bluuebunny/crossref_metadata_2025 huggingface dataset:
Fine structure of the vomeronasal and septal olfactory epithelia and of glandular structures
Authors: Adams Donald R.
Published: February 2005

The vomeronasal and septal olfactory organs are two neurosensory structures in the mammalian nasal septum which are poorly understood relative to the main olfactory system. The vomeronasal organ is a paired, blind‐ending tubular structure that opens rostrally into the nasal cavity in some species and into the incisive ducts in others. When present in mammals, the septal olfactory organ is an island of olfactory mucosa positioned such that it is in the primary air pathway in the caudal portion of the nasal cavity. Mammalian nasal glands, with a diverse histochemical and ultrastructural morphology, secrete a variety of substances onto the mucosal surface. One of these substances, odorant binding protein, localized in bovine nasal glands and lateral nasal glands of rodents may be important in the capture and conveyance of odorant molecules to olfactory receptors. The objectives of this paper are to present original data while reviewing the literature on the ultrastructure of vomeronasal and septal olfactory neuroepithelia, and of vomeronasal, bovine nasal, and lateral nasal glands. Nasal tissues from pigs, calves, and hamsters were prepared for electron microscopy. Neurosensory epithelia of the porcine vomeronasal organ and the hamster septal olfactory organ are similar to that described for the vomeronasal and septal olfactory organs of other mammals. Bovine nasal and rodent lateral nasal glands consist of subregions which differ morphologically; the most abundant acinar cell type in the bovine nasal gland contains lightly electron dense secretory granules while that of the rodent lateral nasal gland contains both small electron dense and large, electron lucent granules. The porcine vomeronasal gland contains numerous small, dense granules of a diverse morphology. © 1992 Wiley‐Liss, Inc.
Fine structural aspects of secretion and extrinsic innervation in the olfactory mucosa
Authors: Getchell Marilyn L. , Getchell Thomas V.
Published: February 2005

The mucus at the surface of the olfactory mucosa constitutes the milieu in which perireceptor events associated with olfactory transduction occur. In this review, the ultrastructure of olfactory mucus and of the secretory cells that synthesize and secrete olfactory mucus in the vertebrate olfactory mucosa is described. Bowman's glands are present in the olfactory mucosa of all vertebrates except fish. They consist of acini, which may contain mucous or serous cells or both, and ducts that traverse the olfactory epithelium to deliver secretions to the epithelial surface. Sustentacular cells are present in the olfactory epithelium of all vertebrates. In fish, amphibia, reptiles, and birds, they are secretory; in mammals, they generally are considered to be “nonsecretory,” although they may participate in the regulation of the mucous composition through micropinocytotic secretion and uptake. Goblet cells occur in the olfactory epithelium of fish and secrete a mucous product.

Secretion from Bowman's glands and vasomotor activity in the olfactory mucosa are regulated by neural elements extrinsic to the primary olfactory neurons. Nerve fibers described in early anatomical studies and characterized by immunohistochemical studies contain a variety of neuroactive peptides and have several targets within the olfactory mucosa. Ultrastructural studies of nerve terminals in the olfactory mucosa have demonstrated the presence of adrenergic, cholinergic and peptidergic input to glands, blood vessels, and melanocytes in the lamina propria and of peptidergic terminals in the olfactory epithelium. The neural origins of the extrinsic nerve fibers and terminals are the trigeminal, terminal, and autonomic systems. © 1992 Wiley‐Liss, Inc.
Ultrastructural studies of microtubules and microtubule organizing centers of the vertebrate olfactory neuron
Authors: Burton Paul R.
Published: February 2005

The olfactory neuron is specialized along its length into highly determined morphological regions. These regions include the dendritic cilia, dendritic vesicle, dendritic shaft proper, perikaryon, axon, zone of transition where the axon widens as it approaches its termination, and the axon terminal. Except for the zone of transition and the terminal, characteristic populations of microtubules occur in these compartments. In the olfactory vesicle, three discrete microtubule organizing centers (MTOCs) nucleate microtubules: the basal body, the lateral foot associated with the body, and dense masses of nearby material. Little is known about MTOCs elsewhere in the neuron, although the polarity of the axonal microtubules indicate that they originate at or near the perikaryon. An attempt is made to summarize what is known of the origin, structure, distribution, and function of microtubules in vertebrate olfactory neurons, which are useful model systems in which to study microtubules. Information about olfactory neuron microtubules may be applicable to neurons in general (e.g., the discovery that axons contain microtubules of uniform polarity was first made in the olfactory neuron) or to microtubules in other eukaryotic cells. © 1992 Wiley‐Liss, Inc.
Lectins bind differentially to cilia and microvilli of major and minor cell populations in olfactory and nasal respiratory epithelia
Authors: Menco Bert Ph.M.
Published: February 2005

