bluuebunny/crossref_metadata_2025_split huggingface dataset:
Response of rat cerebral cortical astrocytes to freeze‐ or cobalt‐induced injury: An immunocytochemical and gap‐FRAP study
Authors: Anders Juanita J. , Niedermair Suzanne , Ellis Elaine , Salopek Maureen 
Published: January 2005

AbstractAstrocytic response in the immediate vicinity of freeze‐ and cobalt‐induced lesions has been examined at the light and ultrastructural level. However, the temporal and spatial distribution of astrocytic reactivity throughout the rat cerebral cortex, using glial fibrillary acidic protein (GFAP) immunolabeling, has not been examined. The first purpose of this study was to establish the chronological distribution of astrocytic reactivity, as measured by changes in GFAP immunoreactivity, following freeze‐ or cobalt‐induced injury to the rat cerebral cortex. Cobalt metal also has been proposed to have a direct effect on astrocytes and has been shown to stimulate in vitro astrocytes to become reactive. The second purpose of this report was to determine if cobalt had an effect on in vitro astrocytic gap junctional dye coupling as measured by fluorescence recovery after laser‐photobleaching (gap‐FRAP). Although the chronological development of the increased GFAP immunoreactivity was different for the freeze‐ and cobalt‐induced lesions, astrocytes initially showed an increase in GFAP immunoreactivity in the region surrounding these lesions. This initial response was followed by a spread of increased GFAP immunoreactivity throughout certain regions of the ipsilateral cerebral hemisphere and then by a restriction of the increased immunolabeling to the lesion site. Cobalt also had a direct effect on in vitro astrocytes as demonstrated by the inhibition of astrocytic gap junctional dye coupling. Based on gap‐FRAP analysis, cobalt significantly blocked fluorescence recovery (2.5%) as compared to the fluorescence recovery in control astrocytes (26%). It is proposed that the initial increase in GFAP immunoreactivity may be due to decreased gap junctional activity.
Flow cytometric analysis of mammalian glial cultures treated with methotrexate
Authors: Serrano Elba E. , Schimke Robert T. 
Published: January 2005

AbstractMethotrexate (MTX) is an antineoplastic drug that acts by competitive inhibition of the enzyme dihydrofolate reductase (DHFR). MTX treatment of cultured cell lines leads to the emergence of resistant cell populations. Studies using stepwise selection procedures have demonstrated that MTX resistance conferred by overproduction of DHFR can be caused by DHFR gene amplification. We examined the effect of MTX on cells whose origin more closely approximates the in vivo condition by developing a culture system using dissociated brain tissue from 17–19 day old mouse embryos. At the first passage, cultures were divided into control and MTX groups. Cells were treated with the same or successively higher concentrations of MTX at each passage over a 3–4 month period. The first passage eliminated neurons and left a glial culture comprised of approximately 90% astrocytes. We used the Fluorescence Activated Cell Sorter in conjunction with fluorescent dyes to measure DHFR content, DNA content, size, and viability of glial cells following MTX treatment. MTX‐treated cells divided but grew more slowly and were larger than untreated cells. Stepwise selection in 30/60/90 nM or 60/120 nM MTX resulted in significant two‐ to threefold increases in fluorescence, and hence DHFR levels. Slot hybridizations assays demonstrated a threefold increase in DHFR gene copy number in the DNA from the 30/60/90 cultures. Thus, our findings were consistent with the results obtained from somatic cell lines, and lend support to the hypothesis that gene amplification may be a common mechanism for the acquisition of resistance in many types of cells. They also indicate that glial cells may be a specific target for cytotoxic effects of MTX on the central nervous system.
Regeneration of axons from adult rat retinal ganglion cells on cultured schwann cells is not dependent on basal lamina
Authors: Hopkins James M. , Bunge Richard P. 
Published: January 2005

