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Proliferation and differentiation of adult endogenous neural stem cells in response to neurodegenerative process within the striatum.

The ongoing process of neurogenesis in the adult mammalian forebrain suggests the possible capacity for limited self-repair after brain injury. Previously, we have demonstrated that in an animal model of Huntington's disease the neurodegenerative process initiates immediate intensive cell proliferation and differentiation resulting in characteristic enlargement of the subependymal zone (SEZ) of lateral brain ventricles. Now, our interest is focused on the architecture of the neurogenic niche of the SEZ in the identical model, particularly on characteristic features of astrocyte-like cells which are considered to be not only niche cells but also neural stem cells. Our findings prove higher activation of the lateral part of the SEZ (L-SEZ) adjacent to the degenerated striatum compared with the rostral part of the SEZ (R-SEZ). In the activated L-SEZ, niche cells which ensheathe clusters of neural progenitors are of immature astrocytic phenotype because of nestin and vimentin expression (except the expression of glial fibrillary acidic protein). However, the coexpression of all three filaments is not always found. Intermediate filaments also enable us to distinguish the basic shape of astrocytic cells within the SEZ, majority of which resemble protoplasmic rather than fibrillary astrocytes. Furthermore, our results show a wide plasticity of these astrocyte-like cells in immediate response to an extensive pathological process in the brain. These observations are consistent with the fact that adult stem cells undergo different processes in an already mature environment, and therefore can exhibit some specific characteristics unlike the embryonic or fetal neural stem cells.

Adult↗

Induction of metallothionein in astrocytes by cytokines and heavy metals.

Immunohistochemical staining of human and rat brains for metallothionein (MT) using a monoclonal anti-MT antibody (MT45) revealed that protoplasmic astrocytes, which are densely localized in the gray matter, contain high levels of MT. Human U373MG astrocytoma cells were stimulated with interleukin (IL)-1 or heavy metals to produce MT. When expression of MT in U373MG cells was analyzed by Northern blotting or indirect immunofluorescence using the MT45 antibody, it was found that IL-1 (> or = 10 U/ml), CdCl2 (50 microM) and high concentrations of ZnCl2 (500 microM) induced marked biosynthesis of MT.IL-6 (up to 3,000 U/ml) and lower concentrations of ZnCl2 (10-50 microM), however, showed little inducing activity. Hippocampal astrocytes in primary culture produced a relatively high basal level of MT. The MT level increased in response to addition of IL-1 (> or = 10 U/ml), ZnCl2 (50 microM) and CdCl2 (5 microM). However, the increase induced by IL-6 (1,000 U/ml) was not very marked. The in vivo induction of MT in the brain by cytokines is not fully understood. However, our data and other indirect evidence suggest that IL-1 may be a potent stimulator of MT induction in astrocytes. Furthermore, the astrocytoma cell line, U373MG, is a suitable in vitro system to analyze the expression of MT in astrocytes.

Animals↗

Metallothionein in amyotrophic lateral sclerosis.

Amyotrophic lateral sclerosis (ALS) is a neurodegenerative disorder characterized by loss of motor neurons in the spinal cord, brainstem and motor cortex. Ten percent of the cases are familial and these have been linked to point mutations in the gene coding for cytosolic copper, zinc superoxide dismutase. The etiology of sporadic ALS is unknown. To further investigate the possible role of metals in causing the disease, we investigated metallothionein (MT) levels in ALS organs and serum. We previously reported significantly increased MT levels in ALS liver and kidney. These are not reflected in serum MT levels, which are normal in ALS. In ALS spinal cord, MT is expressed in gray matter protoplasmic astrocytes. Induction of MT synthesis in ALS may denote increased metal exposure or may result from increased oxidative stress, as in familial ALS.

Aged↗

Glutamine synthetase: a marker of an astroglial subpopulation in primary cultures of defined brain areas.

