bluuebunny/crossref_metadata_2025_split huggingface dataset:
Glutamate receptor‐linked changes in membrane potential and intracellular Ca2+ in primary rat astrocytes
Authors: Enkvist M.O. Kristian , Holopainen Irma , Åkerman Karl E.O. 
Published: January 2005

AbstractKainate‐, quisqualate‐ and glutamate‐induced depolarization and mobilization of intracellular Ca2+ was determined in primary cultured astrocytes using the fluorescent probes DiBa‐C4‐(3) and fura‐2, respectively. All three receptor agonists depolarized the cells in a Na+‐dependent manner and increased the intracellular Ca2+ concentration. The glutamate‐ and quisqualate‐induced increase in cytosolic Ca2+ was only partially inhibited by removal of extracellular Ca2+, whereas the response to kainate was totally dependent on extracellular Ca2+. The mechanisms for depolarization and increases in cytosolic Ca2+ appeared to be independent of each other, as extracellular Ca2+ removal or intracellular Ca2+ buffering with entrapped BAPTA did not affect the depolarization. Removal of extracellular Na+ did not affect the agonist‐induced increase in Ca2+. If quisqualate was added after kainate, the cells were hyperpolarized in a Ca2+‐ and K+‐dependent manner. This could be due to effects on a Ca2+‐dependent K+ channel, the effects of which are normally hidden by the greater effect on Na+ channels as a response to quisqualate.
C17, a retrovirally immortalized neuronal cell line, inhibits the proliferation of astrocytes and astrocytoma cells by a contact‐mediated mechanism
Authors: Weinstein David E. , Shelanski Michael L. , Liem Ronald K. H. 
Published: January 2005

AbstractWe have investigated the ability of various cell lines to effect contact‐mediated inhibition of astrocytic cells. Of the lined tested, only C17, a mouse cell line from postnatal day 0 cerebellum immortalized by infection with a retroviral construct containing the avian myc gene, and U251, a human astrocytoma line, were able to inhibit the proliferation of astrocytic cells. With co‐cultured with either primary astrocytes from rat cerebellum or the U251 line, the C17 cells induced a rapid cessaion of glial cell division as well as complex astrocytic process extension. The effects on glial mitosis were cell‐dose‐dependent, with ten C17 cells/glial cell being the optimal ratio. At this ratio [3H]thymidine incorporation into the U251 cells was reduced by greater than 80% and there was a virtual stasis in glial cell number at 48 hours. Fixed C17 cells as well as partially purified C17 membranes were also potent inhibitors of astrocytic proliferation, suggesting that the gliastatic effect of the C17 cell line is membrane associated. However, neither of these preparations induced astrocytic process formation. We also confirmed earlier reports that U251 cells inhibited their own proliferation in a density‐dependent manner but at a lower fficiency than the C17 cells.
Electrophysiological measurements of volume changes in leech neuropile glial cells
Authors: Ballanyi K. , Grafe P. , Serve G. , Schlue W.‐R. 
Published: January 2005

AbstractDouble‐barrelled microelectrodes, sensitive to quaternary ammonium ions, were used for simultaneous measurements of the intracellular free concentrations of choline ([Ch]i) or tetramethylammonium [TMAi] as well as membrane potential (Em) in neuropile glial cells of the leech, Hirudo medicinalis. Bath application of Ch or TMA (5 mM, 1 min) resulted in a transient membrane depolarization accompanied by a long‐ lasting (0.5–1 h) intracellular accumulation of these compoundsto levels of between 5 and 15 mM. Changes in [Ch]i or [TMA]i were used for the calculation of changes in relative cell volume. Elevation of the extracellular K+ concentration ([K+]e) from 4 to 9,15,21,27.5, or 40 mM elicited a membrane depolarization and a reversible cell swelling by about 7.5,14, 18.5, 27 and 50%, whereas reduction of [K+]e to 1.5 mM as well as bath application of serotonin (5‐HT) produced a membrane hyperpolarization and a concomitant shrinkage by about 6 and 14.3%, respectively. The measured alterations in cell volume were compared with calculated data based on the assumption of an osmotic equilibrium disturbed by potential‐dependent changes of the intracellular Cl− concentration. The results indicate, that K+‐ and serotonin‐induced changes in the cell volume of the neuropile glial cells are due to passive KCl and water fluxes.
Stimulation of thromboxane release from primary cell cultures derived from human astrocytic glioma biopsies
Authors: Murphy Sean , Welk Greg , Thwin Soe Soe 
Published: January 2005

