MODELING ATROPHY THROUGH FASTING IN AN IN VITRO SYSTEM REVEALS POSSIBLE CELLULAR SELF-RENEWAL.

Zebrafish (Danio rerio) remains the teleost fish of choice for biological investigations due to the plethora of molecular tools available for use in this system. Adult zebrafish possess a rather limited growth potential compared to many commercially important fish species. However, the pathways regulating muscle growth are conserved across vertebrate taxa regardless of adult growth potential. In vertebrate skeletal muscle, growth is largely regulated by protein turnover, with the balance between protein synthesis and degradation governing whether myofibers hypertrophy or atrophy. To better describe the potentially divergent mechanisms of skeletal muscle atrophy in this species, we developed an atrophy-induction regime to apply to a primary myotube culture system generated from isolated myogenic precursor cells (MPCs) from adult zebrafish.   

By culturing myotubes in a media devoid of amino acids, several genes necessary for autophagosome formation and function are upregulated (Figure 1a). Additionally, Murf1b, an ubiquitin-like ligase that contributes to muscle atrophy, is increased following myotube starvation, suggesting that amino acid-induced starvation increases autophagy and proteasome activity. Surprisingly, as autophagy and atrophy-related genes are upregulated following cell starvation, pax3-a and -b (Figure 1b), pax7, and Mef2ca were also upregulated. These data suggest that removing amino acids from media in this primary culture system activates the autophagy pathway, but also induces the upregulation of a biomarker known to be important in self-renewal of muscle stem cells.

Overall, these data demonstrate that adult zebrafish primary myoblast cultures can serve as an excellent model to study muscle growth in relation to the balance between atrophy and hypertrophy in relation to specific nutrients, like amino acids.