AIMS: We previously showed that pharmacological activation of adenosine monophosphate activated protein kinase (AMPK) inhibited amiloride-sensitive Na⁺ transport across H441 human airway epithelial cells. We have tested the hypothesis that metabolic stresses such as anoxia and glucose deprivation activate AMPK to decrease Na⁺ transport in the airway epithelium. METHODS: We examined phosphorylation of AMPK and acetyl-CoA carboxylase (ACC) (downstream target of AMPK) by Western blotting in H441 cell monolayers cultured at air-liquid interface, which were exposed to anoxia (0.2% O₂, 5% CO₂) or glucose deprivation. The effects of anoxia and glucose deprivation on transepithelial Na⁺ transport were studied using Ussing chamber experiments. RESULTS: Exposure to an hour of anoxia and 4 hours of glucose deprivation induced a 6.0 ±0.7 (P < 0.01, n = 4) and 4.0 ±0.9 (P < 0.05, n = 3) fold increase in phospho/total AMPK respectively, compared to control. Phospho/total ACC was also increased by 2.2 ±0.2 (P < 0.01, n = 4) and 1.8 ±0.3 (P < 0.05, n = 3) fold. Anoxia and glucose deprivation reduced amiloride-sensitive (10mM) short circuit current (Isc) by 52% from 18.9 ±0.9 to 9.0 ±0.9 mA/cm² (P < 0.01, n = 5) and by 27% from 25.0 ±3.3 to 17.7 ±1.7 mA/cm² (P < 0.05, n = 4) respectively. In the presence of Compound C, a pharmacological inhibitor of AMPK, anoxia induced a 33.3 ±1.7 % (P < 0.01, n = 3) decrease in amiloride-sensitive current. However, no significant effect of glucose deprivation was observed and levels remains similar to control (112 ±16.7 %, n = 3). CONCLUSIONS: Glucose deprivation inhibits amiloride-sensitive transepithelial Na⁺ transport via activation of AMPK. Anoxia also activated AMPK, however, alternative pathways may be involved in its inhibition of amiloride-sensitive Na⁺ transport. Supported by the BBSRC and St George's University of London.