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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

Akira Sawaguchi, Kent L. McDonald and John G. Forte

Department of Molecular and Cell Biology (AS,JGF) and Electron Microscopy Lab (KLM), University of California, Berkeley, California

Correspondence to: Prof. John G. Forte, Dept. of Molecular and Cell Biology, University of California, 241 LSA, Berkeley, CA 94720-3200. E-mail: jforte{at}uclink.berkeley.edu

High-pressure freezing (HPF) is currently the most reliable method to obtain an adequately frozen sample for high-resolution morphological evaluation. Here we applied the HPF technique to isolated rabbit gastric glands to reveal structural evidence that may be correlated with functional activity of gastric parietal cells. This approach provided well-preserved fine structure and excellent antigenicity of several parietal cell proteins. Microtubules were abundant in the cytoplasm and frequently appeared to be associating with tubulovesicles. Interestingly, many electron-dense coated vesicles were apparent around the intracellular canaliculi (IC) of resting parietal cells, consistent with active membrane retrieval from the apical membranes. Immunolabeling of H⁺/K⁺-ATPase was evident on the endocytic components (e.g., multivesicular bodies) and tubulovesicles. After histamine stimulation, the parietal cells characteristically showed expanded IC membranes with varied features of their apical microvilli. The labeling density of H⁺/K⁺-ATPase was four-fold higher on the IC membrane of stimulated parietal cells than on that of resting parietal cells. Immunolabeling of ezrin was clearly identified on the IC and basolateral membranes of parietal cells, corresponding to their F-actin-rich sites. The present findings provide a new insight into the correlation of cell structure and function in gastric parietal cells.

(J Histochem Cytochem 52:77–86, 2004)

Key Words: high-pressure freezing • electron microscopy • isolated gastric gland • parietal cell • transformation • immunogold labeling • H⁺/K⁺-ATPase • ezrin

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