering organelle in skeletal and heart muscle. Calsequestrin (CSQ) is a glycoprotein with a molecular weight of 42,000. This protein is located in the lumen of the SR and it binds Ca/sup 2 +/ to maintain the concentration of this cation in the lumen at 10/sup /minus/3/M. Highly purified coated vesicles (CVs) were isolated from chick muscle and a Western blot using polyclonal anti-CSQ revealed the presence of CSQ within the CVs. Another major protein in the SR, the Ca/sup 2 +/-ATPase, was not contained in CVs suggesting different routes of insertion into the SR. Cultured chick myotubes were labelled with Trans/sup 35/S-label contain (/sup 35/S)-methionine and (/sup 35/S)-cysteine to follow the transport of CSQ. Labelled CSQ remained high in the CVs until after 45 minutes of chase, then declined. The amount of labelled CSQ in the SR continued to rise over the chase period. No CSQ was secreted. All the CSQ in CVs and SR was sensitive to the activity of endoglycosidase H, and a significant fraction also bound wheat germ agglutinin. A small amount of CSQ was also co-transported with the muscle protein acetylcholinesterase within CVs. Non-secreted forms of acetylcholinesterase had the same carbohydrate structure as CSQ and were shown to be degraded in the SR.

  8. A mutation in the CASQ1 gene causes a vacuolar myopathy with accumulation of sarcoplasmic reticulum protein aggregates.

    PubMed

    Rossi, Daniela; Vezzani, Bianca; Galli, Lucia; Paolini, Cecilia; Toniolo, Luana; Pierantozzi, Enrico; Spinozzi, Simone; Barone, Virginia; Pegoraro, Elena; Bello, Luca; Cenacchi, Giovanna; Vattemi, Gaetano; Tomelleri, Giuliano; Ricci, Giulia; Siciliano, Gabriele; Protasi, Feliciano; Reggiani, Carlo; Sorrentino, Vincenzo

    2014-10-01

    A missense mutation in the calsequestrin-1 gene (CASQ1) was found in a group of patients with a myopathy characterized by weakness, fatigue, and the presence of large vacuoles containing characteristic inclusions resulting from the aggregation of sarcoplasmic reticulum (SR) proteins. The mutation affects a conserved aspartic acid in position 244 (p.Asp244Gly) located in one of the high-affinity Ca(2+) -binding sites of CASQ1 and alters the kinetics of Ca(2+) release in muscle fibers. Expression of the mutated CASQ1 protein in COS-7 cells showed a markedly reduced ability in forming elongated polymers, whereas both in cultured myotubes and in in vivo mouse fibers induced the formation of electron-dense SR vacuoles containing aggregates of the mutant CASQ1 protein that resemble those observed in muscle biopsies of patients. Altogether, these results support the view that a single missense mutation in the CASQ1 gene causes the formation of abnormal SR vacuoles containing aggregates of CASQ1, and other SR proteins, results in altered Ca(2+) release in skeletal muscle fibers, and, hence, is responsible for the clinical phenotype observed in these patients.

  9. 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.

  10. 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

  11. 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

  12. 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.

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

    PubMed

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

    2016-09-01

    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. To investigate the geniculocalcarine (GCT) and striate areas in primary glaucoma patients using region of interest (ROI) analysis of magnetization transfer ratio (MTR). 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. 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). 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.

  14. 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.

  15. Glycogen hyperaccumulation in white muscle fibres of RN- carrier pigs. A biochemical and ultrastructural study.

    PubMed

    Estrade, M; Vignon, X; Rock, E; Monin, G

    1993-02-01

    1. The dominant RN- gene affects meat quality of pigs by increasing the glycogen content of muscle. Glycogen localization was studied in Longissimus dorsi muscle from RN- carrier pigs (RN- pigs) and rn+ rn+ homozygous pigs (rn+ pigs). 2. Ultrastructural study showed an excess of glycogen in the sarcoplasm of white fibres from RN- pigs as compared to rn+ pigs. 3. Lysosome-enriched fractions extracted from muscles contained 6% of the tissue glycogen content in both types of pigs. The distribution of the glycogen particles between sarcoplasm and lysosomes appeared to be similar in both RN- and rn+ pig tissues. 4. White fibres from RN- pigs with an increased glycogen level showed two ultrastructural alterations: the sarcoplasmic compartment was abnormally enlarged and a large proportion of mitochondria was morphologically modified. 5. The RN- gene seems, therefore, to be associated with alterations in the glycolytic metabolism, in the distribution of proteic compartments and in the oxidative metabolism of white muscle fibres.

  16. Dystrophy of the diaphragmatic muscles in Holstein-Friesian steers.

    PubMed

    Nakamura, N

    1996-01-01

    Diaphragmatic muscles in two slaughtered Holstein-Friesian revealed slightly pale color, swelling, and stiffness on palpation. Histologically the muscle fibers showed internal nuclei, fiber-splitting, variation in diameter, central core-like structures, sarcoplasmic masses, and vacuolar degeneration. These lesions were the same as those in dystrophy of the diaphragmatic muscles in Holstein-Friesian cows. It was demonstrated that muscular dystrophy of the diaphragm in Holstein-Friesian cattle occurred also in males, probably by inheriting an autosomal recessive trait.

  17. Calcium Movements Inside the Sarcoplasmic Reticulum of Cardiac Myocytes

    PubMed Central

    Bers, Donald M.; Shannon, Thomas R.

    2013-01-01

    Sarcoplasmic reticulum (SR) Ca content ([Ca]SRT) is critical to both normal cardiac function and electrophysiology, and changes associated with pathology contribute to systolic and diastolic dysfunction and arrhythmias. The intra-SR free [Ca] ([Ca]SR) dictates the [Ca]SRT, the driving force for Ca release and regulates release channel gating. We discuss measurement of [Ca]SR and [Ca]SRT, how [Ca]SR regulates activation and termination of release, and how Ca diffuses within the SR and influences SR Ca release during excitation-contraction coupling, Ca sparks and Ca waves. The entire SR network is connected and its lumen is also continuous with the nuclear envelope. Rapid Ca diffusion within the SR could stabilize and balance local [Ca]SR within the myocyte, but restrictions to diffusion can create spatial inhomogeneities. Experimental measurements and mathematical models of [Ca]SR to date have greatly enriched our understanding of these [Ca]SR dynamics, but controversies exist and may stimulate new measurements and analysis. PMID:23321551

