Death in the taste bud: engulfment of dying taste receptor cells by glial-like Type I cells.

Taste buds comprise 50-100 epithelial derived cells, including glial-like cells (Type I) and two types of receptor cells (Types II and III). All of these taste cells are renewed throughout the life of an organism from a pool of uncommitted basal cells. Immature cells enter the bud at its base, maturing into one of the three mature cell types.

Taste Bud Connectome: Implications for Taste Information Processing.

Taste buds contain multiple cell types, two of which mediate transduction of specific taste qualities: Type III cells transduce sour while Type II cells transduce either sweet, or bitter or umami. In order to discern the degree of interaction between different cell types and specificity of connectivity with the afferent nerve fibers (NFs), we employed serial blockface scanning electron microscopy (sbfSEM) through five circumvallate mouse taste buds. Points of contact between Type II and Type III cells are rare and lack morphologically identifiable synapses, suggesting that interaction between these cell types does not occur via synapses.

Three-dimensional reconstructions of mouse circumvallate taste buds using serial blockface scanning electron microscopy: I. Cell types and the apical region of the taste bud.

Taste buds comprise four types of taste cells: three mature, elongate types, Types I-III; and basally situated, immature postmitotic type, Type IV cells. We employed serial blockface scanning electron microscopy to delineate the characteristics and interrelationships of the taste cells in the circumvallate papillae of adult mice. Type I cells have an indented, elongate nucleus with invaginations, folded plasma membrane, and multiple apical microvilli in the taste pore.

Chemical synapses without synaptic vesicles: Purinergic neurotransmission through a CALHM1 channel-mitochondrial signaling complex.

Conventional chemical synapses in the nervous system involve a presynaptic accumulation of neurotransmitter-containing vesicles, which fuse with the plasma membrane to release neurotransmitters that activate postsynaptic receptors. In taste buds, type II receptor cells do not have conventional synaptic features but nonetheless show regulated release of their afferent neurotransmitter, ATP, through a large-pore, voltage-gated channel, CALHM1. Immunohistochemistry revealed that CALHM1 was localized to points of contact between the receptor cells and sensory nerve fibers.

Quantitative analysis of cardiac tissue including fibroblasts using three-dimensional confocal microscopy and image reconstruction: towards a basis for electrophysiological modeling.

Electrophysiological modeling of cardiac tissue is commonly based on functional and structural properties measured in experiments. Our knowledge of these properties is incomplete, in particular their remodeling in disease. Here, we introduce a methodology for quantitative tissue characterization based on fluorescent labeling, 3-D scanning confocal microscopy, image processing and reconstruction of tissue micro-structure at sub-micrometer resolution.

Electrical stimulation directs engineered cardiac tissue to an age-matched native phenotype.

Quantifying structural features of native myocardium in engineered tissue is essential for creating functional tissue that can serve as a surrogate for in vitro testing or the eventual replacement of diseased or injured myocardium. We applied three-dimensional confocal imaging and image analysis to quantitatively describe the features of native and engineered cardiac tissue. Quantitative analysis methods were developed and applied to test the hypothesis that environmental cues direct engineered tissue toward a phenotype resembling that of age-matched native myocardium.

Three-dimensional modeling and quantitative analysis of gap junction distributions in cardiac tissue.

Gap junctions play a fundamental role in intercellular communication in cardiac tissue. Various types of heart disease including hypertrophy and ischemia are associated with alterations of the spatial arrangement of gap junctions. Previous studies applied two-dimensional optical and electron-microscopy to visualize gap junction arrangements.

Intracellular calcium transients evoked by pulsed infrared radiation in neonatal cardiomyocytes.

Neonatal rat ventricular cardiomyocytes were used to investigate mechanisms underlying transient changes in intracellular free Ca2+ concentration ([Ca2+]i) evoked by pulsed infrared radiation (IR, 1862 nm). Fluorescence confocal microscopy revealed IR-evoked [Ca2+]i events with each IR pulse (3-4 ms pulse⁻¹, 9.1-11.

Design and characterization of a modified T-flask bioreactor for continuous monitoring of engineered tissue stiffness.