Binding of colloidal gold‐conjugated lectins was studied in cilia and microvilli of rat olfactory and respiratory epithelia. This was done in sections of rapidly frozen, freeze‐substituted specimens embedded in Lowicryl K11M or, for wheat germ agglutinin (WGA) alone, in deep‐etched replicas. Olfactory dendritic endings and cilia labeled with WGA and faintly with soybean agglutinin (SBA); olfactory supporting cell microvilli bound only Dolichos biflorus agglutinin (DBA). Microvilli of an infrequent cell bound peanut agglutinin (PNA), SBA, and WGA. These microvilli labeled more strongly with the last two lectins than the olfactory cilia. Respiratory cilia bound WGA and, somewhat more weakly, PNA; microvilli of ciliated respiratory cells bound all four lectins. Visualization of specific labeling improved after preincubation of sections with neuraminidase, except for DBA where lectin binding was abolished. PNA labeling was seen only after neuraminidase preincubation. The densities of membrane surface particles that labeled with WGA corresponded with those of fracture plane particles in a quantitative freeze‐fracture, deepetch analysis. Therefore, a considerable fraction of the WGA‐bound particles could reflect transmembrane proteins in olfactory dendritic endings and cilia and in respiratory cilia. The possible nature of these particles is discussed. © 1992 Wiley‐Liss, Inc.
Photoconversion of diaminobenzidine with different fluorescent neuronal markers into a light and electron microscopic dense reaction product
Authors: Lübke Joachim
Published: February 2005

This article describes methods for photoconverting diaminobenzidine (DAB) into a stable, light and electron microscopically visible dark reaction product in neurons which contain a fluorescent dye. Photoconversion of DAB has been achieved so far with the following fluorescent dyes: rhodamine labeled latex microspheres (RLM), 4,6‐diamidino‐2‐phenylindole (DAPI), 5,7‐di‐hydroxytryptamine (5,7‐DHT), Fast Blue (FB), Nuclear Yellow (NY), Diamidino Yellow (DY), Evans Blue (EB), acridine orange (AO), ethidium bromide (EBR),1,1′‐dioctadecyl‐3,3,3′,3′‐tetramethylindolcarbocyanine perchlorate, D‐282 (DiI), propidium iodide (PI), and intracellularly injected Lucifer Yellow (LY). The dye is introduced into the neurons by tinctorial staining, retrograde transport, or intracellular injection. Photoconversion is conducted by incubating the tissue with the fluorescent substance‐containing cells in a DAB solution under simultaneous strong illumination with ultraviolet (UV) light. During the formation of the reaction product, the fluorescence disappears from the cell. In all cases, photoconversion provided a stable, nonfading DAB reaction product for light microscopy. In addition, at the electron microscopic level, it appeared that the photoconversion results in a homogeneously distributed, fine granular, dark, intracellularly located reaction product. With most of the retrograde tracers tested, photoconversion led only to staining of the cell bodies and the proximal portions of primary dendrites. Following photoconversion with intracellularly LY‐filled neurons and cells labeled retrogradely with DiI, DiO, and 5,7‐DHT, the reaction product was present throughout the cells, extending from the cell bodies into dendrites and dendritic appendices, and into axons. The high selectivity and methodological simplicity of photoconversion of DAB with fluorescent dyes into a stable, light and electron microscopical dense reaction product provide a promising alternative to classical neuroanatomical techniques and a new useful application of fluorescent neuronal tracers to light and electron microscopy. © 1993 Wiley‐Liss, Inc.
Ultrastructural and functional connectivity of intracellularly stained neurones in the vertebrate retina: Correlative analyses
Authors: Djamgoz Mustafa B. A. , Kolb Helga
Published: February 2005

A variety of intracellular recording and staining techniques has been used to establish structure‐function and, in some cases, structure‐function‐neurochemical correlations in fish, turtle, and cat retinae. Cone photoreceptor‐horizontal cell connectivity has been studied extensively in the cyprinid fish retina by intracellular staining with horseradish peroxidase (HRP) and subsequent electron microscopy. The available data suggest that horizontal cell dendrites around the ridge of the synaptic ribbon are postsynaptic, whilst finger‐like extensions (“spinules”) of lateral dendrites function as inhibitory feedback terminals. An interesting feature of this inter‐action is its plasticity: the feedback pathway is suppressed in the dark and becomes potentiated by light adaptation of the retina.

Intracellular recordings and stainings of ganglion cells in both turtle and cat retinae have been possible. Prelabelling of ganglion cells by retrograde transport of rhodamine from the tectum allows ganglion cells to be stained under visual control, and their synaptic inputs determined by electron microscopy. Such studies have been extended to double labelling by using autoradiography or postembedding immunohistochemistry to identify the neurotransmitter content of the labelled cell and/or the neurotransmitter(s) converging upon it. It is envisaged that further applications of intracellular staining followed by double‐ or even triple‐labelling will continue to enhance greatly our understanding of the functional architecture of the vertebrate retina. © 1993 Wiley‐Liss, Inc.
Programmable freeze‐substitution and cryo‐embedding device
Authors: Trachtenberg Shlomo
Published: February 2005

The construction and performance of a modular and fully controlable freeze‐substitution device are described. The core of the device consists of a heavy brass block providing a large, stable thermal mass. The block is composed of two perforated plates and a base plate forming nine deep wells, which enable the concomitant substitution of several samples in various substitution fluids, and in large volumes. The wells are surrounded by an isometric network of tunnels through which either liquid nitrogen or hot air can flow. The isometric network enables heat transfer across short uniform distances throughout the entire block's volume, thus minimizing temperature gradients and differences. The temperature of the substitution fluid, rather than that of the metal block, is monitored by a programmable controller, enabling the presetting of any freeze‐substitution regime. © 1993 Wiley‐Liss, Inc.
Improved TEM samples of semiconductors prepared by a small‐angle cleavage technique
Authors: McCaffrey John P.
Published: February 2005

