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1. Mitochondrial phosphagen kinases support the volatile power demands of motor nerve terminals

   https://doi.org/10.1113/JP284872  

   Justs, Karlis A. ; Sempertegui, Sergio ; Riboul, Danielle V. ; Oliva, Carlos D. ; Durbin, Ryan J. ; Crill, Sarah ; Stawarski, Michal ; Su, Chenchen ; Renden, Robert B. ; Fily, Yaouen ; et al ( November 2023 , The Journal of Physiology)

   Motor neurons are the longest neurons in the body, with axon terminals separated from the soma by as much as a meter. These terminals are largely autonomous with regard to their bioenergetic metabolism and must burn energy at a high rate to sustain muscle contraction. Here, through computer simulation and drawing on previously published empirical data, we determined that motor neuron terminals inDrosophila larvae experience highly volatile power demands. It might not be surprising then, that we discovered the mitochondria in the motor neuron terminals of bothDrosophila and mice to be heavily decorated with phosphagen kinases ‐ a key element in an energy storage and buffering system well‐characterized in fast‐twitch muscle fibres. Knockdown of arginine kinase 1 (ArgK1) inDrosophilalarval motor neurons led to several bioenergetic deficits, including mitochondrial matrix acidification and a faster decline in the cytosol ATP to ADP ratio during axon burst firing.image

   Neurons commonly fire in bursts imposing highly volatile demands on the bioenergetic machinery that generates ATP.

   Using a computational approach, we built profiles of presynaptic power demand at the level of single action potentials, as well as the transition from rest to sustained activity.

   Phosphagen systems are known to buffer ATP levels in muscles and we demonstrate that phosphagen kinases, which support such phosphagen systems, also localize to mitochondria in motor nerve terminals of fruit flies and mice.

   By knocking down phosphagen kinases in fruit fly motor nerve terminals, and using fluorescent reporters of the ATP:ADP ratio, lactate, pH and Ca²⁺, we demonstrate a role for phosphagen kinases in stabilizing presynaptic ATP levels.

   These data indicate that the maintenance of phosphagen systems in motor neurons, and not just muscle, could be a beneficial initiative in sustaining musculoskeletal health and performance.

    

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2. The impact of measurement technique and sampling on estimates of skeletal muscle fibre architecture

   https://doi.org/10.1002/ar.25415  

   Taylor, Andrea B. ; Terhune, Claire E. ; Ross, Callum F. ; Vinyard, Christopher J. ( February 2024 , The Anatomical Record)

   Skeletal muscle fibre architecture provides important insights into performance of vertebrate locomotor and feeding behaviours. Chemical digestion and in situ sectioning of muscle bellies along their lengths to expose fibres, fibre orientation and intramuscular tendon, are two classical methods for estimating architectural variables such as fibre length (L_f) and physiological cross‐sectional area (PCSA). It has recently been proposed thatL_festimates are systematically shorter and hence less accurate using in situ sectioning. Here we addressed this hypothesis by comparingL_festimates between the two methods for the superficial masseter and temporalis muscles in a sample of strepsirrhine and platyrrhine primates. Means or single‐specimenL_festimates using chemical digestion were greater in 17/32 comparisons (53.13%), indicating the probability of achieving longer fibres using chemical digestion is no greater than chance in these taxonomic samples. We further explored the impact of sampling on scaling ofL_fand PCSA in platyrrhines applying a bootstrapping approach. We found that sampling—both numbers of individuals within species and representation of species across the clade significantly influence scaling results ofL_fand PCSA in platyrrhines. We show that intraspecific and clade sampling strategies can account for differences between previously published platyrrhine scaling studies. We suggest that differences in these two methodological approaches to assessing muscle architecture are relatively less consequential when estimatingL_fand PCSA for comparative studies, whereas achieving more reliable estimates within species through larger samples and representation of the full clade space are important considerations in comparative studies of fibre architecture and scaling.

