To determine whether exotoxin A may affect the transport of fluid across the lung epithelium, two isogenic strains of Pseudomonas aeruginosa PA103 (108 colony-forming units), one (PA103 toxΩ) with a structural gene mutation in exotoxin A, were instilled into the distal airspaces of anesthetized rats. PA103 parental strain, but not its mutant, stimulated the removal of fluid from the distal airspaces of the lung. Instillation of exotoxin A alone caused a dose-dependent increase in the fluid transport across the lung epithelium. Instillation of amiloride (10-3 M) with exotoxin A demonstrated that this effect partially depended on increased uptake of sodium across the lung epithelium. The absence of stimulation after instillation of an exotoxin A mutant (PEΔGlu553) without ADP- ribosyltransferase activity demonstrated that the effect of exotoxin A depended on its ADP-ribosyltransferase activity. Finally, the instillation of exotoxin A in rats depleted of macrophages indicated that the effect of exotoxin A was not secondary to the activation of alveolar macrophages by this toxin. In conclusion, these results indicate that the in vivo release of exotoxin A by live airspace P. aeruginosa directly stimulates the fluid removal from the airspaces by the lung epithelium. This may alter the volume or composition of airway secretions, and may contribute to the lung disease in patients infected with P. aeruginosa.