A small‐angle cleavage technique has been developed that produces superior transmission electron microscope (TEM) samples of semiconductors and related materials. The technique involves back‐thinning the sample to approximately 100 μm, then scribing a groove on this back face at a specified small angle to a standard cleavage plane. The sample is cleaved along this scribe line followed by cleaving along the standard cleavage plane to produce a thin wedged sample. Samples prepared by this method are characterized and compared with conventional and low‐angle ion milled samples. The technique is illustrated, and the characteristic geometry of the cleaved sample is explained in terms of a simplified cleavage model. © 1993 Wiley‐Liss, Inc.
Tracing neural pathways in snail olfaction: From the tip of the tentacles to the brain and beyond
Authors: Chase Ronald , Tolloczko Barbara
Published: February 2005

The anatomical organization of the olfactory system of terrestrial snails and slugs is described in this paper, primarily on the basis of experiments using the African snail Achatina fulica. Behavioral studies demonstrate the functional competence of olfaction in mediating food finding, conspecific attraction, and homing. The neural substrate for olfaction is characterized by an extraordinarily large number of neurons relative to the rest of the nervous system, and by the fact that many of them are unusually small. There exist multiple serial and parallel pathways connecting the olfactory organ, located at the tip of the tentacle, with integrative centers in the central nervous system. Our methods of studying these pathways have relied on the selective neural labels horseradish peroxidase and hexamminecobaltous chloride. One afferent pathway contains synaptic glomeruli whose ultrastructure is similar to that of the glomeruli seen in the mammalian olfactory bulb and the insect olfactory lobe. All of the olfactory neuropils, but especially the tentacle ganglion, contain large numbers of morphologically symmetrical chemical synapses. The procerebrum is a unique region of the snail brain that possesses further features analogous with olfactory areas in other animal groups. Olfactory axons from the tentacle terminate in the procerebrum, but the intrinsic neurons do not project outside of it. An output pathway from the procerebrum to the pedal ganglion has been identified and found to consist of inter‐ganglionic dendrites. The major challenge for future studies is to elucidate the pattern of connectivity within, rather than between, the various olfactory neuropils. © 1993 Wiley‐Liss, Inc.
Synaptic organization and development of the antennal lobe in insects
Authors: Boeckh J. , Tolbert L. P.
Published: February 2005

Many insects possess a highly developed sense of smell. This paper summarizes the cellular and synaptic organization of the antennal (olfactory) lobe of the insect brain and then reviews morphological and fine‐structural aspects of the development of the lobe. Visualization of synapses between classes of neurons identified by physiological, morphological, or transmitter‐cytochemical properties has provided insights into arrangements of contacts and their possible roles in information processing. Studies of development have revealed the requirement for afferent axons from the antenna for the formation of olfactory glomeruli, where virtually all of the synapses in the lobe occur, and have suggested the possibility that glial cells play a role in the instructive influence of the axons on their target neurons in the lobe. The findings reviewed in this paper are primarily from one representative hemimetabolous insect, the American cockroach, and one representative holometabolous insect, a hawkmoth, and comparisons are made with vertebrate systems when appropriate. © 1993 Wiley‐Liss, Inc.
Microscopy in solid state science
Authors: McGibbon A. J. , Brown L. M. , Bleloch A. L. , Browning N. D. , Cadete Santos Aires F. , Fallon P. J. , Gaskell P. H. , Gilkes K. W. R. , Hansen P. L. , Howie A. , Maynard A. D. , McComb D. W. , McMullan D. , Müllejans H. , Murooka Y. , Paterson J. H. , Perovic D. D. , Pike W. T. , Rauf I. A. , Rodenburg J. M. , Saeed A. , Stelmashenko N. , Tu K. N. , Walls M. G. , Walsh C. A. , Yuan J. , Zhao J.
Published: February 2005

The Microstructural Physics group at the Cavendish Laboratory is actively involved in a considerable number of research projects which cover a broad range of materials science. In this paper, we describe briefly several such projects, with particular emphasis given to the application of parallel‐detection electron energy loss spectroscopy (PEELS) on a scanning transmission electron microscope (STEM) to the analysis of materials such as stainless steels, catalysts, and high temperature superconductors. In addition, we describe a number of related projects that are currently being carried out in the group, particularly those which utilise and develop novel STEM imaging and analytical techniques. © 1993 Wiley‐Liss, Inc.
Instrumentation, techniques, and applications of electron microscopy in the solid state physics group at Glasgow university
Authors: Craven A. J. , McVitie S. , Chapman J. N.
Published: February 2005