AbstractStimulation of primary cultures of rat astrocytes with appropriate agents results in the mobilization of arachidonic acid from intracellular lipid pools and the synthesis of eicosanoids. Thromboxane A2 is one of the major prostanoids released upon stimulation with calcium ionophore, phorbol esters, and ATP; but a number of other predicted effectors are inactive. In an attempt to understand the pathophysiological significance of eicosanoid release from astrocytes, primary cultures have been derived from human astrocytic glioma biopsies. The majority of cells in the cultures expressed glial fibrillary acidic protein (GFAP), frequently in conjunction with vimentin and fibronectin. Cell sorting revealed that a significant proportion of cells in the cultures from the high‐grade (malignant) tumors expressed epidermal growth factor receptor, indicative of neoplastic cells. Both effective and ineffective agents in rat cultures were tested for their ability to stimulate release of thromboxane from these gliomas, and also from cultures of medulloblastoma and ependymoma which contained significant numbers of GFAP‐positive cells. Only cells from the high‐grade tumors released thromboxane in response to the known effective stimuli. While the muscarinic agonist carbachol was ineffective, norepinephrine evoked thromboxane release from malignant astrocytomas. These data show that cells derived from malignant human gliomas retain the ability to release thromboxane upon stimulation and suggest that a transformation in receptor coupling might accompany neoplasia, such that the cells now respond to a previously ineffective agonist.
Investigation of dysmyelinogenesis in caprine β‐mannosidosis: In vitro characterization of oligodendrocytes
Authors: Boyer Philip J. , Lovell Kathryn L. 
Published: January 2005

AbstractCentral nervous system myelin deficiency is a consistent feature of caprine β‐mannosidosis, and autosomal recessive neurovisceral lysosomal storage disease. To investigate the possibility of an intrinsic oligodendrocyte defect in β‐mannosidosis, oligodendrocyte‐enriched glial cultures from the cerebral hemisphere white matter of two affected and six control goats were compared with respect to culture yield and morphology. Fewer oligodendrocytes were cultured per gram of white matter from affected animals than from control animals. Galactocerebroside‐positive oligodendrocytes from all animals were similar morphologically at all stages of culture by phase contrast and fluorescence microscopy. These findings are consistent with in vivo morphological observations and suggest that differentiated oligodendrocytes from affected animals do not show morphological abnormalities in culture. However, increased numbers of galactocerebroside‐negative bipolar cells, which may be glial progenitor cells, were present in cultures from affected animals. This observation suggests the possibility of a defect in differentiation to mature oligodendrocytes, with persistence of the undifferentiated glia during late stages of development.
Sorbitol pathway activity and utilization of polyols in astroglia‐rich primary cultures
Authors: Wiesinger Heinrich , Thiess Ulrike , Hamprecht Bernd 
Published: January 2005

AbstractAstroglia‐rich rat primary cultures can be grown in a glucose‐free medium containing 25 mM sorbitol. After 10 days under these conditions, the total number of cells and DNA content are reduced to 50–60% of those of control cultures, but remain constant thereafter. The specific activities of the sorbitol pathway enzymes, sorbitol dehydrogenase and aldose reductase, are increased 2.5‐fold and unchanged, respectively, if the cells are grown in the presence of sorbitol instead of glucose. Treatment with the aldose reductase inhibitor sorbinil does not decrease the number of cells cultured in the glucose‐free medium in the presence of sorbitol. Fructose is as good a substrate for the glial cells as sorbitol, whereas out of a number of other polyols tested only xylitol can support the primary cultures for more than 3 days. Neither neuron‐rich rat brain primary cultures nor rat glioma cells can be cultured in the sorbitol‐containing medium in the absence of glucose. With sorbitol substituting for glucose in the culture medium, effects of glucose deprivation on cellular functions like sugar transport and metabolism can be investigated in glial cultures for an extended period of time.
Remote astrocytic response as demonstrated by glial fibrillary acidic protein immunohistochemistry in the visual cortex of dorsal lateral geniculate nucleus lesioned rats
Authors: Hajós Ference , Kálmán Mihály , Zilles Karl , Schleicher Axel , Sótonyi Peter 
Published: January 2005

