Muscle-specific expression of PPARgamma coactivator-1alpha improves exercise performance and increases peak oxygen uptake.

The induction of peroxisome proliferator-activated receptor-gamma coactivator-1alpha (PGC-1alpha), a key regulator of mitochondriogenesis, is well-established under multiple physical exercise regimens, including, endurance, resistance, and sprint training. We wanted to determine if increased expression of PGC-1alpha in muscle is sufficient to improve performance during exercise in vivo. We demonstrate that muscle-specific expression of PGC-1alpha improves the performance during voluntary as well as forced exercise challenges. Additionally, PGC-1alpha transgenic mice exhibit an enhanced performance during a peak oxygen uptake exercise test, demonstrating an increased peak oxidative capacity, or whole body oxygen uptake. This increased ability to perform in multiple exercise paradigms is supported by enhanced mitochondrial function as suggested by increased mitochondrial gene expression, mitochondrial DNA, and mitochondrial enzyme activity. Thus this study demonstrates that upregulation of PGC-1alpha in muscle in vivo is sufficient to greatly improve exercise performance under various exercise paradigms as well as increase peak oxygen uptake.

A naturally occurring calcineurin variant inhibits FoxO activity and enhances skeletal muscle regeneration.

The calcium-activated phosphatase calcineurin (Cn) transduces physiological signals through intracellular pathways to influence the expression of specific genes. Here, we characterize a naturally occurring splicing variant of the CnAbeta catalytic subunit (CnAbeta1) in which the autoinhibitory domain that controls enzyme activation is replaced with a unique C-terminal region. The CnAbeta1 enzyme is constitutively active and dephosphorylates its NFAT target in a cyclosporine-resistant manner. CnAbeta1 is highly expressed in proliferating myoblasts and regenerating skeletal muscle fibers. In myoblasts, CnAbeta1 knockdown activates FoxO-regulated genes, reduces proliferation, and induces myoblast differentiation. Conversely, CnAbeta1 overexpression inhibits FoxO and prevents myotube atrophy. Supplemental CnAbeta1 transgene expression in skeletal muscle leads to enhanced regeneration, reduced scar formation, and accelerated resolution of inflammation. This unique mode of action distinguishes the CnAbeta1 isoform as a candidate for interventional strategies in muscle wasting treatment.

Loaded wheel running and muscle adaptation in the mouse.

Voluntary cage wheel exercise has been used extensively to determine the physiological adaptation of cardiac and skeletal muscle in mice. In this study, we tested the effect of different loading conditions on voluntary cage wheel performance and muscle adaptation. Male C57Bl/6 mice were exposed to a cage wheel with no-resistance (NR), low-resistance (LR), or high-resistance (HR) loads for 7 wk. Power output was elevated (3-fold) under increased loading (LR and HR) conditions compared with unloaded (NR) exercise training. Only unloaded (NR) exercise induced an increase in heart mass, whereas only loaded (LR and HR) exercise training induced an increase in skeletal (soleus) muscle mass. Moreover, unloaded and loaded exercise training had a differential impact on the cross-sectional area of muscle fibers, depending on the type of myosin heavy chain expressed by each fiber. The biochemical adaptation of the heart was characterized by a decrease in genes associated with pathological (but not physiological) cardiac hypertrophy and a decrease in calcineurin expression in all exercise groups. In addition, transcriptional activity of myocyte enhancer factor-2 (MEF-2) was significantly decreased in the hearts of the LR group as determined by a MEF-2-dependent transgene driving the expression of beta-galactosidase. Phosphorylation of glycogen synthase kinase-3beta, protein kinase B (Akt), and p70 S6 kinase was increased only in the hearts of the NR group, consistent with the significant increase in cardiac mass. In conclusion, unloaded and loaded cage wheel exercise have a differential impact on cage wheel performance and muscle (cardiac and skeletal) adaptation.

Development of a mammalian series-fibered muscle.

