Stimulation of mucosal alkaline secretion represents an opportunity for discovering novel drugs of potential benefit in maintenance therapy of duodenal ulcer disease. We screened over 200 agents representing the full spectrum of pharmacological categories in order to characterize stimulatory pathways and identify mechanistic leads. A variety of eicosanoids, phospho-diesterase inhibitors and adrenoreceptor agonists together with forskolin, 6-hydroxy-dopamine, 2-chloroadenosine, diazepam, testosterone, dipyridamole and dihydropyridazinone caused a reproducible increase in the metabolism-dependent component of alkaline secretion in bullfrog proximal duodenum. PGE2 (ED50 0.02 microgram/ml) was the most potent agent in vitro and was also the most effective stimulant of duodenal alkalinization in vivo in an anaesthetized cat preparation. Agents without effect on spontaneous alkaline secretion by amphibian duodenum included agonists and antagonists of histamine, 5-hydroxy-tryptamine, gamma-aminobutyric acid, dopamine, muscarinic and nicotinic receptors, inhibitors of amine uptake, monoamine oxidase and cholinesterase, plus various corticoids, diuretics, oestrogens, chemotherapeutic (anticancer) and antimicrobial agents. The major mechanism of stimulating alkaline secretion in the isolated duodenum is by increasing intracellular cyclic AMP levels. This may occur by either inhibiting metabolism of the nucleotide or by stimulating its formation. Additionally, many stimulants appear to act indirectly via liberation of endogenous prostaglandins as judged from the marked attenuation of responses in the presence of indomethacin to all agonists apart from exogenous PGE2, forskolin, ICI 63197 (PDE inhibitor), 2-chloroadenosine and diazepam. Whether purinergic agonists and benzodiazepines act directly on the enterocyte or by releasing other paracrine mediators is unknown.