Abnormally hyperphosphorylated tau is the major protein component of neurofibrillary tangles, the characteristic lesion of Alzheimer's disease (AD). Protein phosphatases (PP) type 1 (PP-1), type 2A (PP-2A) and type 2B (PP-2B) appear to be involved in the regulation of tau phosphorylation. The incidence of neurofibrillary tangles is higher in brains of schizophrenic patients treated with neuroleptics than in those without this treatment. We have found that the commonly used neuroleptics chlorpromazine, trifluoperazine and clozapine inhibit PP-2B but not PP-1 or PP-2A activity towards [32P]phosphorylase kinase as a substrate. When AD abnormally hyperphosphorylated tau is used as a substrate, PP-2B activity is inhibited by trifluoperazine > chlorpromazine > clozapine. Using phosphorylation-dependent monoclonal antibodies, tau-1, AT8 and PHF-1, we have found that the dephosphorylation of the abnormal tau by PP-2B is inhibited at all the sites recognized by these antibodies. The IC50 of the inhibition of dephosphorylation at tau-1 site is approximately 20 microM for trifluoperazine and approximately 120 microM for chlorpromazine. These two neuroleptics inhibit tau dephosphorylation by PP-2B through antagonizing calmodulin as well as directly interacting with PP-2B. The inhibition of the dephosphorylation of abnormally hyperphosphorylated tau by neuroleptics raises an intriguing possibility that the chronic use of these drugs might contribute to neurofibrillary degeneration in schizophrenic and AD patients.