Stress fibers in the mesenteric mesothelial cells of the large intestine of the bullfrog,Rana catesbeiana

Summary Actin-containing cytoplasmic fibers were visualized in the mesenteric mesothelial cells of the large intestine of bullfrog tadpoles by rhodamine-phalloidin staining of en face preparations of mesothelial cells. These fibers were concurrently stained by immunofluorescence using antibodies to myosin or -actinin. Electron microscopy showed the presence of bundles of microfilaments in the basal cytoplasm of the cells. Such fibers in the mesothelial cells may be comparable to the stress fibers present in cultured cells. The mesothelial cells initially formed axially oriented stress fibers when they changed from a rhombic to a slender spindle-like shape. On the other hand, stress fibers disappeared as cells transformed from elongated to polygonal shapes during the period of metamorphic climax. Expression of stress fibers in these cells appears to be related to the degree of tension loaded on the mesentery, which may be generated by mesenteric winding. These stress fibers in the mesothelial cells may serve to regulate cellular transformation. They may also help to maintain cellular integrity by strengthening the cellular attachment to subepithelial tissue against tensile stress exerted on the mesentery.     

Effects of lead on hepatocyte ultrastructure in Carassius carassius (L.) var. auratus

Subcellular modifications in hepatocytes of Carassius carassius var. auratus subjected to 24 hr and 48 hr sublethal acute lead (5mg·1⁻¹) exposure were studied by electron microscopy. Cytological alterations were observed after 24 hr of treatment and became more evident after 48 hr. Lead induced an increase in nuclear heterochromatin and alterations in mitochondria, endoplasmic reticulum and Golgi complex ultrastructure. Glycogen granula decreased, and secondary lysosomes and lipid droplets increased. Furthermore, intracytoplasmic lumina with microvilli-bearing surfaces and numerous autophagic vacuola were observed after 48 hr of exposure.     

Digestion of prey in foraminifera is not anomalous: A correlation of light microscopic, cytochemical, and hvem technics to study phagotrophy in two allogromiids

Correlative light, high-voltage electron and conventional electron microscopic methods were used to investigate digestion in two allogromiid foraminiferans, Allogromia sp., strain NF, and A. laticollaris Arnold. Microscopic observations showed that bacterial prey are phagocytosed by reticulopodia and are transported to the allogromiid cell body within blister-like phagosomes. Larger prey (algae, diatoms) are transported along the reticulopodial surface and are either stored extrathalamously or phagocytosed at the oral opening (peduncle). Studies of allogromiids optimally fixed and labeled with an extracellular-space label (colloidal thorium) showed that phagocytosed prey are completely enclosed by a plasma membrane envelope; this finding was corroborated by a serial-section three-dimensional reconstruction of the oral zone of one allogromiid. Cytochemical staining for acid phosphatase showed that lysosomes are absent from reticulopods but abundant in the cell body, particularly in the oral zone cytoplasm. We conclude that digestion in allogromiid foraminiferans is accomplished by a vacuole-based digestive apparatus and not by extracellular digestion within a lacunary system, as has been suggested in earlier studies.     

Morphohistochemical changes in hepatocytes during the life cycle of the European eel

The comparative analysis of morphological, histochemical and cytochemical patterns of eel (Anguilla anguilla L.) hepatocytes reveals clear differences between two stages of its life cycle, i.e. the trophic stage (yellow eel) and reproductive stage (silver eel). The storage of glycogen prevails in the yellow eel, whilst lipids appear to be remarkably increased in the silver eel, in which some hepatocytes also show glycogen-rich areas. Generally, in the silver eel dehydrogenase and acid phosphatase activities seem greater and different distribution of the reaction products is present; on the contrary, a lower G6PDH activity is observed. The electron microscopy characteristics and distribution of both cellular organelles and reserve materials reflect the modifications found at light microscopy. The ultrastructural patterns provide further evidence for the heterogeneity of liver parenchyma in silver eel. In particular, the coexistence of nuclei showing a different degree of chromatin compactness is also accounted for by the quantitative cytochemical data on the nuclear DNA after Feulgen reaction and intercalation with propidium iodide at low and high concentrations. With regard to the DNA content, the hepatocytes in the silver eel as well as in the yellow eel are mainly 2c. However, some 4c values are also found, which according to the literature can be ascribed to cells in G2 phase. The present data may express the onset of different functional requirements during the reproductive stage in comparison with the trophic one. Moreover, our results are consistent with modifications found by other authors as a consequence of interruption of nourishment and during gonad maturation, i.e. two phenomena characterizing the transition from yellow to silver eel.     

