The neuronal Cl- -ATPase/pump is a candidate for an outwardly directed active Cl- transport system, which requires phosphatidylinositol-4-monophosphate (PI4P) for its optimal activity. We previously reported that low concentrations (1-10 nM) of amyloid beta proteins (Abetas, Abeta1-42, Abeta25-35), the neurotoxic peptides in Alzheimer's disease, reduced Cl- -ATPase activity in cultured rat hippocampal neurons without any changes in the activities of Na+/K+-ATPase or anion-insensitive Mg(2+)-ATPase, and decreased PI, PIP, and PIP2 levels in neuronal plasma membranes (Journal of Neurochemistry 2001, 78, 569-579). In this study, we examined the effects of exogenously applied PI and PI4P on the Abeta25-35-induced changes in Cl- -ATPase activity, the intracellular concentration of Cl- ([Cl- ]i), and glutamate neurotoxicity using primary cultured rat hippocampal neurons. The Abeta decreased Cl- -ATPase activity to 47% of control and increased [Cl- ]i in hippocampal pyramidal cell-like neurons to a level 3 times higher than the control. The addition of PI (50-750 nM) or PI4P (50-150 nM) dose-dependently blocked the inhibitory effects of Abeta on Cl- -ATPase activity. High doses of PI (750 nM) and PI4P (100-150 nM) reduced Na+/K+-ATPase activity to 41% and 35% of control, respectively, but this inhibition was attenuated by the co-application of phosphatidylserine (PS, 1 micro M). PI or PI4P (75 nM each) reversed the Abeta-induced increase in [Cl-]i. In the Abeta-exposed culture, stimulation by glutamate (10 micro M, 10 min) resulted in an increase in DNA fragmentation and decreases in cell viability. Addition of PI or PI4P prevented the Abeta-induced aggravation of glutamate neurotoxicity. Thus, PI and PI4P were demonstrated to prevent Abeta-induced decreases in Cl- -ATPase activity and increases in neuronal [Cl- ]i in parallel with the attenuation of Abeta-induced aggravation of glutamate neurotoxicity.