1. Examination has been made of the effects of epithelium removal on the reactivity of guinea-pig trachealis to methacholine at 37 degrees C and 22 degrees C, and on responses to activation of the Na+/K(+)-pump by abrupt temperature increase from 22 degrees C to 37 degrees C. 2. At 37 degrees C, epithelium removal increased the sensitivity of isolated tracheal strips to methacholine without affecting the maximum isometric contractile response. Epithelium removal resulted at 22 degrees C in a decrease in sensitivity to methacholine, i.e. an effect opposite to that seen at 37 degrees C. While the maximum response of intact strips to methacholine was enhanced at 22 degrees C, the maximum response of denuded preparations was decreased. 3. The increase in sensitivity to methacholine at 37 degrees C after epithelium removal was mimicked in intact preparations by indomethacin (1 microM). Indomethacin did not mimic the decrease in methacholine sensitivity and maximum response caused by epithelium removal at 22 degrees C. 4. Following incubation at 22 degrees C, abrupt increase in temperature to 37 degrees C elicited relaxation in both epithelium-containing and epithelium-denuded tracheal strips. In epithelium-containing preparations the relaxation was more pronounced and followed by contraction. Ouabain (1 microM) converted the relaxation of denuded preparations to contraction, but was ineffective in intact strips. The relaxation of intact strips was, however, inhibited by a greater ouabain concentration (10 microM). 5. These findings indicate that the modulatory effect of the epithelium is temperature-dependent. In cooled preparations, the epithelium enhances reactivity. At 37 degrees C, an epithelium-derived factor reduces reactivity, and this may partially be due to activation of the electrogenic Na+/K+-pump.