AbstractThe reaction of astroglia was investigated after unilateral destruction of the dorsal lateral geniculate nucleus in the primary visual cortex of adult albino rats. The destruction of the dorsal lateral geniculate nucleus was performed by stereotaxic injections of ibotenic acid, and the location was verified in Nissl stained sections in each animal. Electron microscopic observations demonstrated the presence of degenerating axon terminals surrounded by hypertrophic astroglial processes mainly in layers III and IV of the ipsilateral primary visual cortex. The ipsilateral (impaired) and contralateral (control) sides of the primary visual cortex showed light microscopically a clearly differing appearance and distribution of glial fibrillary acidic protein (GFAP) immunoreactivity 7 to 11 days after the unilateral injection of ibotenic acid into the dorsal lateral geniculate nucleus. Whereas the control side of the primary visual cortex showed GFAP staining only in the subpial zone of layer I and close to the white matter, all layers of the impaired cortex showed an intense GFAP immunoreactivity. The increase in immunoreactivity was confined to the primary visual cortex. The extent of and increase in immunoreactivity was corroborated by image analysis. These findings were interpreted as a localized hypertrophy of astroglia caused by the anterograde degeneration of geniculocortical terminals. This hypertrophy is accompanied by an increase in GFAP, which may represent the stabilization of the cytoskeleton of newly formed glial processes involved in the rearrangement of the impaired neuropil.
Müller cell changes precede vascularization of the pigment epithelium in the dystrophic rat retina
Authors: Roque Rouel S. , Caldwell Ruth B. 
Published: January 2005

AbstractIn the Royal College of Surgeons rat with inherited retinal dystrophy, photoreceptor cell degeneration is accompanied by retinal pigment epithelial (RPE) cell alterations and Müller cell changes such as increased expression of glial fibrillary acidic protein (GFAP). Vascular changes such as vascularization of the RPE, vascular proliferation, and formation of vitreoretinal membranes (VRMs) are observed later. To study the relationship of Müller cell changes to the vascular alterations in the dystrophic retina, we used immunoperoxidase techniques and antibodies against GFAP and vimentin. Our study showed that during photoreceptor degeneration, Müller cells expressed small amounts of GFAP. As degeneration progressed, GFAP expression increased and morphological alterations occurred in Müller cells. Müller cell apical processes extended and proliferated in the subretinal space and contacted the apical surface of duplicated RPE cells. Later, GFAP reactive fibers surrounded retinal vessels apposed to the RPE. As the vessels became enmeshed within the RPE, the GFAP‐positive perivascular processes disappeared. Eventually, the RPE‐associated vessels became displaced into the inner retina where VRMs were sometimes observed. Immunoblots showed increased GFAP in dystrophic as compared with control retinas. Studies of vimentin distribution in the dystrophic retina showed results similar to the GFAP study. Moreover, the vimentin study suggested increased number of Müller cell processes in the dystrophic as compared with control retinas. The close temporal and anatomical relationships among Müller cell, RPE, and vascular changes in the dystrophic rat suggest a role for Müller cells in retinal neovascularization and proliferative retinopathy.
Norepinephrine‐evoked calcium transients in cultured cerebral type 1 astroglia
Authors: Salm A. K. , McCarthy Ken D. 
Published: January 2005

AbstractWe have used the CA++ indicator dye fura‐2 AM and computerized imaging systems to investigate adrenergic regulation of intracellular calcium in cultured cerebral type 1 astroglia. We have found that norepinephrine (NE) and other adrenergic agonists stimulate increases in intracellular calcium in over 80% of type 1 astroglia tested. A wide range in effective NE concentrations was seen. With sufficient agonist concentrations the calcium response was biphasic, exhibiting an initial sharp peak followed by a sustained calcium elevation. This secondary component was sensitive to reductions in extracellular calcium concentrations and dependent on the continued presence of agonist. Pharmacological studies indicated that astroglial calcium responses were mediated by α1‐ and α2‐adrenergic receptors. At times these two receptor subtypes appeared to underlie calcium responses by the same cells, whereas other cells only responded to stimulation of one or the other subtypes of α‐adrenergic receptor. Finally, we have also observed spontaneous and agonist‐evoked oscillations in astroglial calcium levels. The major findings of these studies indicate that 1) astroglial cells respond to α‐adrenergic receptor stimulation with increased intracellular calcium, 2) these responses can be mediated by α1‐and/or α2‐adrenergic receptors, and 3) subpopulations of cerebral type 1 astroglia exist with respect to α‐adrenergic receptor expression.
Retinal neurite growth on astrocytes is not modified by extracellular matrix, anti‐L1 antibody, or oligodendrocytes
Authors: Ard March D. , Bunge Mary Bartlett , Wood Patrick M. , Schachner Melitta , Bunge Richard P. 
Published: January 2005

