We examined whether lipopolysaccharide (LPS)-treatment affects reactivity of guinea-pig airway smooth muscle to the contractile effects of methacholine (MCh), and the relaxant effects of epithelium-derived relaxing factor (EpDRF), the release of which is stimulated by elevated intraluminal osmolarity. Contractile and relaxant responses of the smooth muscle were measured in vitro using the isolated, perfused trachea apparatus which allows pharmacological agents to be added separately to the mucosal or serosal surfaces. Due to the epithelium, serosal reactivity to MCh is greater than mucosal reactivity. Eighteen hours after LPS (4 mg/kg, i.p.) there were no differences in the EC~ and maximum response values for contraction to serosally- and mucosally applied MCh between saline- and LPS-treated guinea-pigs. After precontraction of the smooth muscle with 3 x 10-7 M MCh, EpDRF-induced relaxation was initiated with cumulative additions of NaCl to the mucosal bath. The EC50 for relaxation remained unchanged; however, the maximum relaxation response to NaCl was increased in LPS-treated animals. These results suggest that LPS-treatment does not alter reactivity of the smooth muscle to MCh, but does enhance responsiveness to intraluminal hypertonicity by increasing the synthesis and/or release of EpDRF, or altering EpDRF's relaxant effect on the smooth muscle.