Patients with diabetes mellitus present muscle mass loss and muscle atrophy that activate signaling pathways of protein degradation, such as MURF‐1 and MAF‐bx. The aim of this study was to evaluate the effects of glutamine (Gln) supplementation on signaling pathways of protein synthesis (Akt, GSK3, p70s6K, 4E‐BP1, mTOR) and degradation (MuRF‐1 e MAF‐bx) in rat soleus muscles in vitro and in vivo, by using Western blotting. In the in vitro experiments, skeletal muscle cells were cultured in the absence and in the presence of Gln (4 and 16 mM) for seven days; whereas, in the in vivo experiments, streptozocin‐induced diabetic rats were supplemented with glutamine (1g/kg bw for 15 days. Protein synthesis was determined by measuring the incorporation of labeled leucine into muscle cells. In cultured skeletal muscle cells, Gln (at 4 and 16 mM) increased L‐[U‐14C]‐leucine incorporation, Akt, mTOR and GSK3 phosphorylation, as well as p70s6k protein content phosphorylation, and decreased 4E‐BP1 and MuRF‐1 expression. In soleus muscle from diabetic rats there was an increase of Akt, GSK3 and mTOR phosphorylation, followed by a reduction of 4E‐BP1, MuRF‐1 and MAF‐bx. In conclusion, Gln stimulated the signaling pathway of protein synthesis and inhibited that of protein degradation in skeletal muscle.Financial Support: Fapesp, Capes and CNPq