  18. The sarcoplasmic calcium pump - a most efficient ion translocating system.

    PubMed

    Hasselbach, W

    1977-04-21

    In contrast to the sodium-potassium transporting plasma membranes, the sarcoplasmic membranes (SR) are highly specialized structures into which only two major intrinsic proteins, a calcium transporting protein and a calcium binding protein are embedded. The calcium transporting protein is a highly asymmetric molecule. It binds two calcium ions with a very high affinity at its external, and two calcium ions with low affinity at the internal section of the molecule. ATP is bound with high afffinity to an external binding site, inducing a conformational change. When the vesicular membranes are exposed to solutions containing Ca++, Mg++ and ATP, ATP is hydrolyzed and simultaneously calcium ions are translocated from the external medium into the vesicular space. When calcium ions are translocated in the opposite direction, ATP is synthesized. The calcium-ATP ratio for ATP cleavage as well as for ATP synthesis is 2. Thus, the SR membranes can transform reversibly chemical into osmotical energy. Inward and outward movements of calcium ions are relatively slow processes connected with the appearance and disappearance of different phosphorylated intermediates. One phosphorylated intermediate is formed by phosphoryltransfer from ATP when calcium ions are present in the medium. In contrast, when calcium ions are absent from the external medium, two different intermediates can be formed by the incorporation of inorganic phosphate. Only when calcium ions present in the internal space of the vesicles are released, the incorporation of inorganic phosphate gives rise to an intermediate who phosphoryl group can be transferred to ADP.

  19. 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.

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

    PubMed Central

    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; Bruton, Joseph; Truffert, Andre; Lanner, Johanna T.; Skurvydas, Albertas; Westerblad, Håkan

    2015-01-01

    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) Ca2+ 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 Ca2+ leak at rest, and depressed force production due to impaired SR Ca2+ 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 Ca2+-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

  1. A region of the myosin rod important for interaction with paramyosin in Caenorhabditis elegans striated muscle.

    PubMed Central

    Hoppe, P E; Waterston, R H

    2000-01-01

    The precise arrangement of molecules within the thick filament, as well as the mechanisms by which this arrangement is specified, remains unclear. In this article, we have exploited a unique genetic interaction between one isoform of myosin heavy chain (MHC) and paramyosin in Caenorhabditis elegans to probe the molecular interaction between MHC and paramyosin in vivo. Using chimeric myosin constructs, we have defined a 322-residue region of the MHC A rod critical for suppression of the structural and motility defects associated with the unc-15(e73) allele. Chimeric constructs lacking this region of MHC A either fail to suppress, or act as dominant enhancers of, the e73 phenotype. Although the 322-residue region is required for suppression activity, our data suggest that sequences along the length of the rod also play a role in the isoform-specific interaction between MHC A and paramyosin. Our genetic and cell biological analyses of construct behavior suggest that the 322-residue region of MHC A is important for thick filament stability. We present a model in which this region mediates an avid interaction between MHC A and paramyosin in parallel arrangement in formation of the filament arms. PMID:11014812

  2. Calcium binding to Procambarus clarkii sarcoplasmic calcium binding protein splice variants.

    PubMed

    E Rohrback, Suzanne; Wheatly, Michele G; Gillen, Christopher M

    2015-01-01

    Sarcoplasmic calcium binding protein (SCP) is a high-affinity calcium buffering protein expressed in muscle of crayfish and other invertebrates. In previous work, we identified three splice variants of Procambarus clarkii SCP (pcSCP1a, pcSCP1b, and pcSCP1c) that differ in a 37 amino acid region that lies mainly between the 2nd and 3ed EF-hand calcium binding domain. To evaluate the function of the proteins encoded by the pcSCP1 transcripts, we produced recombinant pcSCP1 and used tryptophan fluorescence to characterize calcium binding. Tryptophan fluorescence of pcSCP1a decreased in response to increased calcium, while tryptophan fluorescence of the pcSCP1b and pcSCP1c variants increased. We estimated calcium binding constants and Hill coefficients with two different equations: the standard Hill equation and a modified Hill equation that accounts for contributions from two different tryptophans. The approaches gave similar results. Steady-state calcium binding constants (Kd) ranged from 2.7±0.7×10(-8)M to 5.6±0.1×10(-7)M, consistent with previous work. Variants displayed significantly different apparent calcium affinities, which were decreased in the presence of magnesium. Calcium Kd was lowest for pcSCP1a and highest for pcSCP1c. Site-directed mutagenesis of pcSCP1c residues to the amino acids of pcSCP1b decreased the calcium Kd, identifying residues outside the EF-hand domains that contribute to calcium binding in crayfish SCP.

  3. Alignment of sarcoplasmic reticulum-mitochondrial junctions with mitochondrial contact points.

    PubMed

    García-Pérez, Cecília; Schneider, Timothy G; Hajnóczky, György; Csordás, György

    2011-11-01

    Propagation of ryanodine receptor (RyR2)-derived Ca(2+) signals to the mitochondrial matrix supports oxidative ATP production or facilitates mitochondrial apoptosis in cardiac muscle. Ca(2+) transfer likely occurs locally at focal associations of the sarcoplasmic reticulum (SR) and mitochondria, which are secured by tethers. The outer mitochondrial membrane and inner mitochondrial membrane (OMM and IMM, respectively) also form tight focal contacts (contact points) that are enriched in voltage-dependent anion channels, the gates of OMM for Ca(2+). Contact points could offer the shortest Ca(2+) transfer route to the matrix; however, their alignment with the SR-OMM associations remains unclear. Here, in rat heart we have studied the distribution of mitochondria-associated SR in submitochondrial membrane fractions and evaluated the colocalization of SR-OMM associations with contact points using transmission electron microscopy. In a sucrose gradient designed for OMM purification, biochemical assays revealed lighter fractions enriched in OMM only and heavier fractions containing OMM, IMM, and SR markers. Pure OMM fractions were enriched in mitofusin 2, an ~80 kDa mitochondrial fusion protein and SR-mitochondrial tether candidate, whereas in fractions of OMM + IMM + SR, a lighter (~50 kDa) band detected by antibodies raised against the NH(2) terminus of mitofusin 2 was dominating. Transmission electron microscopy revealed mandatory presence of contact points at the junctional SR-mitochondrial interface versus a random presence along matching SR-free OMM segments. For each SR-mitochondrial junction at least one tether was attached to contact points. These data establish the contact points as anchorage sites for the SR-mitochondrial physical coupling. Close coupling of the SR, OMM, and IMM is likely to provide a favorable spatial arrangement for local ryanodine receptor-mitochondrial Ca(2+) signaling.

