We have previously shown that endogenous IGF-I regulates human intestinal smooth muscle cell proliferation by activation of PI 3-kinase-dependent and Erk1/2-dependent pathways that jointly regulate cell cycle progression and cell division. While IGF-I stimulates PI 3-kinase-dependent activation of Akt, expression of a kinase-inactive Akt did not alter IGF-I stimulated proliferation. In other cell types, Akt-dependent phosphorylation of glycogen synthase kinase -3 (GSK-3), inhibits its activity and its ability to stimulate apoptosis. The aim of the present study was to determine whether endogenous IGF-I regulates Akt-dependent, GSK-3 phosphorylation and activity, and regulates apoptosis in human intestinal muscle cells. IGF-I elicited time-dependent and concentration-dependent GSK-3 phosphorylation (inactivation) that was measured by Western blot using a phosphor-specific GSK-3 antibody. Endogenous IGF-I stimulated GSK-3 phosphorylation and inhibited GSK-3 activity (measured by in vitro kinase assay) in these cells. IGF-I-dependent, GSK-3 phosphorylation and the resulting GSK-3 inactivation were mediated by activation of a PI 3-kinase-dependent, phosphoinositide-dependent kinase -1 (PDK-1)-dependent and Akt-dependent mechanism. Deprivation of serum induced -catenin phosphorylation, increase in caspase 3 activity and apoptosis of muscle cells that was inhibited by either IGF-I or a GSK-3 inhibitor. Endogenous IGF-I inhibited -catenin phosphorylation, caspase 3 activation and apoptosis induced by serum deprivation. IGF-I-dependent inhibition of apoptosis, like GSK-3 activity, was mediated by a PI 3-kinase-dependent, PDK-1-dependent and Akt-dependent mechanism. We conclude that endogenous IGF-I exerts two distinct but complementary effects on intestinal smooth muscle cell growth, it stimulates proliferation and inhibits apoptosis. The growth of intestinal smooth muscle cells is regulated jointly by the net effect of these two processes.