Bioelectric properties and ion transport of excised human segmental/subsegmental bronchi were measured in specimens from 40 patients. Transepithelial electric potential difference (PD), short-circuit current (Isc), and conductance (G), averaged 5.8 mV (lumen negative), 51 microA X cm-2, and 9 mS X cm-2, respectively. Na+ was absorbed from lumen to interstitium under open- and short-circuit conditions. Cl- flows were symmetrical under short-circuit conditions. Isc was abolished by 10(-4) M ouabain. Amiloride inhibited Isc (the concentration necessary to achieve 50% of the maximal effect = 7 X 10(-7) M) and abolished net Na+ transport. PD and Isc were not reduced to zero by amiloride because a net Cl- secretion was induced that reflected a reduction in Cl- flow in the absorptive direction (Jm----sCl-). Acetylcholine (10(-4) M) induced an electrically silent, matched flow of Na+ (1.7 mueq X cm-1 X h-1) and Cl- (1.9 mueq X cm-12 X h-1) toward the lumen. This response was blocked by atropine. Phenylephrine (10(-5) M) did not affect bioelectric properties or unidirectional ion flows, whereas isoproterenol (10(-5) M) induced a small increase in Isc (10%) without changing net ion flows significantly. We conclude that 1) Na+ absorption is the major active ion transport across excised human bronchi, 2) Na+ absorption is both amiloride and ouabain sensitive, 3) Cl- secretion can be induced by inhibition of the entry of luminal Na+ into the epithelia, and 4) cholinergic more than adrenergic agents modulate basal ion flow, probably by affecting gland output.