bluuebunny/crossref_metadata_2025 huggingface dataset:
A new insect virus?
Authors: Letendre Michel , Lu Jean , Beaulieu Jacques , Bédard Guy
Published: June 2020

A new virus—like particle has been observed in haemolymph from Pseudaletia unipuncta (Haw. )

A high degree of mortality was noticed on an epidemic population of the true army worm, Pseudaletia unipuncta (Haw. ) in La Sarre, Abitibi, Quebec.

In a first approach, a few specimens of larvae were collected. Haemolymph was extracted for a preliminary observation under the electron microscope. The preparation technique was very simple. A 400 mesh formvar grid was put on top of a drop of haemolymph for a few seconds and was negatively stained with PTA 2% pH 7. 00.

A huge number of spherical particles ( a count of nearly 2, 000 on one grid quadrat) were observed in this way.

Conscious of our inexperience with insect viruses we consulted three specialists in comparative virology:
Spatial structure of the tails of some bacterial viruses
Authors: Vainshtein B. K. , Mikhailov A. M.
Published: June 2020

Phages are complex viruses having a special apparatus for injection of genetic material into the cell. The phage has a head representing an RNA-containing protein coat, and a tail with a baseplate and filamants. Bodies in which the packing of asymmetric units (as. u. ) is described by helical symmetry group can be represented as a pile of elementary disks (the height of a disk is c/p) rotated with respect to each other at an angle of 2πq/p where p is the number of as. u. and q- the number of turns of the continuous helix that fall into the translation period c. On the micrograph the helical object is presented by pN/2 (at N -the order of the rotation axis- even) or pH (at N odd) projections of an elementary disk. The availability of these projections makes it possible to restore from one micrograph of an object its three dimensional structure at a resolution defined by the number of projections, resolution of micrographs or calculation methods.
Lomasome biogenesis and function inarmillaria mellea
Authors: Behboodi B. Ch.
Published: June 2020

Introduction

Border bodies or lomasomes are the aggregation of membranes and vesicles located between the plasma membrane and the cell wall of many fungi, algae, and higher plants. Despite extensive investigation, the biogenesis as well as function of these structures is not yet known. The purpose of this investigation was to describe the biogenesis of lomasomes in Armillaria mellea and to provide some observations on their function related to cell wall formation.

Materials and Methods

Various thalli of fungi as non-aggregated hyphae, pseudosclerotes, rhizomorphs and carpophores were grown either on orange or synthetic media as described previously. The thalli were fixed in 4% glutaraldehyde buffered with 0.1 M sodium cacodylate (pH 7.4), and 0.15 M sucrose for 4 h at 4°. They were postfixed with 1% osmium tetroxide in the same buffer for 2 h at 4° and embedded in Epon according to the Luft procedure. Cytochemical studies using thiocarbohydrazide-silver proteinate were performed according the Thiéry.
Complex electron microscopic study of structural and functional arrangement of chloroplast membranes
Authors: Tageeva S. V. , Popov V. I. , Allakhverdov B. L.
Published: June 2020

Supramolecular arrangement of photosynthetic membranes of cyanobacteria and higher plant chloroplasts has been studied by means of electron microscopy, ultrathin sections, freeze-substitution and freeze-fracture technique. Principal similarity between the structural arrangement of the photosynthetic apparatus of higher plants and cyanobacteria has been established. It consists in a continuity of the photosynthetic membranes and their relation to the inner membrane of chloroplast envelope or to plasmalemma in the cyanobacteria; in the ability of thylakoids stacking; in an asymmetrical distribution of principal number and sizes on complemetary faces of membrane fractures revealed by the freeze-fracture (Fig. 1a, b, c, d; Fig. 2a).

A comparative analysis of particle morphology was carried out by means of angular and circular shading of fracture faces with Pt, “direct” and “inverse” replicas being obtained.
Ultrastructural aspects of the first postmeiotic mitosis and cytokinesis in the pollen grain ofallium cepal
Authors: Rodriguez-Garcia M.I. , Risueno M.C.
Published: June 2020

This report deals with research work on first pollen grain mitosis and the formation of vegetative and generative cells in Allium cepa L.

The haploid nucleus of young microspores has a long interphase period which starts when the microspores begin to free themselves from the surrounding special callose wall and finishes when a central vacuole is formed in their cytoplasm. The pollen wall (Exine and Intine) is formed during this period.

Before the mitosis, a large vacuole is formed in the cytoplasm. The microspore nucleus that previously occupied a central position in the cell can now be found in a lateral position generally on the opposite side of the porus. At this moment the nucleus enters prophase and the mitotic division takes place. This is characterized by the formation of an asymmetric spindle and as a result two unequal daughter cells are formed (Fig. 1) : the vegetative cell is larger and its nucleus remains near the centre of the pollen grain, whilst the generative cell is smaller and at the beginning lies close to the pollen wall (Fig. 2, 3).
The ultrastructure of modified root-tip cells induced by the feeding of an ectoparasitic nematode
Authors: Lehmann Heiner , Wyss Urs
Published: June 2020

It is well known that sedentary endoparasitic root nematodes (Fam. Heteroderidae) induce highly modified ‘nurse cells’ in their host plants. The induction and maintenance of such cells is a precondition for the reproduction of these nematodes. The modified cells, generally called giant cells, are multinucleate, rich in cytoplasm and, concerning cell wall structure, show characteristic features of transfer cells.

