Isoprostanes comprise a class of membrane lipid metabolites produced during oxidative stress, including asthma, chronic obstructive pulmonary disorder and cystic fibrosis. They are now widely recognized to evoke a variety of biological responses in airway and pulmonary vascular smooth muscle, lymphatics and innervation. However, their effects on airway epithelium are largely unstudied. We examined the electrophysiological responses evoked by several different isoprostane species in bovine airway epithelium using the Ussing chamber technique. The E-ring isoprostanes 15-E_1t-IsoP and 15-E_2t-IsoP evoked a substantial increase in short-circuit current (I_SC), whereas four different F-ring isomers were ineffective. 15-E_2t-IsoP-evoked I_SC was mimicked by the EP-agonist PGE₂, but not by agonists of EP₁/EP₃-, FP- or TP-receptors (sulprostone, fluprostenol and U46619, respectively). On the other hand, this response was significantly reduced by the EP₄-receptor blocker GW627386 but not by blockers of various other prostanoid receptors (ICI 192,605 [TP-selective]; SC19220 [EP₁-selective]; AH6809 [DP/EP₁/EP₂-selective]; AL8810 [FP-selective]). 15-E_2t-IsoP-evoked I_SC was reduced by blockers of Cl^- channels (niflumic acid; NPPB), of Na⁺/K⁺/2Cl^- co-transport (furosemide; bumetanide), of adenylate cyclase (MDL 12,330A) or of guanylate cyclase (ODQ), but not by blockers of Na⁺ conductances (amiloride). We conclude that 15-E_2t-IsoP activates a transepithelial Cl^- conductance in bovine airway epithelium through an EP₄ receptor coupled to adenylate cyclase as well as soluble guanylate cyclase.