Diffusion barriers in the vaginal epithelium during the estrous cycle in guinea pigs

Summary In the present study the permeability barriers of the multilayered vaginal epithelium were examined using tracer perfusion techniques, freeze-fracture and thin sectioning. During diestrus and proestrus the upper layers of mucified epithelial cells exhibit tight-junctional belts, which restrict tracer molecules such as lanthanum and horseradish peroxidase. When the highly mucified cells begin to degenerate toward the end of proestrus the underlying epithelium is already keratinized as typical for estrus. The keratinized epithelial cells have a tight-junctional network that joins the basal plasma membranes with the apical membranes of subjacent cells and blocks paracellular diffusion of the tracer molecules. During conversion of the cornified epithelium to a mucified epithelium in metestrus the intercellular space of the epithelium is stained by tracer molecules even though tight-junctional belts can be observed.These results indicate that during cyclic changes of the vaginal epithelium tight junctions can, in general, be considered for the restriction of paracellular diffusion. In metestrus, however, junctions become functionally leaky although they remain morphologically intact.Intercellular lipids, which are normally common in cornified epithelia, are extremely rare and cannot constitute an effective barrier to diffusion in the vagina of the guinea pig. The significance of a strategy that bases the regulation of the permeability on tight junctions rather than on intercellular lipids is discussed.     

Macromolecules can pass through occluding junctions of rat ileal epithelium during cholinergic stimulation

Summary Crypt, but not villus, goblet cells in the ileum accelerate their secretion of mucus within 5 min following cholinergic stimulation. This study was done to determine whether the macromolecular permeability and structure of occluding junctions in the ileum are altered during accelerated secretion. Rats were injected intravenously with horseradish peroxidase followed by carbachol (250 g/kg, subcutaneous) and the intestinal mucosa was fixed 3–12 min later. In control mucosa (saline-injected), peroxidase filled lateral intercellular spaces up to the occluding junctions of both crypt and villus epithelium, but did not enter occluding junctions or pass into the lumen. In 3 of 8 carbachol-stimulated rats, peroxidase was present within occluding junctions in crypt epithelium and in the crypt lumen, although all intermembrane junctional fusion sites appeared intact. Villus epithelial occluding junctions, in contrast, continued to exclude peroxidase. In freeze-fracture replicas of crypt cells prepared after carbachol stimulation, we detected no structural changes in strand networks of occluding junctions that could account for increased paracellular permeability.     

Cell junctions in the cyst envelope in the silkworm testis,Bombyx mori linné

Summary The cell junctions of the cyst envelope in the testes of Bombyx mori were examined by electron microscopy utilizing a thin-sectioning technique following conventional fixation, tannic acid fixation and lanthanum tracer study, and also using a freeze-fracture technique. There are three kinds of junctions; septate junctions, gap junctions and tight junctions. Septate junctions are of the pleated type. Gap junctions are characterized by four electron-dense lines and three electronlucent lines in the reduced intercellular spaces seen by thin-sectioning. They are of the E type, having clusters of intramembraneous particles on the E-fracture face. The most striking finding is the frequent presence of tight junctions on the fracture planes, while focally fused outer leaflets of the junctional unit membranes are rarely detected on thin-sectioned preparations. Tight junctions are characterized by branching zigzag ridges on the P-fracture face and complementary grooves on the E-fracture face. It is proposed that tight junctions are new morphological evidence of blood-germ cell barrier in an insect. Acknowledgements: For helpful assistance the authors are indebted to their colleagues Miss N. Minemoto, Miss H. Kiyotake and Mr. Y. Goto     

Junctions between interstitial cells of the renal medulla: A freeze-fracture study

Summary Cell junctions between interstitial cells of the renal medulla were studied in freeze-fracture replicas of kidneys from rat, rabbit, hamster and the tree-shrew Tupaia belangeri. In all species studied a composite type of intercellular junction was found comprising elements of tight junctions and irregular gap junctions of highly variable size and shape. The number of these junctions increased towards the tip of the papilla.Our findings suggest that the composite junctions observed play a role in the maintainance of the ladder-like arrangement of the interstitial cells in the inner zone. The existence of irregular gap junctions raises the possibility that the functions of the interstitial cells are coordinated, especially during alterations of the functional state of the kidney.Supported by the Deutsche Forschungsgemeinschaft     

Ultrastructure of perfusion-fixed fetal capillaries in the human placenta

Summary The ultrastructure of human placental capillaries was investigated using perfusion fixation and the freeze-fracturing technique. The capillaries have a continuous endothelium especially rich in microfilaments, whereas micropinocytotic vesicles are exceedingly scarce. The endothelial cells are connected by three types of junctions: (1) zonulae occludentes characterized by 2 to 4 focal regions of membrane contact in thin-sectioned specimens and an equal number of ridges on the membrane E-face in freeze-fractured specimens; (2) small gap junctions associated with the zonula occludens. (3) attachment plaques resembling zonulae adhaerentes in their fine structure. Endothelial cells are provided with long, circularly oriented pseudopodial extensions, which may be responsible for intermittent constrictions of the vessel lumen. These findings indicate that diaplacental transport at the level of the fetal capillary is controlled by the cytoplasm of the endothelial cells and probably occurs only to a very limited extent by way of micropinocytotic vesicles.Dedicated to Prof. Dr. Drs. h.c. W. Bargmann on his 70. birthdayWith the support of the Deutsche ForschungsgemeinschaftThe authors acknowledge the technical help of Mrs. E. Benecci, and the criticism and discussion of Drs. D.W. Fawcett and S. Ito. We are also indebted to Mr. R. Partsch (Zeiss, Inc. New York) for helping us with the goniometric study     