In contrast to sedentary endoparasitic nematodes, ectoparasitic nematodes are not known to induce giant cells. However, it was recently shown that root-tip feeding sites of the ectoparasite Xiphinema index contain enlarged multinucleate cells in the two host plants examined (Vitis vinifera and Ficus carica). Root-tips of Ficus carica seedlings - grown in aseptic agar culture at low light intensity - start to swell about 10 h after the first attack by this nematode and gradually become transformed into a proliferating gall as long as the parasite continues its feeding activities.
Studies on tubular myelin figures of lining layer of lung alveoli
Authors: Groniowski Janusz , Walski MichaŁ
Published: June 2020

Extracellular alveolar lining layer of mammalian lungs is rich in phospholipids associated or bound to proteins and carbohydrates. There are implications that all the molecular components aggregate in an obligatory sequence and that protein has an organizing role. In electron microscopy alveolar lining layer is composed of a surface film and a liquid subphase. The latter is marked with tubular myelin figures /TMF/. In this report some attempts to characterize more precisely the TMF are presented.

Prefixation of tissue samples in osmium or glutaraldehyde vapor was followed by fixation in diluted Kamovsky′ s fixative. The TMF is composed of lamellae showing the unit membrane structure. On cross section the TMF reveal a square lattice pattern. The centre of each square contains fragments of a somewhat circular profile /Fig. 1/, suggesting also their linear continuity along the axis of each tubule. The tridimensional reconstruction reveals that the fragments may belong to a randomly coiled formation located in the centre of tubule.
Plasmalemma localisation of inorganic phosphate in B cells pancreas
Authors: Wooding F. B. P. , Pedley K. , Freinkel N. , Dawson R. M. C.
Published: June 2020

Freinkel et al (1974) demonstrated that isolated perifused rat pancreatic islets reproduceably release up to 50% of their total inorganic phosphate when the concentration of glucose in the perifusion medium is raised.

Using a slight modification of the Libanati and Tandler (1969) method for localising inorganic phosphate by fixation-precipitation with glutaraldehyde-lead acetate we can demonstrate there is a significant deposition of lead phosphate (identified by energy dispersive electron microscope microanalysis) at or on the plasmalemma of the B cell of the islets (Fig 1, 3). Islets after incubation in high glucose show very little precipitate at this or any other site (Fig 2). At higher magnification the precipitate seems to be intracellular (Fig 4) but since any use of osmium or uranyl acetate to increase membrane contrast removes the precipitate of lead phosphate it has not been possible to verify this as yet.
Electron microscope study of the microgranules in human vaginal epithelium
Authors: Campos A. , Vilches J. , Gomez J.
Published: June 2020

Microgranules have been described with different names in keratinized and in nonkeratinized epithelium. In keratinized epithelium it seems clear that the microgranules are lamellated bodies bounded by a membrane which empty their contents into the intercellular space. Their existence in nonkeratinized epithelium is more debatable. Until now the so-called microgranules have been described in nonkeratinized bucal, lingual and cervical epithelium. In the present work we describe the morphology and nature of such structures in human vaginal epithelium.

Biopsies from the midlevel of the vaginal mucosa were taken from voluntary fertile women. The specimens were divided into three groups with four vaginal specimens. The first group was obtained in the folicular phase; those of the second in the postovulatory phase and, finally, the last group corresponded to the secretory phase.
Dense core tubuli—a new structural element of the spermatophores of pogonophora
Authors: Flügel H.
Published: June 2020

Pogonophora are tube-dwelling marine invertebrates without a digestive tract. They feed entirely on dissolved organic matter which occurs in traces in the sea water. Their phylogenetic position is still controversial. In spite of numerous anatomical investigations using the light microscope our knowledge of their reproduction is still limited (Ivanov, 1963; Webb, 1963a, b; Southward, 1975). It is well established that the sexes are separate. The sperm is discharged in the form of elongated spermatophores and the eggs are fertilized in the forepart of the tube, at least in the families Siboglinidae and Oligobrachiidae.

But the mechanism of fertilization has not yet been observed. Electron microscopic investigations of the spermatophores may help to elucidate the biology of reproduction. Investigations on the spermiogenesis and the spermatophores of Siboglinum ekmani were carried out recently (Franzén, 1 973; Flügel, 1977).
Surface morphology and secretory activity of rat anterior pituitary cells in primary culture
Authors: Antakly T. , Zeytinoglu F. , Pelletier G. , Labrie F.
Published: June 2020

So far, there has been no report concerning the surface morphology of cultured secretory cells in different states of activity, as observed by scanning electron microscopy. The anterior pituitary cells in monolayer culture offer an unique system to study the modifications of cell conformation in relation with changes of activity since these cells can be specifically modulated by stimulating or inhibiting factors. Scanning electron microscopy of anterior pituitary cells in primary culture was thus performed in different states of secretory activity. Adult female Sprague-Dawley rats at random stage of the estrous cycle, were used for the preparation of the primary cultures of anterior pituitary cells as previously described (Endocrinology 98: 1528, 1976). The cells 7. 5 x 105 in 1. 5 ml of Dulbecco's modified Eagle's medium containing 10% horse serum and 0. 25% foetal calf serum were plated in 3. 5 x 10 mm Petri dishes and were used six days after plating.
Ultrastructural study of the adrenocortical homologue in freshwater and seawater adapted eels
Authors: Kumar Bhattacharyya Tapan , Gordon Butler David
Published: June 2020