This review discusses some of the work performed by the Solid State Physics Group at Glasgow University. A major aim of the group is to obtain quantitative information with high spatial resolution and to do this reliably requires a thorough understanding of both the instrumentation and the interactions between the electron beam and the specimen. Thus the first part of the review discusses those aspects of instrumentation and techniques that the group has considered in detail while the final part deals with applications which involve the study of a wide range of materials covering metallurgical, semiconductor, organic, and magnetic systems. In all these applications, the results from a range of techniques have been required to provide as complete a picture of the material as possible. © 1993 Wiley‐Liss, Inc.
Microanalysis near particles and voids at grain boundaries
Authors: Tatlock G. J. , Partridge A. , Beahan P. G.
Published: February 2005

Some of the problems of using high spatial resolution microanalysis in the vicinity of particles and voids in metal grain boundaries are discussed. New analytical results are presented for nickel based alloys, which suggest that elemental distributions in the region immediately adjacent to growing grain boundary particles and voids are often anisotropic. © 1993 Wiley‐Liss, Inc.
Ultrastructure of early chick embryo tissues after high pressure freezing and freeze substitution
Authors: Allenspach Allan
Published: February 2005

Early chick embryos (stages 11–29), or parts thereof, were frozen by the high pressure technique and subjected to a variety of freeze substitution protocols. In addition to hexadecene, unincubated albumen was used as a cryoprotectant to surround the specimens during the freezing event. The quality of freeze ranged from poor to very good in these highly aqueous specimens; very good freeze quality was observed minimally to depths of 30–40 μ. Freeze substitution was carried out with a custom‐made aluminum chamber, automatic freeze substitution machine, or a combination of the two, in acetone containing (1) uranyl acetate; (2) acrolein and/or tannic acid; or (3) a sequence of (2), osmium, and glutaraldehyde. Excellent quality of morphology was cryopreserved with all solutions, given optimally cryoimmobilized tissue. Mitotic and pericentriolar microtubules and cytoskeletal elements were cryopreserved with all methods; membranous organelles were better preserved with osmium. Outstanding preservation of morphology of the ground substance of very hydrated embryonic cells, though limited in depth, was accomplished by high pressure cryoimmobilization and cryopreservation, demonstrating that the technique can be successfully applied to highly aqueous tissues. © 1993 Wiley‐Liss, Inc.
Impact of freeze substitution on biological electron microscopy
Authors: Hippe‐Sanwald Sigrun
Published: February 2005

Considering the increasing necessity for improved preparation techniques in biological electron microscopy as a basis for the identification and localization of cellular substances within the compartments of the cell, this review is focussed on the method of freeze substitution as an important link between the cryofixation (ultrarapid freezing) and resin embedding of biological specimens. The theory and practice of freeze substitution is summarized with particular interest in the physical and thermodynamic as well as in the chemical basis of this technique. A survey of practical aspects of the technical process of freeze substitution concerning the equipment and various protocols successfully applied in biological systems is also given. The main advantage of freeze substitution versus conventional chemical fixation is seen in the maintenance of the hydration shell of molecules and macromolecular structures. This results in an improved fine structural preservation, superior retention of the antigenicity of proteins and decreased loss of unbound, diffusible cellular components. Examples of excellent visualization of the ultrastructure of macro‐molecular complexes (nucleic acids, extracellular material, membranes etc.), small organisms (bacteria, algae, cyanobacteria and fungi) and large biological samples such as plant and animal tissue as well as the plant‐pathogen (fungus) interface and infection structures are presented. Recent data on the molecular characterization of freeze‐substituted biological tissue are exemplified with special emphasis on the subcellular detection of soluble components (elements, lipids, proteins and drugs) and the inter‐/intracellular localization of proteins including foreign proteins in transgenic plants. The molecular analysis of freeze‐substituted specimens is achieved by the combination of low temperature preparation techniques in biological electron microscopy with various detection methods such as X‐ray microanalysis, immunocytochemistry and high resolution autoradiography. © 1993 Wiley‐Liss, Inc.
Sequential observations followed by acid etching on the enamel surfaces of human teeth under scanning electron microscopy at low vacuum
Authors: Kodaka Tetsuo , Mori Ryoichi , Miyakawa Michiyo
Published: February 2005

A scanning electron microscope equipped with a low vacuum specimen chamber and a Robinson's backscattered electron detector was employed to observe the natural surfaces of human buccal enamel before and after 30 percent phosphoric acid etching sequentially up to 90 sec at the same sites with no coatings. Furthermore, successive etching patterns were compared between deciduous and permanent teeth. On the imbrication lines of young permanent teeth, prismend pits surrounded with a “prismless” structure occasionally disappeared after acid etching and became a prismless enamel. Sequential etching caused the prismless areas and the areas of a type 1 etching pattern to decrease, and a cone‐shaped prism structure and a complex type of the type 1 and type 2 etching pattern (type 1–2) to appear. The former was a transitional type between the prismless enamel and type 2 prisms. These etched surfaces show type 2 prisms after deeper etching. Small dome‐shaped structures, slightly elevated on the attrited enamel surfaces, were found only in deciduous teeth. After acid etching, such areas which retained the prismless enamel rose to the underlying surfaces of cone‐shaped prisms. © 1993 Wiley‐Liss, Inc.
Cross‐sectional AEM preparation technique for ceramic‐coated WC‐Co cutting tools
Authors: Ostreicher Kim , Sung Changmo
Published: February 2005