Lectin cytochemical evaluation of somatosensory neurons and their peripheral and central processes in rat and man

Summary Paraffin sections of the trigeminal nerve root of the rat, and human spinal nerve root and trigeminal ganglion were stained with a battery of lectin-horseradish peroxidase conjugates to localize and characterize glycoconjugate (GC) in situ. In the rat the myelin sheath of the peripheral segment contained GC with sialic acid most probably linked to the penultinate disaccharide galactose(1 4)-N-acetylglucosamine (Gal(1 )-GlcNAc), and complex type N-glycosidic side chains. The myelin sheath in the central segment differed in containing little if any of the GC named above and in containing GC with terminal -Gal linked to N-acetylgalactosamine (GalNAc), terminal GalNAc and fucose. Schwann cells stained for GC with GlcNAc or mannose whereas oligodendroglia stained for GC with the terminal disaccharide Gal-(1 3)-GalNAc and N-glycosidic side chains, especially in presumed Golgi zones, but also in processes continued as the outer myelin sheath. The human myelin sheath in the central segment differed from that of the rat in not staining with lectins specific for fucose and terminal GalNAc. Sialic acid and terminal -Gal were seen in the human central segment but these sugars appeared to bind to astroglial structures rather than to the myelin sheath as in the rat. Astrocytes in both rat and man were stained by two fucose-binding lectins. Several lectins revealed affinity for GC in the neurilemmal sheath, and staining of this structure was stronger in the human specimens. Neurons in the human trigeminal ganglion ranged from unstained to strongly positive for fucoconjugate in cytoplasmic bodies and plasmalemma. Positive ganglion cells gave rise to unmyelinated fibers which also stained for fucoconjugate. Remak fibers and their extensions into the substantia gelatinosa of the human spinal cord stained strongly for content of fucose.The stronger lectin affinity for N-glycosidic core sugars in the peripheral as compared with the central segment suggests that lectins localize P_o protein in peripheral myelin. The reactivity for several sugars in the central segment can possibly be attributed to myelin-associated glycoprotein (MAG) of central myelin, but lectin staining for GalNAc shows in addition a biochemically unrecognized GC with O-glycosidic linked oligosaccharides in myelin. The lectin cytochemistry indicates that the 170 K Dalton glycoprotein with PNA affinity obtained from rat sciatic nerves occurs in nodes of Ranvier.This research was supported by NIH Grants AM-10956, HL-29775 and United Health and Medical Research Foundation of South Carolina, Inc. Grant No. 79     

On catecholamine-containing cells in the rat interatrial septum

Summary The interatrial septum of the rat heart contains cells which show a strong intensive-yellow paraformaldehyde-induced fluorescence. By electron microscopy these cells are characterized by an abundance of dense-core vesicles.Cholinergio axons form axo-somatic synaptic contacts with the catecholamine-containing cells. These cells, packed with dense-core vesicles, are frequently interdigitated and interconnected by zonulae and maculae adhaerentes and occludentes. The catecholamine-containing cells are surrounded by satellite cells either individually or in groups.The catecholamine-containing cells, which bear blunt, plumpish processes, can be subdivided, on the basis of position and morphology into two types. One class of cells lies within the fibroblast capsule of the intra-atrial ganglion (van der Zypen, Hasselhorst, Merz and Fillinger, 1974). A second aggregation of catecholamine-containing cells occurs outside the ganglia in close proximity to capillaries. The capillaries exhibit pores in the area of contact with the catecholaminergic cells. The structure of these catecholamine-containing cells is described and their possible function discussed.

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Zusammenfassung Im Septum interatriale des Rattenherzens treten Zellen in Erscheinung, die nach Paraformaldehyd-Bedampfung eine intensive hellgelbliche Fluoreszenz zeigen. Diese Zellen zeichnen sich durch einen großen Reichtum an dense-core vesicles aus. Cholinerge Axone bilden axo-somatische Synapsen an den katecholaminhaltigen Zellen aus. Die mit dense-core vesicles angefüllten Zellen sind oft ineinander verzahnt und durch Zonulae adhaerentes verbunden. Einzeln oder in Gruppen werden die katecholamin-enthaltenden Zellen von Satelliten-Zellen umgeben.Die mit kurzen plumpen Fortsätzen versehenen katecholaminhaltigen Zellen lassen aufgrund ihrer Lage und eines andersartigen Baues zwei Typen erkennen. Eine Gruppe von Zellen liegt innerhalb der Fibrozytenkapsel des Ganglion intraatriale (van der Zypen, Hasselhorst, Merz und Fillinger, 1974). Eine zweite Ansammlung von Katecholamin enthaltenden Zellen findet sich außerhalb der Ganglien in engem Kontakt zu Kapillaren. Die Kapillaren weisen im Bereich des Kontaktes mit den katecholaminergen Zellen Poren auf. Die Struktur dieser Zellen wird geschildert und ihre mögliche Funktion diskutiert.

