FAST FREEZE-FIXATION FREEZE-SUBSTITUTION REVEALS THE SECRETORY MEMBRANES OF THE GASTRIC PARIETAL-CELL AS A NETWORK OF HELICALLY COILED TUBULE - A NEW MODEL FOR PARIETAL-CELL TRANSFORMATION

The parietal cell of the gastric mucosa undergoes rapid morphological transformation when it is stimulated to produce hydrochloric acid. In chemically fixed cells, this process is seen as a reduction in number of cytoplasmic 'tubulovesicles' as the apical surface of the cell prog ressively invaginates to increase the secretory surface area. It is wi dely believed that the tubulovesicles represent stored secretory membr ane in the cytoplasm of the unstimulated cell, which is incorporated i nto the apical membrane upon stimulation, because they share H+,K+-ATP ase activity with the apical membrane. However, fusion of tubulovesicl es with the apical membrane concomitant with parietal cell activation has never been convincingly demonstrated. We have used fast freeze-fix ation and freeze-substitution to study stages of morphological transfo rmation in these cells. Tubulovesicles were not seen in the cytoplasm of any of our cryoprepared cells. Instead, the cytoplasm of the unstim ulated cell contained numerous and densely packed helical coils of tub ule, each having an axial core of cytoplasm. The helical coils were li nked together by connecting tubules, lengths of relatively straight tu bule. Lengths of straight connecting tubule also extended from coils l ying adjacent to the apical and canalicular surfaces and ended at the apical and canaliculus membranes. Immunogold labelling with alpha- and beta-subunit-specific antibodies showed that the gastric H+,K+-ATPase was localized to the membranes of this tubular system, which therefor e represented the configuration of the secretory membrane in the cytop lasm of the unstimulated parietal cell. Stimulation of the cells with histamine and isobutylmethylxanthine lead to modification of the tubul ar membrane system, correlated with progressive invagination of the ap ical membrane. The volume of the tubule lumen increased and, as this o ccurred, the tight spiral twist of the helical coils was lost, indicat ing that tubule distension was accounted for by partial unwinding. Thi s exposed the cores of cytoplasm in the axes of the coils as rod-shape d elements of a three-dimensional reticulum, resembling a series of mi crovilli in random thin sections. Conversely, treatment with the H-2 a ntagonist cimetidine caused severe contraction of the tubular membrane system and intracellular canaliculi. Our results indicate that tubulo vesicles are an artifact of chemical fixation; consequently, they cann ot have a role in parietal cell transformation. From our findings we p ropose an alternative model for morphological transformation in the pa rietal cell. This model predicts cytoskeleton-mediated control over ex pansion and contraction of the tubular membrane network revealed by cr yopreparation. The model is compatible with the localization of cytosk eletal components in these cells.