Primary cultures from various areas of newborn mouse brain were developed and characterized. Enriched astroglial cultures of the cerebral hemispheres, cerebellum, medulla oblongata and olfactory bulbs contained about 80-90% glial fibrillary acidic protein (GFA) immunolabelled cells. These cultures were composed of a majority of flat, 'protoplasmic-like' cells. The aim of this culture model was used: (1) to study glutamine synthetase (GS) activity during in vitro astroglial development; (2) to consider the hydrocortisone effect on GS activity during both growth and maturation periods, and (3) to determine the development of GS immunoreactivity in the cells and eventual GFA and GS expression in these cells. We observed that GS increased during brain maturation in vivo and in vitro, and that addition of hydrocortisone (1 microM, 48 h) to the culture medium induced varying GS activity depending on the developmental stage and the area. In the four areas studied, the number and intensity of GS-immunolabelled cells reached an optimum between 18 and 30 days in vitro. Only about 50-70% of the cell population was GS positive. Double-labelling experiments showed that three groups of cells coexist whatever the considered area. Two expressed both GFA and GS proteins, the last marker at either a low or a high level, and the third was devoid of GS immunoreactivity. Regional differences in GS-specific activity, GS inducibility and GS immunoreactivity exist in the astroglial population, but the factors responsible for these variations are not yet known.

Animals↗

Plasticity of astrocytes in primary cultures: an experimental tool and a reason for methodological caution.

Astrocytes in primary cultures constitute an exceedingly useful preparation for studies of astroglial development and function. These cells, however, demonstrate a pronounced plasticity in their reactions to culturing conditions. Thus, species and spatiotemporal region of CNS chosen for source of cells, dissociation procedures used, cell density in culture, culture medium chosen, type and/or concentration of serum used (if any) and exposure to dibutyryl cyclic AMP (dBcAMP) may all markedly affect the epigenotype of the cells. This provides an experimental tool for studies of astroglial development and function, but is at the same time a reason for caution in the interpretation of data when such cultured cells are used as models of their in vivo counterparts. About 95% of the cells in cultures used by the authors are positive for astrocyte-specific markers (glial fibrillary acidic protein and glutamine synthetase), and the cells possess presumably astrocytic characteristics such as high potassium permeability. This latter characteristic may be drastically altered by minor changes in culturing conditions. It is likely that an overwhelming majority of the astrocytes in such cultures are protoplasmic in nature. Addition of dBcAMP to the culture medium results in a pronounced morphological and a more modest functional differentiation.

Animals↗

A light and electron microscopic study of the pineal body of the nutria (Myocastor coypus).

Light and electron microscopic studies of the nutria (Myocastor coypus) pineal gland revealed pinealocytes interspersed among glial, vascular, and neuronal elements. Each pinealocyte possessed a single process that terminated within the parenchyma near the perivascular region. The eccentrically located nucleus in these cells contained euchromatic chromatin, a prominent nucleolus, and a highly infolded nuclear envelope. The cytoplasm was rich in mitochondria, Golgi complexes, and glycogen particles. The smooth endoplasmic reticulum (SER) was better developed thant he rough endoplasmic reticulum (RER) and polyribosomes were not abundant. Long profiles of subsurface cisterns constituted prominent cytoplasmic features, and these were most conspicuous in the regions of soma-somatic apposition. The bulbous endings of the pinealocyte processes were filled with clear, round, secretory vesicles. Dense-cored vesicles were rarely observed. Glia reminiscent of protoplasmic astrocytes displayed cytoplasmic processes that enveloped blood vessels, invested the pineal periphery, and intervened among the pinealocytes. They thus seemed to form a barrier between the meningeal capsule and vascular space on the one hand and the parenchyma on the other.

Animals↗

[Etiology and pathogenesis of recurrent erosions of the cornea. A scanning and transmission electron microscopy study].

