Receptors In Skeletal Muscle Research Articles

Disturbed adiponectin – AMPK system in skeletal muscle of patients with metabolic syndrome

Patients with metabolic syndrome are characterized by low circulating adiponectin levels and reduced adiponectin sensitivity in skeletal muscles. Through binding on its main skeletal muscle receptor AdipoR1, adiponectin activates AMP-activated protein kinase (AMPK), a key player in energy homeostasis. Fourteen metabolic syndrome patients and seven healthy control subjects were included. Blood samples were taken to determine insulin resistance, adiponectin, lipoproteins, and C-reactive protein. Muscle biopsies (m. vastus lateralis) were obtained to assess mRNA expression of AdipoR1 and both AMPKα1 and AMPKα2 subunits, as well as downstream targets in lipid and glucose metabolism. Skeletal muscle mRNA expression of AMPKα1 and AMPKα2 was lower in metabolic syndrome patients (100 ± 6 vs. 122 ± 8 AU, p = 0.030 and 64 ± 4 vs. 85 ± 9 AU, p = 0.044, respectively), whereas the expression of AdipoR1 was upregulated (138 ± 9 vs. 105 ± 7, p = 0.012). AMPKα1 and AdipoR1 correlated positively in both the control (r = 0.964, p < 0.001) and the metabolic syndrome group (r = 0.600, p = 0.023). However, this relation was shifted upwards in metabolic syndrome patients, indicating increased AdipoR1mRNA expression for a similar AMPKα1 expression. Previously, a blunted stimulatory effect of adiponectin on AMPK activation has been shown in metabolic syndrome patients. The present data suggest that the disturbed interaction of adiponectin with AMPK is located downstream of the AdipoR1 receptor.

M.I.3 The role of laminins in myomatrix assembly and skeletal muscle stability

The basement membrane of skeletal muscle (myomatrix) consists of a complex network of highly glycosylated proteins. Important components of the myomatrix are the laminins and the collagens. During development, the myomatrix is assembled by interactions with cell surface receptors and by self-polymerization, which results in an extracellular scaffold important for skeletal muscle stability. Mutations in the α2 chain of laminin-211 impinge on myomatrix assembly and its interaction with cell surface receptors, and result in one of the most severe forms of congenital muscular dystrophy (MDC1A). Based on a detailed mechanistic understanding of the function of laminin-211, we have designed artificial linker molecules derived from agrin or perlecan with the aim to bridge the remaining myomatrix to cell surface receptors. In the conducted preclinical, proof-of-concept studies we were able to show that they can substantially ameliorate the disease phenotype in a mouse model of MDC1A. Subsequent approaches to further improve the treatment have focused on testing interventions that affect downstream pathways, such as apoptosis and inflammation, using pharmacological and genetic tools. All those treatments show additional beneficial effects on disease progression. However, semi-quantitative comparison of the treatments indicates that interventions that target early events in the disease are more efficacious than those targeting further downstream pathways. Finally, we will also report on the ongoing efforts to combine interventions that boost myomatrix assembly with those that connect it to skeletal muscle receptors. In summary, we will report on proof-of-concept studies that provide mechanistic insights and unequivocal preclinical evidence for the potential use of those therapies. As the MDC1A mice used in our studies share many hallmarks with the human disease, these preclinical data may be of high predictive value for future clinical studies.

Quantitative analysis of locomotor defects in neonatal mice lacking proprioceptive feedback

Proprioceptive feedback derived from specialized receptors in skeletal muscle is critical in forming an accurate map of limb position in space, and is used by the central nervous system to plan future movements and to determine accuracy of executed movements. Knockout mouse strains for genes expressed by proprioceptive sensory neurons have been generated that result in generalized motor deficits, but these deficits have not been quantitatively characterized. Here we characterize a conditional knockout mouse model wherein proprioceptive sensory neuron synaptic transmission has been blocked by selective ablation of munc18-1, a synaptic vesicle associated protein required for fusion of synaptic vesicles with the plasma membrane. Proprioceptive munc18-1 conditional mutants are impaired in surface righting – a dynamic postural adjustment task – and display several specific deficits in pivoting, an early locomotor behavior. Before the emergence of forward locomotion during postnatal development, animals explore their surroundings through pivoting, or rotating the upper torso around the relatively immobile base of the hind limbs. 3-D kinematic analysis was used to quantitatively describe this pivoting behavior at postnatal days 5 and 8 in control and munc18-1 conditional mutants. Mutant animals also pivot, but demonstrate alterations in movement strategy and in postural placement of the forelimbs during pivoting when compared to controls. In addition, brief forelimb stepping movements associated with pivoting are altered in mutant animals. Step duration and step height are increased in mutant animals. These results underscore the importance of proprioceptive feedback even at early stages in postnatal development.

