Interaction between substance P (SP) and PGE(2) on Cl(-) secretion in the guinea pig distal colonic epithelia was investigated. A short-circuit current (I(sc)) was measured as an index of ion transport. Mucosa preparations deprived of muscle and submucosa of distal colon were mounted in the Ussing flux chamber and treated with TTX and piroxicam to remove the influences of neuronal activity and endogenous PG synthesis, respectively. Although SP (10(-7) M) itself evoked little increase in I(sc), exogenous PGE(2) concentration dependently enhanced the response of SP. The effect of PGE(2) on the SP-evoked response was mimicked by forskolin and 8-bromoadenosine cAMP. Depletion of Ca2+ from the bathing solution reduced the PGE(2)-dependent response of SP. Effects of PGE(2), SP, and SP in the presence of PGE(2) on intracellular Ca2+ concentration ([Ca2+](i)) in isolated crypt cells were measured by the confocal microscope fluorescence imaging system. SP, but not PGE(2), temporally evoked an increase in [Ca2+](i) but declined to the baseline within 3 min. A return of the SP-evoked increase in [Ca2+](i) was slower in the presence of PGE(2) than SP alone. These results suggest that PGE(2) synergistically enhances SP-evoked Cl(-) secretion via an interaction between the intracellular cAMP and [Ca2+](i) in the epithelial cells. In conclusion, SP and PGE(2) could cooperatively induce massive Cl(-) secretion in guinea pig distal colon at epithelial levels.