bluuebunny/crossref_metadata_2025_split huggingface dataset:
Transport of Connexins and Gap Junction Turnover in Living Cells Visualized Using Green Fluorescent Protein Tagged Connexins.
Authors: Laird DW , Jordan K , Fistouris P , Solan JL , Lampe PD , Shao C 
Published: July 2020

Abstract
Connexins oligomerize into hemichannels, traffic to the cell surface and dock with hemichannels from an adjacent cell to form intercellular gap junction channels. Pulsechase studies have revealed that connexins are subject to a short half-life ranging from 1- 5 hrs. To examine the mechanisms involved in connexin trafficking, gap junction assembly and internalization in living cells we fused green fluorescent protein (GFP) to the amino or carboxyl terminus of full length connexins (Cx). When cDNA encoding Cx43-GFP was transfected into communication-competent NRK cells, Cx43-negative MDCK cells, or communication-deficient Neuro2A or HeLa cells, the fusion protein of predicted length was expressed, transported, and assembled into functional gap junctions (Figure 1). Most notably, the fusion of GFP to the amino terminal of Cx43 (GFP-Cx43) or Cx32 (GFP-Cx32) did not inhibit the co-translational insertion of these polytopic type IV connexins into the membrane and as a result gap junction plaques assembled at cellcell interfaces.
Clathrin-Mediated Endocytosis in the Giant Reticulospinal Synapse in Lamprey
Authors: Shupliakov O 
Published: July 2020

Abstract
The chemical synapse is a key structure used by neurons to communicate with neighboring cells. This is an asymmetric structure composed of two elements. The presynaptic part contains small synaptic vesicles filled with neurotransmitter. In an active synapse synaptic vesicles fuse with the presynaptic membrane and release neurotransmitter into the synaptic cleft. This results in activation of receptor molecules and respective ion channels on the postsynaptic cell. After a fusion event, the vesicle membrane is retrieved to form a new vesicle. Clathrin-mediated endocytosis is one of the main pathways for synaptic vesicle recycling. It involves a series of steps that starts with a flat plasma membrane and ends with an internalized coated vesicle. Four distinct stages have been defined: (1) the assembly of the clathrin coat, (2) the invagination of the coated membrane into a pit, (3) the constriction of the coated pit, and (4) the fission reaction that liberates the coated vesicle (e.g. De Camilli and Takei, 1996; Brodin et al., 1997). Analysis of mutant yeast and animal cells have linked a number of proteins to each of these stages. Taken together with data from in vitro binding studies, these observations indicate the existence of a complex molecular network that regulate the endocytic reactions.
Localization of Nicotinic Acetylcholine Receptors on Somatic Spines of Chick Ciliary Ganglion Neurons
Authors: Shoop R , Martone M , Yamada N , Ellisman M , Berg D 
Published: July 2020

Abstract
Nicotinic acetylcholine receptors (nAChRs) are pentameric membrane proteins that function as cation selective, ligand-gated ion channels and are widely distributed throughout the vertebrate nervous system. One of the most abundant nAChRs is a species that contains the α7 gene product (α7-AChRs), binds the snake venom oc-bungarotoxin, and rapidly desensitizes. These receptors have been shown to function at presynaptic sites to modulate transmitter release, as well as on postsynaptic cells where they mediate transmission. Interestingly, these receptors have an exceptionally high relative permeability to calcium.
In the chick ciliary ganglion, the α 7-AChRs play a prominent role, by generating large synaptic currents, but the receptors appear to be excluded from postysnaptic densities on the cell. Immunohistochemical studies have shown that the receptors form large clusters on the surface of the ciliary ganglion neurons. We have recently shown that the α 7-containing receptors are concentrated on mats of somatic spines in close proximity to putative sites of presynaptic transmitter release.
Molecular Mechanisms of Self-Assembly and Polymorphic Switching of the Bacterial Flagellum
Authors: Hasegawa Kazuya , Yamashita Ichiro , Mimori-Kiyosue Yuko , Vonderviszt Ferenc , Namba Keiichi 
Published: July 2020