Scratched off corneae of 2 patients were examined under the scanning electron and the transmission electron microscope. Whereas in the scanning electron microscope the expected picture of the destruction of the epithelium in the upper zones of the cornea could be presented in the 1st case, the histological picture of the 2nd case shows in the transmission electron microscope the reaction of the influence of the noxa to the texture of the epithelial cells, to the protoplasm, and to the nucleus of the concerned epithelial cell.

Adult↗

Intravitreal application of ganciclovir in rabbits: ERG and electron-microscopic findings.

The retinal toxic effect after intravitreal use of ganciclovir has been the subject of our experimental investigation on 20 New Zealand rabbits. Different doses of ganciclovir ranging from 200 to 600 micrograms/0.1 ml were injected intravitreously, and ERG changes were recorded at different time points after drug administration. Finally the animals were killed and the retinal lesions were studied by electron microscopy. According to our experiments, ganciclovir doses of 300-600 micrograms/0.1 ml have a clearly toxic effect on the retina as shown by ERG changes. One month after the injection, the electroretinogram is either extinguished or clearly affected. Noteworthy is the fact that in a dose of 200 micrograms/0.1 ml, the ERG b-wave shows a 20% reduction of its normal amplitude 4 months after ganciclovir delivery. Electron microscopy of the retina revealed in the inner segment of the receptor cells the existence of dilatation and vesiculation of the endoplasmic reticulum in the surrounding nuclear cytoplasm and swelling of the mitochondria with fragmentation of the cristae in several of them. In the outer segment of the photoreceptor cells, degeneration and destruction of the lamellae accompanied by disintegration of the adjacent protoplasm in parts of certain rods and cones appeared. Finally large vacuoles of irregular shape in the synapses and degenerative mitochondria and empty vacuoles in the optic nerve fibers were shown.

Animals↗

Substructure of intercellular junctions in freeze-fractured alveolar-capillary membranes of mouse lung.

The purpose of this study was to examine by freeze-fracture the ultrastructure of intercellular junctions between mouse pulmonary endothelial and epithelial cells, and to relate this fine structure to that deduced from previous physiological and ultrastructural studies using tracer techniques. Junctions between capillary endothelial cells consist of one to three interconnected rows of particles which show occasional discontinuities. Small gap junctions are associated with these rows of particles in the arteriolar end of the capillary bed. At the venular end, the junctions consist of low profile ridges on the protoplasmic fracture (PF) face or complimentary grooves on the exoplasmic fracture (EF) face some of which have a sparse number of associated particles. The vascular junctions are similar to those of vessels in rat omentum and mesentery, and resemble "leaky" junctions described in renal proximal convoluted tubular cells. Tight junctions (zonulae occludentes) between type I pneumocytes or between types I and II pneumocytes consist of a band of interconnecting ridges on the PF face and complimentary interconnecting grooves on the EF face. These continuous epithelial junctions have a structure that is typical of tight occluding junctions. Occasionally zonulae occludentes between type I and type II pneumocytes are discontinuous; this may be the result of cell translocation as pneumocytes are shed into the alveolus. Intravascular perfusion fixation at high pressure (140 cm H2O) had no discernible effect on the structure of endothelial or epithelial junctions. The appearance in our study of freeze-fractured pulmonary endothelial and epithelial junctions reveals clearly the physical basis for the results of ultrastructural tracer and physiological studies which have suggested that it is the alveolar epithelium rather than the endothelium that is the chief permeability barrier to small, water-soluble molecules.

Animals↗

Nanostructures in diatom frustules: functional morphology of valvocopulae in Cocconeidacean monoraphid taxa.