The teleost fishes have been extensively utilized to explore the biochemical nature of adrenocortical steroids, pathways of steroidogenesis and the metabolic role of adrenocortical steroids. The adrenocortical homologue (AH) has been localized in more than two hundred species of teleosts, although the fine structure of teleost AH has been studied only in the goldfish. It seemed to be of interest to investigate the normal fine structure of this endocrine organ in another zoologically important teleost species, the north American eel (Anguilla rostrata) as a baseline for experimental studies. This animal being a euryhaline species, the AH ultrastructure of animals exposed to artificial seawater for a long period (1 1/2 years) was also examined.

Mature female silver eels held in freshwater(FW) and seawater(SW) aquaria were anesthetized and perfused with glutaraldehyde in phosphate buffer or formaldehyde-glutaraldehyde in cacodylate buffer, and glutaraldehyde containing 0. 2% digitonin for demonstration of cholesterol.
A study of synaptic morphology in the trigeminal motor nucleus utilizing the HRP method
Authors: Konishi A. , Mizuno N. , Nomura S. , Itoh K. , Nakamura Y.
Published: June 2020

The synaptic organization of the motor nucleus of the trigeminal nerve in the cat was examined electron microscopically after labelling the motoneurons or the axon terminals on the motoneurons by retrograde or anterograde axonal transport of horseradish peroxidase (HRP).

The experiments were carried out on young adult cats (1. 5 to 3. 0 kg) anesthetized with intraperitoneal Nembutal (35 mg/kg). For labelling the motoneurons, 2% solution of HRP (Toyobo Grade-I-C, RZ: 3. 4) dissolved in sterile 0. 9% saline was injected within the muscle belly of the temporal, masseter or anterior digastric muscle with a fine caliber intradermic needle attached to a syringe. For labelling axon terminals of the commissural fibers arising contralaterally from the commissural interneurons around the trigeminal motor nucleus, a single injection of 0. 05 μ l of 50% HRP dissolved in 2% dimethylsulfoxide or in sterile 0. 9% saline was performed stereotaxically within the trigeminal motor nucleus and its adjacent regions.
Characterization of the synaptic membranes in the central nervous system through electron microscopic immunocytochemistry of the botulinum toxin
Authors: Toxin Botulinum , Iiirokawa Nobutaka , Kitamura Masaru
Published: June 2020

Synapses are established between neurites of nerve cells and target cells (e. g. neurons, muscles, sensory cells). Synaptic membranes differentiate to show two main characteristics: one is for release of neurotransmitters and the other is for recognition of its partner during development. Many investigations have been undertaken to visualize the characteristics of the synaptic membranes by lectins, PTA staining and freeze fracture technique. The precise substantial nature of the synaptic membranes, however, has little been known till now. In the present study we intend to characterize synaptic membranes through electron microscopic immunocytochemistry of the botulinum toxin. The binding site of this neurotoxin is supposed to be specific gangliosides and it is well known that the toxin blocks the transmitter release at the neuromuscular junction and gangliosides may contribute much in neuronal recognition.

The botulinum toxin type A cristalline toxin was supplied by Dr. E. J. Schantz(Wiseonsin Univ. ).
Rod and cone-type photoreceptors in the post-equatorial region of the albino rat retina: a morphometric analysis of their incidence and fine structure
Authors: Jonson K. M. , Champion R. A. , Shorey C. D. , Beaumont P. E.
Published: June 2020

The existence of cones in the rat retina is controversial. Although the rat has been thought to possess an exclusive rod retina (Lashley, 1932; Detwiler, 1943), Walls (1934) reported the existence of “typical cones” essentially similar to those of the primate. More recent light and electron microscope studies have also reported on the presence of cones in the rat retina (Leure-duPree, 1974; LaVail, 1976). A quantitative analysis of their incidence and fine structure has not previously been reported.

The results of the present investigation demonstrate that although the rat photoreceptor layer is rod-dominant, approximately 17% of its cells would, on morphological grounds, qualify as “cone-type” receptors. These, however, are not typical cones, as their inner and outer segments do not show the characteristic conical taper and appear morphologically similar to the slender cylindrical-shaped rod inner and outer segment. Apart from this, the rat cone-type receptor cell does resemble the cone typically observed in other species.
Cell differentiation in the olfactory pit of the chick embryo
Authors: Kasuga S. , Okano M. , Sugawa Y.
Published: June 2020

It is well known that the development of the olfactory organ derives from the epithelium lining the surface of the olfactory pit which developed from the ectodermal olfactory placode. The development of the olfactory epithelium has been explored in the chick embryo(Robecchi; 1972, Breipohl and Fernandez;1977) and the mouse embryo(Cuschieri and Bannister;1975, Graziadei and Okano;1978) to a certain extent.

In order to investigate the cellular differentiation of chemoreceptor cells in the olfactory organ proimordium, the olfactory placode of the chick embryo(50-53 incubation hours, Hamburger and Hamilton, 1951) and the olfactory pit of the chick embryo(52-69 incubation hours, HH) are utilized in this study with the aid of a scanning and a transmission electron microscope.