Abstract
Bacterial flagellum is a helical filament by means of which bacteria swim. Each filament rotated by the motor at its base works as a screw that propels the cell, but it is not simply a rigid propeller. The filament is normally in a left-handed supercoiled form and several of them form a bundle when bacteria swim. Upon quick reversal of the motor rotation, which occurs every few seconds, the filament switches into a right-handed supercoil, making the filament bundle fall apart and enabling the cell to tumble for its tactic behavior. The filament is a tubular structure formed by helical assembly of single protein, flagellin, whose molecular mass is 51.5 kDa in the case of Salmonella typhimurium, which we study. The supercoiling of the filament is thought to involve two distinct subunit conformations and/or packing, whose mechanism is interesting in terms of conformational distinctness and adaptability of flagellin.
To understand the mechanisms of self-assembly and polymorphism of the filament, electron cryomicroscopy (EM) and X-ray fiber diffraction have been used to analyze the structures of two straight filaments with distinct helical symmetries.
Requirement of Prolonged Calcium Increases For the Induction of Nuclear Camca4 Signals
Authors: Teruel Mary N , Meyer Tobias 
Published: July 2020

Abstract
Calmodulin is thought to be the most important calcium sensor in signal transduction. When calcium binds to its 4 calcium binding sites, CaMCa4 becomes a multifunctional activator that regulates a large number of critical cell functions. In the nucleus, CaMCa4 controls gene transcription and other important nuclear processes. Here, using confocal microscopy to measure changes in fluorescence resonance energy transfer (FRET) of an indicator composed of two GFP-variants (CFP and YFP) linked by a calmodulin-binding sequence, we show that free cytosolic CaMCa4 concentration increased rapidly after a cell-wide increase in calcium concentration while the initial free CaMCa4 in the nucleus remained much lower and only equilibrated to the cytosolic CaMCa4 concentration after a delay of 1-2 minutes. Thus, immediately after a cell-wide calcium increase, a significant gradient in free CaMCa4 concentration exists between the cytosol and nucleus.
Increase In Intracellular Sodium Evoked by Glutamate Transporter Activation In Cortical Astrocytes: A Fluorescence Microscopy Analysis
Authors: Chatton J-Y , Marquet P , Magistretti PJ 
Published: July 2020

Abstract
Primary cultures of mouse cortical astrocytes were used to investigate the changes in intracellular Na+ concentration ([Na+]i) following exposure to the excitatory neurotransmitter glutamate. The fluorescent probe sodium-binding benzofuran isophthalate (SBFI) was used to measure [Na+]i by epifluorescence microscopy imaging. Detection of the low light level fluorescent signal was accomplished using a Gen III+ intensified CCD camera. Fluorescence excitation ratio images of dye-loaded cells were obtained after sequential illumination at 340nnm and 380nm, with an emission observed at &gt;520nm. Ratio images were proportional in intensity to [Na+]i. In situ calibration of the fluorescent signals was obtained for each experiment and each cell under study by equilibrating [Na+]i with external Na+ after treatment with the cation ionophores monensin and gramicidin, and with ouabain, a specific inhibitor of the Na+/K+ ATPase. Cells on glass coverslips were perfused at 35°C in a closed microscope chamber using solutions buffered with 25 mM bicarbonate/5%CO2.
Quantitative Imaging of Metabolism by Two-Photon Excitation Microscopy
Authors: Piston David W , Knobel Susan , Patterson George 
Published: July 2020

Abstract
Two-photon excitation microscopy provides attractive advantages over confocal microscopy for three-dimensionally resolved fluorescence imaging. Because of the intensity-squared dependence of the two-photon absorption, the excitation is limited to the focal volume. This inherent localization minimizes photobleaching and photodamage - the ultimate limiting factors in fluorescence microscopy of living cells. One of the most powerful applications of two-photon excitation microscopy is imaging from the naturally occurring reduced pyridine nucleotides (NAD(P)H). NAD(P)H is a useful indicator of cellular metabolism, but it is not a “good“ fluorophore (it has a small absorption cross-section and a low quantum yield). Two-photon excitation of NAD(P)H yields minimal photodamage, thus allowing time-resolved threedimensional metabolic mapping of cellular redox state. We have used two-photon excitation microscopy to examine glucose metabolism in pancreatic and muscle cells. As glucose is metabolized by these cells, intermediate metabolism results in an increase in the reduced-tooxidized NAD(P)H/NAD(P)+ ratio, and a concomitant increase in autofluorescence.
Phytofluors: Phytochrome-Based Orange Fluorescent Protein Probes
Authors: Clark Lagarias J , Montgomery Beronda L , Murphy John T , Wu Shu-Hsing 
Published: July 2020

