A flowcytometric method for evaluation of acid secretion from isolated rat gastric mucosal cells

Abstract

Studies on the gastric proton pump are mostly performed on the H⁺, K⁺-ATPase enzyme in the microsomal preparation or by aminopyrine accumulation in the gastric parietal cells. H⁺, K⁺-ATPase activity is estimated by both spectrophotometric and fluorimetric methods. In the present study, quenching or augmentation in acridine orange (AO) fluorescence was monitored on a flowcytometer in rat gastric mucosal cells. Rat gastric mucosal cells were isolated by the standard pronase-EDTA method. The effect of oleic acid, a proton pump inhibitory was evaluated on gastric parietal cell activity and was compared with its effect on proton transport, H⁺, K⁺-ATPase, and p-nitrophenyl phosphatase (p-NPPase) activity in gastric microsomes. In addition, the effect of histamine and carbachol, gastric acid release inducers, was also investigated by flowcytometry in isolated parietal cells. Histamine and carbachol, in a dose-dependent manner, stimulated acid release from isolated gastric cells. Oleic acid also dose-dependently inhibited the basal and stimulated acid release from the cells, as well as in all three enzyme preparations associated with gastric proton pump activity. Thus, the results suggest that flowcytometric method might be used to study basal, as well as stimulated, proton pump activity in isolated gastric parietal cells.