Olfactory placode; the placode is recognized on the antero-lateral aspect of the head of the chick embryo(50-53 incubation hours, HH), and distingwished from the general surrounding ectoderm as a circular territory of 140 μm in diameter, and the cellular arrangement at the placode surface observed by a scanning electron microscope is characteristic and recognized as a rosette consisted from several oval swollen cellular endings (olfactory. vesicles) which have a single cilium and few microvilli, and the other intercalated cellular elements(supporting cells) (Fig. 1).
Calcium localization in fast frog muscle fibers at rest and during potassium contracture
Authors: Samosudova N.V. , Ludkovskaya R.G.
Published: June 2020

There is abundant data in the muscle literature referring to Ca as the activator of the contractile system. The intracellular calcium localization in the fast frog muscle fibers at rest and during K-contrac-ture was studied by electron microscopy combined with pyroantimonate- osmium technique. It appears that the increased Ca-concentration in the myofibrillar space has to be maintained for the development of the prolonged contraction. The fast fibers are not capable of maintaining the prolonged contraction in solution with a high K-concentration. however the prolonged potassium contracture of fast fibers was obtained in response to the action of the KSCN-solution (20 mM). Anion SCN¯ inhibits Ca - uptake by sarcoplasmic reticulum and thus slows down the relaxation.

In the present study we have given the main attention to localization of myofibrillar calcium.

Three Ca-accumulating lines, corresponding to the N-lines, were observed in the I band of stretched fibers (the sarcomere length 3.2 μm) at a distance of N1 - 0.44, N2 - 0.55, N3 - 0.67μ from the Z line (fig. 1).
X-ray microanalytical and enzyme-cytochemical study on vesical malacoplakia
Authors: Hokano Masami , Oh-I Tsunao , Narita Yoshie , Sassa Hiroshi , Saburo Suzuki
Published: June 2020

Malacoplakia is a kind of granulomatous inflammation and characterized by the presence of calcium-stain positive granules (Michaelis-Gutmann bodies, hereinafter abbreviated as M-G bodies ) in the macrophages.

In this report we want to say about the following articles:the ultrastruc tural findings in four cases of vesical malacoplakia;the ultrastructural morphogenesis of the M-G bodies;X-ray microanalytical studies which examine the change of the chemical component of M-G bodies, according to their developing stages; andacid phosphatase activity of lysosome within the malacoplakic macrophages.

Biopsy materials taken from four cases with vesical malacoplakia were divided into 2 parts; one for a light microscopy and the other for an electron microscopy. The specimen for an electron microscopy was fixed in glutaraldehyde and osmium tetroxide, dehydrated in ethanol and then embedded in Epon 812. Ultrathin sections were doublestained with lead citrate and uranyl acetate and then subjected to routin TEM observation. For X-ray microanalysis was mounted an energy dispersive X-ray microanalyzer on a JEM-100 C type electron microscope. Ultrathin sections for microanalysis were cut 100-200nm thich and not stained.
New aspects of articular cartilage transplantation with TEM and SEM
Authors: Hesse W. , Hesse I.
Published: June 2020

Much experimental work has been done in the field of transplantation of articular cartilage. The data are very conflicting. As these investigations were usually performed with routine histological techniques, the ultrastructure and functions of the chondrocytes of the grafts were not examined. Likewise the morphology of the surface, which is the important factor in the function of the joint, was not studied in previous experiments. The purpose of this presentation is to describe the ultrastructure of osteo-chondral grafts to obtain new evidence on the healing process of cartilage transplants.

In our experiment osteo-chondral grafts were implanted in 150 animals. The animals were divided into four groups: 1) 30 sheep with fresh homologous grafts, 2) 15 sheep with autologous grafts. 3) 20 rabbits with stored homologous grafts, 4) 35 rabbits with autologous grafts. The grafts consisting of the articular cartilage and a narrow margin of cancellous bone measured 3. 5 mm in thickness.
Compositional imaging in the Electron Microscope
Authors: Lyman C. E.
Published: June 2020

Imaging of elemental distributions on a fine scale is one of the triumphs of electron microscopy. Compositional imaging frees the operator from the necessity of making decisions about which features contain the elements of interest. Elements in unexpected locations, or in unexpected association with other elements, may be found easily without operator bias as to where to locate the electron probe for compositional data collection. This technique may be applied to bulk or thin specimens using a variety of composition-sensitive signals as shown in Figure 1.

Cosslett and Duncumb obtained the first such compositional image in an electron microprobe modified to scan the electron beam and collect a characteristic x-ray signal as a function of beam position. Early images of this type were called x-ray “dot maps” and provided a qualitative indication of the location of elements on a flat polished bulk specimen to a spatial resolution of about 1 μm.
Scleroderma crest autoantibodies as fluorescent and Immuno-Electron Microscopic probes: Keys to a chromosomal black box
Authors: Brinkley B. R. , Zinkowski R. P.
Published: June 2020

The mammalian kinetochore is a highly differentiated structure found at the centromere (primary constriction) of chromosomes that serves as an attachment site for spindle microtubules. Ultrastructurally, the kinetochore typically appears as a tri-layered plate or disc situated at the sides of the centromere (Fig.1). Recent evidence demonstrates that kinetochores have the ability to capture and stabilize microtubules that grow from the spindle poles. Moreover, the motor(s) for chromosome movement appear to be located in or near the kinetochore which actively participates in the generation of forces necessary for chromosome movement in mitosis and meiosis. To understand how the precise ballet-like movements of chromosomes on the mitotic spindle occur, attention has focused on the “black box” of the chromosome; the centromere-kinetochore complex.

