The contribution of the surface mucous gel to the stomach's hydrophobic luminal properties and how these properties are affected by both damaging and cytoprotective agents were studied. Canine gastric mucosa, determined to be hydrophobic in nature by contact angle analysis, had an adherent periodic acid-Schiff-reactive mucous gel layer over 85% of its luminal surface, as observed under light microscopy. Extracellular structures reactive with the phospholipid-selective stain, iodoplatinate, were observed ultrastructurally in these tissues, within and at the luminal interface of the surface mucous gel. Incubating the luminal surface of gastric mucosa mounted in Ussing chambers in acidified aspirin promoted the exfoliation of surface epithelium and markedly reduced surface hydrophobicity, surface periodic acid-Schiff reactivity, and transmucosal potential difference. Addition of 16,16-dimethyl prostaglandin E2 to the nutrient compartment of these chambers maintained surface hydrophobicity at control levels but did not prevent aspirin-induced reductions in potential difference or cellular damage to the surface epithelium. However, prostaglandin did attenuate exfoliation of aspirin-damaged surface mucous cells and preserved the surface mucous gel. These results indicate that the stomach's hydrophobic lining is closely associated with the presence of a surface mucous gel layer, is not an effective barrier to the penetration of lipid-soluble damaging agents such as acidified aspirin, and is maintained by exogenous prostaglandin as is the mucous gel layer, even in the presence of luminal aspirin. The ability of prostaglandin to maintain a hydrophobic mucous gel layer over compromised tissue may, in part, explain its ability to limit aspirin-induced injury and promote the recovery and restitution of the surface epithelium.