Acid secretion and membrane reorganization in single gastric parietal cell in primary culture

Abstract

A digitally-enhanced videomicroscopy study of rabbit gastric parietal cells in primary culture was performed using alternate observations with differential interference contrast and fluorescence optics of cells mounted and perfused on a temperature-controlled microscope stage. The effect of histamine, a physiological effector of acid secretion, was followed. Isolated parietal cells possess an internal apical vacuole, which kept the cell in a pseudopolarized state. This apical vacuole is a site of acid secretion. This was demonstrated by the direct visualization of the uptake of the fluorescent weak base 9-amino acridine and of the concomitant enormous swelling of the acid vacuole which reached an estimated size of 3-7 times the normal cell volume. This morphological change of shape and acidification of apical vacuoles was fully reversible and cells could respond to successive stimulations. A quantitative study of these events provided a value of the acid accumulation index for each single cell in response to histamine. Individual cell response varied within a factor of 7. The cellular localization of the proton pump complex responsible for acid secretion and of the major components of the secretory microvilli, actin and ezrin, a histamine-dependent phosphorylation target of protein kinase A, were detected by indirect immunofluorescence microscopy in resting and stimulated cells. Both actin and ezrin colocalized at the apical vacuole membrane in resting and stimulated cells, whereas the proton pump shifted from an intracytoplasmic pool to the apical vacuole membrane upon stimulation.

High-pressure Freezing of Isolated Gastric Glands Provides New Insight into the Fine Structure and Subcellular Localization of H+/K+-ATPase in Gastric Parietal Cells

“…For some systems it is believed that vesicular transport is directed by microtubular motor proteins, such as kinesin, in an ATP-dependent manner (for review see McNiven and Marlowe 1999), but it still remains to be clarified whether and which motor proteins exist and operate on the microtubules in gastric parietal cells. Stimulation-associated volume expansions have been previously observed in the internalized apical membrane vacuoles of cultured parietal cells (Chew et al 1989;Mangeat et al 1990). It has been proposed that the volume expansion was due to the concomitant secretion of hydrochloric acid and water (Mangeat et al 1990).…”

“…Stimulation-associated volume expansions have been previously observed in the internalized apical membrane vacuoles of cultured parietal cells (Chew et al 1989;Mangeat et al 1990). It has been proposed that the volume expansion was due to the concomitant secretion of hydrochloric acid and water (Mangeat et al 1990). In the present study, extensive volume expansion was also observed in the glandular lumen of some histaminestimulated glands, suggesting an accumulation of glandular secretory products, e.g., hydrochloric acid, mucin, zymogen.…”

“…It could be postulated that the occluded IC were formed in recovering parietal cells induced by cimetidine treatment in this study. To date, surprisingly, there have been relatively few studies concerning the morphological changes in the recovering state (Helander and Hirschowitz 1972;Forte et al 1977;Schofield et al 1979;Mangeat et al 1990), even though there is general acceptance of the membrane recruitment and "recycling" hypothesis. It will be of great interest to morphologically examine the recovering state of parietal cells using HPF and to elucidate the physiological significance of the occluded structures as well as other forms of membrane retrieval in relation to the regulation of acid secretion.…”

Phosphorylation of ezrin on threonine 567 produces a change in secretory phenotype and repolarizes the gastric parietal cell

“…2A,B. Several laboratories commonly report that cultured parietal cells take on a different morphological form than in situ (Agnew et al, 1999;Chew, 1994;Mangeat et al, 1990). After a few hours in culture the apical membrane is sequestered in the form of easily identified vacuoles (2-5 m in diameter), which we refer to as apical membrane vacuoles.…”

“…Because the surrounding plasma membrane continues to serve as the site of receptor-mediated cell activation events, we refer to it as the basolateral membrane. Cultured parietal cells continue to maintain a secretagogue-dependent phenotype whereby H,KATPase-rich tubulovesicles are translocated to, and fuse with, apical membrane vacuoles and the resulting secretion of HCl produces gross vacuolar enlargement (Agnew et al, 1999;Mangeat et al, 1990;Zhou et al, 2003b). From the images of resting cells (i.e.…”