AbstractTwo factors that may influence the course of axonal regeneration in the central nervous system (CNS) are extracellular matrix (ECM) and cell surface molecules that may enhance or inhibit neurite outgrowth. Whereas cultured astrocytes have been reported to be a good substratum for neurite outgrowth, there is recent evidence that cultured oligodendrocytes are inhibitory. To test the influences of (1) ECM components, (2) the L1 adhesion molecule, and (3) the inhibitory potential of mature oligodendrocytes in the astrocytic environment, we have utilized a culture system in which neurites from embryonic rat retina grow vigorously on astrocyte monolayers. The major ECM components were assembled in neonatal rat cortical astrocyte‐retina co‐cultures only when the medium contained serum. In electron microscopic studies of serum containing cultures, retinal neurites were seen to be related to astrocyte surfaces but rarely were found in contact with ECM; in serum‐free medium the association between neurites and astrocytes was similar. In addition, the growth of neurites was vigorous whether ECM was present or absent. Presence of antibodies against the cell surface adhesion molecule L1 did not inhibit retinal neurite elongation on glial fibrillary acidic protein‐positive astrocytes, When oligodendrocytes from adult rat spinal cord were combined with the astrocytes, retinal neurites grew as well on the mixed glial population as on astrocytes alone. Immunostaining for galactocerebroside showed many oligodendrocyte processes to be aligned in the direction of neurite growth, suggesting association between the two cell types. This association was verified by electron microscopy. Furthermore, retinal explants extended neurites among myelin basic protein‐positive oligodenrocytes cultured without astrocytes.Thus, the astrocyte surface is a strong promoter of neurite growth from embryonic rat retina. This growth did not depend upon either ECM or the L1 adhesion molecule. Because neurites grew on astrocytes in the presence of mature oligodendrocytes or among oligodendrocytes alone, we conclude that oligodendrocytes do not inhibit neurite growth under certain conditions.
GABA release triggered by the activation of neuron‐like non‐NMDA receptors in cultured type 2 astrocytes is carrier‐mediated
Authors: Gallo Vittorio , Patrizio Mario , Levi Giulio 
Published: January 2005

AbstractKainate (KA), quisqualate (QA), and α‐amino‐3‐hydroxy‐5‐methyl‐4‐isoxazolepropionic acid (AMPA) stimulated γ‐aminobutyric acid [3H]γ‐aminobutyric acid (GABA) release from cultured cerebellar type 2 astrocytes and from their bipotential precursors. The evoked release was prevented by the antagonist 6‐cyano‐2,3‐dihydroxy‐7‐nitro‐quinoxaline (CNQX). AMPA and QA applied together with KA at concentrations around or above their EC50s (20–50 μM) antagonized the stimulatory effect of KA on [3H]GABA release. On the other hand, the releasing action of KA was potentiated by concentrations of QA in the low micromolar range (2–5 μM), particularly when the concentration of KA was at the borderline of effectiveness (10 μM). KA and QA did not elevate intracellular cyclic GMP levels in astrocyte cultures, although guanylate cyclase was present in both type 2 and type 1 astrocytes. The inability of KA to elevate cyclic GMP levels in astrocytes was the only major difference in the behavior of this glutamate agonist between astroglial and neuronal cultures. The GABA transport inhibitor nipecotic acid or replacement of NaCl with LiCl abolished [3H]GABA uptake and also KA‐ and QA‐induced release of preaccumulated [3H]GABA. Therefore, [3H]GABA was released from type 2 astrocytes and their progenitors through its Na+‐dependent transport system, operating in an outward direction when the cells were depolarized by non‐NMDA receptor agonists.
Effect of thyroid deficiency on glial fibrillary acidic protein (GFAP) and GFAP‐mRNA in the cerebellum and hippocampal formation of the developing rat
Authors: Faivre‐Sarrailh Catherine , Rami Abdelhaq , Fages Christiane , Tardy Marcienne 
Published: January 2005

