It has been shown that phospholipase D (PLD) activity is stimulated by the beta-amyloid protein in neuronal cells. The aim of this study was to determine whether overexpression of the amyloid precursor protein (APP) affects the activity and the level of PLD expression in P19 embryonic carcinoma cells. We observed that the unstimulated basal PLD activity was higher in wild-type APP(695)-transfected cells than in non-transfected control cells. The protein kinase C (PKC) activator, phorbol 12-myristate 13-acetate (PMA), has been shown to activate PLD. PMA-stimulated PLD activity was 3-fold higher in the APP overexpressing cells than in the control cells. P19 cells express two distinct PLD isozymes, PLD1 and PLD2. The level of PLD2 expression was increased by APP overexpression. Although the PKC inhibitor, GF109203X, inhibited PMA-stimulated PLD activity, it did not affect the high basal PLD activity induced by APP overexpression. Neuronal differentiation of the P19 cells by retinoic acid did not affect the basal or PMA stimulated-PLD activity. Interestingly, APP overexpression in the differentiated P19 cells also led to an increase in PLD activity. The PLD activity of the P19 cells is apparently regulated by amyloid protein through both PKC-dependent and -independent mechanisms.