Three-dimensional reconstruction of cytoplasmic membrane networks in parietal cells

Abstract

There is general agreement that stimulation and consequent secretion of gastric parietal cells result in a great expansion of the apical canalicular membrane at the expense of an extensive intracellular network of membranes rich in the gastric proton pump (H,K-ATPase). However, there is ongoing controversy as to the precise nature of the intracellular membrane network,conventionally called tubulovesicles. At the heart of this controversy lies the question of whether tubulovesicles are a distinct membrane compartment or whether they are continuous with the apical plasma membrane.To address this controversy we used high-pressure, rapid freezing techniques to fix non-stimulated (resting) rabbit gastric glands for electron microscopy. Ultra-thin (60-70 nm) serial sections were used for conventional TEM; 400-500 nm sections were used for tomography. Images were digitized and models constructed using Midas and Imod software(http://bio3d.colorado.edu). Images were aligned and contours drawn on specific cellular structures. The contours from a stack of serial sections were arranged into objects and meshed into 3D structures. For resting parietal cells our findings are as follows:(1) The apical canaliculus is a microvilli-decorated, branching membrane network that extends into and throughout the parietal cell. This agrees well with a host of previous studies. (2) The plentiful mitochondria form an extensive reticular network throughout the cytoplasm. This has not previously been reported for the parietal cell, and the significance of this observation and the dynamics of the mitochondrial network remain unknown. (3)H,K-ATPase-rich membranes do include membrane tubules and vesicles; however,the tubulovesicular compartment is chiefly comprised of small stacks of cisternae. Thus a designation of tubulocisternae seems appropriate; however,in the resting cell there are no continuities between the apical canaliculus and the tubulocisternae or between tubulocisternae. These data support the recruitment-recycling model of parietal cell stimulation.