Deletion of Growth Hormone Receptors in Postnatal Skeletal Muscle of Male Mice Does Not Alter Muscle Mass and Response to Pathological Injury

In this study, we investigated whether loss of GH receptor (GHR) signaling in postnatal skeletal muscle alters muscle mass and regenerative ability in adult mice and whether this was dependent on IGF-1 receptor (IGF-1R) signaling. To do so, we used mouse models with skeletal muscle-specific loss of GHR signaling (mGHRKO), IGF-1R and insulin receptor signaling (MKR), or both GHR and IGF-1R/insulin receptor signaling (mGHRKO/MKR). We did not find a reduction in muscle cross-sectional area, fiber type composition, or response to pathological muscle injury in male mGHRKO and mGHRKO/MKR mice when compared with control and MKR mice, respectively. This could potentially be explained by unchanged skeletal muscle Igf-1 expression in mGHRKO and mGHRKO/MKR mice relative to control and MKR mice, respectively. Furthermore, MKR and mGHRKO/MKR mice, but not mGHRKO mice, demonstrated reduced fiber fusion after cardiotoxin injection, suggesting that IGF-1, and not GH, promotes fiber fusion in adult mice. In summary, our data suggest that GHR signaling in postnatal skeletal muscle does not play a significant role in regulating muscle mass or muscle regeneration. Additionally, in our model, muscle Igf-1 expression is not dependent on GHR signaling in postnatal skeletal muscle.

Identification and characterization of a taxon-specific three-finger toxin from the venom of the Green Vinesnake (Oxybelis fulgidus; family Colubridae)

Snake venoms contain a variety of protein and peptide toxins, and the three-finger toxins (3FTxs) are among the best characterized family of venom proteins. The compact nature and highly conserved molecular fold of 3FTxs, together with their abundance in many venoms, has contributed to their utility in structure-function studies. Although many target the nicotinic acetylcholine receptor of vertebrate skeletal muscle, often binding with nanomolar Kds, several non-conventional 3FTxs show pronounced taxon-specific neurotoxic effects. Here we describe the purification and characterization of fulgimotoxin, a monomeric 3FTx from the venom of Oxybelis fulgidus, a neotropical rear-fanged snake. Fulgimotoxin retains the canonical 5 disulfides of the non-conventional 3FTxs and is highly neurotoxic to lizards; however, mice are unaffected, demonstrating that this toxin is taxon-specific in its effects. Analysis of structural features of fulgimotoxin and other colubrid venom 3FTxs indicate the presence of a "colubrid toxin motif" (CYTLY) and a second conserved segment (WAVK) found in Boiga and Oxybelis taxon-specific 3FTxs, both in loop II. Because specific residues in loop II conventional α-neurotoxic 3FTxs are intimately associated with receptor binding, we hypothesize that this loop, with its highly conserved substitutions, confers taxon-specific neurotoxicity. These findings underscore the importance of rear-fanged snake venoms for understanding the evolution of toxin molecules and demonstrate that even among well-characterized toxin families, novel structural and functional motifs may be found.

Egr3‐dependent muscle stretch receptor morphogenesis and innervation homeostasis

Muscle spindles are skeletal muscle receptors that mediate stretch reflexes and proprioception in vertebrates. Proprioception abnormalities are observed in many human neuropathies with unknown etiology and factors involved in sensory nerve‐stretch receptor homeostasis are poorly understood. During skeletal muscle development, sensory (Ia‐afferent) innervation is restricted to only a few myotubes which induces their transformation into intrafusal muscle fibers that form the stretch receptor core. The transcriptional regulator Egr3 is induced by sensory innervation and it has an essential role in stretch receptor morphogenesis, but whether its function is restricted to developing receptors and whether it has a role in innervation homeostasis is not known. Ablation of Egr3 using skeletal muscle specific knockout mice demonstrated its autonomous function in developing stretch receptors. Lineage tracing experiments using Egr3‐Cre knock‐in mice showed that stretch receptor differentiation is blocked in the absence of Egr3 and innervation fails to be maintained. Thus, Egr3 has an essential function in intrafusal muscle fiber differentiation and gene regulation required to maintain innervation homeostasis. (Support: NIH R01‐NS063078 and K26‐OD010945).

