W1553 Glutamate Reduces Helicobacter pylori-Induced Gastric Atrophy in Rodents - Possible Involvement of Chief Cell Protection via Glutamate Metabolism-

Abstract

The pH of the alkaline microclimate overlying the duodenal enterocyte brush border is regulated by an ecto-purinergic signaling system consisting of intestinal alkaline phosphatase (IAP), extracellular ATP and P2Y receptors. This system regulates the rate of duodenal bicarbonate secretion (DBS). IAP inhibition increases DBS and non-lytic ATP release into the lumen, the mechanism of which has not been clarified. Ecto-F1F0-ATP synthase has been localized to the plasma membrane of several non-epithelial cell types. We hypothesized that ecto-ATP synthase generates extracellular ATP, regulating DBS as part of the ectopurinergic signaling system. We measured DBS with flow-through pH and CO2 electrodes with the perfusate ATP content measured by luciferin-luciferase bioassay. We tested the effect of perfusion of the competitive AP inhibitor glycerol phosphate (GP, 10 mM) with or without the addition of ATP synthase inhibitors oligomycin (Om, 5 μg/ml), piceatannol (Pic, 20 μM) or resveratorol (Res, 20 μM), or the mitochondrial uncoupler carbonyl cyanide 3-chlorophenylhydrazone (CCCP, 1 μM) on DBS and ATP output. GP increased DBS and ATP output. Pic or Res inhibited GP-induced augmented DBS and ATP output. GP, or Pic or Res with GP had no effect on LDH release, confirming the absence of lytic cellular injury. Om or CCCP had a lesser effect on GP-induced DBS, but inhibited GP-induced ATP output accompanied by increased LDH release, consistent with cellular injury. Furthermore, α and β subunits of F1 complex of ATP synthase were immunolocalized to the brush border membranes (α β) in the lamina propria mucosa. Furthermore, phosphate buffer saline (pH 7.0, 10 mM) increased ATP output, suggesting that excess supply of inorganic phosphate (Pi) may enhance ATP synthesis and/or inhibit ATP degradation. Luminal ATP release unmasked by IAP inhibition is dependent on extracellular ATP synthesis, consistent with the presence of ecto-ATP synthase on the brush border. Extracellular ATP synthesis and ATP degradation by IAPmay coordinately regulate ATP-P2Y signaling for DBS as part of a novel ecto-purinergic signaling system.