Our laboratory has shown that tumor necrosis factor-α (TNFα) can regulate normal mammary epithelial cell (MEC) growth, morphogenesis, and, under certain circumstances, functional differentiation in a manner similar to epidermal growth factor (EGF). As TNFα has been shown to up-regulate EGF receptor (EGFR) expression and function in other systems, the present studies were undertaken to determine whether TNFα action in MEC was indirect through stimulation of the EGFR. An inhibitor of EGFR tyrosine kinase activity, PD158780, failed to block proliferation induced by 40 ng/ml TNFα and only partially inhibited growth in response to 2 ng/ml TNFα. PD158780 was also unable to suppress the extensive morphological development induced by either TNFα concentration. In contrast, the effects of TNFα and PD158780 on functional differentiation (i.e. casein accumulation) were time dependent. When measured on day 7 after 48 h of treatment, casein accumulation was unaffected by either concentration of TNFα or by PD158780. When assessed on day 21 after 16 days of treatment, however, casein levels were decreased by 40 ng/ml TNFα and increased by PD158780. Significantly, this PD158780- induced increase in casein was not observed in MEC that had been treated with both PD158780 and TNFα. These results thus suggest that EGFR tyrosine kinase activity is not necessary for TNFα action in normal MEC.