Hyperphosphorylated tau is a major component of the neurofibrillary tangles found in Alzheimer's disease (AD) brains. Glycogen synthase kinase 3 beta (GSK3b) and cyclin-dependent protein kinase 5 (Cdk5) are two kinases known to phosphorylate tau at AD related epitopes. Here, we examined the role of these two kinases in phosphorylating tau at the AT8 epitope in cultured rat cortical neurons. We found that several Cdk5 inhibitors (butyrolactone I, purvalanol A and roscovitine) were ineffective in blocking the AT8 signal in the cultured neurons. However, these same inhibitors were effective in inhibiting phosphorylation of a histone peptide by purified p25/Cdk5 and, more importantly, the AT8 signal in intact CHO cells inducibly expressing human recombinant p25/Cdk5/tau (Cook et al., SFN abstract 447.16, 1999). In contrast, a GSK3b inhibitor, LiCl, inhibited the AT8 signal in cultured cortical neurons and in CHO cells expressing GSK3b/tau with similar potencies. These data suggest that basal tau phosphorylatio...