  4. [Effects of hypoxic acclimatization on myocardial sarcoplasmic reticulum ATPase and 45Ca2+ uptake in rats].

    PubMed

    Long, Chao-liang; Zhang, Yan-fang; Yin, Zhao-yun; Wang, Hai

    2005-08-01

    To study the effect of acute hypoxia and hypoxic acclimatization on myocardial function of rats. Eighteen male Wistar rats were randomly divided into three groups: normoxic control, acute hypoxia and intermittent hypoxic acclimatization group (n=6). After being exposed to hypoxia (8000 m) for 4 h before and after intermittent hypoxic acclimatization (3000 m and 5000 m, 14 d respectively, 4 h/d), the rats were decapitated and then myocardial sarcoplasmic reticulum (SR) were derived from cardiac muscles. Activities of Na+, K(+)-ATPase, Ca2+, Mg2(+)-ATPase in SR, phosphorylation of phospholamban (PLB) and the ability of 45Ca2+ uptake in SR were observed in all these three groups. 1) Hypoxia had no effects on the activity of Na+, K(+)-ATPase in rats myocardial SR of rats. 2) Compared with normoxic control rats, the activity of Ca2+, Mg2(+)-ATPase in myocardial SR of rats after acute hypoxia was reduced significantly (P<0.01). After intermittent hypoxic acclimatization, its activity increased significantly as compared with that of acute hypoxic rats (P<0.01). 3) The phosphorylation of PLB in acute hypoxic rats was reduced significantly compared with normoxic control rats. After intermittent hypoxic acclimatization, its phosphorylation was increased significantly compared with that of acute hypoxic rats. It suggests that hypoxic acclimatization could alleviate the inhibition of calcium pump. 4) The ability of 45Ca2+ uptake of SR in acute hypoxic rats was decreased significantly. After hypoxic acclimatization, its ability was strengthened significantly. These results suggest that the increased function of myocardial SR calcium pump, the strengthened phosphorylation of PLB to alleviate the inhibition of calcium pump and the increased function of Ca2+ transport in SR are the mechanisms of hypoxic acclimatization protecting cardiac functions from injury induced by severe hypoxia.

  5. 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

  6. 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.

  7. Affinity-chromatographic isolation and some properties of troponin C from different muscle types.

    PubMed Central

    Head, J F; Weeks, R A; Perry, S V

    1977-01-01

    1. The formation of a complex between troponin I and troponin C that is stable in 6M-urea and dependent on Ca2+ was demonstrated in extracts of vertebrate striated and smooth muscles. 2. A method using troponin I coupled to Sepharose is described for the rapid isolation of troponin C from striated and smooth muscles of vertebrates. 3. Troponin C of rabbit cardiac muscle differs significantly in amino acid composition from troponin C of skeletal muscle. The primary structures of troponin C of red and white skeletal muscle are very similar. 4. The troponin C-like protein isolated from rabbit uterus muscle has a slightly different amino acid composition, but possess many similar properties to the forms of troponin C isolated from other muscle types. 5. The electrophoretic mobilities of the I-troponin C complexes formed from components isolated from different muscle types are determined by the troponin I component. Images PLATE 2 PLATE 1 PMID:851428

  8. 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.

  9. Sarcolipin provides a novel muscle-based mechanism for adaptive thermogenesis.

    PubMed

    Gamu, Daniel; Bombardier, Eric; Smith, Ian C; Fajardo, Val A; Tupling, A Russell

    2014-07-01

    The sarco(endo)plasmic reticulum Ca-ATPase (SERCA) transports Ca into the sarcoplasmic reticulum lumen and contributes significantly to skeletal muscle metabolic rate. Sarcolipin (SLN) has been shown recently to uncouple Ca transport from adenosine triphosphate hydrolysis by SERCA. We have hypothesized that SLN provides a novel mechanism of adaptive thermogenesis within skeletal muscle and protects against diet-induced obesity.

  10. Hypernitrosylated ryanodine receptor calcium release channels are leaky in dystrophic muscle.

    PubMed

    Bellinger, Andrew M; Reiken, Steven; Carlson, Christian; Mongillo, Marco; Liu, Xiaoping; Rothman, Lisa; Matecki, Stefan; Lacampagne, Alain; Marks, Andrew R

    2009-03-01

    Duchenne muscular dystrophy is characterized by progressive muscle weakness and early death resulting from dystrophin deficiency. Loss of dystrophin results in disruption of a large dystrophin glycoprotein complex, leading to pathological calcium (Ca2+)-dependent signals that damage muscle cells. We have identified a structural and functional defect in the ryanodine receptor (RyR1), a sarcoplasmic reticulum Ca2+ release channel, in the mdx mouse model of muscular dystrophy that contributes to altered Ca2+ homeostasis in dystrophic muscles. RyR1 isolated from mdx skeletal muscle showed an age-dependent increase in S-nitrosylation coincident with dystrophic changes in the muscle. RyR1 S-nitrosylation depleted the channel complex of FKBP12 (also known as calstabin-1, for calcium channel stabilizing binding protein), resulting in 'leaky' channels. Preventing calstabin-1 depletion from RyR1 with S107, a compound that binds the RyR1 channel and enhances the binding affinity of calstabin-1 to the nitrosylated channel, inhibited sarcoplasmic reticulum Ca2+ leak, reduced biochemical and histological evidence of muscle damage, improved muscle function and increased exercise performance in mdx mice. On the basis of these findings, we propose that sarcoplasmic reticulum Ca2+ leak via RyR1 due to S-nitrosylation of the channel and calstabin-1 depletion contributes to muscle weakness in muscular dystrophy, and that preventing the RyR1-mediated sarcoplasmic reticulum Ca2+ leak may provide a new therapeutic approach.

  11. Inhibition of mouth skeletal muscle relaxation by flavonoids of Cistus ladanifer L.: a plant defense mechanism against herbivores.

