Concentration-relaxation profiles for pinacidil, verapamil, terbutaline and theophylline were studied in guinea-pig trachealis contracted by two commonly applied techniques for K+ depolarization. All drugs were much less effective on contractions induced by hyperosmolar 124 mMn K+ solution (added KCl) than on contractions elicited by an isoosmolar 124 mM K+ Krebs solution (substituted KCl). The maximal relaxant responses were (isoosmolar K+/hyperosmolar K+): pinacidil 100%/40%, verapamil 100%/60%, theophylline 100%/0%, terbutaline 50%/0%. Addition of mannitol to establish the same hyperosmolarity as with 124 mM KCl also produced contraction of guinea-pig trachealis. Concentration-relaxation curves for the drugs on mannitol-induced contractions had close resemblance to those obtained in hyperosmolar 124 mM K+ solution. When contraction was elicited by 30 mM K+, pinacidil showed seven times higher relaxant potency in hyperosmolar compared to isoosmolar solution, whereas the relaxant responses to verapamil, theophylline and terbutaline were not influenced by osmolarity. When K+ depolarization is used as a tool for evaluation of drug action in airway smooth muscle, the two different techniques produce dissimilar results. The influence of hyperosmolarity per se appears to be an important and unwanted feature when K+ depolarization is produced by addition of KCl.