Controlling environmental conditions, such as mechanical stimuli, is critical for directing cells into functional tissue. This study reports on the development of a bioreactor capable of controlling the mechanical environment and continuously measuring force-displacement in engineered tissue. The bioreactor was built from off the shelf components, modified off the shelf components, and easily reproducible custom built parts to facilitate ease of setup, reproducibility and experimental flexibility.

The mechanically enhanced phase separation of sprayed polyurethane scaffolds and their effect on the alignment of fibroblasts.

This paper reports a method to fabricate anisotropic scaffolds of tunable porosity and mechanical properties. Scaffolds were fabricated using a computer controlled sprayed phase separation technique. Following fabrication, the sheets were elongated 0, 35 or 70% of their original length to induce varying degrees of scaffold alignment and anisotropy.

Towards modeling of cardiac micro-structure with catheter-based confocal microscopy: a novel approach for dye delivery and tissue characterization.

This work presents a methodology for modeling of cardiac tissue micro-structure. The approach is based on catheter-based confocal imaging systems, which are emerging as tools for diagnosis in various clinical disciplines. A limitation of these systems is that a fluorescent marker must be available in sufficient concentration in the imaged region.

Defining and evaluating wrapping surfaces for MRI-derived spinal muscle paths.

Muscle paths can be approximated in biomechanical models by wrapping the path around geometric objects; however, the process for selecting and evaluating wrapping surface parameters is not well defined, especially for spinal muscles. In this study, we defined objective methods to select the shape, orientation, size and location of wrapping surfaces and evaluated the wrapping surfaces using an error metric based on the distance between the modeled muscle path and the centroid path from magnetic resonance imaging (MRI). We applied these methods and the error metric to a model of the neck musculature, where our specific goals were (1) to optimize the vertebral level at which to place a single wrapping surface per muscle; and (2) to define wrapping surface parameters in the neutral posture and evaluate them in other postures.

Maintenance of rat taste buds in primary culture.

The differentiated taste bud is a complex end organ consisting of multiple cell types with various morphological, immunocytochemical and electrophysiological characteristics. Individual taste cells have a limited lifespan and are regularly replaced by a proliferative basal cell population. The specific factors contributing to the maintenance of a differentiated taste bud are largely unknown.

Sodium channel Na(v)1.6 is localized at nodes of ranvier, dendrites, and synapses.

Voltage-gated sodium channels perform critical roles for electrical signaling in the nervous system by generating action potentials in axons and in dendrites. At least 10 genes encode sodium channels in mammals, but specific physiological roles that distinguish each of these isoforms are not known. One possibility is that each isoform is expressed in a restricted set of cell types or is targeted to a specific domain of a neuron or muscle cell.

Hepatitis B virus X protein colocalizes to mitochondria with a human voltage-dependent anion channel, HVDAC3, and alters its transmembrane potential.

Understanding the mechanism(s) of action of the hepatitis B virus (HBV)-encoded protein HBx is fundamental to elucidating the underlying mechanisms of chronic liver disease and hepatocellular carcinoma caused by HBV infection. In our continued attempts to identify cellular targets of HBx, we have previously reported the identification of a novel cellular protein with the aid of a yeast two-hybrid assay. This cellular gene was identified as a third member of the family of human genes that encode the voltage-dependent anion channel (HVDAC3).

Localization of varicella-zoster virus gene 21 protein in virus-infected cells in culture.

Although four varicella-zoster virus (VZV) genes have been shown to be transcribed in latently infected human ganglia, their role in the development and maintenance of latency is unknown. To study these VZV transcripts, we decided first to localize their expression products in productively infected cells. We began with VZV gene 21, whose open reading frame (ORF) is 3,113 bp.

ETO and AML1 phosphoproteins are expressed in CD34+ hematopoietic progenitors: implications for t(8;21) leukemogenesis and monitoring residual disease.

