Microtubules in myelinated and unmyelinated axons of rat sciatic nerve

Summary Tannic acid in glutaraldehyde was used to stain microtubules in myelinated and unmyelinated axons of rat sciatic nerve. In the majority of areas the tannic acid failed to penetrate the unmyelinated axons whilst penetrating neighbouring myelinated axons, suggesting a difference in the ability of the two types of nerves to exclude tannic acid. Where tannic acid had penetrated the unmyelinated axons the 13 protofilament substructure and size of the microtubules appeared identical to those seen in the myelinated axons.     

Demonstration of microtubules in the terminal web of mature absorptive cells from the small intestine of the rat

Summary The terminal web (TW) region of mature absorptive cells in the small intestine of the rat contains an elaborate cytoskeleton which supports the apical microvillus membrane. In studies regarding the structural organization of the cytoskeleton and associated proteins in the small intestine, microtubules have not been mentioned as components of the TW. By transmission electron microscopy of conventional resin-embedded sections of rat small intestine, we observe many microtubule profiles in the TW of mature absorptive cells. These microtubules are found in various orientations, although most course parallel to the long axis of the cell, and many microtubule profiles are seen in close association with smooth-surfaced vesicles.     

Linkage of manchette microtubules to the nuclear envelope and observations of the role of the manchette in nuclear shaping during spermiogenesis in rodents.

Structural features of the mouse and rat manchette and the role of the manchette in shaping the spermatid nucleus were investigated. Rod-like elements about 10 nm in diameter and 40-70 nm in length were seen linking the innermost microtubules of the manchette and the outer leaflet of the nuclear envelope in step 8 through step 11 rat and mouse spermatids that either had been routinely fixed for electron microscopy or had been isolated and detergent extracted. Rod-like linkers were also seen joining the nuclear ring to the plasma membrane and nuclear envelope. These linkers may ensure that under normal conditions the manchette remains in a defined position relative to these membranous components. A variety of compounds (taxol, cytoxan, and 5-fluorouracil) were found to perturb the manchette and to affect nuclear shaping. In addition, sys and azh mutant mice were used to determine the consequences of defective manchette formation. These genetic conditions and chemical treatments either produced manchettes that were not in their normal position (azh, sys, and taxol) and/or caused the manchette to appear abnormal (azh, sys, cytoxan, 5-fluorouracil, and taxol), and all resulted in a deformation of the step 9-11 spermatid nucleus. In all instances where the manchette was present, either in normal or ectopic locations, the sectioned nuclear envelope was parallel to the long axis of the microtubules of the manchette. In general, areas of the nuclear envelope where the manchette was not present, or where it was expected to be present but was not, were rounded (normal animals, sys, cytoxan). In addition, there are indications using certain compounds (cytoxan and 5-fluorouracil) as well as in the azh and sys mouse that the manchette may exert pressure to deform the nucleus. It is suggested that the rod-like linkages of the manchette ensure that the nuclear envelope remains at a constant distance from the manchette microtubules and that this is a major factor acting to impart nuclear shape changes on a region of the head caudal to the acrosome during the early elongation phase of spermiogenesis. The manchette microtubules, which are also known to be linked together, may act as a scaffold to deform this part of the nucleus from its spherical shape, perhaps in concert with forces initiated by other structural elements. Evidence from sys animals indicates that structural elements, such as the acrosomal complex over the anterior head (acrosome-actin-nuclear envelope), may affect nuclear shaping over the acrosome-covered portion of the spermatid head.(ABSTRACT TRUNCATED AT 400 WORDS)     

Effects of antimicrotubular agents on the fine structure of the Golgi complex in embryonic chick osteoblasts

Embryonic chick frontal bones were cultured in the presence of colchicine or vinblastine and subsequently examined by tranmission electron microscopy. In control cultures the osteoblasts showed a large Golgi complex consisting of dictyosomes arranged in a well-defined juxtanuclear area. Microtubules were particularly numerous within this Golgi area although they could be observed throughout the cytoplasm. Colchicine and vinblastine caused the disappearance of cytoplasmic microtubules, while bundles of 10 nm diameter filaments appeared more frequently. In addition, cell polarity was lost and the Golgi complex became disorganized, with the dictyosomes randomly dispersed in the cytoplasm and showing a decreased number of cisternae and an increased number of vacuoles, the latter generally lacking stainable material. Increased number of autophagosomes were also noted. These findings indicate that microtubules function in the organization of the Golgi complex in osteoblasts. In view of the well documented role of this organelle system in collagen secretion it is suggested that previously observed secretory disturbances produced by antimicrotubular drugs may be due to a defective transfer of material to the dictyosomes and/or a defect in the packaging and transport of such material away from them.     

