Previous reports suggest that burn-induced muscle proteolysis can be inhibited by treatment with GSK-3 inhibitors, suggesting that burn injury may be associated with increased GSK-3b activity. The influence of burn injury on muscle GSK-3 kinase activity, however, is not known. We determined the effect of a 30% total body surface full-thickness burn injury in rats on muscle GSK-3 activity by measuring GSK-3 kinase activity and tissue levels of serine 9 phosphorylated GSK-3, p(Ser9)-GSK-3, by Western blotting and immunohistochemistry. Because burn-induced muscle wasting is, at least in part, mediated by glucocorticoids, we used dexamethasone-treated cultured muscle cells in which GSK-3 expression had been reduced with siRNA to further assess the role of GSK-3b in muscle atrophy. Burn injury resulted in a 7-fold increase in GSK-3 kinase activity in skeletal muscle. This effect of burn was accompanied by reduced tissue levels of p(Ser9)-GSK-3, suggesting that burn injury stimulates GSK-3 in skeletal muscle secondary to inhibited phosphorylation of the enzyme. In addition, burn injury resulted in inhibited phosphorylation and activation of Akt, an upstream regulatory mechanism of GSK-3 activity. Reducing the expression of GSK-3 in cultured muscle cells with siRNA inhibited dexamethasone-induced protein degradation by approximately 50%. The results suggest that burn injury stimulates GSK-3 activity in skeletal muscle and that GSK-3 may, at least in part, regulate glucocorticoid-mediated muscle wasting.