Tau protein kinase I(TPKI)/glycogen synthase kinase (GSK)-3beta is abundant in the developing rat brain. The highly phosphorylated juvenile form of tau is present during the same developmental period. To study the role of TPKI/ GSK-3beta in neuronal growth, we examined the effects of lithium, a direct inhibitor of TPKI/GSK-3beta, using primary cultures of rat hippocampal neurons. Immunohistochemical staining of the neurons indicates that in the presence of lithium (2-10 mM), neurite growth becomes inhibited in a dose-dependent manner. Western blot analyses of the cell extracts revealed that the presence of lithium in the culture medium increased the amount of dephosphorylated tau while decreasing phosphorylation at Ser199 and Ser396, both of which are TPKI/GSK-3beta phosphorylation sites on tau. The inhibition by lithium is reversible. Although the amount of TPKI/GSK-3beta remained constant, the amount of tau decreased in a dose-dependent manner in the presence of lithium. TPKI/GSK-3beta was distributed in the somata and proximal neurites of the cultured hippocampal neurons. These results therefore suggest that TPKI/GSK-3beta plays an important role in the axonal growth of neurons during synapse formation in the developing brain.