Abstract
Plants sense the light environment using pigment-protein complexes that discriminate light color, intensity, duration and direction. The most well-studied of these photoreceptors are the phytochromes, a family of soluble biliproteins found in plants, algae and cyanobacteria. Owing to the linear tetrapyrrole pigment phytochromobilin (PΦB) or phycocyanobilin (PCB) that is covalently linked to a large polypeptide via a thioether linkage, phytochromes perceive differences in the quality and quantity of light via their ability to photointerconvert between red (λmax660 nm) and far-red (λmax730 nm) light absorbing forms. Due to an efficient Z,E photoisomerization of the double bond between the C and D-ring pyrroles, phytochromes are nonfluorescent proteins with fluorescent quantum yields less than 10“3 at room temperature (Figure 1).
Phytochrome genes have been cloned from a wide variety of photosynthetic organisms.
Submembrane Events In Triggerable Cells Studied by Total Internal Reflection Fluorescence Microscopy.
Authors: Axelrod D , Johns LM , Levitan ES , Omann GM , Holz RW 
Published: July 2020

Abstract
We have studied the dynamics of certain key processes near the plasma membrane inside two types of chemically-triggerable living cells using total internal reflection fluorescence microscopy (TIRFM). In TIRFM, a laser beam is incident upon the cell/glass-substrate interface from the glass side at an angle greater than the critical angle for total internal reflection. This creates an exponentially decaying evanescent field in the cell medium (with a characteristic depth of &gt; 100 nm) capable of exciting fluorescence selectively from the membrane-proximal regions at cell/substrate contacts. Various ways of setting up the optics for such a system are discussed, involving the use of either prisms or very high aperture objectives.
In one application of TIRFM, the motion of adrenalin-containing secretory granules in the immediate submembrane region of chromaffin cells is examined before and after chemical stimulation that causes the granules to release their contents to the cell exterior.
Imaging Subcellular Changes in Living Mammalian Embryos Using 1047 Nm Two Photon Excitation Fluorescence Microscopy
Authors: Squirrell J M , Wokosin D L , Bavister B D , White J G 
Published: July 2020

Abstract
A major challenge for fluorescence imaging of living cells is maintaining viability during and following prolonged exposure to excitation illumination, especially when imaging over hours or even days, as when studying mammalian embryonic development. The use of specific fluorescently labeled components in living embryos promises to reveal the roles of organelles and molecules in a native and reproducible context. However, to gain a thorough understanding of dynamic biological systems, events of interest must be recorded as they occur, while limiting perturbations caused by the observation technique. Therefore, establishing long-term fluorescence imaging methods that maintain viability is critical for advancing our understanding of cell and developmental biology.
One promising technique for imaging living cells is two photon laser scanning microscopy (TPLSM). The lower energy per photon and the restriction of fluorophore excitation to the imaged focal plane should reduce the total photodamage to thick specimens when compared to conventional laser scanning confocal microscopy (LSCM).
Two-Photon Microscopy as a Useful Tool for Elucidating the Mode of Action of Novel Potential Cancer Therapeutics.
Authors: Navara C S , Uckun F M 
Published: July 2020

Abstract
Multiphoton microscopy allows us to follow specific cellular structures (ie. nuclei, mitochondria, and the plasma membrane) in a high resolution three dimensional and time resolved fashion. We have used multiphoton confocal microscopy to evaluate the cellular effects of potential cancer therapeutics. Cancer cells were plated onto coverslips and labelled with either the vital DNA dye Hoechst to image the nuclei and chromosomes or the lipid dye DiA to label the plasma membrane. Both of these dyes can be imaged with several sections without affecting cell viability as assessed by cell motility and the ability to undergo mitosis and cytokinesis.
We have used this technology to show that one compound (DDE 131) rapidly induces in cancer cell lines the hallmarks of apoptosis; DNA hypercondensation, nuclear fragmentation and rapid membrane blebbing. In a leukemic cell line these changes take place within 20 minutes of treatment.
Imaging Techniques Combined With Enzyme-Based Procedures That Facilitate the Simultaneous Imaging of Nerves, Neurons, Cartilage And Bones During Development In Gambusia
Authors: Rosa-Molinar E 
Published: July 2020