AbstractThe concentrations of glial fibrillary acidic protein (GFAP) and its encoding mRNA in the cerebellum and hippocampal formation were assayed during the development of normal and hypothyroid rats. Neonatal hypothyroidism induced a significant reduction in the GFAP concentration in both regions from day 14. The reduction was especially marked on day 35 in the cerebellum (−43%) and the hippocampal formation (−55%). The immunocytochemical study of vimentin showed that the developmental disappearance of this protein from the Bergmann and internal astroprcytes is greatly delayed in the cerebellum of the hypothyroid rats. The reduction in GFAP concentration together with the delayed vimentin‐GFAP transition could explain how astrocyte morphogenesis is impaired by neonatal thyroid deficiency.The GFAP‐mRNA concentration in the hippocampal formation was reduced throughout the development of thyroid‐deficient rats, while the GFAP‐mRNA concentration in the cerebellum first increased between birth and day 14 to reach a peak well above the normal value (+78%) and decreased thereafter to reach 53% of the normal value by day 35. This transient increase in the cerebellar GFAP‐mRNA concentration may be related to the astroglial hyperplasia that occurs in these animals. The difference between the developmental profile of GFAP and its encoding mRNA, especially under pathological conditions, indicates that two distinct mechanisms control the synthesis or stability of the protein and its messenger RNA, as was previously found in the forebrain of the developing normal rat.
Characterization of glycosaminoglycans produced by primary astrocytes in vitro
Authors: Johnson‐Green P. C. , Dow K. E. , Riopelle R. J. 
Published: January 2005

AbstractQuantitative biosynthetic studies with cultures highly enriched for glial fibrillary acidic protein (GFAP+) cells of neonatal mammalian brain demonstrated production of four proteoglycans: hyaluronate (HA), heparan sulphate (HS), chondroitin sulphate (CS), and dermatan sulphate (DS). The glycosaminoglycans were present in cell conditioned medium and in the cellular compartment. There were qualitative differences in the subcellular disposition of the various proteoglycans. The ratio of HS to CS/DS in cell extracts was 1:1, while in medium this ratio was 1:6. All of the glycosaminoglycans were associated with core proteins that were integral to the cell membrane and associated with the cell surface by non‐covalent interactions involving glycosaminoglycans. Less than 20% of the HS was non‐covalently associated with the astrocyte cell surface reflecting in part the proportionately smaller amounts of this proteoglycan released to astrocyte conditioned medium. HS released to medium was undersulphated relative to that associated with cells.The astrocyte can contribute proteoglycans to the extracellular milieu and displays cell surface proteoglycans that have the potential to provide appropriate substrates for neuron adhesion, process extension, and other cell‐cell interactions.
Effects of dexamethasone on the differentiation of membrane structure in cultured astrocytes
Authors: Landis Dennis M. D. , Weinstein Lori A. , Skordeles Christine J. 
Published: January 2005

AbstractAstrocytic processes investing vascular structures or forming the surface of mammalian brain have large numbers of orthogonally packed aggregates of intramembrane particles, termed “assemblies”. Similar particle aggregates are expressed by astrocytes derived from neonatal rat forebrain in secondary culture, but they are much more uniformly distributed across the membranes of the cultured cells. Dexamethasone, a potent glucocorticoid, affects the differentiation of astrocyte membrane structure in two patterns, depending on the rate of proliferation in the culture. When confluent secondary cultures of astrocytes are exposed to 5 μM dexamethasone, the densities of assemblies increase, and in some cells approach the values present in the glial limitans in vivo. However, when rapidly proliferating astrocytes are exposed to dexamethasone during the first week of secondary culture, most of the astrocytes fail to express any assemblies. The rate of astrocyte proliferation is slowed, and a lower cell density is reached during the first 2 weeks of secondary culture in dexamethasone. The suppression of assemblies is transient: as the cultures approach confluence, the proportion of cells expressing assemblies increases to nearly control levels, and the density of assemblies increases to greater than control values in some astrocytes. Certain of the effects of dexamethasone on cultured astrocytes may have relevance for understanding the mechanism(s) of its action in treating cerebral edema.