Porphyrins affect the self-assembly of tubulin in solution

Self-assembly of tubulin heterodimers in solution has been studied in the past to predict the effects that ligands and/or conformational changes have on the formation of tubulin filaments. Self-assembly of tubulin in solution has produced formations similar to cellular microtubules (MTs). The present study reports on the effects that two porphyrins (protoporphyrin IX, PPIX and tetrakis(4-sulfonatophenyl)porphyrin, TPPS) produce on the self-assembly of tubulin α,β-heterodimers in buffer solution. The study shows that, when incubated simultaneously with MT-stabilizing ligands (i.e., paclitaxel and guanosine triphosphate, GTP), porphyrins do not affect the ability of tubulin to form MT. However, if paclitaxel and GTP are added after tubulin has been allowed to self-assemble in the presence of either porphyrin, the ability to form MT-like structures is reduced or suppressed. We suggest that this effect is due to the formation of porphyrin-mediated aggregates that cannot be broken or elongated by the addition of GTP or paclitaxel.     

Final stages of erythrophagocytosis in the sheep placenta

Summary In trophoblastic epithelial cells of the sheep placenta the final stages of erythrocyte breakdown within the lysosomal apparatus were studied at the ultrastructural level.As a result of hemoglobin digestion lysosomes containing hemoglobin-derived pigments (HDP) were formed. The HDP-lysosomes were acid phosphatase-positive, highly electron-dense bodies of round to irregular shape containing whorled membranous formations. The accumulation of these lysosomes in epithelial cells led to fusion resulting in the formation of conglomerates. At the end of the gestation period the amount of HD Plysosomes and their conglomerates markedly increased.In addition to erythrocytes the trophoblastic epithelial cells in the erythrophagocytic regions phagocytosed maternal leukocytes and neighbouring epithelial cells and giant cells.By gradual accumulation of HDP-lysosomes and remnants of phagocytosed cells, highly electron-dense acid phosphatase-positive residual bodies of variable appearance were formed within the epithelial cells.At the end of pregnancy the spaces between juxtaposed villi of the trophoblastic epithelium in the erythrophagocytic zones were occluded by apposition of the epithelial cells. In these occluded regions an increase in highly electron-dense large-sized residual bodies (15–22 m of dimension) occurred as a result of multiple cell phagocytosis in combination with fusion. In these residual bodies the numerous incorporated HDP-lysosomes and the remnants of phagocytosed cells could still be recognized.     

Ultrastructural studies on the secretory process in the epithelioid cells of the juxtaglomerular apparatus

The epithelioid cells of the juxtaglomerular apparatus have been studied with respect to the release mechanism of the secretory granules. Invaginations of the plasma membrane into the interior of the epithelioid cells are interpreted as stages before or after an exocytotic process. Granules are sometimes observed in close contact with the plasma membrane, and material with electron density similar to that of the granules can also be observed in the invaginations. These morphological features suggest that the granular material of the epithelioid cells is extruded into the texture of the basal lamina. Furthermore, a dense network of microtubules and microfilaments is described and the functional role of this system in exocytosis is discussed.     

Development ot the granule population in heterophil granulocytes from rat bone marrow

Summary The development of the heterophil granulocyte in the bone marrow of the rat is described, and an electron-microscopical analysis of the changes in the cytoplasm as well as in the granule population in several stages of maturation is reported. Three types of granule originate in consecutive stages of heterophil maturation. Granules with an internal fine structure (nucleated granules) are the first to be formed, i.e., in early promyelocytes; azurophil granules are formed in late promyelocytes; and specific granules appear in myelocytes. Quantitative analysis showed that the granule population in mature cells, i.e., about 160 granules per electron micrograph, is composed of roughly 14% nucleated granules, 10% azurophil granules, and 76% specific granules. Three cell stages were observed in mitosis: the early promyelocyte, the late promyelocyte, and the myelocyte. Granule counts in non-dividing cells confirmed the occurrence of mitosis in the late promyelocyte and myelocyte.     

Ultrastructural localization of radiolabelled L-dopa in the endocrine hypothalamus of the rat

Summary Light and electron microscopic autoradiography has been employed to define the neuroanatomical patterns of uptake and binding of radiolabelled L-dopa in the endocrine hypothalamus of the rat. A dorsomedial continuum of arcuate and periventricular neurons selectively sequester ³H L-dopa 20 min following its intraventricular infusion. By 40 and 60 min following the infusion labelling of neurons is minimal and supports the notion of rapid degradation. Other cell compartments such as tanycytes demonstrate uptake of ³H L-dopa. The ultrastructural localization and distribution of radiolabelled L-dopa (or its metabolites) in the rodent hypothalamus is discussed with respect to mechanisms and cell compartments involved in neuroendocrine regulatory processes.Supported by USPHS Program Project Grant NS-11642-04 (DES) and RR-05403Career Development Awardee RO4GM-70001     