    

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3. Lipin1 plays complementary roles in myofibre stability and regeneration in dystrophic muscles

   https://doi.org/10.1113/JP284085  

   Jama, Abdulrahman ; Alshudukhi, Abdullah A. ; Burke, Steve ; Dong, Lixin ; Kamau, John Karanja ; Voss, Andrew Alvin ; Ren, Hongmei ( February 2023 , The Journal of Physiology)

   Duchenne muscular dystrophy (DMD) is a severe muscle wasting disorder caused by dystrophin mutations, leading to the loss of sarcolemmal integrity, and resulting in progressive myofibre necrosis and impaired muscle function. Our previous studies suggest that lipin1 is important for skeletal muscle regeneration and myofibre integrity. Additionally, we discovered that mRNA expression levels of lipin1 were significantly reduced in skeletal muscle of DMD patients and the mdx mouse model. To understand the role of lipin1 in dystrophic muscle, we generated dystrophin/lipin1 double knockout (DKO) mice, and compared the limb muscle pathology and function of wild‐type B10, muscle‐specific lipin1 deficient (lipin1^Myf5cKO), mdx and DKO mice. We found that further knockout of lipin1 in dystrophic muscle exhibited a more severe phenotype characterized by increased necroptosis, fibrosis and exacerbated membrane damage in DKO compared to mdx mice. In barium chloride‐induced muscle injury, both lipin1^Myf5cKOand DKO showed prolonged regeneration at day 14 post‐injection, suggesting that lipin1 is critical for muscle regeneration.In situcontractile function assays showed that lipin1 deficiency in dystrophic muscle led to reduced specific force production. Using a cell culture system, we found that lipin1 deficiency led to elevated expression levels of necroptotic markers and medium creatine kinase, which could be a result of sarcolemmal damage. Most importantly, restoration of lipin1 inhibited the elevation of necroptotic markers in differentiated primary lipin1‐deficient myoblasts. Overall, our data suggests that lipin1 plays complementary roles in myofibre stability and muscle function in dystrophic muscles, and overexpression of lipin1 may serve as a potential therapeutic strategy for dystrophic muscles.image

   We identified that lipin1 mRNA expression levels are significantly reduced in skeletal muscles of Duchenne muscular dystrophy patients and mdx mice.

   We found that further depletion of lipin1 in skeletal muscles of mdx mice induces more severe dystrophic phenotypes, including enhanced myofibre sarcolemma damage, muscle necroptosis, inflammation, fibrosis and reduced specific force production.

   Lipin1 deficiency leads to elevated expression levels of necroptotic markers, whereas restoration of lipin1 inhibits their expression.

   Our results suggest that lipin1 is functionally complementary to dystrophin in muscle membrane integrity and muscle regeneration.

    

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4. A chloride channel blocker prevents the suppression by inorganic phosphate of the cytosolic calcium signals that control muscle contraction

   https://doi.org/10.1113/JP279917  

   Ferreira, Juan J. ; Pequera, Germán ; Launikonis, Bradley S. ; Ríos, Eduardo ; Brum, Gustavo ( October 2020 , The Journal of Physiology)

    Accumulation of inorganic phosphate (P_i) may contribute to muscle fatigue by precipitating calcium salts inside the sarcoplasmic reticulum (SR). Neither direct demonstration of this process nor definition of the entry pathway of P_iinto SR are fully established.

    We showed that P_ipromoted Ca²⁺release at concentrations below 10 mmand decreased it at higher concentrations. This decrease correlated well with that of [Ca²⁺]_SR.

    Pre‐treatment of permeabilized myofibres with 2 mmCl⁻channel blocker 9‐anthracenecarboxylic acid (9AC) inhibited both effects of P_i.

    The biphasic dependence of Ca²⁺release on [P_i] is explained by a direct effect of P_iacting on the SR Ca²⁺release channel, combined with the intra‐SR precipitation of Ca²⁺salts. The effects of 9AC demonstrate that P_ienters the SR via a Cl⁻pathway of an as‐yet‐undefined molecular nature.