    PubMed

    Sosa, T; Chaves, N; Alias, J C; Escudero, J C; Henao, F; Gutiérrez-Merino, C

    2004-06-01

    Cistus ladanifer exudate is a potent inhibitor of the sarcoplasmic reticulum Ca2+-ATPase (Ca2+-pump) of rabbit skeletal muscle, a well-established model for active transport that plays a leading role in skeletal muscle relaxation. The low concentration of exudate needed to produce 50% of the maximum inhibition of the sarcoplasmic reticulum Ca2+-ATPase activity, 40-60 microg/ml, suggests that eating only a few milligrams of C. ladanifer leaves can impair the relaxation of the mouth skeletal muscle of herbivores, as the exudate reaches up to 140 mg/g of dry leaves in summer season. The flavonoid fraction of the exudate accounts fully for the functional impairment of the sarcoplasmic reticulum produced by the exudate (up to a dose of 250-300 microg/ml). The flavonoids present in this exudate impair the skeletal muscle sarcoplasmic reticulum function at two different levels: (i) by inhibition of the Ca2+-ATPase activity, and (ii) by decreasing the steady state ATP-dependent Ca2+-accumulation. Among the exudate flavonoids, apigenin and 3,7-di-O-methyl kaempferol are the most potent inhibitors of the skeletal muscle sarcoplasmic reticulum. We conclude that the flavonoids of this exudate can elicit an avoidance reaction of the herbivores eating C. ladanifer leaves through impairment of mouth skeletal muscle relaxation.

  12. Gender differences in sarcoplasmic reticulum calcium loading after isoproterenol.

    PubMed

    Chen, Jarvis; Petranka, John; Yamamura, Ken; London, Robert E; Steenbergen, Charles; Murphy, Elizabeth

    2003-12-01

    Males exhibit enhanced myocardial ischemia-reperfusion injury versus females under hypercontractile conditions associated with increased sarcoplasmic reticulum (SR) Ca2+. We therefore examined whether there were gender differences in SR Ca2+. We used NMR Ca2+ indicator 1,2-bis(2-amino-5,6-difluorophenoxy)-ethane-N,N,N',N'-tetraacetic acid to measure SR Ca2+ in perfused rabbit hearts. Isoproterenol increased SR Ca2+ in males from a baseline of 1.13 +/- 0.07 to 1.52 +/- 0.24 mM (P < 0.05). Female hearts had basal SR Ca2+ that was not significantly different from males (1.04 +/- 0.03 mM), and addition of isoproterenol to females resulted in a time-averaged SR Ca2+ (0.97 +/- 0.07 mM) that was significantly less than in males. To confirm this difference, we measured caffeine-induced release of SR Ca2+ with fura-2 in isolated ventricular myocytes. Ca2+ release after caffeine in untreated male myocytes was 377 +/- 41 nM and increased to 650 +/- 55 nM in isoproterenol-treated myocytes (P < 0.05). Ca2+ release after caffeine addition in untreated females was 376 +/- 27 nM and increased to 503 +/- 49 nM with isoproterenol, significantly less than in male myocytes treated with isoproterenol (P < 0.05). Treatment of female myocytes with NG-nitro-l-arginine methyl ester, an inhibitor of nitric oxide synthase (NOS), resulted in higher SR Ca2+ release than that measured in females treated only with isoproterenol and was not significantly different from that measured in males with isoproterenol. Female myocytes also have significantly higher levels of neuronal NOS. This gender difference in SR Ca2+ handling may contribute to reduced ischemia-reperfusion injury observed in females.

  13. Molecular dynamics in mouse atrial tumor sarcoplasmic reticulum.

    PubMed Central

    Voss, J C; Mahaney, J E; Jones, L R; Thomas, D D

    1995-01-01

    We have determined directly the effects of the inhibitory peptide phospholamban (PLB) on the rotational dynamics of the calcium pump (Ca-ATPase) of cardiac sarcoplasmic reticulum (SR). This was accomplished by comparing mouse ventricular SR, which has PLB levels similar to those found in other mammals, with mouse atrial SR, which is effectively devoid of PLB and thus has much higher (unregulated) calcium pump activity. To obtain sufficient quantities of atrial SR, we isolated the membranes from atrial tumor cells. We used time-resolved phosphorescence anisotropy of an erythrosin isothiocyanate label attached selectively and rigidly to the Ca-ATPase, to detect the microsecond rotational motion of the Ca-ATPase in the two preparations. The time-resolved phosphorescence anisotropy decays of both preparations at 25 degrees C were multi-exponential, because of the presence of different oligomeric species. The rotational correlation times for the different oligomers were similar for the two preparations, but the total decay amplitude was substantially greater for atrial tumor SR, indicating that a smaller fraction of the Ca-ATPase molecules exists as large aggregates. Phosphorylation of PLB in ventricular SR decreased the population of large-scale Ca-ATPase aggregates to a level similar to that of atrial tumor SR. Lipid chain mobility (fluidity), detected by electron paramagnetic resonance of stearic acid spin labels, was very similar in the two preparations, indicating that the higher protein mobility in atrial tumor SR is not due to higher lipid fluidity. We conclude that PLB inhibits by inducing Ca-ATPase lateral aggregation, which can be relieved either by phosphorylating or removing PLB. Images FIGURE 1 FIGURE 2 PMID:7612820

  14. Dynamic Calcium Movement inside Cardiac Sarcoplasmic Reticulum during Release

    PubMed Central

    Picht, Eckard; Zima, Aleksey V.; Shannon, Thomas R.; Duncan, Alexis M.; Blatter, Lothar A.; Bers, Donald M.

    2011-01-01

    Rationale Intra-sarcoplasmic reticulum (SR) free [Ca] ([Ca]SR) provides the driving force for SR Ca release and is a key regulator of SR Ca release channel gating during normal SR Ca release or arrhythmogenic spontaneous Ca release events. However, little is known about [Ca]SR spatiotemporal dynamics. Objective To directly measure local [Ca]SR with subsarcomeric spatiotemporal resolution during both normal global SR Ca release and spontaneous Ca sparks, and to evaluate the quantitative implications of spatial [Ca]SR gradients. Methods & Results Intact and permeabilized rabbit ventricular myocytes were subjected to direct simultaneous measurement of cytosolic [Ca] and [Ca]SR and fluorescence recovery after photobleach (FRAP). We found no detectable [Ca]SR gradients between SR release sites (junctional SR) and Ca uptake sites (free SR) during normal global Ca release, clear spatio-temporal [Ca]SR gradients during isolated Ca blinks, faster intra-SR diffusion in the longitudinal vs. transverse direction, 3-4 fold slower diffusion of fluorophores in the SR than in cytosol, and that intra-SR Ca diffusion varies locally, dependent on local SR connectivity. A computational model clarified why spatiotemporal gradients are more detectable in isolated local releases vs. global releases and provides a quantitative framework for understanding intra-SR Ca diffusion. Conclusions Intra-SR Ca diffusion is rapid, limiting spatial [Ca]SR gradients during excitation-contraction coupling. Spatiotemporal [Ca]SR gradients are apparent during Ca sparks, and these observations constrain models of dynamic Ca movement inside the SR. This has important implications for myocyte SR Ca handling, synchrony and potentially arrhythmogenic spontaneous contraction. PMID:21311044

  15. Comparative Biomechanics of Thick Filaments and Thin Filaments with Functional Consequences for Muscle Contraction

    PubMed Central

    Miller, Mark S.; Tanner, Bertrand C. W.; Nyland, Lori R.; Vigoreaux, Jim O.

