We measured in vivo in human subjects the airway transepithelial electric potential difference (PD), a parameter that reflects passive ion permeabilities and active ion transport across respiratory epithelia. A high impedance voltmeter was used to measure PDs between a fluid-filled and perfused recording bridge located on the airway surface and a reference bridge in the subcutaneous space. Tracheal PD was a relatively high 32 +/- 5 mV (lumen negative) in subjects younger than 35 yr of age (n = 23). Tracheal PD declined after 35 yr of age at a rate of 6 mV per decade. The PDs in bronchi were lower than in the trachea in subjects of all age groups. Neither the magnitude nor the regional variation of the PD appeared influenced by the chemical agents used in general anesthesia. However, superfusion of the airway surface with a pharmacologic inhibitor of Na+ absorption, amiloride, significantly reduced PD in the trachea (-38.5 +/- 7.2%, n = 7) and bronchi (-37.6 +/- 6.3%, n = 6). Both cigarette smoking and tracheostomy appear to reduce tracheal PD when the patients were compared with age-matched control subjects. We conclude that (1) the tracheal PD in young human subjects is relatively high, (2) substantial regional variations in the magnitude of human airway PDs can be demonstrated, (3) active Na+ absorption contributes to the generation of airway PD, and (4) measurements of PD in vivo may help detect early epithelial dysfunction.