This in vitro study investigated the relationship between rabbit colonic muscularis mucosae motor activity and changes in transmucosal potential difference. Spontaneous muscle contractions and potential difference oscillations occurred independently and were not neurally driven. ACh and histamine directly stimulated the muscularis mucosae, but their mucosal effects were largely indirect, suggesting that muscularis mucosae contractions promote epithelial secretion. 1,1-Dimethyl-4-phenyl-piperazinium iodide and vasoactive intestinal polypeptide induced large potential difference changes but small muscularis mucosae contractions, demonstrating mucosal secretion without significant muscle activity. Lowered intraluminal pH directly stimulated the muscle, whereas a bile salt-lipid mixture evoked TTX- and atropine-sensitive increases in its contractile activity. Increased intraluminal pressure and hypertonic luminal perfusion did not elicit muscularis mucosae excitation. Thus under basal conditions muscle and mucosal activities are independent, but evoked muscularis mucosae contractions can stimulate epithelial secretion. In response to specific luminal stimuli, muscularis mucosae motor activity is increased via the activation of cholinergic nerves. These data suggest that muscularis mucosae and mucosal functions are physiologically linked and that their activities can be coordinated by multiple mechanisms.