Regulated alkali secretion acts in tandem with unstirred layers to regulate mouse gastric surface pH

Background&Aims: The physical basis for the protective pH gradient at the gastric surface is unconfirmed. This study examined the role of mucus, the unstirred layer, and acid/alkali secretion in controlling gastric surface pH in vivo. Methods: Stomachs of anesthetized mice were exteriorized, and exposed gastric mucosa was imaged by confocal microscopy. Results: Accessibility of molecules at the gastric surface was determined by monitoring the decrease in probe fluorescence over time after dyes were removed from perfusate. On dye removal, CI-NERF (400 molecular weight) fluorescence decreased more slowly at the gastric surface in the presence of mucus (rate constant [k] = 0.08 +/- 0.02 per second) than in the absence of mucus (k = 0.15 +/- 0.02 per second) or 90 mum distant from the surface (k = 0.22 +/- 0.03 per second). In contrast, 70-kilodalton Cl-NERF/dextran washed from the gastric surface more slowly in the presence (k = 0.05 +/- 0.01 per second) or absence (k = 0.09 +/- 0.01 per second) of mucus compared with 90 mum from the tissue surface (k = 0.36 +/- 0.08 per second). Two-photon uncaging of fluorescein near nonsuperfused gastric surface showed that diffusion was not slowed at the gastric surface compared with diffusion in free solution. Surface pH was measured by CI-NERF ratio imaging. Increasing the superfusion rate decreased the thickness of the surface pH gradient without significantly changing surface pH values, suggesting a pH set point of approximately 4 for control of surface pH. Increasing perfusate pH buffers decreased surface pH toward perfusate values. Conclusions: Proton concentration at the gastric surface is the result of regulating acid/alkali secretion to a set point in combination with an unstirred layer and not by trapping of proton or small-molecular-weight buffers in the unstirred layer.