    2010-01-01

    The scaffold of striated muscle is predominantly comprised of myosin and actin polymers known as thick filaments and thin filaments, respectively. The roles these filaments play in muscle contraction are well known, but the extent to which variations in filament mechanical properties influence muscle function is not fully understood. Here we review information on the material properties of thick filaments, thin filaments, and their primary constituents; we also discuss ways in which mechanical properties of filaments impact muscle performance. PMID:20625489

  16. DisAp-dependent striated fiber elongation is required to organize ciliary arrays.

    PubMed

    Galati, Domenico F; Bonney, Stephanie; Kronenberg, Zev; Clarissa, Christina; Yandell, Mark; Elde, Nels C; Jerka-Dziadosz, Maria; Giddings, Thomas H; Frankel, Joseph; Pearson, Chad G

    2014-12-22

    Cilia-organizing basal bodies (BBs) are microtubule scaffolds that are visibly asymmetrical because they have attached auxiliary structures, such as striated fibers. In multiciliated cells, BB orientation aligns to ensure coherent ciliary beating, but the mechanisms that maintain BB orientation are unclear. For the first time in Tetrahymena thermophila, we use comparative whole-genome sequencing to identify the mutation in the BB disorientation mutant disA-1. disA-1 abolishes the localization of the novel protein DisAp to T. thermophila striated fibers (kinetodesmal fibers; KFs), which is consistent with DisAp's similarity to the striated fiber protein SF-assemblin. We demonstrate that DisAp is required for KFs to elongate and to resist BB disorientation in response to ciliary forces. Newly formed BBs move along KFs as they approach their cortical attachment sites. However, because they contain short KFs that are rotated, BBs in disA-1 cells display aberrant spacing and disorientation. Therefore, DisAp is a novel KF component that is essential for force-dependent KF elongation and BB orientation in multiciliary arrays. © 2014 Galati et al.

  17. Chronic electrical stimulation drives mitochondrial biogenesis in skeletal muscle of a lizard, Varanus exanthematicus.

    PubMed

    Schaeffer, Paul J; Nichols, Scott D; Lindstedt, Stan L

    2007-10-01

    We investigated the capacity for phenotypic plasticity of skeletal muscle from Varanus exanthematicus, the savannah monitor lizard. Iliofibularis muscle from one leg of each lizard was electrically stimulated for 8 weeks. Both stimulated and contralateral control muscles were collected and processed for electron microscopy. We used stereological analysis of muscle cross-sections to quantify the volume densities of contractile elements, sarcoplasmic reticulum, mitochondria and intracellular lipids. We found that mitochondrial volume density was approximately fourfold higher in the stimulated muscle compared to controls, which were similar to previously reported values. Sarcoplasmic reticulum volume density was reduced by an amount similar to the increase in mitochondrial volume density while the volume density of contractile elements remained unchanged. Intracellular lipid accumulation was visibly apparent in many stimulated muscle sections but the volume density of lipids did not reach a significant difference. Although monitor lizards lack the highly developed aerobic metabolism of mammals, they appear to possess the capacity for muscle plasticity.

  18. 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.

  19. Muscle spindles in the human bulbospongiosus and ischiocavernosus muscles.

    PubMed

    Peikert, Kevin; May, Christian Albrecht

    2015-07-01

    Muscle spindles are crucial for neuronal regulation of striated muscles, but their presence and involvement in the superficial perineal muscles is not known. Bulbospongiosus and ischiocavernosus muscle specimens were obtained from 31 human cadavers. Serial sections were stained with hematoxylin and eosin, Sirius red, antibodies against Podocalyxin, myosin heavy chain isoforms (MyHC-slow tonic, S46; MyHC-2a/2x, A4.74), and neurofilament for the purpose of muscle spindle screening, counting, and characterization. A low but consistent number of spindles were detected in both muscles. The muscles contained few intrafusal fibers, but otherwise showed normal spindle morphology. The extrafusal fibers of both muscles were small in diameter. The presence of muscle spindles in bulbospongiosus and ischiocavernosus muscles supports physiological models of pelvic floor regulation and may provide a basis for further clinical observations regarding sexual function and micturition. The small number of muscle spindles points to a minor level of proprioceptive regulation. © 2014 Wiley Periodicals, Inc.

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

    PubMed

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

    2016-09-01

    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 Ca(2+)-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 Ca(2+) 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.

  1. 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

  2. Differential mechanism of the effects of ester-type local anesthetics on sarcoplasmic reticulum Ca-ATPase.

    PubMed

    Sánchez, G A; Di Croce, D E; de la Cal, C; Richard, S B; Takara, D

    2013-12-01

    The effect of the local anesthetics procaine and tetracaine on sarcoplasmic reticulum membranes isolated from two masticatory muscles, masseter and medial pterygoid, was tested and compared to fast-twitch muscles. The effects of the anesthetics on Ca-ATPase activity, calcium binding, uptake, and phosphorylation of the enzyme by inorganic phosphate (Pi) were tested with radioisotopic methods. Calcium binding to the Ca-ATPase was non-competitively inhibited, and the enzymatic activity decreased in a concentration-dependent manner. The inhibition of the activity depended on pH, calcium concentration, the presence of the calcium ionophore calcimycin, and the membrane protein concentration. Unlike fast-twitch membranes, the pre-exposure of the masseter and medial pterygoid membranes to the anesthetics enhanced the enzymatic activity in the absence of calcimycin, supporting their permeabilizing effect. Procaine and tetracaine also interfered with the calcium transport capability, decreasing the maximal uptake without modification of the calcium affinity for the ATPase. Besides, the anesthetics inhibited the phosphorylation of the enzyme by Pi in a competitive manner. Tetracaine revealed a higher inhibitory potency on Ca-ATPase compared to procaine, and the inhibitory concentrations were lower than usual clinical doses. It is concluded that procaine and tetracaine not only affect key steps of the Ca-ATPase enzymatic cycle but also exert an indirect effect on membrane permeability to calcium and suggest that the consequent myoplasmic calcium increase induced by the anesthetics might account for myotoxic effects, such as sustained contraction and eventual rigidity of both fast-twitch and masticatory muscles.