Electron microscopical investigations of the intercellular contacts during the early cleavage stages ofLymnaea stagnalis (Mollusca,Gastropoda)

Summary In early cleavage stages ofLymnaea stagnalis, three kinds of intercellular junctions could be distinguished up to the sixth cleavage: intermediate, septate and gap junctions. The first two form junctional belts located on the cell border at the periphery of the embryo. For the purpose of our study we were most interested in gap junctions as they are alleged to be structures that allow cell-to-cell communication. Gap junctions first appear at the four cell stage. Up to the sixth cleavage no difference in the distribution pattern could be found between and within each of the four quadrants of the embryo. Some of the cell tiers along the animal-vegetal axis lack gap junctions either between the blastomeres within the tier or between the blastomeres from adjacent tiers. All gap junctions observed in freeze fracture replicas show plaques with an irregular IMP pattern. The average IMP diameter measures 12 nm (SD±2 nm). In stages fixed after the fifth cleavage, gap junctions are found between micromeres at the animal pole and the central 3D macromere. This is in agreement with the presumed interaction between these cells at this stage. The possibility of a transition of non-functional into functional gap junctions after the fifth cleavage is discussed.     

Cells and intercellular contacts in glomeruli and tubules of the frog kidney

Summary By the use of thin sections and freeze-fracture replicas the glomerular and tubular structures of the kidney of the frog (Rana esculenta) were studied with special reference to intercellular junctions.In the glomerulus the filtration barrier is of very variable thickness, and frequent tight and gap junctional contacts occur between podocyte processes.Although structurally less elaborate, the proximal tubule resembles its mammalian counterpart. In the initial part the tight junctions are relatively shallow but become very broad in the mid and distal portions of the proximal tubule. The proximal tubular cells are extensively linked by gap junctions. In some animals the shapes of the cells in the proximal and distal portions of the proximal tubule were markedly different.The distal tubule consists of two segments which differ mainly in the pattern of interdigitations and the structure of the zonulae occludentes. Similarities with the tight junctional morphology of the mammalian distal tubule are striking. In the first part of the distal tubule (diluting segment) a narrow band of parallel tight junctions is found closely resembling that found in the mammalian straight distal tubule; in the more distal part of the distal tubule, however, a broad band of anastomosing tight junctional strands exists, like the zonula occludens of the mammalian convoluted distal tubule.The connecting tubule displays cellular dimorphism: its wall contains a mixture of light and dark (flask) cells. The luminal and basolateral membranes of the flask cells are covered with numerous rod-shaped particles. The tight junctions of the connecting tubule are broad and increase in depth and number of strands along its length; they are typical of a very tight epithelium.In spite of several dissimilarities with phylogenetically younger kidneys our findings suggest that many structural principles of the mammalian kidney are also represented in the kidneys of amphibians. The structural-functional relationships are discussed.     

A novel type of cell-cell cooperation between epithelial cells

Ma104 cells (renal, epithelial) have a peculiar way of resisting ouabain: their Na⁺,K⁺-pumps bind the drug with high affinity, cellular K⁺ is lost and cell division arrested, but cells do not detach as most cell types do. Then, if up to 4 days later the drug is removed, Ma104 cells recover K⁺ and resume proliferation (Contreras et al., 1994). In the present work, we investigate whether Ma104 cells are able to protect ouabain-sensitive MDCK cells in co-culture. The main finding is that they do, but in this case protection is not elicited by the usual mechanism of maintaining the K⁺ content of neighboring cells through cell-cell communications. Ma104 cells treated with ouabain simply remain attached to the substrate and to their MDCK neighbors, and both cells lose K⁺. This attachment includes tight junctions, because the transepithelial electrical resistance of the monolayers is not abolished by ouabain. Although the -subunit of the Na⁺,K⁺-ATPase is known to possess molecular characteristics of cell-cell attachment molecules, attachment between Ma104-MDCK cells does not seem to be mediated by this enzyme, as immunofluorescence analysis reveals that Na⁺,K⁺-ATPase is only inserted in the plasma membrane facing a neighboring cell of the same type.We wish to thank Dr. Enrique Rodríguez-Boulan (Cornell University Medical College) for the generous supply of Ma104 cells, as well as the generous economic support of COSBEL, SA de CV and the CONACYT of Mexico and the National Institutes of Health. Confocal experiments were performed in the Physiology Department's confocal microscopy unit, CINVESTAV.     

Serum Opens Tight Junctions and Reduces ZO-1 Protein in Retinal Epithelial Cells

Abstract: We have shown previously that serum inhibits tight junction formation in a retinal epithelial cell culture model for the blood-brain barrier. We have now examined in detail the effects of serum on the tight junctions. Our data show that serum induces a breakdown in tight junction function as indicated by decreased transepithelial electrical resistance and increased permeability. Rat serum had effects similar to those of bovine serum, indicating that the activity is species-independent. The effect is concentration-dependent, reversible, and specific for the apical surface, suggesting the involvement of a specific receptor-ligand interaction. Differences in the time course, response magnitude, and structural manifestations between the serum-induced breakdown and that induced by switching the cultures to a low-calcium medium suggest fundamental differences in their mechanisms. The calcium switch results in an immediate and complete junctional breakdown with cell retraction and perinuclear translocation of both actin and the tight junction protein zonula occludens-1. The serum-induced breakdown occurs slowly, is incomplete, and is manifested structurally by decreases in zonula occludens-1 protein, whereas actin organization is unchanged. Thus, serum induces a specific breakdown in retinal epithelial cell tight junctions that may be mediated by effects on the expression of zonula occludens-1.