Electron microscopic and cytochemical studies of the mouse subcommissural organ

Summary In mice most of the ependymal cells of the subcommissural organ (SCO cells) are densely packed with dilated cisternae of the endoplasmic reticulum (ER) containing either finely granular or flocculent materials. The well developed supra-nuclear Golgi apparatus consists of stacks of flattened saccules and small vesicles; the two or three outer Golgi saccules are moderately dilated and exhibit numerous fenestrations; occasional profiles suggesting the budding of coated vesicles and formation of membrane-bound dense bodies from the ends of the innermost Golgi saccules are seen. A few coated vesicles and membrane-bound dense bodies of various sizes and shapes are also found in the Golgi region.The contents of the dilated ER cisternae are stained with periodic acid-silver methenamine techniques. In the Golgi complex the two or three inner saccules are stained as deeply as the dense bodies, and the outer saccules are only slightly stained. The stained contents of ER cisternae are more electron opaque than those of the outer but less opaque than those of the inner Golgi saccules and the dense bodies.Acid phosphatase activities are localized in the dense bodies, some of the coated vesicles in the Golgi region, and in the one or two inner Golgi saccules.On the basis of these results the following conclusions have been reached: (1) In mouse SCO cells the finely granular and the flocculent materials in the lumen of ER cisternae contain a complex carbohydrate(s) which is secreted into the ventricle to form Reissner's fiber; (2) the secretory substance is assumed to be synthesized by the ER and stored in its cisternae, and the Golgi apparatus might play only a minor role, if any, in the elaboration of the secretory material; (3) most of the dense bodies in the mouse SCO cells are lysosomal in nature instead of being so-called dark secretory granules.Sponsored by the National Science Council, Republic of China.     

Electron cytochemical study of carbohydrate components in cultured nerve and glial cells of the snail Helix pomatia

• 1.1. A variety of colloidal gold-labelled lectins with different sugar specificities to determine whether different nerve and glial cells of the snail Helix pomatia cultured in vitro, can be distinguished by the carbohydrates that they express was screened. The analysis of lectin binding has shown substantial differences in the carbohydrate pattern between nerve and glial cells and between the soma of monoaminergic and peptidergic neurons.
• 2.2. The surface of monoaminergic and peptidergic neurons contains N-acetylglucosamine and N-acetyllactosamine determinants, and does not exhibit neuraminic acid and complex branched N-glycosyl chains. Moreover, N-acetylgalactosamine can be detected on peptidergic neuron membranes only.
• 3.3. N-Acetylglucosamine residues are not present on the surface of the glial cells, and the density of the N-acetyllactosamine and/or terminal β-galactose residues is much higher here than on the surface of the nerve cells.
• 4.4. These results suggest that nerve cells in the snail brain can be distinguished from glial cells by the presence of a cell-surface glycoconjugate containing terminal N-acetyl-d-glucosamine residues, whereas peptidergic neurons can be distinguished from monoaminergic neurons by the presence of a surface glycoconjugate containing terminal α-linked N-acetyl-d-galactosamine residues.

     

Electron microscopic research on the mitochondria-rich bladder cells of the frog

The ultrastructural peculiarities of mitochondria-rich cells of the frog urinary bladder are analysed using three electron microscopic methods: ultrathin sections, scanning electron microscopy, freeze fracture. The mitochondria and tubular and vesicular structures are most abundant in the apical region of cytoplasm. The P-face (PF) of the apical plasma membrane is characterized by the presence of rod-shaped intramembrane particles (IMP), whereas the E-face (EF) possesses complementary pits. Depending on the distribution density of the rod-shaped IMP, three types of cells are described. The apical plasma membrane has an invert distribution of the globular IMP: a great quantity of IMP on the EF and a few particles on the PF. This structure of the apical plasma membrane is supposed to correlate with its very low water permeability. Using filipin as a marker of cholesterol localization, it has been shown that the mitochondria-rich cell apical membrane contains more cholesterol than that of the granular cells. The nature of the rod-shaped IMP and their role in the transmembrane ion transport have been discussed.     

