BACKGROUND The mechanism by which topical isoproterenol protects the gastric mucosa from ethanol-induced injury is unclear; previous studies suggest that enhancement of mucosal blood flow may be contributory. Therefore, we investigated the role of isoproterenol pretreatment on rat gastric mucous gel layer thickness, mucosal blood flow, and surface cell intracellular pH. METHODS A portion of the gastric corpus of an anesthetized rat was isolated in a superfused chamber. Intracellular pH was measured with the fluorescent dye 5(6)-carboxyfluorescein diacetate, mucous gel thickness was measured by quantifying the distance between the focal planes of the epithelial cell and the gel surfaces, and mucosal blood flow was measured in separate experiments with a laser-Doppler flowmeter. RESULTS Pretreatment with topical isoproterenol markedly increased gastric mucous gel layer thickness, decreased surface cell initial acidification rates, and significantly enhanced the recovery of intracellular pH toward baseline values. Gastric mucosal blood flow was significantly higher in the isoproterenol-treated group compared with untreated controls, initially increasing, decreasing, and then increasing a second time during superfusion with acidic solutions. CONCLUSIONS Isoproterenol preserves gastric surface cell intracellular pH during acid superfusion. This effect may be a result of an increased thickness of the mucous layer, which delays proton entry into the surface cell, and an increase in mucosal blood flow in response to luminal acid, which appears to enhance recovery from intracellular acidosis.