AMP-activated protein kinase (AMPK) is activated by increases in the intracellular AMP:ATP ratio and plays a central role in cellular responses to metabolic stress. While activation of AMPK has been shown to have anti-inflammatory effects, there is little information concerning the role that AMPK may play in modulating neutrophil function and neutrophil-dependent inflammatory events, such as acute lung injury. To examine these issues, we determined the effects of pharmacological activators of AMPK, 5-amino-4-imidazole carboxamide riboside (AICAR) and barberine, on TLR4 induced neutrophil activation. AICAR and barberine dose dependently activated AMPK in murine bone marrow neutrophils. Exposure of LPS-stimulated neutrophils to AICAR or barberine inhibited release of TNF- and IL-6, as well as degradation of IB and nuclear translocation of NF-B, as compared to findings in neutrophil cultures that contained LPS without AICAR or barberine. Administration of AICAR to mice resulted in activation of AMPK in the lungs and was associated with decreased severity of LPS-induced lung injury, as determined by diminished neutrophil accumulation in the lungs, reduced interstitial pulmonary edema, and diminished levels of TNF- and IL-6 in bronchoalveolar lavage fluid. These results suggest that AMPK activation reduces TLR4 induced neutrophil activation and diminishes the severity of neutrophil driven proinflammatory processes, including acute lung injury.