The fortuitous discovery that serum from individuals with the CREST variant of scleroderma contain autoantibodies that bind to components of the centromere-kinetochore complex has led to major advancements in the understanding of this chromosomal black box. Indirect immunofluorescence has demonstrated the presence of paired fluorescent structures (Fig.2) at the centromeres of both mammalian and plant chromosomes.
Autoantibodies as probes for the molecular architecture of the nucleus
Authors: Spector David L. , Lark Gayle , Sovak Mika , Huang Sui
Published: June 2020

Autoantibodies to a variety of nuclear components including DNA, histones, DNA polymerases, DNA topoisomerases, lamins, coiled bodies, RNA polymerase I, and several classes of ribonucleoproteins have been identified in the sera of individuals with autoimmune disorders. We have taken advantage of autoantibodies to several major classes (U1, U2, U4/U6, U5) of small nuclear ribonucleoprotein particles (snRNPs) which play a crucial role in the processing of pre-mRNA molecules. We have used these antibodies to evaluate the organization of factors which participate in pre-mRNA splicing in the mammalian cell nucleus.

We have previously shown snRNPs to be localized to discrete immunostained regions, “speckles”, which form a latticework within the interphase nuclei of several mammalian cell types. Since RNA polymerase II, which is involved in the synthesis of pre-mRNA, is highly sensitive to a-amanitin we were interested in the effect of an inhibition of pre-mRNA synthesis on the snRNP distribution pattern. Cells incubated in drug for 5 hours (Fig. 1d) showed no detectable effect on the organization of snRNPs nor any effect on the level of transcription.
Three-dimensional light microscopy for the analysis of chromosome organization in Drosophila embryo
Authors: Hiraoka Yasushi , Dernburg Abby F. , Parmelee Susan J. , Rykowski Mary C. , Agard David A. , Sedat John W.
Published: June 2020

We are interested in the three-dimensional physical relationships among chromosomes in mitotically active nuclei. Using high-resolution, three-dimensional optical sectioning microscopy incorporating a charge-coupled device (CCD), we have analyzed the spatial arrangements of chromosomes in nuclei from Drosophila melanogaster embryos. During early stages of nuclear division, these embryos exist as syncytial blastoderms, with up to 5000 nuclei forming a single layer near the embryo's surface and dividing essentially synchronously. This greatly facilitates the analysis of chromosome organization because of the simple monolayer arrangement of nuclear structures at defined mitotic stages.

From prophase to anaphase, three-dimensional paths of condensed chromosomes stained with a DNA-specific fluorescent dye can be traced using an interactive computer program. We have used this method to ask the specific biological question of whether pairs of homologous chromosomes have a consistent, ordered arrangement in diploid nuclei. Our results have shown that homologous chromosomes are not associated with each other from prophase to anaphase in syncytial blastoderm stage embryos.
Molecular morphology of hepatic glycogen metabolism
Authors: Cardell Robert R.
Published: June 2020

For over two decades we have studied morphological aspects of hepatic glycogen metabolism, particularly the role of smooth endoplasmic reticulum in this process. Recently investigators have emphasized the role of hepatic gluconeogenesis (formation of glucose from non-carbohydrate precursors) in glycogen synthesis. To contribute new morphological information to this discussion we have developed probes for the detection of the relevant gluconeogenic enzymes by immunocytochemistry and the expression of the genes for the enzymes by in situ hybridization histochemistry. In this report we present: our work on the expression of a gene for the major rate limiting enzyme in hepatic gluconeogenesis, phosphoenolpyruvate carboxykinase (PEPCK).
In situlocalization of mRNAs and proteins with both light and Electron Microscopy
Authors: Pollock J. A. , Benzer Seymour , Deerinck T. , Martone M. , Ellisman M. H.
Published: June 2020

Cell biological analysis through direct microscopic observation has adopted molecular biological techniques to study the expression of particular genes. With antibodies and other molecular probes, the specific subcellular localization of a given type of protein can readily be visualized using conventional light microscopes, Confocal microscopes and electron microscopes. A tremendous amount of information has been reported on the synthesis, processing and transport of many classes of proteins resulting from the direct visualization of protein expression in the tissue. Recently, the specific subcellular localization of mRNA transcripts has been reported in several systems relating the localization of the RNA to the localization and function of the proteins they encode (Garner, et al., 1988; Macdonald and Struhl, 1988; Singer, et al., 1989; Yisraeli and Melton, 1988, 1990; Pollock et al., 1990). The success of these studies have relied on the large physical dimensions of the cells used so that the non-isotopic, light microscopic techniques employed could resolve distinct regional differences in the RNA signal.
Cryogenic atomic force microscopy
Authors: Fisher K. A. , Gustafsson M. G. L. , Shattuck M. B. , Clarke J.
Published: June 2020

