We studied the effect of corticotropin-releasing factor (CRF) on airway smooth muscle functions in isolated rabbit tracheal segments under isometric conditions in vitro. The addition of ovine CRF synthesized by solid-phase methods did not cause muscle contraction, but it potentiated the contractile response to electrical field stimulation (EFS) at 5 Hz in a dose-dependent fashion, the maximal increase from the baseline response being 43.9 +/- 6.1% (mean +/- SE, p less than 0.001). This effect was not influenced by propranolol, phentolamine, indomethacin, pyrilamine, or (D-Pro2,D-Trp7.9)-substance P, but was completely inhibited by alpha-helical CRF, a CRF receptor antagonist. CRF (10(-7) M) also increased the responses to EFS at all frequencies of stimulation (1 to 40 Hz), so that the stimulus frequency required to produce a half-maximal contraction (ES50) decreased from 7.7 +/- 1.0 to 3.8 +/- 0.6 Hz (p less than 0.01). In contrast, contractile response to administered acetylcholine was not affected by CRF. CRF-induced potentiation of the response to EFS (5 Hz) was further increased from 44.5 +/- 5.4 to 144.6 +/- 11.1% in the presence of physostigmine and was abolished by atropine. These results suggest that CRF prejunctionally potentiates the vagally mediated contraction of airway smooth muscle through activation of CRF receptors on the cholinergic nerve terminals, likely involving the accelerated release of acetylcholine.