Abstract
A persistent problem in elucidating the anatomy of the peripheral nervous system has been an inability to stain both myelinated and unmyelinated nerve fibers. To overcome this problem, our laboratory developed two workmg protocols for reliably and differentially labeling and staining the peripheral nervous system and combined them with an enzyme clearing and staining procedure for the simultaneous visualization of bone and cartilage.
One protocol uses anti-acetylated α-tubulin immunohistochemistry to follow the course, peripheral branching, and origin of the ventral spinal nerve innervating the axial musculature and a second uses anterograde and retrograde transport of selectively applied 3000 molecular weight (MW) biotin dextran amines and/or biocytin to identify specific afferent and efferent projections and their cell bodies. Both procedures can be combined with an enzyme clearing and staining procedure for the simultaneous visualization of bone (alizarin red S) and cartilage (alcian blue) in whole-mount preparations.
Approaches for Understanding Dynamic Cell Movements, Cell-Cell Interactions and Tissue Shaping During Embryogenesis of the Vertebrate Body Plan
Authors: Schoenwolf GC 
Published: July 2020

Abstract
The early vertebrate embryo develops a characteristic tube-within-a-tube body plan. This plan is realized through a series of cell movements and cell-cell interactions that collectively result in tissue shaping and the formation of the three-dimensional body plan. Tissue shaping is a highly choreographed process that is under the control of the organizer--a specialized region of the embryo that is both sufficient and required for formation of the body plan. Recent technical advances have greatly increased our understanding of the role of the organizer in vertebrate embryogenesis. Such advances include the use of new cellular, molecular, genetic, and embryological approaches.
A hallmark of embryogenesis is its dynamic nature. Classically, embryos were studied in three major ways. 1) With morphological/descriptive analysis, initially involving histological procedures (stained whole mounts and serial sections cut in the three cardinal axes) and more recently electron microscopy.
Microscopic Characterization of Ocular Development and Disease
Authors: Potts J , Conley P , Champion R 
Published: July 2020

Abstract
The embryonic lens contains two populations of cells, lens epithelial cells and lens fiber cells. As the growth of the human lens is linear from the age of 10 to 90, lens epithelial cells divide and differentiate into fiber cells at a uniform rate during adulthood. Any defects that arise during fiber formation will remain in the lens. We postulate that regulation of early lens growth is critical for the onset of cataracts that usually occur later in life. Slowing the growth of the lens could therefore provide a strategy to suppress the formation of cataracts. However, the factor(s) that control lens growth in vivo have not been identified.
We have identified 18 receptor and cytoplasmic kinases present in lens epithelial cells. Using these as potential targets for factors controlling growth, we tested corresponding ligands in a lens epithelial cell proliferation assay. Platelet-derived growth factor (PDGF) proved to promote a growth activity similar to, but distinct from, that of embryo serum.
Microscopy as a Tool in Understanding the Role of Growth Factors in Cardiac Development
Authors: Price Robert L , Thielen Thomas E , Borg Thomas K , Terracio Louis 
Published: July 2020

Abstract
Over the past several years the central roles of several different growth factors in the normal development of the embryonic heart have been identified through a variety of techniques involving microscopy. Initially, most studies consisted of descriptions of gross changes in cardiac morphology associated with naturally occurring mutations that affected growth factor function. More recently the development of specific probes for growth factor receptors that can be used in confocal microscopy have aided in the identification of changes in the temporal and spatial distributions of receptors at various stages of development. The correlation of these changes with developmental events such as valve formation and trabeculation in the heart, in conjunction with biochemical studies and blocking agents for the growth factors have significantly increased our understanding of growth factor function in cardiac development.
Confocal Laser Scanning Microscopy of Whole Mouse and Human Embryos: Apoptosis and Morphology.
Authors: Robert M. Zucker , Kathy Sulik , Price Owen T 
Published: July 2020