The atomic force microscope (AFM) is capable of imaging electrically conductive and non-conductive surfaces at atomic resolution. When used to image biological samples, however, lateral resolution is often limited to nanometer levels, due primarily to AFM tip/sample interactions. Several approaches to immobilize and stabilize soft or flexible molecules for AFM have been examined, notably, tethering coating, and freezing. Although each approach has its advantages and disadvantages, rapid freezing techniques have the special advantage of avoiding chemical perturbation, and minimizing physical disruption of the sample. Scanning with an AFM at cryogenic temperatures has the potential to image frozen biomolecules at high resolution. We have constructed a force microscope capable of operating immersed in liquid n-pentane and have tested its performance at room temperature with carbon and metal-coated samples, and at 143° K with uncoated ferritin and purple membrane (PM).
A plunge-freezing method for imaging mammalian tissue culture cells
Authors: O'Toole E.T. , McIntosh J.R.
Published: June 2020

Ultrarapid freezing of tissue culture cells followed by freeze substitution has been used a method for the optimum fixation of cytoskeletal components that are often sensitive to routine chemical fixation. This is due to the fact that freezing methods such as plunge freezing, result in the almost instantaneous fixation of all cellular components without alteration of the cell's morphology. In addition, we have found that the plunge freezing method is useful for obtaining thin frozen cells for direct cryoimaging. Here we describe how the plunge freezing method can be applied both for freeze substitution analysis and for direct cryoimaging of frozen tissue culture cells.

PTK, cells were grown to confluence on formvar coated, carbon stabilized gold grids. Prior to freezing, the grids were blotted in a 37°C, high humidity chamber so that a minimum of culture medium remained on the grid. This blotting step was critical to obtain a sample thin enough for optimum cryopreservation and subsequent cryoimaging.
Ultrastructural observations of human monocytes infected with HIV-1
Authors: Koehler James K. , Reed Steven G. , Silva Joao S.
Published: June 2020

As part of a larger study involving the co-infection of human monocyte cultures with HIV and protozoan parasites, electron microscopic observations were made on the course of HIV replication and infection in these cells. Although several ultrastructural studies of the cytopathology associated with HIV infection have appeared, few studies have shown the details of virus production in “normal,” human monocytes/macrophages, one of the natural targets of the virus, and suspected of being a locus of quiescent virus during its long latent period. In this report, we detail some of the interactions of developing virons with the membranes and organelles of the monocyte host.

Peripheral blood monocytes were prepared from buffy coats (Portland Red Cross) by Percoll gradient centrifugation, followed by adherence to cover slips. 90-95% pure monocytes were cultured in RPMI with 5% non-activated human AB serum for four days and infected with 100 TCID50/ml of HIV-1 for four hours, washed and incubated in fresh medium for 14 days.
Neural activity triggers Ca waves in hippocampal astrocyte networks
Authors: Dani J.W. , Chernjavsky A. , Smith S.J.
Published: June 2020

Recent findings that the neurotransmitter glutamate can trigger cytoplasmic Ca waves in networks of cultured astrocytes suggest a new range of possible mechanisms for neural-glial interaction and for signalling over distance within the brain. To explore some of these possibilities, we have used confocal microscopy and the Ca indicator fluo-3 to study organotypically cultured slices of rat hippocampus where astrocytes and neurons are intermingled in their normal tissue relationships. We observe that electrical stimulation of the dentate gyrus induces Ca signals in astrocytes of region CA3, acting via a TTX-sensitive neuronal fiber projection that releases glutamate as its major neurotransmitter. Astrocyte Ca transients can occur within two seconds of the onset of neural stimulation, and waves and oscillations result from sustained stimulation at frequencies as low as 2 Hz. Neurally-evoked astrocyte Ca waves propagate both within and between individual hippocampal astrocytes at velocities of 10 - 20 um/sec, while the oscillations have periods of approximately 20 sec (at 21 °C).
Correlative light microscopy, electron microscopy, and cytochemistry of cytoskeletal structures in motile fibroblasts
Authors: Heath Julian P. , Turner Donna , Holifield Bruce F.
Published: June 2020

Contrast-enhanced video differential interference contrast microscopy (VDICM) is revealing new details about the dynamics of F-actin assemblies in motile cells. We are using correlative single cell light and electron microscopy and immunocytochemistry to understand the dynamic interactions of the actin cytoskeleton in lamellipodia and the leading lamella.

Fibroblasts were cultured on carbon-coated glass #1 coverslips and examined on a Zeiss Axiophot. Cells were perfused with 1% glutaraldehyde (GA) in PIPES buffer and fixed for 15 min., quenched with 1 mg/ml borohydride/PBS and extracted with 0.1% Triton X-100 in PBS for 1 min. Cell postions were marked with a diamond objective, and the cells were incubated for 30 min. with 0.6 uM rhodamine-conjugated phalloidin to stain for F-actin. Light and fluorescence micrographs were taken on fine-grained film. After photography, cells were further fixed in 2.5% GA, postfixed in 1% osmium, dehydrated in ethanol and embedded in Spurrs resin. The glass was removed by scoring with a diamond pencil followed by immersion in liquid N2. Cells were relocated in the blocks by phase-contrast microscopy. Thin (80 nm) sections were examined in Philips EM 410 at 60 kV on a dicentric goniometric stage.
Three-dimensional imaging of the living eye
Authors: Masters Barry R.
Published: June 2020

The structure of the in situ rabbit cornea can be observed at high resolution and contrast with reflected light confocal microscopy. In vivo confocal images of the living cornea have been made at lower resolution and lower contrast using a SIT video camera together with a real-time Nipkow disk confocal microscope adapted for in vivo observations. This paper describes the three dimensional reconstruction of the in situ cornea from an enucleated rabbit eye with confocal reflected light microscopy and volume rendering computer techniques.

