We have shown previously that the lung surfactant inhibits LPS-stimulated nitric oxide (NO) production in alveolar macrophages (Miles et al., Am.J.Physiol. 276:Ll86, 1999). But lung surfactant had no effect on NO production in two macrophage cell lines RAW 264.7 and J-774. Stimulation of primary macrophages and cell lines by LPS has been shown to activate MAPKs. To determine if differences in MAPK activation may explain the difference in response to lung surfactant between primary macrophages and the cell lines, we compared the activation of MAPKs, p38, extracelluar-regulated kinase (ERK), and c-Jun N-terminal kinase (JNK/SAPK), in rat alveolar macrophages and three macrophage cell lines, RAW 264.7, J-774, and NR8383. In alveolar macrophages there was basal phosphorylation of p38 and ERK which did not significantly increase on LPS stimulation. There was no detectable JNK phosphorylation in unstimulated or LPS-stimulated alveolar macrophages. In contrast, in all three cell lines LPS induced a significant increase in phosphorylation of p38. Differences in activation of MAP kinases may determine the response to lung surfactant.