AbstractThe ability of sciatic nerve grafts to support in vivo regeneration of retinal ganglion cell axons in the adult rat raises the question of which peripheral nerve constituents may be required to promote this unexpected central regenerative response. Prime candidates for this role include the surface of the Schwann cell and components of extracellular matrix present in peripheral nerve trunks. To determine the relative importance of Schwann cells and their basal lamina in promoting retinal ganglion cell axon regeneration in the mammalian visual system, we have used an in vitro model. This approach allowed analysis of the abilities of defined peripheral nerve constituents to promote in vitro outgrowth of neurites from explants of adult rat retina harvested 7 to 10 days after in vivo optic nerve crush. Neurite outgrowth was assessed by neurofilament immunofluorescence after 3 to 20 days in vitro. Culture substrata, consisting of isolated Schwann cells (SC), Schwann cells with their assembled extracellular matrix (SC + ECM), or isolated extracellular matrix from which the Schwann cells had been removed (ECM), were prepared by first co‐culturing rat Schwann cells with embryonic dorsal root ganglion neurites on a layer of type I collagen, and then manipulating the cultures to produce the desired substrata. Type I collagen alone did not support neurite growth from adult rat retina. SC and SC + ECM supported regeneration of axons from retinal explants at average growth rates of 18 and 30 μm/h, respectively. Isolated ECM was a poor substrate for retinal neurite growth; the few neurites that gained access to this material grew at rates averaging less than 3 μm/h. These observations suggest that regeneration of adult mammalian retinal ganglion cell axons through peripheral nerve grafts (in vivo) is primarily dependent on neurite‐promoting factors present on the surface of Schwann cells and does not require organized extracellular matrix.
Schwann cell precursors and their development
Authors: Jessen Kristjan R. , Mirsky Rhona 
Published: January 2005

AbstractDuring development of peripheral nerves, an apparently homogeneous pool of embryonic Schwann cells gives rise to two morphologically and antigenically distinct mature Schwann cell types. These are the myelin‐forming cells associated with axons of larger diameter and the non‐myelin‐forming cells associated with axons of smaller diameter.The development of these cells from precursors that can be identified in early embryonic nerves can be followed with the help of antigenic differentiation markers. This development depends on Schwann cells retaining a close association with axons. The effect of axons can be mimicked in vitro by agents that elevate cAMP levels. This has given rise to the idea that the effects of axon‐associated signals in Schwann cell development are to a significant extent mediated via elevation in Schwann cell cAMP levels. In vitro, the cAMP induced progression of cells from a premyelination state to a myelination state depends on withdrawal from the cell cycle. It is therefore possible that in vivo, the timing of myelin formation by individual Schwann cells is determined by signals that suppress proliferation.
The origin of remyelinating cells in the adult central nervous system: The role of the mature oligodendrocyte
Authors: Wood Patrick M. , Bunge Richard P. 
Published: January 2005

AbstractThe sequence of events by which new oligodendrocytes are generated in the adult mammalian central nervous system has not been clearly defined. Here we review old evidence that remyelinating cells can arise from the division of mature oligodendrocytes. In addition, we report the results of a tissue culture study comparing oligodendrocytes and immature progenitors with regard to their capacity for proliferation, for the generation of new oligodendrocytes and for myelination. Monoclonal antibodies 04 and 01 were used to distinguish oligodendrocytes (04+01+) from progenitors (04+01‐). Dissociated cell suspensions from adult rat spinal cord were separated by flow cytometry into 01+ and 01‐ cell fractions, at > 93% purity. The 01+ fraction contained ≈︁ 0.7% 04+01‐ cells while the 01‐ fraction contained ≈︁ 4.4% 04+01‐ cells. Cells from these fractions were plated onto cultures of purified dissociated dorsal root ganglion neurons. The cultures that received 01+ cells developed numerous expanding colonies of cells expressing both 01 and 04, or 04 only, by 8 days and were essentially covered by oligodendrocytes by 16 days. In marked contrast, oligodendrocyte colonies were rare in cultures receiving 01‐ cells. By 24 days, myelination was extensive in cultures receiving 01+ cells; in contrast, only a few myelin segments were observed in cultures receiving the 01‐ fraction. Thus, oligodendrocytes (01+ cells) appear more capable than progenitors (04+01‐) of generating new myelinating oligodendrocytes in this culture system.
Potassium channels in crustacean glial cells
Authors: Erxleben Christian 
Published: January 2005