Electron cytochemical study of carbogydrate components of surface membrane in cultured glial cells of the snailHelix pomatia

Using a variety of colloidal gold-labelled lectins, the structure and topography of carbohydrate determinants of the surface membrane in different types of cultured glial cells of the snailHelix pomatia have been electron cytochemically investigated. Analysis of lectin binding having different sugar specificities have shown heterogeneity of carbohydrate pools between glial and nerve cells and among different types of glial cells. It was found that satellite glial cells displaying ultrastructural traits of intensive metabolism (type II cells) selectively bindGNA, which is specific for terminal -D-mannose residues, and do not interact (Con A) or slightly interact (LCA) with other mannose-specific lectins.GNA determinants remain during the whole period of cell growth and are absent in satellite type-I glial cells, fibrous glial cells, microglia, and neurons.LTA, PVA, andLABA do not bind to any glial cells.WGA determinants, which are abundant on the neurons, are completely absent onGNA-binding glial cells and single on other types of glial cells. The density ofPNA determinants on microglial cells is the highest, as compared with other types of glial cells or neurons. It is concluded that some lectin determinants (forRCA-1, PNA, LPA) are present on all types of glial cells, while another determinant (GNA) is specific for a certain type of glial cells only and can serve as a marker of these cells. The role of specific carbohydrate determinants for neuron-glia interaction in mature brain is discussed.Neirofiziologiya/Neurophysiology, Vol. 26, No. 3, pp. 177–189, May–June, 1994.     

Electron microscopic cytochemical localization of Ca-ATPase in toad urinary bladder

The calcium-regulating enzyme calcium adenosine triphosphatase (Ca-ATPase) was localized in the epithelium of amphibian urinary bladder by the one-step electron microscopic cytochemical procedure. The enzyme was identified along the basolateral border of the epithelial cells that comprise the bladder mucosa. The electron-dense precipitate indicating Ca-ATPase activity was seen in association with the outer leaflet of the basolateral plasmalemmae. Intracellularly, Ca-ATPase activity was seen in association with the mitochondrial matrix of the mitochondria-rich cells. Ca-ATPase was not seen along the apical microvillated border. Enzyme activity was also not seen after incubation in substrate-free media, calcium-free media, or incubation in the presence of vanadate. However, Ca-ATPase activity was evident when the calcium in the standard reaction medium was deleted in favor of magnesium. Addition of antidiuretic hormone (ADH; vasopressin) increased both the basolateral Ca-ATPase reaction and the mitochondrial reaction. Such data appear to indicate further that changes in cytosolic calcium ion concentration take place during the response of amphibian urinary bladder to the polypeptide hormone vasopressin.     

Electron microscopic analysis of glial cells in the rat telencephalon stained with the Neo-Timm and selenium methods

Two histochemical methods for visualization of zinc in synaptic vesicles, the Neo-Timm and selenium methods, have been shown to additionally stain glial cells and neuronal somata. In a previous light microscopic study the majority of stained glial cells were seen in the major fiber tracts of the rat telencephalon. The aim of the present study was to further characterize the stained glial cells with respect to glial cell type and ultrastructural localization of the silver grains responsible for the staining. Electron microscopic analysis of brains treated according to either method revealed that the vast majority of stained glial cells belonged to the dark oligodendroglial cell type. However, a smaller number of stained astrocytes was also seen, especially in the grey matter. The silver grains responsible for the staining were located in electron-dense rounded cytoplasmic organelles, suggestive of lysosomes.     

Electron microscopic analysis of glial cells in the rat telencephalon stained with the Neo-Timm and selenium methods

Summary Two histochemical methods for visualization of zine in synaptic vesicles, the Neo-Timm and selenium methods, have been shown to additionally stain glial cells and neuronal somata.In a previous light microscopic study the majority of stained glial cells were seen in the major fiber tracts of the rat telencephalon. The aim of the present study was to further characterize the stained glial cells with respect to glial cell type and ultrastructural localization of the silver grains responsible for the staining.Electron microscopic analysis of brains treated according to either method revealed that the vast majority of stained glial cells belonged to the dark oligodendroglial cell type, However, a smaller number of stained astrocytes was also seen, especially in the grey matter. The silver grains responsible for the staining were located in electron-dense rounded cytoplasmic organelles, suggestive of lysosomes.     