CL316,243, a Selective β3‐Adrenoceptor Agonist Activates Protein Synthesis Through mTOR/p70S6K Signaling Pathway in L6 Cells.

The existence of β3‐adrenergic receptors (AR) in skeletal muscle has been demonstrated by the detection of the β3‐mRNA and protein and by the selective effects β3‐AR agonists on metabolic oxidation and glucose utilization. However, the role of β3‐AR on protein metabolism in skeletal muscle remains still unclear. To determine whether β3‐AR regulates protein translation we evaluated in L6 skeletal muscle cells the effect of CL316,243, a selective β3‐AR agonist, on the rate of protein synthesis and on the activation of the Akt/mTOR/ p70S6K signaling pathway which is known to support the protein synthesis requirement during muscle growth. When L6 cells were incubated in the presence of CL 316,243, we observed a net increase of [35S] methionine incorporation into muscle cell proteins such myosin heavy chain and myosin light chain at 24 hr. The anabolic effect of CL 316,243 was associated with an increase of p70S6K which was inhibited by the β3‐AR antagonist, SR 59230A, but not by the β2‐AR antagonist ICI 118551. In addition, the increase in p70S6K levels induced by CL 316,243 was inhibited by wortmannin, a PI3K inhibitor, and by rapamycin, mTOR inhibitor suggesting that β3‐AR regulates protein synthesis via PI3K/Akt/ p70S6K signaling pathway. This finding unveils a new cellular mechanism by which β‐AR system exerts anabolic actions on skeletal protein metabolism and therefore regulates skeletal muscle mass.

Metabolic functions of glucocorticoid receptor in skeletal muscle

Glucocorticoids (GCs) exert key metabolic influences on skeletal muscle. GCs increase protein degradation and decrease protein synthesis. The released amino acids are mobilized from skeletal muscle to liver, where they serve as substrates for hepatic gluconeogenesis. This metabolic response is critical for mammals’ survival under stressful conditions, such as fasting and starvation. GCs suppress insulin-stimulated glucose uptake and utilization and glycogen synthesis, and play a permissive role for catecholamine-induced glycogenolysis, thus preserving the level of circulating glucose, the major energy source for the brain. However, chronic or excess exposure of GCs can induce muscle atrophy and insulin resistance. GCs convey their signal mainly through the intracellular glucocorticoid receptor (GR). While GR can act through different mechanisms, one of its major actions is to regulate the transcription of its primary target genes through genomic glucocorticoid response elements (GREs) by directly binding to DNA or tethering onto other DNA-binding transcription factors. These GR primary targets trigger physiological and pathological responses of GCs. Much progress has been made to understand how GCs regulate protein and glucose metabolism. In this review, we will discuss how GR primary target genes confer metabolic functions of GCs, and the mechanisms governing the transcriptional regulation of these targets. Comprehending these processes not only contributes to the fundamental understanding of mammalian physiology, but also will provide invaluable insight for improved GC therapeutics.

Transforming growth factor type beta 1 increases the expression of angiotensin II receptor type 2 by a SMAD‐ and p38 MAPK‐dependent mechanism in skeletal muscle