   Fatiguing exercise causes hydrolysis of phosphocreatine, increasing the intracellular concentration of inorganic phosphate (P_i). P_idiffuses into the sarcoplasmic reticulum (SR) where it is believed to form insoluble Ca²⁺salts, thus contributing to the impairment of Ca²⁺release. Information on the P_ientrance pathway is still lacking. In amphibian muscles endowed with isoform 3 of the RyR channel, Ca²⁺spark frequency is correlated with the Ca²⁺load of the SR and can be used to monitor this variable. We studied the effects of P_ion Ca²⁺sparks in permeabilized fibres of the frog. Relative event frequency (f/f_ref) rose with increasing [P_i], reaching 2.54 ± 1.6 at 5 mm,and then decreased monotonically, reaching 0.09 ± 0.03 at [P_i] = 80 mm. Measurement of [Ca²⁺]_SRconfirmed a decrease correlated with spark frequency at high [P_i]. A large [Ca²⁺]_SRsurge was observed upon P_iremoval. Anion channels are a putative path for P_iinto the SR. We tested the effect of the chloride channel blocker 9‐anthracenecarboxylic acid (9AC) on P_ientrance. 9AC (400 µm)applied to the cytoplasm produced a non‐significant increase in spark frequency and reduced the P_ieffects on this parameter. Fibre treatment with 2 mm9AC in the presence of high cytoplasmic Mg²⁺suppressed the effects of P_ion [Ca²⁺]_SRand spark frequency up to 55 mm[P_i]. These results suggest that chloride channels (or transporters) provide the main pathway of inorganic phosphate into the SR and confirm that P_iimpairs Ca²⁺release by accumulating and precipitating with Ca²⁺inside the SR, thus contributing to myogenic fatigue.

    

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5. Scaling of axial muscle architecture in juvenile Alligator mississippiensis reveals an enhanced performance capacity of accessory breathing mechanisms

   https://doi.org/10.1111/joa.13523  

   Rose, Kayleigh A. R. ; Tickle, Peter G. ; Elsey, Ruth M. ; Sellers, William I. ; Crossley, II, Dane A. ; Codd, Jonathan R. ( July 2021 , Journal of Anatomy)

   Quantitative functional anatomy of amniote thoracic and abdominal regions is crucial to understanding constraints on and adaptations for facilitating simultaneous breathing and locomotion. Crocodilians have diverse locomotor modes and variable breathing mechanics facilitated by basal and derived (accessory) muscles. However, the inherent flexibility of these systems is not well studied, and the functional specialisation of the crocodilian trunk is yet to be investigated. Increases in body size and trunk stiffness would be expected to cause a disproportionate increase in muscle force demands and therefore constrain the basal costal aspiration mechanism, necessitating changes in respiratory mechanics. Here, we describe the anatomy of the trunk muscles, their properties that determine muscle performance (mass, length and physiological cross‐sectional area [PCSA]) and investigate their scaling in juvenileAlligator mississippiensisspanning an order of magnitude in body mass (359 g–5.5 kg). Comparatively, the expiratory muscles (transversus abdominis,rectus abdominis,iliocostalis), which compress the trunk, have greater relative PCSA being specialised for greater force‐generating capacity, while the inspiratory muscles (diaphragmaticus,truncocaudalis ischiotruncus,ischiopubis), which create negative internal pressure, have greater relative fascicle lengths, being adapted for greater working range and contraction velocity. Fascicle lengths of the accessorydiaphragmaticusscaled with positive allometry in the alligators examined, enhancing contractile capacity, in line with this muscle's ability to modulate both tidal volume and breathing frequency in response to energetic demand during terrestrial locomotion. Theiliocostalis, an accessory expiratory muscle, also demonstrated positive allometry in fascicle lengths and mass. All accessory muscles of the infrapubic abdominal wall demonstrated positive allometry in PCSA, which would enhance their force‐generating capacity. Conversely, the basal tetrapod expiratory pump (transversus abdominis) scaled isometrically, which may indicate a decreased reliance on this muscle with ontogeny. Collectively, these findings would support existing anecdotal evidence that crocodilians shift their breathing mechanics as they increase in size. Furthermore, the functional specialisation of thediaphragmaticusand compliance of the body wall in the lumbar region against which it works may contribute to low‐cost breathing in crocodilians.

    

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