Effects of eicosanoids on the canine proximal colonic mucosa were examined. Both arachidonic acid (AA) and prostaglandin E2 (PGE2) increased short-circuit currents. Tetrodotoxin did not affect these responses, suggesting that functioning nerves are not required. Indomethacin abolished responses to AA, indicating that the cyclooxygenase pathway is the primary metabolic pathway. Indomethacin significantly potentiated responses to PGE2, suggesting that in the presence of cyclooxygenase inhibition either 1) a normally inhibitory cyclooxygenase product is not present or 2) a potentiating lipoxygenase product is being produced in greater amounts. PGE2 is produced in significant quantities, whereas leukotriene B4 (LTB4) is produced in smaller amounts. Cyclooxygenase inhibitors significantly decreased PGE2 production but had no effect on LTB4. This suggests that an inhibitory PG may be opposing the response to PGE2. Therefore, we tested the effects of several cyclooxygenase products on PGE2 responsiveness. PGD2 alone significantly reduced responses to PGE2. In the canine proximal colon the response to AA is apparently the algebraic sum of the opposing responses of PGE2 and PGD2.