The preparation of cross‐sectional specimens for AEM studies of materials such as ceramic coated tungsten carbide presents some unique problems. Pieces joined by the use of epoxies often separate at the interface between the WC and ceramic coating during the initial mechanical grinding and subsequent thinning process as a result of the vibration and physical strain placed on the sample. These problems have been overcome through the use of a preparation process which essentially encapsulates the sample within the confines of an epoxy filled quartz tube. This preparation process has allowed for facile AEM cross‐sectional analysis of TiN/TiCN coatings on WC‐Co substrates, and has revealed two distinct grain morphologies within the TiCN coating. © 1993 Wiley‐Liss, Inc.
Discrepancies in kinematic calculations of HOLZ lines
Authors: Eades J. A. , Moore S. , Pfullmann T. , Hangas Jon
Published: February 2005

We have found significant differences between the results of computer simulations of HOLZ line patterns. The computations in question are made in the kinematical approximation. After trivial errors are eliminated the programs fall into two groups. There is a discrepancy between the two that increases with distance from the zone axis. The difference is small but not negligible at the level of precision used in determining lattice parameters or strain.

We show which of the two is correct in the kinematic approximation and that the discrepancy between the two groups is of the order of the error introduced by dynamical interaction. © 1993 Wiley‐Liss, Inc.
Quantitative analysis and cartography in scanning electron microscopy: Application to the study of bacterial adhesion to respiratory epithelium
Authors: Colliot G. , De Bentzmann S. , Plotkowski M. C. , Lebonvallet S. , Puchelle E. , Bonnet N.
Published: February 2005

This paper describes the coupling between a scanning electron microscope (SEM) and an image analysis workstation. The system was designed in order to drive the SEM and to analyse any sample. It allows automatic (edge detection) or semiautomatic (pointing, marking, drawing) object detection. Two types of data can be obtained: (1) topographical information, such as the location of the object within a region of interest drawn at any magnification of the microscope, or (2) quantitative data, such as morphometric characteristics of objects. In addition, high resolution maps of the section, regions of interest, and objects can be obtained with a laser printer. This software was first applied to quantitate the adhesion of the bacteria Pseudomonas aeruginosa to human respiratory epithelial cells in culture. P. aeruginosa was shown associated with ciliated cells. The second application concerned the study of the distribution of specific carbohydrate residues at the surface of the respiratory cells. The gal residues were revealed using the lectin Ricinas communis agglutinin II, adsorbed to colloidal gold particles. A relationship between the presence of adherent bacteria and labelling was shown. © 1993 Wiley‐Liss, Inc.
mRNA structure, in situ, as assessed by microscopic techniques
Authors: Eberwine James H. , Spencer Corinne , Newell David , Hoffman Andrew R.
Published: February 2005

The secondary and tertiary structure of RNA, in situ, is thought to be involved in distinct functions such as directing association of the RNA with the cytoskeleton, enzymatic activity of some RNAs, and the control of translation. In situ transcription (IST), a procedure by which cDNA is synthesized in situ, has been used to assess mRNA structure in situ using fixed cells or tissues. Distinct banding patterns were noted for mouse and rat POMC. Unique IST banding patterns were observed when an oligonucleotide complementary to a putative POMC stem‐loop structure was used to prime IST. Indeed local changes in banding patterns could be elicited by pharmacological agents which modulate POMC translation. Inhibition of POMC synthesis with NaF or dexamethasone decreased the number of POMC mRNAs in the polysome fractions and increased the intensity of high molecular weight IST‐derived bands. Forskolin, a stimulator of POMC synthesis, had the opposite effect. One mechanism by which translational control is thought to occur is by regulation of ribosome movement down the mRNA by specific binding of cytosolic proteins to RNA structure. Cytosolic protein fractions from AtT20 pituitary cells have been shown to specifically bind to the IST‐predicted RNA structure. These findings suggest that 1) mRNA structure can be assessed in situ, 2) translation may be altered by the secondary and tertiary structure of mRNAs, and 3) a predicted stem‐loop structure exists in situ in the 5′‐end of POMC mRNA. © 1993 Wiley‐Liss, Inc.
Assessment of gene expression and peptide secretion from individual cells
Authors: Wise Phyllis M. , Scarbrough Kathryn , Larson Gregg , Chiu Sufen , Weiland Nancy G. , Lloyd Jonathan M. , Hinkle David A. , Cai Aihua
Published: February 2005

We have developed an assay that allows one to monitor gene expression in and peptide secretion from individual cells. By combining the reverse hemolytic plaque with in situ hybridization, investigators can quantitate simultaneously the level of gene expression and the level of secretion of a peptide. The method can be used in any system in which an appropriate antibody for the reverse hemolytic plaque assay and probes complementary to the mRNA of interest are available. It can be used to monitor the level of mRNA and secretion of the peptide product, or expression of one gene and the secretion of another peptide. In this paper we will describe the major steps of the method. We have used the pituitary lactotroph as a model to demonstrate the power of this technique. However, we believe that this method may be an important approach to answer many questions regarding the cellular and molecular mechanisms that regulate the coupling of peptide secretion and gene expression at the single cell level. © 1993 Wiley‐Liss, Inc.
In situ hybridization in HIV research
Authors: Fox Cecil H. , Cottler‐Fox M.
Published: February 2005

