The possibility that prostaglandin E2 (PGE2) increases endometrial vascular permeability and initiates decidualization in sensitized rat uteri by stimulation of adenosine 3':5'-cyclic monophosphate (cAMP) synthesis was investigated. Immature rats, pretreated so that they were sensitized for the decidual cell reaction, were used. Following the unilateral intrauterine injection of 50 microliters phosphate-buffered saline containing gelatin (PBS-G), a deciduogenic stimulus, uterine concentrations of both PGE and cAMP were elevated as early as 1 min after the intrauterine treatment. To determine if uterine stimuli which increase endometrial vascular permeability also increase uterine cAMP concentrations, rats, treated with or without indomethacin, an inhibitor of PG synthesis, received unilateral intrauterine injections of 50 microliters PBS-G with and without 10 micrograms PGE2 and were killed 15 min later. Uterine cAMP concentrations were elevated in all injected horns except in those of indomethacin-treated rats receiving PBS-G intraluminally, thus paralleling the expected changes in endometrial vascular permeability. As indicated by radioactivity levels in the stimulated horn 15 min after the i.v. injection of 125I-labeled bovine serum albumin, the intrauterine injection of dibutyryl cAMP, with or without theophylline, did not increase endometrial vascular permeability in indomethacin-treated animals. In contrast, cholera toxin, an activator of adenylate cyclase activity, markedly elevated permeability and induced decidualization. Except for the lack of a permeability response to the cAMP analogue, these data are consistent with the hypothesis that the effect of PGE2 on endometrial vascular permeability is mediated by cAMP.