Nanopositioning is one of the most intriguing challenges in nanoelectronics and photonics. As the optoelectronic circuit components become more and more complex, the importance of nanopositioning becomes critical. For example, in single-mode fiber telecommunications it is easy to show that an alignment deviation as small as a few tens of nanometers can yield to 50 percent of loss performance. Diatoms are unicellular plants occurring in almost every aquatic environment. Their main morphological character is the frustule, a silica cell wall that consists of two valves, encasing the protoplasm, joined together by a girdle. The girdle is composed of a series of silica bands (copulae) linked together along their margins. In several diatom species, the first girdle bands (valvocopulae), which associate the valves with the rest of the girdle, appear different in shape and bring specific nanostructures devoted to facilitate this linkage. The species of the diatom family Cocconeidaceae show elaborate linkage systems between the valves in which functionally complex valvocopulae are involved. Different levels of complexity and functional efficiency are recognizable in the valve-valvocopula and valvocopula-valvocopula linkages of different cocconeidacean taxa. This work briefly reviews the morphological characteristics and the functional role of the nanostructures involved in the linkages; in fact, we firmly believe that they are stimulating models for design and manufacturing engineers working in the technological nanoworld.

Diatoms↗

Responsiveness of growth hormone-deficient children to human growth hormone. Effect of replacement therapy for one year.

Previous studies have shown that growth hormone (GH)-deficient children are more responsive to exogenous human growth hormone (HGH) than non-GH-deficient children. In six GH-deficient children, velocity of linear growth was less than 2.5 cm/yr. By the metabolic balance study technique, anabolic responses (increments in elemental balances) were measured to a 7 day course of 0.0532 U HGH/kg body weight (BW)(3/4) per day (dose B) and to 0.168 U/kg BW(3/4) per day (dose C). They were then treated for 1 yr with HGH at a dose intermediate between B and C. Velocity of linear growth accelerated to 15-25 cm/yr for the first 4-7 mo, then declined to 0-8 cm/yr. At 12 mo, responsiveness to doses B and C was measured again; the responses were only 20-60% as great as before treatment. After 3 mo without HGH treatment, responsiveness to the anabolic effects of doses B and C returned to the magnitudes observed before treatment. A low titer of plasma antibodies to HGH was detected in two of the six children at the end of the year's treatment; these titers showed little change after 3 mo without HGH. Thus the hyperresponsiveness of GH-deficient subjects to exogenous HGH, compared to non-GH-deficient individuals, declines during long-term HGH treatment and is restored by 3 mo interruption of treatment. These changes in peripheral responsiveness may be related to the decline in velocity of linear growth which occurs after 4-7 mo of continuous treatment. When HGH was withdrawn after 12 mo, all six patients exhibited negative balances of N, P, Na, and K and loss of BW. Ratios of elemental balances showed about half the weight loss to represent protoplasm, and about half extracellular fluid. These observations indicate a role of GH in the continuing regulation of nitrogen and mineral metabolism in addition to its function as a growth-promoting hormone.

Adenoma, Chromophobe↗

Anabolic actions of reduced and S-carbamidomethylated human growth hormone and its plasmin digest in man.

Six children aged 12-15 yr, deficient in endogenous growth hormone, were each treated, after a 7-day control period, for 7 days with 0.0168, 0.052, and 0.168 U/kg body wt3/4 human growth (hGH) (doses A, B, and C, respectively) in separate metabolic balance studies. Doses B and C caused a dose-related retention of N, P, K, Na, and Cl in ratios of 1/0.069/4.5/7.5/5.6. These ratios indicate increments in masses of protoplasm/extracellular fluid (ECF)/bone in ratios of 1/2.0/ less than 0.001. Three of the children were also treated with doses A, B, and C of reduced and carbamidomethylated hGH (RCAM-hGH). Doses B and C caused 1.2-2.8 times as much retention of N, P, and K, and 0.3-0.5 times as much retention of Na and Cl, as did the corresponding doses of hGH. The plasmin digest of RCAM-hGH gave results generally similar to RCAM-hGH. For RCAM-hGH and its plasmin digest, N, P, K, Na, and Cl were retained in ratios of about 1/0.14/5.4/2.2/2.1, indicating increments of protoplasm/ECF/bone of about 1/0.8/0.05. These findings indicate that reduction and carbamidomethylation alter the anabolic actions of hGH in man in both quantitative and qualitative manner. RCAM-hGH is more potent in stimulating enlargement of protoplasm and bone, and less potent in stimulating expansion of ECF, than is the native hormone. The profile of anabolic actions of RCAM-hGH in man does not appear to be further altered by digestion with plasmin.