In situ hybridization (ISH) for HIV is an arduous, demanding means of detecting viral genetic material in cells and tissues. Good ISH requires broad technical skills and devotion to controls for every step of the process as well as a critical eye when interpreting results. ISH may be used to detect HIV in three ways: by hybridizing to viral RNA, by hybridizing to proviral mRNA being produced for virion packaging, and by hybridizing to proviral DNA in the cytoplasm or integrated in the nucleus of an infected cell. Here we discuss the technical considerations involved and the problems encountered in using ISH to study the pathobiology of HIV infection. Published 1993 Wiley‐Liss, Inc.
Characteristics of uterine aging
Authors: Mulholland Joy , Jones Carolyn J. P.
Published: February 2005

Uterine aging is in part responsible for a decline in fecundity which begins in middle age in most mammals. Characteristics of uteri from a variety of animals in middle age and old age are reviewed and the factors which may be responsible for this decline discussed. These include age‐related changes in the hypothalamus, pituitary, and ovaries; loss of number or function of steroid hormone receptors; morphological changes in the uterine epithelium; the accumulation of collagen fibrils in the uterine stroma; and loss or impairment of the decidual response. The ultra‐structural morphology of uterine epithelial, stromal, and decidual tissue from 4 month old and 18 month old Fischer 344 rats is presented and compared. © 1993 Wiley‐Liss, Inc.
Cytokines and growth factors in implantation
Authors: Haimovici Florina , Anderson Deborah J.
Published: February 2005

A variety of cell types at the blastocyst implantation site produce cytokines and growth factors that could play an important role in the implantation process. Furthermore, receptors for cytokines and growth factors have been detected on embryonic and trophoblastic cells. The purpose of the article is to review the published literature on the effect of cytokines and growth factors on implantation events, and to present recent data from our laboratory on effects of growth factors and cytokines on mouse blastocyst implantation events in vitro. © 1993 Wiley‐Liss, Inc.
A method to detect variations in the wetting properties of microporous polymer membranes
Authors: de la Parra Robin E.
Published: February 2005

With the ability to perform dynamic experiments in the environmental scanning electron microscope (ESEM), the evaluation of microporous polymer membranes via a scanning electron microscope has advanced beyond morphological and elemental analysis. By adjusting sample temperature and environmental chamber pressure, the process of condensing water onto the porous membrane surface can be achieved. In doing so, assessments about the uniformity of wetting in hydrophilic membranes can be obtained based on how the liquid water spreads. Variations in the shape of condensed water droplets formed on non‐water wetting structures will reflect the degree of hydrophobicity. This technique has proven useful in the characterization of hydrophobic spots on chemically modified hydrophilic structures and the dynamic examination of irregular wetting patterns in naturally hydrophilic membranes. © 1993 Wiley‐Liss, Inc.
The esem used to image crystalline structures of polymers and to image ink on paper
Authors: Rask James H. , Flood John E. , Borchardt John K. , York Greg A.
Published: February 2005

This article describes two cases in which the advantages of the ESEM have been exploited in unanticipated ways. First, we have found that etching occurs as the electron beam scans the surface of uncoated polymers in the ESEM. The surface topography caused by this etching, as seen in ESEM images, reflects the morphology of crystalline structures in the polymers. This technique has been valuable in the study of such textures in polymers. The second application is related to our use of the ESEM in support of research on the deinking of paper. In this effort we have learned that an unconventional contrast mechanism can be used during ESEM imaging to distinguish between inked and non‐inked areas of newsprint. Under usual operating conditions, ESEM imaging does not distinguish between inked and non‐inked areas. However, at relatively low sample chamber pressures the non‐inked areas appear brighter than inked areas in ESEM images. © 1993 Wiley‐Liss, Inc.
Environmental scanning electron microscope (ESEM) evaluation of crystal and plaque formation associated with biocorrosion
Authors: Geiger Steve L. , Ross Timothy J. , Barton Larry L.
Published: February 2005

The biofilm attributed to Desulfovibrio vulgaris growing in the presence of ferrous metals was examined with an environmental scanning electron microscope. This novel microscope produced images of iron sulfide colloids and other iron containing structures that had not been reported previously. A plaque composed of iron sulfide enveloped the surface of the corroding metal while crystals containing magnesium, iron, sulfur, and phosphorus were present in the culture where corrosion was in progress. A structure resembling the tubercule found in aerobic corrosion was observed on stainless steel undergoing biocorrosion and the elements present in this structure included sulfur, iron, chloride, calcium, potassium, and chromium. © 1993 Wiley‐Liss, Inc.
Copper/Solder intermetallic growth studies
Authors: Kirchner Kevin W. , Lucey George K. , Geis James
Published: February 2005