Electron microscopic cytochemical and morphometric study of the cerebral cortex synapses in postmortem autolysis

Brain tissue staining with phosphotungstic acid was performed to assay neurofilament accumulations in synapses in the molecular layer of the rat cerebral cortex at different intervals after the animals' death. It was found that autolysis began in the dense projections of presynaptic grid. Within 30 min autolysis developed in mature and very young (immature) synapses. By the 90th min autolysis in asymmetric synapses was considerably enhanced. 6 hours later autolysis involved mature and indefinite synapses.     

Electron microscopic cytochemical characterization of bile canaliculi and bile ducts in vitro.

Electron microscopic cytochemical localization of Mg++-activated adenosine triphosphatase (Mg++-ATPase) and 5-nucleotidase (AMPase) was investigated in bile canaliculus-rich and bile duct-containing fractions isolated from rat liver. Comparative cyochemical studies between prefixed and non-prefixed fractions revealed that the activity of both enzymes could be detected in the fractions under appropriate experimental conditions. However, the cytochemical activity of AMPase was much more sensitive to glutaraldehyde than that of Mg++-ATPase. Mg++-ATPase and AMPase reaction products were localized primarily on bile canalicular microvilli, that is, along the outer (luminal) surface of canalicular plasma membranes, but they were never observed on bile ductal microvilli. AMPase was also detectable on lateral hepatic plasma membranes. Mg++-ATPase demonstrated by the cytochemical technique described is a reliable enzyme marker for isolated bile canalicular membranes. At high magnification, Mg++-ATPase reaction product was also observed on the microfilaments surrounding isolated bile canaliculi. The possibility that the reaction product on the pericanalicular microfilaments may result from the hydrolysis of ATP byan actomyosin ATPase-like enzyme associated with these filaments is briefly discussed.     

Electron microscopic research on the mechanism of intestinal secretion

Electron microscopic investigation of the rat small intestine revealed a great number of vesicles 50-75 nm in diameter with enterocyte microvilli. The number of vesicles increased with the increase of digestive activity in the small intestine. Vesicles were formed by gemmation of enterocyte microvilli from the lateral membrane in contraction of microvillous actin skeleton. Simultaneously with the production of exocytotic vesicles, the formation of pinocytotic vesicles in the base of microvilli was observed. There is a supposition that the vesicle gemmation is a natural process of the intestinal secretion to fulfil numerous important function: it promotes the penetration of enterocyte hydrolases into the parietal layer; equilibrates an increase in the enterocyte volume during absorption. This is a possible way of translocation of synthesized enzymes into the cytoplasm and of transport proteins on the apical surface of epithelial cells.     

Electron microscopic autoradiographic and electron microscopic immunohistochemical studies on the anti-HPO antibody-producing cells

The secondary immune responses in mouse popliteal lymph nodes to horseradish peroxidase (HPO) were studied by a combination of electron microscopic autoradiography and electron microscopic immunohistochemistry in order to clarify the relationship between antibody-producing and DNA-synthesizing capacities of the plasmacytic series. The anti-HPO antibody-containing cells, which increased in number 72 h after the secondary antigenic stimulation, were mainly immunoblasts and immature plasma cells. Immunoblasts containing anti-HPO antibody incorporated [³H]thymidine more actively than did immature plasma cells containing anti-HPO antibody. In 144 h after the secondary antigenic stimulation, antibody containing cells consisted mainly of mature plasma cells and immature plasma cells. Immature plasma cells containing the anti-HPO antibody incorporated a little [³H]thymidine, but mature plasma cells containing anti-HPO antibody did not incorporate any [³H]thymidine.