Excessive deposition of extracellular matrix (ECM) proteins, a condition known as fibrosis, is a hallmark of Duchenne muscular dystrophy. Among the factors that trigger muscle fibrosis are transforming growth factor beta (TGF-β) and angiotensin II (Ang-II). Ang-II belongs to the renin-angiotensin system, and its biological effects are exerted by Ang-II receptors type 1 and type 2 (AT-1 and AT-2, respectively). This study aims to determine the effect of TGF-β1 on the expression of AT-1 and AT-2 receptor in skeletal muscle. C2 C12 myoblasts exposed to TGF-β1 showed a dose-dependent increase in AT-2 expression but with no effect on AT-1 levels. Injection of TGF-β1 in the skeletal muscle of mice increased the levels of AT-2 and ECM protein but unchanged AT-1 levels. We also detected higher expression levels of AT-2 receptor in dystrophic skeletal muscle of mdx mice than in normal mice. The induction of AT-2 was mediated by the canonical TGF-β pathway because under the inhibitory conditions of the kinase activity of TGFβ receptor I or the knockdown of Smad2/3 levels, TGF-β-induced AT-2 receptor increase was strongly inhibited. Furthermore, we demonstrated that p38MAPK activity in response to TGF-β is also required for AT-2 increase as evaluated by a p38MAPK inhibitor. Our results show that the levels of AT-2 but not AT-1 receptor are modulated by the pro-fibrotic factor TGF-β1 in myoblasts and mouse skeletal muscle. This finding suggests that AT-2 might be involved in the physiopathology of fibrosis in dystrophic skeletal muscle.

AAV-mediated Overexpression of Human α7 Integrin Leads to Histological and Functional Improvement in Dystrophic Mice

Duchenne muscular dystrophy (DMD) is a severe muscle disease caused by mutations in the DMD gene, with loss of its gene product, dystrophin. Dystrophin helps link integral membrane proteins to the actin cytoskeleton and stabilizes the sarcolemma during muscle activity. We investigated an alternative therapeutic approach to dystrophin replacement by overexpressing human α7 integrin (ITGA7) using adeno-associated virus (AAV) delivery. ITGA7 is a laminin receptor in skeletal and cardiac muscle that links the extracellular matrix (ECM) to the actin skeleton. It is modestly upregulated in DMD muscle and has been proposed to be an important modifier of dystrophic symptoms. We delivered rAAV8.MCK.ITGA7 to the lower limb of mdx mice through isolated limb perfusion (ILP) of the femoral artery. We demonstrated ~50% of fibers in the tibialis anterior (TA) and extensor digitorum longus (EDL) overexpressing α7 integrin at the sarcolemma following AAV gene transfer. The increase in ITGA7 in skeletal muscle significantly protected against loss of force following eccentric contraction-induced injury compared with untreated (contralateral) muscles while specific force following tetanic contraction was unchanged. Reversal of additional dystrophic features included reduced Evans blue dye (EBD) uptake and increased muscle fiber diameter. Taken together, this data shows that rAAV8.MCK.ITGA7 gene transfer stabilizes the sarcolemma potentially preserving mdx muscle from further damage. This therapeutic approach demonstrates promise as a viable treatment for DMD with further implications for other forms of muscular dystrophy.

Physical activity opposes the age‐related increase in skeletal muscle and plasma endothelin‐1 levels and normalizes plasma endothelin‐1 levels in individuals with essential hypertension

Endothelin-1 has potent constrictor and proliferative activity in vascular smooth muscle, and essential hypertension and aging are associated with increased endothelin-1-mediated vasoconstrictor tone. The aim of this study was to investigate the effect of physical activity, hypertension and age on endothelin-1 levels in plasma and skeletal muscle and endothelin receptors in skeletal muscle in human subjects. In study 1, normotensive (46 ± 1 years, n = 11) and hypertensive (47 ± 1 years, n = 10) subjects were studied before and after 8 weeks of aerobic exercise training. In study 2, young (23 ± 1 years, n = 8), older lifelong sedentary (66 ± 2 years, n = 8) and older lifelong endurance-trained (62 ± 2 years, n = 8) subjects were studied in a cross-sectional design. Skeletal muscle and plasma endothelin-1 levels were increased with age and plasma endothelin-1 levels were higher in hypertensive than normotensive individuals. Eight weeks of exercise training normalized plasma endothelin-1 levels in the hypertensive subjects and increased the protein expression of the ET(A) receptor in skeletal muscle of normotensive subjects. Similarly, individuals that had performed lifelong physical activity had similar plasma and muscle endothelin-1 levels as the young controls and had higher ET(A) receptor levels. Our findings suggest that aerobic exercise training opposes the age-related increase in skeletal muscle and plasma endothelin-1 levels and normalizes plasma endothelin-1 levels in individuals with essential hypertension. This effect may explain some of the beneficial effects of training on the cardiovascular system in older and hypertensive subjects.