Adolescent↗

Virulent strain associated outer membrane proteins of Borrelia burgdorferi.

We have isolated and purified outer membrane vesicles (OMV) from Borrelia burgdorferi strain B31 based on methods developed for isolation of Treponema pallidum OMV. Purified OMV exhibited distinct porin activities with conductances of 0.6 and 12.6 nano-Siemen and had no detectable beta-NADH oxidase activity indicating their outer membrane origin and their lack of inner membrane contamination, respectively. Hydrophobic proteins were identified by phase partitioning with Triton X-114. Most of these hydrophobic membrane proteins were not acylated, suggesting that they are outer membrane-spanning proteins. Identification of palmitate-labeled lipoproteins revealed that several were enriched in the OMV, several were enriched in the protoplasmic cylinder inner membrane fraction, and others were found exclusively associated with the inner membrane. The protein composition of OMV changed significantly with successive in vitro cultivation of strain B31. Using antiserum with specificity for virulent strain B31, we identified OMV antigens on the surface of the spirochete and identified proteins whose presence in OMV could be correlated with virulence and protective immunity in the rabbit Lyme disease model. These virulent strain associated outer membrane-spanning proteins may provide new insight into the pathogenesis of Lyme disease.

Animals↗

Novel neurotransmitters and their neuropsychiatric relevance.

OBJECTIVE: The purpose of this review is to integrate insights regarding novel neurotransmitters or neuromodulators of neuropsychiatric significance. METHOD: Evolving concepts of neurotransmitter criteria are reviewed in light of the unexpected properties displayed by recently identified transmitters. RESULTS: Classic criteria for transmitters were based on the properties of acetylcholine but were markedly revised with the recognition of the catecholamines, serotonin, gamma-aminobutyric acid (GABA), and other amino acid transmitters and neuropeptides. Nitric oxide and carbon monoxide are notably atypical, as they are not stored in synaptic vesicles, are not released by exocytosis, and do not act at postsynaptic membrane receptor proteins. D-Serine, recently appreciated as the endogenous ligand for the glycine site of the glutamate N-methyl-D-aspartate (NMDA) receptor, overturns fundamental axioms of biology as well as those of neuroscience. It is a D-amino acid, and it is synthesized and stored in glia rather than neurons. Released glutamate acts on receptors on the protoplasmic astrocytes closely apposed to the synapse to release D-serine, which coactivates postsynaptic NMDA receptors together with glutamate. D-Serine is formed by serine racemase, which directly converts L-serine to D-serine. Inhibitors of this enzyme should reduce NMDA neurotransmission and might be therapeutic in stroke and other conditions associated with glutamate excitotoxicity. CONCLUSIONS: The diversity of novel neurotransmitters and venues of their activity afford multiple opportunities for therapeutic intervention.

Carbon Monoxide↗

Morphometry of ampullar dark cells.

The morphologic characteristics of rat ampullar dark cells were analyzed quantitatively. Four parameters were measured: cell area per micron baseline length, membrane surface density, volume density of intercellular space, and volume density of vacuoles. No statistically significant difference was observed for any parameter, either between dark cell regions of the superior, lateral, and posterior ampullae or between different regions of dark cells within a given ampulla. The membrane surface density of the ampullar dark cells was similar to that reported for principal cells of the cortical collecting duct in the kidney. The volume density of the intercellular spaces of the dark cells was very constant, unlike that reported for epithelia involved in isotonic fluid transport. Vacuoles were found to comprise only a small portion of protoplasm in the dark cells. This new morphometry provides a basis on which to examine the response of ampullar dark cells to differing experimental conditions.