  3. Residual sarcoplasmic reticulum Ca2+ concentration after Ca2+ release in skeletal myofibers from young adult and old mice.

    PubMed

    Wang, Zhong-Min; Tang, Shen; Messi, María Laura; Yang, Jenny J; Delbono, Osvaldo

    2012-04-01

    Contrasting information suggests either almost complete depletion of sarcoplasmic reticulum (SR) Ca(2+) or significant residual Ca(2+) concentration after prolonged depolarization of the skeletal muscle fiber. The primary obstacle to resolving this controversy is the lack of genetically encoded Ca(2+) indicators targeted to the SR that exhibit low-Ca(2+) affinity, a fast biosensor: Ca(2+) off-rate reaction, and can be expressed in myofibers from adult and older adult mammalian species. This work used the recently designed low-affinity Ca(2+) sensor (Kd = 1.66 mM in the myofiber) CatchER (calcium sensor for detecting high concentrations in the ER) targeted to the SR, to investigate whether prolonged skeletal muscle fiber depolarization significantly alters residual SR Ca(2+) with aging. We found CatchER a proper tool to investigate SR Ca(2+) depletion in young adult and older adult mice, consistently tracking SR luminal Ca(2+) release in response to brief and repetitive stimulation. We evoked SR Ca(2+) release in whole-cell voltage-clamped flexor digitorum brevis muscle fibers from young and old FVB mice and tested the maximal SR Ca(2+) release by directly activating the ryanodine receptor (RyR1) with 4-chloro-m-cresol in the same myofibers. Here, we report for the first time that the Ca(2+) remaining in the SR after prolonged depolarization (2 s) in myofibers from aging (~220 μM) was larger than young (~132 μM) mice. These experiments indicate that SR Ca(2+) is far from fully depleted under physiological conditions throughout life, and support the concept of excitation-contraction uncoupling in functional senescent myofibers.

  4. Sarcoplasmic reticulum-mitochondria communication in cardiovascular pathophysiology.

    PubMed

    Lopez-Crisosto, Camila; Pennanen, Christian; Vasquez-Trincado, Cesar; Morales, Pablo E; Bravo-Sagua, Roberto; Quest, Andrew F G; Chiong, Mario; Lavandero, Sergio

    2017-06-01

    Repetitive, calcium-mediated contractile activity renders cardiomyocytes critically dependent on a sustained energy supply and adequate calcium buffering, both of which are provided by mitochondria. Moreover, in vascular smooth muscle cells, mitochondrial metabolism modulates cell growth and proliferation, whereas cytosolic calcium levels regulate the arterial vascular tone. Physical and functional communication between mitochondria and sarco/endoplasmic reticulum and balanced mitochondrial dynamics seem to have a critical role for optimal calcium transfer to mitochondria, which is crucial in calcium homeostasis and mitochondrial metabolism in both types of muscle cells. Moreover, mitochondrial dysfunction has been associated with myocardial damage and dysregulation of vascular smooth muscle proliferation. Therefore, sarco/endoplasmic reticulum-mitochondria coupling and mitochondrial dynamics are now viewed as relevant factors in the pathogenesis of cardiac and vascular diseases, including coronary artery disease, heart failure, and pulmonary arterial hypertension. In this Review, we summarize the evidence related to the role of sarco/endoplasmic reticulum-mitochondria communication in cardiac and vascular muscle physiology, with a focus on how perturbations contribute to the pathogenesis of cardiovascular disorders.

  5. Diversity of Cnidarian Muscles: Function, Anatomy, Development and Regeneration

    PubMed Central

    Leclère, Lucas; Röttinger, Eric

    2017-01-01

    The ability to perform muscle contractions is one of the most important and distinctive features of eumetazoans. As the sister group to bilaterians, cnidarians (sea anemones, corals, jellyfish, and hydroids) hold an informative phylogenetic position for understanding muscle evolution. Here, we review current knowledge on muscle function, diversity, development, regeneration and evolution in cnidarians. Cnidarian muscles are involved in various activities, such as feeding, escape, locomotion and defense, in close association with the nervous system. This variety is reflected in the large diversity of muscle organizations found in Cnidaria. Smooth epithelial muscle is thought to be the most common type, and is inferred to be the ancestral muscle type for Cnidaria, while striated muscle fibers and non-epithelial myocytes would have been convergently acquired within Cnidaria. Current knowledge of cnidarian muscle development and its regeneration is limited. While orthologs of myogenic regulatory factors such as MyoD have yet to be found in cnidarian genomes, striated muscle formation potentially involves well-conserved myogenic genes, such as twist and mef2. Although satellite cells have yet to be identified in cnidarians, muscle plasticity (e.g., de- and re-differentiation, fiber repolarization) in a regenerative context and its potential role during regeneration has started to be addressed in a few cnidarian systems. The development of novel tools to study those organisms has created new opportunities to investigate in depth the development and regeneration of cnidarian muscle cells and how they contribute to the regenerative process. PMID:28168188

  6. Diversity of Cnidarian Muscles: Function, Anatomy, Development and Regeneration.

    PubMed

    Leclère, Lucas; Röttinger, Eric

    2016-01-01

    The ability to perform muscle contractions is one of the most important and distinctive features of eumetazoans. As the sister group to bilaterians, cnidarians (sea anemones, corals, jellyfish, and hydroids) hold an informative phylogenetic position for understanding muscle evolution. Here, we review current knowledge on muscle function, diversity, development, regeneration and evolution in cnidarians. Cnidarian muscles are involved in various activities, such as feeding, escape, locomotion and defense, in close association with the nervous system. This variety is reflected in the large diversity of muscle organizations found in Cnidaria. Smooth epithelial muscle is thought to be the most common type, and is inferred to be the ancestral muscle type for Cnidaria, while striated muscle fibers and non-epithelial myocytes would have been convergently acquired within Cnidaria. Current knowledge of cnidarian muscle development and its regeneration is limited. While orthologs of myogenic regulatory factors such as MyoD have yet to be found in cnidarian genomes, striated muscle formation potentially involves well-conserved myogenic genes, such as twist and mef2. Although satellite cells have yet to be identified in cnidarians, muscle plasticity (e.g., de- and re-differentiation, fiber repolarization) in a regenerative context and its potential role during regeneration has started to be addressed in a few cnidarian systems. The development of novel tools to study those organisms has created new opportunities to investigate in depth the development and regeneration of cnidarian muscle cells and how they contribute to the regenerative process.