Human Aging and Expression of Proteins Interacting with the Ryanodine Receptor in Skeletal Muscle

Sarcopenia is characterised by reduced muscle mass and reduced force that is only partially attributable to muscle atrophy. Fiber type distribution changes with aging (fast twitch decreases, slow twitch increases). Excitation-contraction coupling (ECC) may be impaired by an uncoupling of the dihydropyridine receptor (DHPR) and the ryanodine receptor (RyR1), possibly due to decreased expression of the DHPR α1s subunit (Delbono et al., J Membr Biol. 1995;148:211-22). The DHPR β1a subunit, which may contribute to ECC (Rebbeck et al., Biophys J. 2011;100:922-30), increases (Taylor et al., Aging cell 2009;8:584-94). The 12KDa FK506 binding protein (FKBP12) stabilizes RyR1 and its dissociation may contribute to decreased ECC (Andersson et al., Cell Metab.2011; 14:196-207). Previous studies have used animal models; here we investigate the expression of these proteins in aging human muscle.Human muscle samples were obtained from 42 male and female donors (age 40-90) undergoing knee (vastus medialis) and hip (gluteus minimus or gluteus medius) replacements. Results of fiber type distribution in muscle homogenates from female subjects shows fast twitch fibers decreasing and slow twitch increasing with age. Preliminary data shows that levels of FKBP12 relative to RyR1, determined in microsomal vesicles using western blot and densitometry decrease with age. Expression levels of the DHPR α1s and β1a subunits relative to that of actin determined in muscle homogenates using western blot and densitometry are being analysed. We find that β1a subunit is mostly in the homogenate and not associated with membrane fractions. The preliminary data indicates that the levels of several of the proteins that associate with RyR1 and modulate its activity change with age and could affect the release of Ca2+ during ECC and muscle function in human sarcopenia.

The role and regulation of erythropoietin (EPO) and its receptor in skeletal muscle: how much do we really know?

Erythropoietin (EPO) primarily activates erythroid cell proliferation and growth and is active in several types of non-hematopoietic cells via its interaction with the EPO-receptor (EPO-R). This review focuses on the role of EPO in skeletal muscle. The EPO-R is expressed in skeletal muscle cells and EPO may promote myoblast differentiation and survival via the activation of the same signaling cascades as in hematopoietic cells, such as STAT5, MAPK and Akt. Inconsistent results exist with respect to the detection of the EPO-R mRNA and protein in muscle cells, tissue and across species and the use of non-specific EPO-R antibodies contributes to this problem. Additionally, the inability to reproducibly detect an activation of the known EPO-induced signaling pathways in skeletal muscle questions the functionality of the EPO-R in muscle in vivo. These equivocal findings make it difficult to distinguish between a direct effect of EPO on skeletal muscle, via the activation of its receptor, and an indirect effect resulting from a better oxygen supply to the muscle. Consequently, the precise role of EPO in skeletal muscle and its regulatory mechanism/s remain to be elucidated. Further studies are required to comprehensively establish the importance of EPO and its function in skeletal muscle health.

Three-Dimensional Localization of the α and β Subunits and of the II-III Loop in the Skeletal Muscle L-type Ca2+ Channel

The L-type Ca(2+) channel (dihydropyridine receptor (DHPR) in skeletal muscle acts as the voltage sensor for excitation-contraction coupling. To better resolve the spatial organization of the DHPR subunits (α(1s) or Ca(V)1.1, α(2), β(1a), δ1, and γ), we created transgenic mice expressing a recombinant β(1a) subunit with YFP and a biotin acceptor domain attached to its N- and C- termini, respectively. DHPR complexes were purified from skeletal muscle, negatively stained, imaged by electron microscopy, and subjected to single-particle image analysis. The resulting 19.1-Å resolution, three-dimensional reconstruction shows a main body of 17 × 11 × 8 nm with five corners along its perimeter. Two protrusions emerge from either face of the main body: the larger one attributed to the α(2)-δ1 subunit that forms a flexible hook-shaped feature and a smaller protrusion on the opposite side that corresponds to the II-III loop of Ca(V)1.1 as revealed by antibody labeling. Novel features discernible in the electron density accommodate the atomic coordinates of a voltage-gated sodium channel and of the β subunit in a single docking possibility that defines the α1-β interaction. The β subunit appears more closely associated to the membrane than expected, which may better account for both its role in localizing the α(1s) subunit to the membrane and its suggested role in excitation-contraction coupling.

