The mechanisms which enable cholera toxin (CT) and the Escherichia coli heat-stable enterotoxins (STa and STb) to stimulate intestinal secretion of water and electrolytes are only partially understood. CT evokes the synthesis of 3',5'-cyclic AMP (cAMP), and STa is known to elevate intestinal levels of 3',5'-cyclic GMP (cGMP). Neither of these recognized second messengers appears to mediate E. coli STb responses. We compared the secretory effects of CT, STa, and STb using the pig intestinal loop model and also measured the effects of toxin challenge on the synthesis of cAMP, cGMP, and prostaglandins (e.g., prostaglandin E2 [PGE2]), as well as on the release of 5-hydroxytryptamine (5-HT) from intestinal enterochromaffin cells. All three enterotoxins elicited fluid accumulation within a 2-h observation period. A combination of maximal doses of STa with STb yielded additive effects on fluid accumulation, which suggested different mechanisms of action for these toxins. Similarly, challenge of pig intestinal loops with a combination of CT and STb resulted in additive effects on fluid accumulation and luminal release of 5-HT. Unlike its effect on intestinal tissues from other animals, CT did not appear to elicit a dose-dependent cAMP response measurable in mucosal extracts from pig small intestine. In contrast, luminal fluid from CT-challenged pig intestinal loops contained dose-related amounts of cAMP and PGE2 that had been secreted from the mucosa. cAMP responses to STa or STb could not be demonstrated in either mucosal tissue or luminal fluid. In contrast, cGMP levels were increased in the intestinal fluid of loops challenged with STa but not in those challenged with STb. While the mechanisms of action of CT and STa are thought to involve impulse transmission via the enteric nervous system, we demonstrated significant stimulation of PGE2 synthesis and 5-HT release for CT and STb but very little for STa. We conclude from these data that the mechanisms of action of STa, STb, and CT are distinct, although the mode of action of STb may have some similarity to that of CT. Since STb stimulated the release of both PGE2 and 5-HT from the intestinal mucosa, the data suggested the potential for an effect of STb on the enteric nervous system.