An ultrastructural and autoradiographic study of the immune response in Hyalophora cecropia pupae

Summary Membrane-bounded spherical vesicles found in rat Sertoli cells have been examined quantitatively during the cycle of the seminiferous epithelium. Most of the vesicles were localized to the basal and columnar portions of the Sertoli cell cytoplasm. The thin lateral projections of the Sertoli cells contained very few vesicles. Morphometric analysis of the basal portion of the Sertoli cell cytoplasm revealed that the volume density (V _v ) of the vesicles changed markedly during the cycle. The V _v was at its minimum (0.036) at stage VII and maximum (0.117) at stages XI-I. The vesicles were also smaller at stage VII compared to the vesicles at stages IX-V. The stage-dependent difference in the size of the vesicles was found both in the basal and the columnar portions of the Sertoli cells. At stage VII some of the vesicles appeared to be elongated much like the tubular elements of the smooth endoplasmic reticulum (SER) from which they are probably derived. The stage-dependent differences in volume density and size of the Sertoli cell vesicles may be related to cyclic biochemical variations in the Sertoli cells, and are further indications of a variation in Sertoli cell function during the cycle of the seminiferous epithelium. Whether or not this is due to an internal cycle of the Sertoli cell or to influences from adjacent germ cells remains to be determined.     

Organisation of microtubules and actin filaments in the cortex of differentiating Selaginella guard cells

Summary Using fluorescent probes and confocal laser scanning microscopy we have examined the organisation of the microtubule and actin components of the cytoskeleton in kidney-shaped guard cells of six species of Selaginella. The stomata of Selaginella exhibit novel cytoskeletal arrangements, and at different developmental stages, display similarities in microtubule organisation to the two major types of stomata: grass (dumbbell-shaped) and non-grass (kidney-shaped). Initially, cortical microtubules and F-actin radiate from the stomatal pore and extend across the external and internal periclinal cell surfaces of the guard cells. As the stomata differentiate, the cytoskeleton reorients only along the internal periclinal walls. Reorganisation is synchronous in guard cells of the same stoma. Microtubules on the inner periclinal walls of the guard cells now emanate from areas of the ventral wall on either side of the pore and form concentric circles around the pore. The rearrangement of F-actin is similar to that of microtubules although F-actin is less well organised. Radial arrays of both microtubules and F-actin are maintained adjacent to the external surfaces. Subsequently, in two of the six species of Selaginella examined, microtubules on both the internal and external walls become oriented longitudinally and exhibit no association with the ventral wall. In the other four species, microtubules adjacent to the internal walls revert to the initial radial alignment. These findings may have implications in the development and evolution of the stomatal complex.Abbreviations GC guard cell - MT microtubule     

Fibroblasts as a part of the contractile system in duodenal villi of rat

Summary The stroma of duodenal villi of rats was studied by light- and electron microscopy. Fibroblasts are rather evenly distributed within the villus. Their branched processes embrace all blood vessels, the lacteal and the bundles of smooth muscle cells. They are connected to each other and to smooth muscle cells by close contacts. Unmyelinated axons are found close to the fibroblasts where they may show synapse-like formations.The fibroblasts within intestinal villi contain many dilated cisterns of rER similar to normal fibroblasts. In contrast to the latter, there are many aggregated, contractile filaments, being situated mainly below the plasma membrane and within the processes. It is suggested that fibroblasts representing a 3-dimensional contractile network may be activated by smooth muscle cells and/or by innervation. So, they seem to be involved in the diminution of the vascular and stromal spaces within the villus.     

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.

     

Über den Peroxydasetransport im Darmepithel der Ratte

Zusammenfassung Bei Ratten tritt 3 min nach intravenöser Injektion von Peroxydase elektronenmikroskopisch ein entsprechendes Reaktionsprodukt im Kapillarlumen der Lamina propria des Dünndarms und an der Basalmembrangrenze der Saumepithelzellen auf. 5 min nach der Injektion finden sich im basalen Abschnitt des Darmepithels pinozytotische Bläschen mit dem Peroxydase-Reaktionsprodukt. — 10–30 min nach der Injektion erreichen die Partikel die apikalen Teile der Zelle. Sie dringen in den interzellulären Spalten bis zu den Haftplatten vor, erreichen jedoch nie das Darmlumen. Im Dünndarm existiert vermutlich auch ein der Resorption entgegengesetzter Saftstrom, der durch Peroxydase markiert werden kann.