     

Sites of Tubulin Polymerization in PC 12 Cells

The site at which tubulin enters into polymer in the neuritic process is a very important datum in terms of our understanding of the mechanism of transport of the microtubular cytoskeleton out the axon. If the form of tubulin being transported out the axon is the microtubule, then assembly of tubulin into microtubules should occur at or near the cell body; if, however, the form of tubulin transported is free tubulin dimer, then assembly can occur at any free microtubule end out the neurite. We have injected a fluorescent analog of tubulin into differentiated PC 12 cells and used differential extraction protocols to extract free dimer but not microtubules. We have imaged these cells before and after extraction by low-light-level video fluorescence microscopy and have used image analysis to examine the sites of tubulin incorporation into polymer or other unextracted components as a function of time. We find that tubulin in the distal reaches of the neurite is found initially as monomer and that its appearance in the unextracted component occurs later. This pattern of appearance of fluorescent tubulin initially in the soluble fraction and later in the unextractable component is qualitatively similar to that reported by other workers for biotinylated tubulin, but we see a larger gap between the rates of appearance in soluble fraction and in polymer. Quantitative analysis of fluorescence intensities in the two compartments with distance out the neurite reveals substantial variation between different neurites: In some neurites, the pattern of variation of unextracted/total tubulin suggests that tubulin enters into the unextracted component primarily near the cell body and that this unextracted component moves out the neurite with time, and in other neurites it suggest that monomer adds into microtubule ends staggered out the neurite. In no case do we see a pattern suggesting that distal addition predominates. These analyses of fluorescence intensities in extracted and unextracted neurites suggest that both transport of polymerized microtubules and monomer addition onto staggered microtubule ends occur in PC12 neurites and that in individual neurites one or the other of these two behaviors may predominate.     

Cellulose and xylans in the interface capsule in symbiotic cells of actinorhizae

Summary Enzyme-gold affinity labeling was used to show that in mature infected cells of actinorhizal symbioses the capsule on the plant host side of the symbiotic interface contained cellulose and xylans. Host species examined for cellulose wereAlnus rubra, Casuarina equisetifolia, C. glauca, Ceanothus cuneata, C. velutinus, Elaeagnus pungens, andMyrica cerifera.. Cellulose was in the capsule throughout the infected cell, implying that during development cellulose synthase was present in the host cell membrane component of the symbiotic interface. Any possible degradation of capsule cellulose by the microsymbiont was either incomplete or transient, because the polymer was present in mature infected cells. Cellulose labeling inCeanothus andElaeagnus was less consistent than in the other species. Dual labeled capsules inCasuarina glauca andAlnus rubra showed a similar distribution of xylans and cellulose. Cytochemical studies indicate that the capsule contains three major classes of cell wall polysaccharides: cellulose, hemicellulose (xylans), and pectins (shown previously). This suggests that the capsule is essentially a thin, internal, tubular plant cell wall.Abbreviations Au₅ Au₁₅ colloidal gold particles with mean diameter of 5 and 15 nm, respectively - CBHI cellobiohydrolase I - CBHII cellobiohydrolase II - PBS phosphate-buffered saline     

New cellulose synthesizing complexes (terminal complexes) involved in animal cellulose biosynthesis in the tunicateMetandrocarpa uedai

Summary Subcellular compartments comprising the endomembrane system in filamentous fungi are poorly characterized with most showing significant morphological differences from eukaryotic cells. For example, many filamentous fungi lack stacked Golgi-body cisternae, but contain Golgi equivalents — single cisternae or tubules which appear to serve the same functions. To help identify fungal endomembrane compartments and interrelationships between them we used a pharmacological agent, brefeldin A, known to affect specific endomembrane organelles in other organisms, most prominently the Golgi apparatus. At 10 g/ml brefeldin A, radial hyphal growth of the rice blast pathogenMagnaporthe grisea on solid agar medium was reduced by 96% over an initial 48 h, but recovered and was reduced by only 20% over a subsequent 72 h exposure. Light microscopic examination of individual living hyphae showed that apical elongation generally halted within 1 min after exposure to brefeldin A. Acute effects of 14 g/ml brefeldin A were characterized ultrasiructurally in cells prepared by freeze substitution. These included the appearance of two types of cisternae with unusual morphology, associated with ca. 45 nm diameter vesicles, as well as the unexpected persistence and increase in complexity of the Golgi equivalents. Also observed were (1) reduced numbers of apicale vesicles and disruption of Spitzenkörper organization, (2) apical clusters of 30–35 nm diameter microvesicles and associated tubular arrays, (3) dilation of rough endoplasmic reticulum, (4) packets of membrane-bounded electron-opaque cell wall inclusions, and (5) altered morphology of some vacuolar compartments. The distribution of concanavalin A binding sites, previously mapped to particular endomembrane compartments, was documented to aid the interpretation of these results. We conclude that brefeldin A effects on cells ofM. grisea differ from those reported with plant and animal cells, perhaps reflecting underlying differences in the endomembrane systems among these eukaryotes.Abbreviations BFA brefeldin A - ConA concanavalin A - ER endoplasmic reticulum - PDA potato dextrose agar - RER rough endoplasmic reticulum