To study acute myelogenous leukemia 1 (AML1) transcription factor, ETO protein, and t(8;21) AML chimeric AML1/ ETO protein in normal hematopoiesis and in leukemia, we raised rabbit antisera to a bacterially expressed polypeptide containing amino acid residues 1 to 220 of ETO and to synthetic peptides extending from residues 528 to 548 of ETO and 32 to 50 of AML1. The latter was selected to have little chance of cross-reactivity with other members of the PEBP2 alpha family. With affinity-purified reagents, we observed immunofluorescent staining for both AML1 and ETO in the nucleus of HEL, K562, and Kasumi-1 leukemic cell lines, the last from a t(8;21) AML.

The putative membrane anchor protein for yeast Sec7p recruitment.

Proteins required for yeast secretory pathway function have been identified by genetic selection and characterization of the temperature-sensitive secretory (sec) mutants. The use of genetic and biochemical approaches has expanded the catalog of components of the secretory pathway, yet many proteins, especially membrane and lumenal proteins, remain to be identified. Sec7p, one of the original SEC gene products to be described, is required at multiple stages of the yeast secretory pathway in the coating of transport vesicles.

Identification of breakpoints in t(8;21) acute myelogenous leukemia and isolation of a fusion transcript, AML1/ETO, with similarity to Drosophila segmentation gene, runt.

We have developed a restriction map of the chromosome 21 breakpoint region involved in t(8;21)(q22;q22.3) acute myelogenous leukemia (AML) and have isolated a genomic junction clone containing chromosome 8 and 21 material. Using probes from these regions, rearrangements have been identified in each of nine cases of t(8;21) AML examined.

A developmentally regulated plasmalemmal antigen present in synaptosomes but not in growth cones.

Monoclonal antibody 2L4 was generated against rat synaptosomes but does not cross-react with nerve growth cones. Expression of the 2L4 antigen is developmentally regulated in a manner that is, in part, the opposite of the expression of the 5B4-CAM antigen, a marker of neuronal outgrowth belonging to the N-CAM family. While 5B4-CAM appears and increases during sprouting and then decreases to reach low levels in the adult, the 2L4 antigen appears only late in development, when neuronal outgrowth ceases, at or around the time of synaptogenesis.

Immunocytochemical localization of endopeptidase-24.11 in the nucleus tractus solitarius of the rat brain.

Recently, it has been hypothesized that the N-terminal portion of substance P (SP), SP(1-7), which results from the action of endopeptidase 24.11 (EC3.4.

Kindling induced changes in calmodulin kinase II immunoreactivity.

The distribution of type II calmodulin kinase (CaM kinase) immunoreactivity was studied in control and septally kindled rat brains. CaM kinase was concentrated in limbic structures, such as the hippocampus, lateral septum and amygdala. Within the hippocampus, the molecular layer of the endal limb of the dentate gyrus, the stratum radiatum, and lacunosum moleculare of CA1 were the most heavily stained regions.

A monoclonal antibody against the slow isoform of skeletal muscle Ca2+-ATPase selectively stains a subpopulation of neurons in the central nervous system.

A monoclonal antibody against the slow isoform of chicken skeletal muscle Ca2+-ATPase recognises, in nervous tissue, analogous membrane proteins which are most concentrated in the microsomal fraction. Histochemically, the immunoreactivity of the antibody is restricted to neurones where the staining is most intense in cell bodies and dendrites, weak in axons and absent from cell nuclei. The expression of the antigen varies greatly between different neuronal populations and is developmentally regulated.

Dark-induced changes in activity and compartmentalization of retinal calmodulin kinase in the rat.

Calmodulin kinase (CaM kinase) activity and immunoreactivity were measured in the cytosol and crude synaptic membranes of light- and dark-adapted rat retinas. Dark adaptation increased the calcium-independent CaM kinase activity 2.7 times and calcium-stimulated activity 3.

Localization of retinal calmodulin kinase.

The localization of calmodulin kinase II (CaM kinase) was studied in the retina by light and electron microscopic immunocytochemistry, and by enzymatic and immunoblot assay of cellular and subcellular tissue fractions. By light microscopy, both mono- and polyclonal antibodies revealed CaM kinase-like immunoreactivity in the inner and outer plexiform regions (synaptic layers), retinal pigment epithelium (RPE), and ganglion cells. The inner nuclear layer and photoreceptor outer segments stained much less intensely, and the outer nuclear layer did not stain.