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The transport of horseradish peroxidase in the epithelium of the small intestine

Summary In rats, 3 minutes after intravenous injection of peroxidase the reaction product can be observed electronmicroscopically in the lumina of the capillaries of the small intestine as well as at the border of the basement membrane of the epithelial border cells. Pinocytotic vesicles containing peroxidase particles occur in the basal portion of the epithelium of the small intestine 5 minutes after injection. 10–30 minutes later, the peroxidase reaches the apical region of the cell. The particles infiltrate into the intercellular spaces as far as the tight junctions but never reach the intestinal lumen. In the small intestine there probably exists a flow of fluid in opposite direction to the resorption, which can be marked by peroxidase.

     

Effect of histone on the nucleolar morphology of cells cultured in vitro

Summary The frequency of various types of nucleoli was investigated in tissue cultures of human embryonal lung and HeLa cells cultured in the presence of calf thymus histone. The nucleolar morphology and the frequency of various nucleolar types were dependent on the concentration of histone in the tissue cultures of the human embryonal lung cells. HeLa cells required longer cultivation with histone to manifest some effect on nucleoli. In both cases, the observed nucleolar changes suggest the depression of nucleolar RNA synthesis.     

Carbendazim-induced abnormal development of the acrosome during early phases of spermiogenesis in the rat testis

Effects of a single, high dose of orally administered carbendazim (100 mg/kg) on acrosome formation in the early phases of spermiogenesis were examined by electron microscopy and immunocytochemistry up to day 7.5 post-treatment. No obvious abnormality of acrosome development was noted in the Golgi phase spermatids on day 1.5 post-treatment. On day 3, step 1 spermatids were seen in stage III seminiferous tubules. In stage V tubules at this post-treatment interval, direct connections between the trans-side saccules of the Golgi stacks and the outer acrosomic membranes were observed in step 5 spermatids. Similar direct connections between these two organelles were also observed in the advanced round spermatids in later stages at days 4.5 and 7.5. On day 4.5, step 1 and 3 spermatids were seen in stage V tubules. On day 7.5, round spermatids with various abnormalities of acrosome development were observed in stage VII tubules, in addition to the discontinuous and granular acrosomes reported previously. These features were not observed in testes of control animals. In the immunocytochemical analysis using an antibody mMN7 that recognizes a protein delivered from the Golgi apparatus to the acrosome, spermatids exposed to carbendazim showed various abnormal immunostaining patterns in the acrosomes. On the other hand, strong immunoreactivity was observed in the Golgi saccules connecting to the acrosomes. These results suggest that in testis treated with carbendazim acrosome development is impaired during the early phases of spermiogenesis, and material supply from the Golgi apparatus to the acrosome is perturbed, which is a possible cause of the abnormal development. Received: 31 March 1998 / Accepted: 28 May 1998     

Plasma-membrane glycoproteins during spermiogenesis and in the spermatozoa of some Orthoptera

Summary Orthoptera spermatids and spermatozoa from two species of Tettigoniidae and from one of Acrididae were analysed by means of the fluorescent lectins, concanavalin A or wheat-germ agglutinin, with the aim of finding -D mannose· -D glucose· N-acetylglucosamine and sialic acid sugar residues in their plasmamembrane glycoproteins. Labelling with lectins shows remarkable changes occurring in the plasma membrane during spermiogenesis. In early spermatids, the whole cell surface is labelled, but in mature spermatids and spermatozoa, a noticeable fluorescence is restricted to the membrane that covers the acrosome. The end piece of the tail of Acrididae spermatids and spermatozoa fluoresces after wheat-germ agglutinin labelling. The intense labelling of acrosomal area is independent of acrosomal size and shape, as shown by the marked differences observed in the acrosomes of Tettigoniidae compared with Acrididae: in the former, the acrosome is a well-developed structure with an arrow-like shape, but in Acrididae, the acrosome resembles a small vesicle in the anterior tip of the cell. The large amount of some sugar residues in the plasma membrane covering the acrosome is discussed in relation to the features observed in other species, and also in connection with the physiology of the male gamete prior or during fertilization.     

Actin filaments connected with the microtubules of lipotubuloids, cytoplasmic domains rich in lipid bodies and microtubules

Summary. Lipotubuloids, i.e., cytoplasmic domains containing an agglomeration of lipid bodies surrounded by half-unit membrane, entwined and held together by a system of microtubules, have been found in the ovary epidermis of Ornithogalum umbellatum. Ultrastructural studies demonstrated thin filaments in lipotubuloids that are probably actin filaments arranged parallel to microtubules. It is suggested that interaction of actin filaments with the microtubules determines the driving force for the rotary motion characteristic of lipotubuloids, as this movement is sensitive to cytochalasin B. Correspondence: Department of Cytophysiology, University of Łódź, Pilarskiego 14, 90-231 Łódź, Poland.