A laser scanning confocal microscope (BioRad MRC 600) was used in the reflected light mode to obtain the two-dimensional image stack from the cornea of a freshly enucleated rabbit eye. The eye was maintained in a physiological state with aerated Ringer's solution. The light source was an argon ion laser with a 488 nm wavelength. The microscope objective was a Leitz X25, NA 0.6 water immersion lens. The 400 micron thick cornea was optically sectioned into 133, three micron sections. The optical sectioning was performed perpendicular to the optical axis of the eye globe.
A Transmission electron microscopic study of methods for harvesting epithelial cells from human corneal tissue
Authors: Dimitrijevich S. D. , Reese T. , Gracy R. W. , Oakford L. X. , Howe W. E.
Published: June 2020

A critical aspect of cell harvesting is the separation of cells from the extracellular matrix without damaging the cell attachment structures. Isolation of epithelial cells frequently involves the use of highly skillful microsurgical techniques, and precision instrumentation. The proteolytic detachment (trypsin), or sequestering of membrane bound calcium (EDTA) are also widely used. These two approaches are particularly difficult when isolating cells from the human eye tissue, often leading to poor attachment, slow growth, and heterogeneous cultures. Initiation of primary cultures by harvesting outgrowths from tissue explants is also unsatisfactory. Recently dispase and thermolysin, have been shown to be preferable for harvesting rabbit corneal epithelial cells and human keratinocytes respectively. We have compared these with the effects of trypsin and EDTA on the human corneal epithelium.
Kalinin: A new epithelial basement membrane adhesion molecule
Authors: Keene Douglas R. , Lunstrum Gregory P. , Rousselle Patricia , Burgeson Robert E.
Published: June 2020

A mouse monoclonal antibody produced from collagenase digests of human amnion was used by LM and TEM to study the distribution and ultrastructural features of an antigen present in epithelial tissues and in cultured human keratinocytes, and by immunoaffinity chromatography to partially purify the antigen from keratinocyte cell culture media.

By immunofluorescence microscopy, the antigen displays a tissue distribution similar to type VII collagen; positive staining of the epithelial basement membrane is seen in skin, oral mucosa, trachea, esophagus, cornea, amnion and lung. Images from rotary shadowed preparations isolated by affinity chromatography demonstrate a population of rod-like molecules 107 nm in length, having pronounced globular domains at each end. Polyacrylamide gel electrophoresis suggests that the size of this molecule is approximately 440kDa, and that it is composed of three nonidentical chains disulfide bonded together.
Expression of laminin and beta-1 integrin in embryonic, neonatal, and adult rat heart
Authors: Price R.L. , Borg T.K. , Terracio L. , Nakagawa M.
Published: June 2020

Little is known about the temporal expression of extracellular matrix components (ECM) and its receptors during development of the heart. Recent reports have shown that ECM components undergo both qualitative and quantitative changes during development, and it is believed that ECM components are important in the regulation of cell migration and cell:cell and cell:ECM recognition and adhesion.

Integrins are transmembrane glycoproteins which bind several ECM components on their external face and cytoskeletal elements on the cytoplasmic face. Laminin is a basement membrane component which has been recognized as an important site for cell adhesion. Both the integrins and laminin are expressed early in development and continue to be expressed in the adult heart. With their documented roles in cell recognition, and cell:cell and cell:ECM migration and adhesion these proteins appear to be important components in development of the heart, and their temporal expression may play a pivotal role in morphogenesis and myofibrillogenesis of the heart.
A study of fibrin clot structure under various ionic conditions using stereoscopic IVEM images
Authors: Baradet T. , Weisel J.W. , Peachey L.D.
Published: June 2020

Fibrin clots are formed by the conversion of fibrinogen into fibrin monomers which assemble to produce two-stranded protofibrils that aggregate to form fibers. These fibers may also aggregate laterally with other fibers to form larger fiber bundles. Investigations of fibrin clot structures under various ionic conditions using SEM showed dramatic differences in fiber morphology and clot structure. Fibrin clots formed under various ionic conditions were investigated by the examination of stereoscopic IVEM images. This technique provides greater depth discrimination and higher resolution images of clot ultrastructure. Details of fiber association and branching are of particular interest.