  7. Sphingosylphosphocholine modulates the ryanodine receptor/calcium-release channel of cardiac sarcoplasmic reticulum membranes.

    PubMed Central

    Betto, R; Teresi, A; Turcato, F; Salviati, G; Sabbadini, R A; Krown, K; Glembotski, C C; Kindman, L A; Dettbarn, C; Pereon, Y; Yasui, K; Palade, P T

    1997-01-01

    Sphingosylphosphocholine (SPC) modulates Ca2+ release from isolated cardiac sarcoplasmic reticulum membranes; 50 microM SPC induces the release of 70 80% of the accumulated calcium. SPC release calcium from cardiac sarcoplasmic reticulum through the ryanodine receptor, since the release is inhibited by the ryanodine receptor channel antagonists ryanodine. Ruthenium Red and sphingosine. In intact cardiac myocytes, even in the absence of extracellular calcium. SPC causes a rise in diastolic Ca2+, which is greatly reduced when the sarcoplasmic reticulum is depleted of Ca2+ by prior thapsigargin treatment. SPC action on the ryanodine receptor is Ca(2+)-dependent. SPC shifts to the left the Ca(2+)-dependence of [3H]ryanodine binding, but only at high pCa values, suggesting that SPC might increase the sensitivity to calcium of the Ca(2+)-induced Ca(2+)-release mechanism. At high calcium concentrations (pCa 4.0 or lower), where [3H]ryanodine binding is maximally stimulated, no effect of SPC is observed. We conclude that SPC releases calcium from cardiac sarcoplasmic reticulum membranes by activating the ryanodine receptor and possibly another intracellular Ca(2+)-release channel, the sphingolipid Ca(2+)-release-mediating protein of endoplasmic reticulum (SCaMPER) [Mao, Kim, Almenoff, Rudner, Kearney and Kindman (1996) Proc.Natl.Acad.Sci. U.S.A 93, 1993-1996], which we have identified for the first time in cardiac tissue. PMID:9078280

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

    USDA-ARS?s Scientific Manuscript database

    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 ...

  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. 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.…

  11. Altered Ca2+ sparks in aging skeletal and cardiac muscle

    PubMed Central

    Weisleder, Noah; Ma, Jianjie

    2008-01-01

    Ca2+ sparks are the fundamental units that comprise Ca2+-induced Ca2+ release (CICR) in striated muscle cells. In cardiac muscle, spontaneous Ca2+ sparks underlie the rhythmic CICR activity during heart contraction. In skeletal muscle, Ca2+ sparks remain quiescent during the resting state and are activated in a plastic fashion to accommodate various levels of stress. With aging, the plastic Ca2+ spark signal becomes static in skeletal muscle, whereas loss of CICR control leads to leaky Ca2+ spark activity in aged cardiomyocytes. Ca2+ spark responses reflect the integrated function of the intracellular Ca2+ regulatory machinery centered around the triad or dyad junctional complexes of striated muscles, which harbor the principal molecular players of excitation-contraction coupling. This review highlights the contribution of age-related modification of the Ca2+ release machinery and the effect of membrane structure and membrane cross-talk on the altered Ca2+ spark signaling during aging of striated muscles. PMID:18272434

  12. Aldolase potentiates DIDS activation of the ryanodine receptor in rabbit skeletal sarcoplasmic reticulum

    PubMed Central

    Seo, In-Ra; Moh, Sang Hyun; Lee, Eun Hui; Meissner, Gerhard; Kim, Do Han

    2006-01-01

    DIDS (4,4′-di-isothiocyanostilbene-2,2′-disulfonate), an anion channel blocker, triggers Ca2+ release from skeletal muscle SR (sarcoplasmic reticulum). The present study characterized the effects of DIDS on rabbit skeletal single Ca2+-release channel/RyR1 (ryanodine receptor type 1) incorporated into a planar lipid bilayer. When junctional SR vesicles were used for channel incorporation (native RyR1), DIDS increased the mean Po (open probability) of RyR1 without affecting unitary conductance when Cs+ was used as the charge carrier. Lifetime analysis of single RyR1 activities showed that 10 μM DIDS induced reversible long-lived open events (Po=0.451±0.038) in the presence of 10 μM Ca2+, due mainly to a new third component for both open and closed time constants. However, when purified RyR1 was examined in the same condition, 10 μM DIDS became considerably less potent (Po=0.206±0.025), although the caffeine response was similar between native and purified RyR1. Hence we postulated that a DIDS-binding protein, essential for the DIDS sensitivity of RyR1, was lost during RyR1 purification. DIDS-affinity column chromatography of solubilized junctional SR, and MALDI–TOF (matrix-assisted laser-desorption ionization–time-of-flight) MS analysis of the affinity-column-associated proteins, identified four major DIDS-binding proteins in the SR fraction. Among them, aldolase was the only protein that greatly potentiated DIDS sensitivity. The association between RyR1 and aldolase was further confirmed by co-immunoprecipitation and aldolase-affinity batch-column chromatography. Taken together, we conclude that aldolase is physically associated with RyR1 and could confer a considerable potentiation of the DIDS effect on RyR1. PMID:16817780

  13. 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

  14. 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. Copyright © 2011 Elsevier Inc. All rights reserved.

  15. Primary structures of cysteine-containing peptides from the calcium ion-transporting adenosine triphosphatase of rabbit sarcoplasmic reticulum.

