Glucocorticoids have been shown to exert a rapid non-genomic effect in airway epithelial. Using an intracellular pH (pHi) imaging system and the NH4Cl pre-pulse technique, we have shown that the synthetic glucocorticoid, dexamethasone stimulated intracellular pH (pHi) regulation in a human bronchial epithelial cell line (16HBE14o- cells). A brief exposure to NH4Cl (20 mM) elicited an intracellular acidification to a pHi of 6.46±0.03 within 1.67±0.04 minutes, followed by a pHi recovery until a plateau value of 7.13±0.04 within 6.9±1.2 min after the acid load (recovery rate in control conditions: 0.046±0.021 pH units/min). Dexamethasone (1nM) accelerated the intracellular pH (pHi) recovery to a rate of 0.107±0.029 pH units/min. Both external Na+ removal (substituted by N-methyl D glucamine) and the Na+/H+ exchanger inhibitor, 5-(n-ethyl-N-isopropyl) amiloride (EIPA) inhibited the basal pHi recovery and the dexamethasone stimulation of pHi recovery. The rapid effect of dexamethasone on pHi was not affected by the inhibitors of transcription, cycloheximide and by the classical glucocorticoid and mineralocorticoid receptors antagonists, RU486 and spironolactone respectively. The rapid dexamethasone response was significantly reduced by EIPA used as an inhibitor of Na+/H+ exchanger activity and completely inhibited by Na+ removal. The dexamethasone effect on pHi regulation was reduced by the inhibitors of adenylated cyclase, cAMP dependant protein kinase and MAP kinase (ERK1/2) activities. Taken together our results provided evidences for the rapid stimulation of the Na+/H+ exchange activity by glucocorticoids in bronchial epithelial cells via a non-genomic mechanism involving a cAMP dependant protein kinase and MAP kinase ERK1/2 pathways.