Copper samples, hot solder (eutectic) dipped and thermally aged, were cross‐sectioned and placed in an environmental scanning electronic microscope (ESEM). While in the ESEM the samples were heated for ∼ 2.5 h at 170°C to stimulate the growth of additional Cu/Sn intermetallic compound. The intent of the study was to obtain a continuous real‐time videotape record of the diffusion process and compare the observations to static SEM images reported to represent long‐term, naturally aged intermetallic growth. The video obtained allows the observation of the diffusion process and relativistic growth phenomena at the Cu, Cu3Sn, Cu6Sn5, and solder interfaces as well as effects on the bulk Cu and solder. Effects contrary to earlier reports were observed; for example, growth rates of Cu3Sn were found to greatly exceed those of Cu6Sn5. © 1993 Wiley‐Liss, Inc.
Copper thick film sintering studies in an environmental scanning electron microscope
Authors: Link Leslie F. , Gerristead William R. , Tamhankar Satish
Published: February 2005

The significance of the ElectroScan environmental scanning electron microscope (ESEM) as a processing tool for studying dynamic morphological changes under controlled temperature/atmosphere conditions was evaluated. The ability to observe dynamic processes in situ, which cannot be achieved by other means, is critical to understanding microstructural formation.

Processing of printed copper thick films on ceramics was used as a test case, wherein morphological changes associated with the steps of organic binder removal and sintering of copper particles were observed/examined in real time. Good agreement was seen between microstructures obtained in the ESEM and those achieved in a belt furnace when similar process variables were used. When processed in atmospheres which were proven to induce sintering in a conventional belt furnace, sintering was evident in both cases, and the microstructural changes were documented on videotapes in real time. Determination of critical event temperatures was achieved—that is, binder burnout occurring between 270° and 350°C, onset of oxidation at 520°C, and sintering starting at 770°C.

It was thus verified that the microstructural changes during the copper thick film sintering process can be observed in situ using an ESEM. © 1993 Wiley‐Liss, Inc.
Evidence for stimulus access to taste cells and nerves during development: An electron microscopic study
Authors: Mbiene Joseph Pascal , Farbman Albert I.
Published: February 2005

We have examined developing taste buds in fungiform papillae of rats from the 18th day of gestation (E18) to the 15th postnatal day (P15). Nerve processes were seen in the epithelium of E18 rats before taste buds were obvious. At E20, early taste buds were visible, but were embedded within the epithelium, i.e., their cells were shielded from the oral cavity by overlying squamous epithelium. At this stage, the epithelium on the lateral aspects of the fungiform papillae was keratinized, but that overlying the taste bud was not. Some taste bud cells at E20 contained synapse‐like structures near their contacts with nerve processes. In postnatal animals, keratinized epithelial cells were seen overlying taste buds, but taste pores were not observed until P10. How, then, do stimuli reach the taste cells and elicit physiological and behavioral responses as reported by others? The keratinized epithelium overlying the buds was unlike that on the lateral aspect of the papilla in at least one significant way. Few lamellated bodies were present in intercellular spaces beneath the stratum corneum, whereas these were abundant in the corresponding location within epithelium on the slope of the papilla. Although some were present within the squamous epithelium overlying the bud, they apparently were not released into the intercellular space. These lipid‐rich lamellated bodies are thought to represent the water barrier of the epithelium, i.e., the barrier which prevents aqueous solutions from passing through the epithelium. We determined that the keratinized epithelium overlying the taste bud was permeable to a tracer, lanthanum nitrate, whereas that on the lateral surface was not. Lanthanum was visualized around taste cells and around nerve profiles within and near the taste bud. We propose that the absence of an aqueous permeability barrier in the epithelium overlying taste buds likely explains the ability of tastants to reach the taste bud cells and nerves in the developmental period before pore formation. © 1993 Wiley‐Liss, Inc.
Pre‐ and postnatal development of rabbit foliate papillae with special reference to foliate gutter formation and taste bud and serous gland differentiation
Authors: Fujimoto Sunao , Yamamoto Koji , Yoshizuka Mitsuaki , Yokoyama Mitsuru
Published: February 2005

The epithelial downgrowth arises in the presumptive foliate papilla region of the tongue approximately at prenatal day 22, and the distal portion of the primary epithelial cell cords transforms to the bifurcated serous gland and excretory duct between prenatal day 30 and postnatal day 2. The excretory ducts extend upward through the primary epithelial cell cords at rather random intervals and are open to the tongue surface between postnatal days 1 and 2. In this process, successive connections between the adjacent ascending excretory ducts occur mainly due to desquamation of the keratinized lining cells of the ducts, resulting in the formation of the foliate gutter.

Taste bud primordia appear in the primary epithelial cell cords by nerve penetration through the basal lamina from prenatal day 30 and nearly fully formed taste buds facing the ascending excretory ducts have been already observed before the foliate gutter formation is completed.