Malignant Hyperthermia: Clinical and Molecular Aspects

Malignant hyperthermia (MH) is a potentially lethal pharmacogenetic disorder that affects genetically predisposed individuals. It manifests in susceptible individuals in response to exposure to Inhalant anesthetics, depolarizing muscle relaxants or extreme physical activity in hot environments. During exposure to these triggering agents, there is a rapid and sustained increase of myoplasmic calcium (Ca(2+)) concentration induced by hyperactivation of ryanodine receptor of skeletal muscle (RyR1), causing a profound change in Ca(2+) homeostasis, featuring a hypermetabolic state. RyR1, Ca(2+) release channels of sarcoplasmic reticulum, is the primary locus for MH susceptibility. Several mutations in the gene encoding the protein RyR1 have been identified; however, other genes may be involved. Actually, the standard method for diagnosing MH susceptibility is the muscle contracture test for exposure to halothane-caffeine (CHCT) and the only treatment is the use of dantrolene. However, with advances in molecular genetics, a full understanding of the disease etiology may be provided, favoring the development of an accurate diagnosis, less invasive, with DNA test, and also will provide the development of new therapeutic strategies for treatment of MH. Thus, this brief review aims to integrate molecular and clinical aspects of MH, gathering input for a better understanding of this channelopathy.

Debut de miastenia gravis en atención primaria. A propósito de un caso

Myasthenia gravis is an autoimmune disorder of neuromuscular transmission involving the production of autoantibodies directed against skeletal muscle receptors, in most cases of acetylcholine.Clinically it is characterized by the appearance of muscle weakness after prolonged activity, which tends to recover after a period of rest, or administration of acetylcholinesterase inhibitors.It is a relatively rare disease, although the prevalence has increased by improved diagnosis and increased longevity of the population. The diagnosis can be based on evidence after it is suspected using pharmacological, immunological or electrophysiology tests.Treatment can be divided into: symptomatic, short term and long term.We report the case of a patient who complained of diplopia, this muscle weakness being the most common initial symptom of the disease.

Role of Ca2 + in proteolysis-inducing factor (PIF)-induced atrophy of skeletal muscle

Proteolysis-inducing factor (PIF) induces muscle loss in cancer cachexia through a high affinity membrane bound receptor. This study investigates the mechanism by which the PIF receptor communicates to intracellular signalling pathways. C2C12 murine myoblasts were used as a model using PIF purified from MAC16 tumours. Calcium imaging was determined using fura-4-acetoxymethyl ester (Fura-4-AM). PIF induced a rapid rise in Ca2+i, which was completely attenuated by a anti-receptor antibody, or peptides representing 20 mers of the N-terminus of the PIF receptor. Other agents catabolic for skeletal muscle including angiotensin II (AngII) tumour necrosis factor-α (TNF-α) and lipopolysaccharide (LPS) also induced a rise in Ca2+i, but this was not attenuated by anti-PIF-receptor antibody. The rise in Ca2+i induced by PIF and AngII was completely attenuated by the Zn2+ chelator D-myo-inositol-1,2,6-triphosphate, and this was reversed by administration of exogenous Zn2+. The Ca2+i rise induced by PIF was independent of the presence of extracellular Ca2+, and attenuated by the Ca2+ pump inhibitor thapsigargin, suggesting that the Ca2+i rise was due to release from intracellular stores. This rise in Ca2+i induced by PIF was attenuated by both the phospholipase C inhibitor U73122 and 2-APB, an inhibitor of the inositol 1,4,5-triphosphate receptor, suggesting the involvement of a G-protein. Binding of the PIF to its receptor in skeletal muscle triggers a rise in Ca2+i, which initiates a signalling cascade leading to a depression in protein synthesis, and an increase in protein degradation.

