The effects of nerve growth factor (NGF) and epidermal growth factor (EGF) on the regulation of phosphatidylinositol 3-kinase (PtdIns 3-kinase) activity were assessed in rat pheochromocytoma (PC12) cells. Both NGF and EGF induced a rapid activation of PtdIns 3-kinase as assessed by a dramatic rise in growth factor-induced PtdIns 3-kinase activity found in antiphosphotyrosine immunoprecipitates. The intracellular levels of two of the lipid products of PtdIns 3-kinase, phosphatidylinositol 3,4,5-trisphosphate (PtdIns(3,4,5)P3) and phosphatidylinositol 3,4-bisphosphate (PtdIns(3,4)P2), also rose dramatically, exhibiting time courses very similar to the appearance of PtdIns 3-kinase in immunoprecipitates. The activation of PtdIns 3-kinase is, therefore, a common event in the signal transduction processes elicited by growth factors stimulating distinct cellular end points in PC12 cells, namely the NGF-induced neuronal differentiation and EGF-stimulated mitogenesis. Thus the intracellular products of this enzyme may function in early biochemical events that are common components of the pathways controlling both differentiation and proliferation.