The aim of this study was to assess the influence of endothelin-1 (ET-1) on cholinergic nerve-mediated contractions in rat isolated tracheal smooth muscle by use of electrical-field stimulation (EFS) and [3H]choline efflux studies. EFS (80 V, 0.5 ms, 0.1-30 Hz for 10 s) evoked transient, frequency-dependent contractions of isolated tracheal preparations. Contractions were abolished in the presence of atropine or tetrodotoxin, which suggests they were mediated by acetylcholine (ACh) release from cholinergic nerves. The ETB receptor-selective agonist sarafotoxin S6c (1 nM) augmented EFS (0.6-1 Hz)-induced contractions by 179%. These effects were significantly attenuated in the presence of the ETB receptor-selective antagonist N-cis-2,6-dimethylpiperidinocarbonyl-L-gamma-methyl-leucyl-D-1- methoxycarbonyltryptophanyl-D-norleucine (BQ-788; 1 microM). ET-1 (1 nM) also markedly potentiated EFS-induced contractions (153%). This was apparently not a postjunctional effect, because ET-1 did not alter contractile responses to exogenously applied ACh. Cyclo[D-Trp-D-Asp-L-Pro-D-Val-L-Leu] (BQ-123;3 microM) and BQ-788 when used alone, failed to inhibit ET-1-induced potentiation of EFS-evoked contractions. However, in their combined presence, BQ-123 and BQ-788 significantly attenuated ET-1-induced potentiation of EFS responses. EFS (100 V, 0.5 ms, 3 Hz for 2 min) applied to tracheal preparations preloaded with [3H]choline, caused airway smooth muscle contraction and an efflux of radioactivity. Both sarafotoxin S6c (10 nM) and ET-1 (10 nM) significantly enhanced the EFS-induced 3H-efflux and the latter was abolished only in the combined presence of BQ-123 and BQ-788. These data indicated that ET-1 enhances cholinergic nerve-mediated contractions in rat isolated trachea via activation of prejunctional ETA and ETB receptors that were linked to increased ACh release from cholinergic nerves.