Abstract
Two novel sample preparation procedures have been developed to visualize morphological structures in embryonic and fetal tissues. In the first, optimization of the sample preparation allows for the visualization of embryos with a thickness in excess of 500 microns. The morphology of the internal structures in these embryos is observed by optical sectioning. The general sample preparation procedures included paraformaldehyde fixation, methanol dehydration and clearing with either benzyl alcohol/benzyl benzoate or methyl salicylate. The dyes (i.e. LysoTracker Red, YoPro) are incorporated into living or fixed embryos to indicate a specific biological function or embryo morphology. Using these stained embryos, the detection of apoptosis and its subsequent quantification were made in addition to the observation of morphology.
In the second technique, the confocal microscope has been adapted to produce images similar to those generated by a scanning electron microscope. For this application, the embryos are coated with a dilute solution of acridine orange resulting in opaque surface fluorescence.
Aids Dementia Complex
Authors: DeArmond SJ 
Published: July 2020

Abstract
The World Health Organization estimates by the year 2000 that the cumulative total of HIV-1 infected adults and children will be about 30 million worldwide. Disease of the central nervous system (CNS) in association with AIDS is common. 20-30% of AIDS patients will develop varying degrees of cognitive and motor deficits designated the “AIDS dementia Complex (ADC)” or “HIV-1-associated cognitive/motor deficit”. Progression of symptoms is usually slow and irregular. In the end-stage, the patient is severely impaired with mutism, incontinence and prominent motor impairment. Neuroradiologic studies show generalized cortical atrophy and diffuse white matter changes. HIV can be cultured from the CSF. Neurologic involvement in pediatric AIDS is very common with CNS complications usually occurring within the first two postnatal years. Given the severeness of the dementia and motor dysfunction in ADC, one is struck by the paucity of neuropathological features. Hematoxylin and eosin stained sections reveal only three neuropathological features. First, there are small numbers of microglial nodules scattered in the white matter and sometimes in grey matter (Fig. 1). However, microglial nodules occur in virtually all viral infections of the CNS, including CMV micronodular encephalitis in AIDS patients.
Aids: Tubuloreticular Structures (Trs), Cylindrical Confronting Cisternae (Ccc), and Related Alterations
Authors: Sidhu Gurdip S , Cassai Nicholas D 
Published: July 2020

Abstract
TRS and CCC are endoplasmic reticulum membrane-derived structures seen in HIV-infected individuals in a variety of cell types. TRS (synonym: tubuloreticular inclusions) are 24-25 nm in diameter, branching tubules which are short or long and associated with the granular or smooth endoplasmic reticulum, the Golgi complex, the perinuclear cistern, and annulate lamellae (Fig. 1-3, 8). The tubules are noticeably more stretched out in Kaposi's sarcoma endothelial cells, presumably by an increase of matrix material within the reticulum sac (Fig. 3). TRS consist of membranous components, including polypeptides, but lack nucleic acid.
CCC (test tube and ring-shaped forms; curvilinear membranes) are formed by a concentric stacking of two or three cisterns of endoplasmic reticulum resembling nuclear membrane in mitosis (Fig. 7), but with the interposition between the opposing membranes of a layer of electron-dense material that is resistant to lipid solvents (Fig. 4,5,8).
Early Apoptotic Changes in the Leukocytes of Rhesus Macaques Challenged With Chimeric Simian/Human Immunodeficiency Virus (Shiv)
Authors: Bryant NJ , Asher LV , Lewis M , Mascola J , Carpenter C , Hanson C 
Published: July 2020

Abstract
Introduction
Chimeric Simian/Human Immunodeficiency Viruses (SHIV) causes persistent infection characterized by CD4+ T cell lymphopenia in rhesus monkeys. The SHIV model infection closely resembles HIV-1 infection and disease in humans, except that it progresses at an accelerated rate. During the early stages of HIV-1 infection, viral replication is at its highest rate and there is a rapid decline of the immune system. Cell death induced by apoptosis, a feature in the pathogenesis of acquired immunodeficiency syndrome (AIDS) and AIDS-like syndromes of many lentiviral infections, is thought to be important in the progressive decline of lymphocytes and the cell mediated immunity of infected individuals. The apoptosis is not specific for lymphocytes, this phenomenon also occurs in numerous cell types of the immune system, including the polymorphonuclear leukocytes. Abnormalities of neutrophil function and number because of HIV-1 and other retroviral infections have been recognized.