AbstractUnitary currents through single ion channels in the glial cells, which ensheath the abdominal stretch receptor neurons of the crayfish, were characterized with respect to their basic kinetic properties. In cell‐attached and excised patches two types of Ca++‐independent K+ channels were observed with slope conductances of 57 pS and 96 pS in symmetrical K+ solution.The 57 pS K+ channel was weakly voltage‐dependent with a slope of the Po vs. membrane potential relationship of +95 mV for an e‐fold change in Po. In addition to the main conductance level, the channel displayed conductance levels of 80 and 109 pS. In excised patches, channel activity of this “subconductance” K+ channel showed “rundown” that could be prevented with 2 mM ATP‐Mg on the cytoplasmic side of the membrane.The 96 pS K+ channel was strongly voltage‐dependent with a slope of +12 mV for an e‐fold change in Po. Averaged single‐channel currents elicited by voltage jumps proved the channel to be of the delayed rectifying type. Channel activity persisted in excised patches with minimal salt solution and in virtually Ca++‐free saline.Because of its dependence on intracellular ATP‐Mg, the subconductance K+ channel is discussed as a target of modulation by transmitters or peptides via phosphorylation of the channel.
Natural and induced cytotoxicity of oligodendrocytes by microglia is inhibitable by TGFβ
Authors: Merrill Jean E. , Zimmerman Robert P. 
Published: January 2005

AbstractBlood macrophages and brain macrophages (microglia) have been implicated in demyelination and destruction of the oligodendrocyte in multiple sclerosis (MS), a disease affecting primarily white matter of the central nervous system (CNS). In this study, we demonstrate that at high effector to target cell ratios, normal rat microglia exhibit a natural cytotoxicity against normal rat oligodendrocytes in vitro. The killing is not mediated by the release of soluble factors. The cytotoxic activity is upregulated by pretreatment of microglia with interferon gamma (IFNγ) or phorbol myristate acetate (PMA). Both the natural and induced cytotoxicities are inhibitable by transforming growth factor beta (TGFβ). The increase in numbers and apposition of primed or activated microglia to oligodendrocytes in MS lesions may give rise to natural or induced killing from which oligodendrocytes may be protected by TGFβ.
Localization and stoichiometry of electrogenic sodium bicarbonate cotransport in retinal glial cells
Authors: Newman Eric A. , Astion Michael L. 
Published: January 2005

AbstractAn electrogenic Na+/HCO cotransport system was identified and characterized in freshly dissociated salamander Müller (glial) cells. Under voltage‐clamp, these cells generated an outward current when external HCO3− concentration ([HCO3−]o) was raise. This current was Na+‐dependent, Cl−‐independent, and was blocked by the stilbenes 4,4′‐diisothiocyanato‐stilbene‐2,2′‐disulfonate (DIDS) and 4,4′‐dinitrostilbene‐2,2′‐disulfonate (DNDS), and by harmaline, demonstrating that the current was generated by a Na+/HCO3− cotransport system. Substantially larger currents were evoked when [HCO3−]o was raised at the Müller cell endfoot as compared to other cell regions, indicating that cotransporter sites are localized preferentially to the endfoot. The reversal potential of the current, which varied as a function of HCO3− and Na+ transmembrane gradients, indicated that the cotransporter has a HCO3−:Na+ stoichiometry of 3:1.
Sexual dimorphism in the hamster cerebellum demonstrated by glial fibrillary acidic protein (GFAP) and vimentin immunoreactivity
Authors: Suárez I. , Bodega G. , Rubio M. , Fernández B. 
Published: January 2005