This study examines the processes by which multiply innervated, serially fibered mammalian muscles are constructed during development. We previously reported that primary myotubes of such a muscle, the guinea pig sternomastoid muscle, span from tendon to tendon and are innervated at each of the muscle's four innervation zones. Secondary myotubes form later, in association with each point of innervation (Duxson and Sheard, Dev. Dyn., 1995; 204:391-405). We now describe the further growth and development of the muscle. Secondary myotubes initially insert onto and grow along the primary myotube. However, as they reach a critical length, they encounter other secondary myotubes growing from serially adjacent innervation zones and may transfer their attachment(s) to these serially positioned secondary myotubes. Other secondary myotubes maintain attachment at one or both ends to their primary myotube. Thus, an interconnected network of primary and secondary myotubes is formed. Patterns of reactivity for cell adhesion molecules suggest that early attachment points between myotubes are the embryonic precursors of adult myomyonal junctions, characterized by the expression of alpha7Bbeta1 integrin. Finally, the results show that secondary myotubes positioned near a tendon are generally longer than those lying in the mid belly of the muscle, and we suggest that the environment surrounding the tendinous zone may somehow stimulate myotube growth.

Intramuscular force transmission.

The architectural form of skeletal muscle, the pattern of activity/usage between neighbouring fibres, and the pathways for lateral and lengthwise tension delivery are all of interest in understanding muscle function and dysfunction. We have attempted to contribute to understanding of intramuscular force transmission by investigating the functional relationships between coactive motor units, and by examining the detailed molecular and morphological features at sites of tension transfer. We found that tension delivery is modulated by interaction between active and inactive fibres, that many muscle fibre terminations feature structural coupling between fibres, and that sites of tension delivery feature a variety of proteins including acetylcholinesterase, NCAM, dystrophin and two splice variants of the alpha7 integrins. We conclude that structural and molecular pathways exist to deliver force within, along, and between muscle fibres, and that the quality/quantity of tension delivered from any single fibre is at least partly a consequence of whether its neighbouring fibres are synchronously coactive.

Localization of alpha 7 integrins and dystrophin suggests potential for both lateral and longitudinal transmission of tension in large mammalian muscles.

Non-primate mammalian muscles with fascicles above 35 mm in length are composed predominantly of arrays of short, non-spanning muscle fibres, which terminate within the belly of the muscle fascicle at one or both ends. We have previously described the morphological form of various muscle-to-muscle and muscle-to-matrix junctions which are likely involved in tension transmission within one such muscle - the guinea pig sternomastoid muscle (Young et al. 2000). Here, we use immunohistochemistry to investigate the cell adhesion molecules present at these junctions. We find strong immunoreactivity against the alpha 7B integrin subunit and dystrophin, and slight reactivity against the alpha 7A integrin at all intrafascicular fibre terminations (IFTs), as well as at the muscle-tendon junction (MTJ). Tenascin, the sole ligand for alpha 9 beta 1 integrin, was absent from IFTs but present at the MTJ, suggesting the two sites are molecularly distinct. In addition to their expression at junctional sites, alpha 7B integrin and dystrophin were also expressed ubiquitously along the non-junctional sarcolemma, suggesting potential involvement in diffuse lateral transmission of tension between adjacent fibres. We conclude that the distribution of alpha 7 beta 1 integrins and dystrophin in series-fibred muscles suggests they are involved in transmission of tension from intrafascicularly terminating fibres to neighbouring fibres lying both in-series and in-parallel, via the extracellular matrix (ECM).

Different modes of hypertrophy in skeletal muscle fibers.

Skeletal muscles display a remarkable diversity in their arrangement of fibers into fascicles and in their patterns of innervation, depending on functional requirements and species differences. Most human muscle fascicles, despite their great length, consist of fibers that extend continuously from one tendon to the other with a single nerve endplate band. Other mammalian muscles have multiple endplate bands and fibers that do not insert into both tendons but terminate intrafascicularly. We investigated whether these alternate structural features may dictate different modes of cell hypertrophy in two mouse gracilis muscles, in response to expression of a muscle-specific insulin-like growth factor (IGF)-1 transgene (mIGF-1) or to chronic exercise. Both hypertrophic stimuli independently activated GATA-2 expression and increased muscle cross-sectional area in both muscle types, with additive effects in exercising myosin light chain/mIGF transgenic mice, but without increasing fiber number. In singly innervated gracilis posterior muscle, hypertrophy was characterized by a greater average diameter of individual fibers, and centralized nuclei. In contrast, hypertrophic gracilis anterior muscle, which is multiply innervated, contained longer muscle fibers, with no increase in average diameter, or in centralized nuclei. Different modes of muscle hypertrophy in domestic and laboratory animals have important implications for building appropriate models of human neuromuscular disease.