    PubMed Central

    Allen, G; Green, N M

    1978-01-01

    A preliminary investigation of the primary structure of the Ca(2+-transporting ATPase (adenosine triphosphatase) protein of rabbit skeletal-muscle sarcoplasmic reticulum is reported. The preparation of derivatives of delipidated protein in a form suitable for sequence analysis is described. Tryptic peptides containing S-carboxymethylcysteine residues were isolated from the reduced carboxymethylated protein, and their sequences were partially determined. The results are consistent with mol.wt. about 105000 for the polypeptide, and the absence of extended repeated lengths of sequence. The distribution of tryptophan and cysteine residues between large, aggregated peptides and soluble tryptic peptides shows that these residues are concentrated in different regions of the primary structure. This observation agrees with other evidence that these residues are, on the whole, widely separated in the native protein. The details of the procedures used to isolate the peptides, and the evidence for the determination of their sequences, are given Supplementary Publication SUP 50085 (30 pages), which has been deposited at the British Library Lending Division, Boston Spa, Wetherby, West Yorkshire LS23 7BQ, U.K., from whom copies can be obtained on the terms indicated in Biochem.J. (1978) 169, 5. PMID:151533

  16. Structural similarities of Na,K-ATPase and SERCA, the Ca(2+)-ATPase of the sarcoplasmic reticulum.

    PubMed Central

    Sweadner, K J; Donnet, C

    2001-01-01

    The crystal structure of SERCA1a (skeletal-muscle sarcoplasmic-reticulum/endoplasmic-reticulum Ca(2+)-ATPase) has recently been determined at 2.6 A (note 1 A = 0.1 nm) resolution [Toyoshima, Nakasako, Nomura and Ogawa (2000) Nature (London) 405, 647-655]. Other P-type ATPases are thought to share key features of the ATP hydrolysis site and a central core of transmembrane helices. Outside of these most-conserved segments, structural similarities are less certain, and predicted transmembrane topology differs between subclasses. In the present review the homologous regions of several representative P-type ATPases are aligned with the SERCA sequence and mapped on to the SERCA structure for comparison. Homology between SERCA and the Na,K-ATPase is more extensive than with any other ATPase, even PMCA, the Ca(2+)-ATPase of plasma membrane. Structural features of the Na,K-ATPase are projected on to the Ca(2+)-ATPase crystal structure to assess the likelihood that they share the same fold. Homology extends through all ten transmembrane spans, and most insertions and deletions are predicted to be at the surface. The locations of specific residues are examined, such as proteolytic cleavage sites, intramolecular cross-linking sites, and the binding sites of certain other proteins. On the whole, the similarity supports a shared fold, with some particular exceptions. PMID:11389677

  17. Model of Ca(2+) Concentration Controlled by Sarcoplasmic Reticulum of Skeletal Muscle, Using the State Transition

    DTIC Science & Technology

    2007-11-02

    SR T tubule half sarcomere CS voltage sensor on TT V-channel Ca2+ pump C-channel Ca2+ nerve impulse A B C D TC LSR Fig.2 Model...Aff = σ /τ. III. MODEL STRUCTURE Let us focus on a half sarcomere . We approximate the form as a cylinder with a height of 1.1 µm, a radius 0.5 µm and...a volume of 0.86 µm3 [5]. The half sarcomere is divided into 4 parts: CS, V-channels, C-channels and Ca2+ pumps as illustrated in Fig.2. The

  18. Fast drum strokes: novel and convergent features of sonic muscle ultrastructure, innervation, and motor neuron organization in the Pyramid Butterflyfish (Hemitaurichthys polylepis).

    PubMed

    Boyle, Kelly S; Dewan, Adam K; Tricas, Timothy C

    2013-04-01

    Sound production that is mediated by intrinsic or extrinsic swim bladder musculature has evolved multiple times in teleost fishes. Sonic muscles must contract rapidly and synchronously to compress the gas-filled bladder with sufficient velocity to produce sound. Muscle modifications that may promote rapid contraction include small fiber diameter, elaborate sarcoplasmic reticulum (SR), triads at the A-I boundary, and cores of sarcoplasm. The diversity of innervation patterns indicate that sonic muscles have independently evolved from different trunk muscle precursors. The analysis of sonic motor pathways in distantly related fishes is required to determine the relationships between sonic muscle evolution and function in acoustic signaling. We examined the ultrastructure of sonic and adjacent hypaxial muscle fibers and the distribution of sonic motor neurons in the coral reef Pyramid Butterflyfish (Chaetodontidae: Hemitaurichthys polylepis) that produces sound by contraction of extrinsic sonic muscles near the anterior swim bladder. Relative to adjacent hypaxial fibers, sonic muscle fibers were sparsely arranged among the endomysium, smaller in cross-section, had longer sarcomeres, a more elaborate SR, wider t-tubules, and more radially arranged myofibrils. Both sonic and non-sonic muscle fibers possessed triads at the Z-line, lacked sarcoplasmic cores, and had mitochondria among the myofibrils and concentrated within the peripheral sarcoplasm. Sonic muscles of this derived eutelost possess features convergent with other distant vocal taxa (other euteleosts and non-euteleosts): small fiber diameter, a well-developed SR, and radial myofibrils. In contrast with some sonic fishes, however, Pyramid Butterflyfish sonic muscles lack sarcoplasmic cores and A-I triads. Retrograde nerve label experiments show that sonic muscle is innervated by central and ventrolateral motor neurons associated with spinal nerves 1-3. This restricted distribution of sonic motor neurons in the

  19. Enhanced dihydropyridine receptor calcium channel activity restores muscle strength in JP45/CASQ1 double knockout mice

    PubMed Central

    Mosca, Barbara; Delbono, Osvaldo; Messi, Maria Laura; Bergamelli, Leda; Wang, Zhong-Min; Vukcevic, Mirko; Lopez, Ruben; Treves, Susan; Nishi, Miyuki; Takeshima, Hiroshi; Paolini, Cecilia; Martini, Marta; Rispoli, Giorgio; Protasi, Feliciano; Zorzato, Francesco

    2016-01-01

    Muscle strength declines with age in part due to a decline of Ca2+ release from sarcoplasmic reticulum calcium stores. Skeletal muscle dihydropyridine receptors (Cav1.1) initiate muscle contraction by activating ryanodine receptors in the sarcoplasmic reticulum. Cav1.1 channel activity is enhanced by a retrograde stimulatory signal delivered by the ryanodine receptor. JP45 is a membrane protein interacting with Cav1.1 and the sarcoplasmic reticulum Ca2+ storage protein calsequestrin (CASQ1). Here we show that JP45 and CASQ1 strengthen skeletal muscle contraction by modulating Cav1.1 channel activity. Using muscle fibres from JP45 and CASQ1 double knockout mice, we demonstrate that Ca2+ transients evoked by tetanic stimulation are the result of massive Ca2+ influx due to enhanced Cav1.1 channel activity, which restores muscle strength in JP45/CASQ1 double knockout mice. We envision that JP45 and CASQ1 may be candidate targets for the development of new therapeutic strategies against decay of skeletal muscle strength caused by a decrease in sarcoplasmi