Recent studies have shown that orally supplied nitrates, which substantially exist in our daily diets, are reduced into nitrites and become significant sources of nitric oxide (NO) especially in hypoxic tissues. However, physiological significance of nitrites in normal tissues has not been elucidated though our serum concentrations of nitrites reach as high as micromolar levels. We investigated effects of nitrite on endothelial NO synthase (eNOS) using human glomerular endothelial cells to reveal potential glomerular-protective actions of nitrites with its underlying molecular mechanism. Here we demonstrate that nitrite stimulation evokes eNOS activation which is dependent on 5′AMP-activated protein kinase (AMPK) activation in accordance with ATP reduction. Thus, nitrites should facilitate AMPK–eNOS pathway in an energy level-dependent manner in endothelial cells. The activation of AMPK–eNOS signals is suggested to be involved in vascular and renal protective effects of nitrites and nitrates. Nitrites may harbor beneficial effects on metabolic regulations as AMPK activators.