Animals↗

A contribution to the histological study of hypercementosis using metal staining.

After studying hypercementosis, in detail using metallic staining procedures, the authors reached the following conclusions: (1) The histology of hypercementosis, observed by metallic staining reveals remarkable and characteristic microscopical alterations. (2) The Rio Hortega12 procedures, the mitochondria technique of Polak,10 and the method of Achucarro-Hortega13 are of great value for the study of the morphology of cementum. (3) Among the technical procedures used, we consider the 1916 method of Achucarro-Hortega as affording the best view of cementocytes. (4) The Sharpey fibers, revealing great irregularity in course, shape and size, and frequent interruptions could be clearly observed; heavily stained lacunae were frequently observed along the cementum-dentin border; a direct deposit of cellular cementum on the dentin in the middle third of the root was frequently observed. Curiously, an acellular cementum deposit next to a cellular cementum layer could be observed; we also noted numerous cementocytes of different shapes and sizes, at times exhibiting alterations of a degenerative type, particularly fragmentation of protoplasmic prolongations. (5) The techniques used also showed the frequency of blood vessels in the texture of hyperplastic cementum and the presence of capillaries near cementum-dentin junction.

Collagen↗

Atrial natriuretic peptide-like immunoreactivity in neurons and astrocytes of human cerebellum and inferior olivary complex.

Atrial natriuretic peptide (ANP) has previously been localized in areas of mammalian brain associated with olfaction, cardiovascular function, and fluid/electrolyte homeostasis. Despite the presence of several types of natriuretic peptide receptors in mammalian cerebellum, neither intrinsic nor extrinsic sources of the natriuretic peptides have been described. In this report we describe the immunohistochemical localization of both intrinsic and extrinsic sources for ANP in human cerebellum. ANP-like immunoreactivity (ANP-LIR) was observed in climbing fibers in the cerebellar molecular layer that probably originated from isolated immunopositive neurons of the inferior olivary complex. Intrinsic sources of ANP-LIR included small subpopulations of protoplasmic and fibrous astrocytes and Bergmann glia, as well as Golgi and Lugaro neurons of the granule cell layer. These results suggest that, in addition to its presumptive roles in local vasoregulation, ANP may serve as a modulator of the activity of Purkinje neurons.

Astrocytes↗

Glioneurocytoma: tumor with glial and neuronal differentiation.

We report two cases of low-grade glioma in which multiple cellular components, including cells with dense-core granules consistent with "neurocytes," were identified on electron microscopic studies. The first patient was an apparently normal boy until the onset of seizures at age 10 months. Initially, the seizures improved with phenobarbital treatment, but good seizure control was never achieved. Computed tomographic scan at age 23 months showed a calcified, nonenhancing left parietal mass. This tumor was composed of sheets of cells with clear cytoplasm and round to oval nuclei. Mucinous intercellular material stained positively with periodic acid-Schiff, mucicarmine, and alcian blue stains. Foci of calcification were evident. The second patient was a 13-year-old boy with a left parasagittal parieto-occipital mass who presented with a 4-month history of seizures and declining school performance. The tumor was composed of sheets of astrocytes with dark, hyperchromatic, pleomorphic nuclei in a fibrillary and microcystic background. The tumor contained the pleomorphism seen in the adult variant of pilocytic astrocytoma, as well as the microcystic component seen in the juvenile variety. Ultrastructurally in both cases, there were occasional tumor cells having round to oval nuclei with moderate amounts of cytoplasm containing 150- to 250-nm-diameter dense-core granules. These cells were admixed with the majority of tumor cells, which in case 1 had the ultrastructural features of astrocytes and oligodendrocytes and in case 2 had features of protoplasmic or pilocytic astrocytes.(ABSTRACT TRUNCATED AT 250 WORDS)

Adolescent↗