In the differentiation process of chemoreceptor (type III) cells from less‐differentiated basal (type IV) cells, subsurface cisterns of endoplasmic reticulum are occasionally present where nerve endings make contact. Since subsurface cisterns have been considered to be involved in reciprocal or efferent synaptic transmission, it is reasonable to consider that these morphological specializations may take a role in neurotrophic signals for differentiation of the chemoreceptor cells. © 1993 Wiley‐Liss, Inc.
Morphometric and immunocytochemical assessment of fungiform taste buds after interruption of the chorda‐lingual nerve
Authors: Oakley Bruce , Lawton Anne , Riddle David R. , Wu Lan‐Hsin
Published: February 2005

Unilateral interruption of the chorda‐lingual nerve led to a loss of most epithelial axons and to the deterioration of fungiform taste buds in the anterior portion of the tongue of albino rats, mongolian gerbils, and golden hamsters. By three weeks after surgery the following percentages of fungiform taste buds had completely disappeared: 71% in gerbils, 28% in rats, and 26% in hamsters. Residual taste buds were classified into two groups: atrophic taste buds and taste bud remnants. Atrophic taste buds were smaller than normal and typically had no visible taste pore, although they retained the characteristic oval shape of a taste bud and numerous elongated cells. Taste bud remnants were non‐oval fragments of taste buds with few elongated cells. Specific markers for elongated taste cells (monoclonal antibodies to keratin 19) confirmed that atrophic taste buds, as well as some taste bud remnants, had elongated taste cells. By 180 days after chorda‐lingual nerve transection, 44% of rat fungiform taste buds had disappeared; morphometric analysis of the 311 residual taste buds established that 241 atrophic taste buds and 69 taste bud remnants were, respectively, 50% and 75% smaller than the average volume of 480 normal taste buds. The aggregate loss of gustatory tissue, calculated from the shrinkage of residual taste buds and the volume lost by the outright disappearance of many taste buds, was 88% for gerbils, 72% for rats, and 65% for hamsters. Evaluation in gerbils of the co‐occurrence of taste buds and axons suggests residual taste buds were neurotrophically supported. Every gerbil fungiform papilla that lacked axons lacked a taste bud. Every fungiform papillae that had a residual taste bud had axons; axons were absent from 22% of empty fungiform papillae. Diminished numbers of gustatory neurotrophic axons could account for both the loss of fungiform taste buds and the reduced volume of residual taste buds. © 1993 Wiley‐Liss, Inc.
Organization of the nucleus of the solitary tract in the hamster: Acetylcholinesterase, NADH dehydrogenase, and cytochrome oxidase histochemistry
Authors: Barry M. A. , Halsell C. B. , Whitehead M. C.
Published: February 2005

The distribution of acetylcholinesterase (AChE), NADH dehydrogenase (NADHd), and cytochrome oxidase (CO) was determined in the nucleus of the solitary tract (NST) in the golden hamster. Histochemical staining was compared to cytoarchitectonic subdivisions of the NST (Whitehead: J. Comp Neurol. 276:547–572, 1988) and to terminal fields of primary afferents of the nerves that innervate the tongue. These three histochemical methods resulted in differential staining patterns within the NST that were related to certain subdivisions. Transganglionic transport of horseradish peroxidase (HRP) was used to determine the central projections of the chorda tympani (CT), the lingual branch of the trigeminal (L‐V), and the lingual‐tonsilar branch of the glossopharyngeal nerves (L‐IX). Alternate or the same brain sections were processed to reveal transported HRP, and NADHd or AChE levels. Increased staining of the neuropil with NADHd and AChE was coincident with the dense part of the afferent terminal fields of all three nerves in the NST and the laterally adjacent dorsomedial part of the spinal trigeminal nucleus. CO showed this pattern only for the most rostral part of the CT field. The densest AChE staining coincided with gustatory afferent terminal fields. The histochemical staining facilitated the interpretation of the organization of the NST. For example, at caudal levels of the gustatory NST, it is suggested that taste processing is localized predominately in the medial part of the rostral central, and somatosensory processing in the rostral lateral subdivision. AChE or NADHd staining should facilitate studies of connections, topography, and neuroplastic changes of the gustatory NST. © 1993 Wiley‐Liss, Inc.
Advantages of digitonin extraction to reveal the intracellular structure of rat glomerular podocytes for high‐resolution scanning electron microscopy
Authors: Temkin Robert J. , So Derek Y. F. , Lea Peter J.
Published: February 2005

Kidneys of anesthetized rats were perfused with digitonin to extract cytosolic proteins of glomerular podocytes so that the remaining intracellular structures could be examined by three‐dimensional stereo high‐resolution scanning electron microscopy (HRSEM). Cytoskeleton, consisting of microtubules and intermediate filaments, was preserved with each applied concentration of digitonin. High concentrations of digitonin (1.0 mg/ml) produced a corrugated appearance in plasma membranes likely due to the formation of digitonin‐cholesterol complexes. At 1.0 mg/ml digitonin, the Golgi complex became vesicularized, and mitochondria were well extracted and their ultrastructure preserved. Lower concentrations of digitonin (0.1 and 0.2 mg/ml) were less disruptive to both the plasma membrane and the Golgi complex. Mitochondria, rough endoplasmic reticulum, coated vesicles, nuclear membrane, and chromatin were well preserved. Extraction with digitonin, at the optimal concentration and perfusion time, simultaneously maintains both the cytoskeleton and membranous organelles inside the cell and provides a method to elucidate the interactions between these two components. Furthermore, digitonin extraction should preserve antigenic sites, thereby allowing the localization of intracellular proteins by backscattered electron imaging of immunogold labels in the scanning electron microscope. © 1993 Wiley‐Liss, Inc.