No evidence for inositol 1,4,5-trisphosphate–dependent Ca2+ release in isolated fibers of adult mouse skeletal muscle

The presence and role of functional inositol 1,4,5-trisphosphate (IP3) receptors (IP3Rs) in adult skeletal muscle are controversial. The current consensus is that, in adult striated muscle, the relative amount of IP3Rs is too low and the kinetics of Ca2+ release from IP3R is too slow compared with ryanodine receptors to contribute to the Ca2+ transient during excitation–contraction coupling. However, it has been suggested that IP3-dependent Ca2+ release may be involved in signaling cascades leading to regulation of muscle gene expression. We have reinvestigated IP3-dependent Ca2+ release in isolated flexor digitorum brevis (FDB) muscle fibers from adult mice. Although Ca2+ transients were readily induced in cultured C2C12 muscle cells by (a) UTP stimulation, (b) direct injection of IP3, or (c) photolysis of membrane-permeant caged IP3, no statistically significant change in calcium signal was detected in adult FDB fibers. We conclude that the IP3–IP3R system does not appear to affect global calcium levels in adult mouse skeletal muscle.

Influence of short-term L-arginine supplementation on carbohydrate balance in rats with ischemia-reperfusion syndrome

BackgroundThere are studies showing stimulative effect of arginine on insulin secretion. This mechanism is not fully explained. The effects of the impact of arginine on carbohydrate balance under the conditions of ischemia and reperfusion remain to be determined. The aim of this study is the evaluation of the influence of short-term L-arginine supplementation on the concentration of glucose and insulin in blood and insulin binding in rat skeletal muscle under the conditions of ischemia and reperfusion. MethodsThe study was conducted on male Wistar rats with average body mass 250±30g. Animals were divided into four groups: Group I – control, Group II – placebo, Group III – L-arginine 500mg/kg/24h for 5 days, Group IV – L-arginine and L-NAME (75μmol/rat/24h) for 5 days. Each group was divided into subgroups depending on duration of ischemia and reperfusion. Acute ischemia of hind limb was induced in each group by putting pneumatic tourniquet on the thigh. Blood samples and skeletal muscles were collected from the rats. Plasma concentrations of glucose and insulin were measured. Insulin binding to insulin receptors was determined in skeletal muscle. ResultsA clear reduction of insulin binding to receptor was found in the group of animals without ischemia and the group supplemented with L-arginine and subjected to 4-h ischemia and 30- and 120-min reperfusion. A significant increase in insulin level was found in groups of animals with L-arginine and/or L-NAME subjected to 4-h ischemia at all times of reperfusion. Supplementation with L-arginine and/or L-NAME decreased levels of glucose in blood serum of animals undergoing ischemia-reperfusion syndrome compared to the control and placebo groups. ConclusionUnder conditions of ischemia-reperfusion, short-term administration of L-arginine causes a decrease in insulin binding capacity of insulin receptors in skeletal muscle, an increase in insulin level and a decrease in the concentration of glucose in blood serum.

Alternatively spliced variants of gamma-subunit of muscle-type acetylcholine receptor in fetal and adult skeletal muscle of mouse.

Gamma-subunit of nicotinic acetylcholine receptor is encoded by chrng gene of mouse. This gene is located on chromosome 1, spans 6.5kb, and contains 12 exons and 11 introns. Previous studies have reported three transcript variants (C1-3) produced by alternative splicing; C1 contains all the 12 reported exons, C2 uses an in-frame alternate splice site in exon-2, and C3 produced by exon-5 skipping. These variants differ in their channel kinetics and opening times. In our study, we report the presence of two new transcript variants (T1 and T2) of chrng expressed in mouse postnatal day 3 and adult skeletal muscles. These transcripts contain novel first coding exon either N1 or N2. N1 is located in the 5' UTR, while N2 is an extended exon-2. 5' extension of exon-2 contains an initiation codon which produces a novel transcript variant. Either of the two exons can splice with the internal exons to produce mature transcripts making different 5' ends of the transcripts. Consequently, the proteins encoded by these two transcripts differ at N-termini. The presence of N2 exon containing transcript was further supported by the availability of EST from the database. These new variants display heterogeneous properties. They differ in the presence of signal peptide, phosphorylation, and acetylation of their amino acid residues of the new N-termini of the gamma subunit.