1. Experiments have been performed in order to analyse the mechanism whereby SCA40, a new imidazo[1,2-a]pyrazine derivative relaxes airway smooth muscle. 2. SCA40 (0.01-10 microM) caused a complete and concentration-dependent relaxation of guinea-pig isolated trachea contracted with 20 mM KCl but failed to inhibit completely the spasmogenic effects of 80 mM KCl. 3. Quinine (30 microM) antagonized the relaxant activity of SCA40 in 20 mM KCl-contracted guinea-pig isolated trachea. The ATP-sensitive K(+)-channel blocker, glibenclamide (3 microM), did not antagonize the relaxant activity of SCA40 in either 20 mM KCl or 1 microM carbachol-contracted isolated trachea. 4. SCA40 (0.01-10 microM) and isoprenaline (0.1 nM-10 microM) caused a complete and concentration-dependent relaxation of guinea-pig isolated trachea contracted with carbachol 1 microM. 5. The large-conductance Ca(2+)-activated K(+)-channel blocker, charybdotoxin (60-180 nM), non-competitively antagonized the relaxant activity of isoprenaline on 1 microM carbachol-contracted trachea. The inhibition was characterized by rightward shifts of the isoprenaline concentration-relaxation curves with depression of their maxima. 6. The relaxant activity of SCA40 in 1 microM carbachol-contracted trachea was antagonized by charybdotoxin (60-600 nM) in an apparently competitive manner. The concentration-relaxation curves to SCA40 were shifted to the right with no significant alteration in the maximum response. 7. It is concluded that SCA40 is a novel potassium channel opener which is a potent relaxant of guinea-pig airway smooth muscle in vitro. The relaxant activity of SCA40 does not involve ATP-sensitive K+-channels but rather large-conductance Ca2'-activated K+-channels or other charybdotoxin sensitive K+-channels.