Activation of Kupffer cells by lipopolysaccharide (LPS) is a critical step in the pathogenesis of alcoholic liver disease. Kupffer cells isolated from rats fed ethanol in their diet for 4 wk accumulated 4.3-fold more tumor necrosis factor (TNF)- in response to LPS compared with pair-fed rats. In contrast, LPS-stimulated interleukin (IL)-1 accumulation was 50% lower after ethanol feeding. LPS-stimulated TNF- mRNA accumulation was twofold higher after ethanol feeding, whereas IL-1 mRNA accumulation was blunted. To understand the mechanisms for this differential response, we investigated the effects of ethanol on LPS-dependent signal transduction. Chronic ethanol feeding increased LPS-stimulated extracellular receptor-activated kinases 1/2 (ERK1/2) activation. Activation of ERK1/2 was required for maximal increases in TNF- and IL-1 mRNA and was associated with increased binding of early growth response-1 (Egr-1) to the TNF- promoter after ethanol feeding. In contrast, ethanol feeding completely abrogated activation of nuclear factor-B DNA-binding activity by LPS and had no effect on AP-1 binding. Together, these data suggest that enhanced activation of ERK1/2 and Egr-1 contributes to increased TNF- production after chronic ethanol feeding.