This in vitro study tested the hypothesis that muscularis mucosae contractile activity contributes to rabbit colonic mucosal function by mechanisms other than simple mechanical deformation of the epithelium. Experiments were performed by using a technique that allows simultaneous recording of muscle activity and transmucosal potential difference, a measure of epithelial ion transport. ATP, bradykinin, histamine, PGE₂, PGF₁, and PGF₂ elicited muscularis mucosae contractions that were resistant to atropine and TTX. Only ATP-induced contractions were indomethacin sensitive, and only those to dimethylphenylpiperazinium iodide (DMPP) were reduced by atropine. All agonist-evoked increases in transmucosal potential difference were atropine resistant, and, with the exception of those to PGE₂, PGF₂, and VIP, they were also TTX sensitive. Mucosal responses to ATP, bradykinin, and histamine were indomethacin sensitive, whereas those to DMPP, the prostaglandins, and VIP were not. When cyclooxygenase activity or the mucosal innervation was compromised, even maximal muscularis mucosae contractions did not produce large secretory responses. It is concluded that contraction-related prostaglandin synthesis and noncholinergic secretomotor neuron stimulation represent the physiological transduction mechanism through which muscularis mucosae motor activity is translated into mucosal secretion.