Interstitial buffer capacity influences Na+/H+ exchange kinetics and oxyntic cell pHi in intact frog gastric mucosa

We studied the influence of interstitial buffer capacity and CO2-HCO3- on oxyntic cell intracellular pH (pHi) in intact frog gastric mucosa. Oxyntic cells in stripped gastric mucosa of Rana esculenta were loaded with the pH-sensitive dye 2',7'-bis(2-carboxyethyl)-5(6)-carboxyfluorescein, and pHi was assessed fluorometrically. In the presence of a constant serosal and luminal pH, oxyntic cell pHi was dependent on the serosal but not the luminal concentration of cell-impermeable buffer ions such as N-2-hydroxyethylpiperazine-N'-2-ethanesulfonic acid (HEPES), tris(hydroxymethyl)aminomethane, N-tris(hydroxymethyl)methyl-2-aminoethanesulfonic acid, and 3-(N-morpholino)propanesulfonic acid. The stepwise increase in oxyntic cell pHi from 6.74 +/- 0.05 with 1 mM HEPES to 7.23 +/- 0.08 was almost completely inhibited by removal of serosal Na+ and by amiloride and dimethyl amiloride, suggesting that it was largely due to Na+/H+ exchange. Increasing the serosal concentration of a CO2-HCO3- buffer from 1% CO2-4 mM HCO3- to 10% CO2-40 mM HCO3- increased oxyntic cell pHi from 7.03 +/- 0.06 to 7.39 +/- 0.07. This CO2-HCO(3-)-dependent pHi increase was also Na+ and amiloride sensitive, but high HCO3- concentrations increased pHi even in the absence of Na+, K+, or Cl-, and in the presence of omeprazole, bafilomycin A1, or acetazolamide. We suggest that in intact frog gastric mucosa, cellular proton extrusion acidifies the interstitial pH immediately adjacent to the basolateral membrane of the oxyntic cells in the absence of a high interstitial buffer capacity.(ABSTRACT TRUNCATED AT 250 WORDS)