Purified human fibrinogen was prepared at a concentration of 0.5 mg/ml in 0.4M, 0.2M, and 0.05M buffers (0.05M Tris-HCI, pH 7.4, with 2mM CaCl2). Fibrinogen solutions were mixed with thrombin to a final concentration of 0.3U/ml and aiiquots placed onto Formvar and carbon coated grids. After clotting for 1 hr. at room temperature, clots were fixed with 3% glutaraldehyde in 0.1 M Na-cacodylate buffer, pH 7.2, for 5 min. Grids were kept constantly moist to avoid collapse or syneresis.
Structure-function analysis of anaphase spindle elongation using isolated diatom spindles
Authors: Cande W. Z. , Hogan C.J. , Lee M.
Published: June 2020

Diatom spindles are important model systems for describing the morphological changes associated with anaphase chromosome movement because the fibrous systems responsible for anaphase A (chromosome-to-pole movement) and anaphase B (spindle elongation) are spatially separate and the central spindle is a paracrystalline array of microtubules. The diatom central spindle, which is responsible for anaphase B, is constructed of two sets of interdigiting microtubules that originate from plate-like spindle poles and display specific near-neighbor interactions in the zone of microtubule overlap. The microtubules of each half-spindle are of relatively unifrom length such that the plus ends are clustered together in narrow zones at each edge of the zone of microtubule overlap. This has allowed us to monitor changes in extent of microtubule overlap in the light microscope with polarization optics. We have isolated spindles from synchronized populations of several species of dividing diatom cells to study the mechanochemistry of anaphase spindle elongation in vitro and to analyze the rearrangement of spindle components by light and electron microscopy during reactivation.
Characterization of male-sterile Arabidopsis thaliana mutants exhibiting altered tapetum development
Authors: Bricker C. S. , Smith-Huerta N. , Makaroff C. A.
Published: June 2020

The tapetum is the tissue in the anther that is in closest contact with the developing microspores. It consists of large and frequently multinucleate, richly cytoplasmic cells that provide most of the nutrients and growth substances necessary for the developing microspore. A great deal of our knowledge on the tapetum has come from studies on malesterile mutants. We are using T-DNA insertional mutants of Arabidopsis thaliana to identify, isolate and characterize the genes that control the differentiation of the tapetum. As a first step we have identified those mutants that exhibit the sporogenous type of male sterility (relatively normal stamen development but alterations in microsporogenesis). Further analyses have identified those mutants that exhibit alterations in tapetum development that have resulted from insertional mutagenesis by the T-DNA. Through the analysis of these mutants we hope to identify the molecular signals that regulate tapetal tissue differentiation and provide molecular evidence on the role of the tapetum in microspore development.
Thylakoid membrane development in pigment-deficient wheat chloroplasts
Authors: Freeman Thomas , Duysen Murray , Eskins Ken , Guikema James
Published: June 2020

The thylakoid membranes of higher plant chloroplasts contain at least two major pigment protein complexes, photosystem I (PSI), and photosystem II (PSII). The mature apoprotein of these complexes (involved in the initial reactions of photosynthesis) bind specific chlorophylls (Chl) and specific carotenoids in an unknown manner. It has been suggested, however, that the synthesis of pigments is normally coordinated with that of apoproteins. We have examined the effect of gabaculine (3-amino-2, 3-dihydrobenzoic acid) on granal thylakoid stacking as well as pigment and apoprotein accumulations for PSI and PSII in wheat.

Gabaculine (0.5mM) was applied with nutrient solution to 6.5 day-old wheat seedlings maintained in a growth chamber at 23C. One seedling lot grown under continuous light (400 μmol photons s-1 m-2) possessed green primary leaves at time of treatment whereas another seedling lot, dark grown, possessed only etiolated primary leaves. Twelve hours after treatment, the etiolated seedlings were transferred into continuous light. The primary and secondary leaves were subsequently harvested from 14 day-old seedlings of both lots.
Occurrence of leucocytes in the intracellular granules in the enterocytes of the jejunum of newborn pigs
Authors: Kömüves Lászlo G. , Heath Julian P. , Nichols Buford L.
Published: June 2020

In some mammals, where the transport of maternal immunoglobulins across the intestine plays an important role in the passive immunity of neonates (such as rat, pigetc), one of the most striking features of the enterocytes in the small intestinal mucosa of suckling newborns is the formation of large cytoplasmic granules shortly after birth. These large granules are believed to be represent specialized lysosomes and/or late endosomes and accumulate maternal immunoglobulins from the colostrum. Here we report our observation that different leucocytes can frequently be found in the subnuclear granules in suckling piglets.

Suckling piglets of various age from three different litters were used in this study. Pieces of the jejunum were collected and processed for Spurr's and LR White embedding. Semithick section were observed with a Zeiss Axiophot microscope. Thin LR White sections were immunolabeled with a rabbit anti-porcine IgG, followed by goat anti-rabbit IgG, conjugated to 10 nm gold. Semithick LR White sections were silver intensified following immunolabeling. Thin sections were observed in a Philips CM 12 electron microscope at 80 kV.
Three-dimensional architecture of Golgi apparatus in the hormone secretory cell
Authors: Tai S. , Albrecht R.M.
Published: June 2020

The Golgi apparatus plays an important role in the process of packaging and sorting of secretory granules in endocrine and exocrine cells. The intra Golgi site for concentrating secretory materials, packaging, and sorting the secretory granules has been widely studied using cytochemical, immunocytochemical and biochemical methods on cells and cell fractions. It is generally accepted that the secretory products fo How the cis to trans pathway across the Golgi stack. Within the Golgi complex, secretory products appear to be concentrated in the dilated rims of the trans-most cisternae which are associated with the formation of secretory granules. In pituitary cells the organization of the Golgi apparatus is not as regular as that described for many other cell types. The cisternae of different Golgi stacks are irregular in size and shape. The cis-trans arrangement is not in a definitive orientation.