AbstractMale and female hamsters aged 1, 4, and 10 postnatal weeks were used to study the distribution of vimentin and glial fibrillary acidic protein (GFAP) in the cerebellum. Vimentin immunoreactivity exceeded that of GFAP during the first postnatal week, although GFAP was also observed in all cerebellar layers. Immunoperoxidase analysis revealed that by the fourth postnatal week vimentin was only detected in Bergmann fibers and the very scarce fibrous astrocytes located in the inner white matter. The Purkinje cell bodies were only coated with GFAP‐immunopositive processes. At 10 weeks, vimentin immunoreactivity was reduced to thin Bergmann glial processes, whereas GFAP immunoreactivity had greatly increased in the whole cerebellum.The GFAP immunostaining was denser in males than in females; however, in females, the Bergmann fibers were heavily immunostained with anti‐vimentin in contrast to the males. The results described in the present paper indicate a sex difference in vimentin and GFAP immunoreactivities in the cerebellar astrocytes at 4 weeks of age, which persisted in the oldest hamsters in this study. The existence of sexual dimorphism might suggest that the expression of both gliofilament proteins could be influenced by circulating sex steroids.
Neurite growth modulation associated with astrocyte proteoglycans: Influence of activators of inflammation
Authors: Johnson‐Green P. C. , Dow K. E. , Riopelle R. J. 
Published: January 2005

AbstractOf the three classes of sulphated proteoglycans produced by type 1 astrocytes in vitro and released into conditioned medium, only heparan sulphate (HS) was associated with enhanced neurite growth by sensory neurons following pretreatment of a laminin substratum. Astrocyte‐conditioned medium (ACM) produced in the presence of certain inflammatory mediators had reduced titers of neurite‐promoting activity. The low activity ACM contained inhibitors of neurite growth. Heparan sulphate proteoglycans may modulate neurite growth when complexed to constituents of the extracellular milieu either directly or by interacting with other growth‐promoting or growth‐inhibitory factors.
Preparation and characterization of type 1 astrocytes cultured from adult rat cortex, cerebellum, and striatum
Authors: Schwarts Joan P. , Wilson Delores J. 
Published: January 2005

AbstractAstrocytes have been prepared from adult rat cortex, cerebellum, and striatum, using a modification of the McCarthy‐DeVellis (J Cell Biol 85:890, 1980)method. The cultures consist of 99% type 1 polygonal astrocytes, which divide more slowly than cells from newborn animals. One day after preparing the cultures, 90% of the cells are glial fibrillary acidic protein (GFAP)‐positive and 80% are tin‐positive by immunohistochemical staining, suggesting that they are present de novo and not derived from precursor cells. The astrocytes from adult brain respond to an elevation of intracel‐lular cyclic AMP, following treatment with forskolin, by becoming more stellate in shape and putting out fine ramified processes. They contain the same amount of GFAP per mg protein, measured by immunoblot, as cells from newborn animals. These cultures thus offer the possibility of comparing the biochemical properties of astrocytes derived from adult animals with those from newborn animals, or with cultures of reactive astrocytes isolated from lesioned brain.
Synthesis and release of neuroactive substances by glial cells
Authors: Martin David L. 
Published: January 2005

AbstractGlia contain, synthesize, or release more than 20 neuroactive compounds including neuropeptides, amino acid transmitters, eicosanoids, steroids, and growth factors. The stimuli that elicit release differ among compounds but include neuropeptides, neurotransmitters, receptor agonists, and elevated external [K+]. The mechanisms of release are poorly understood in most cases. Many of the neuroactive compounds are localized in discrete subpopulations of glia. Thus, glia are equipped to send as well as receive chemical messages and appear to be present as classes of cells with differing abilities to communicate chemically. It is possible that glia are as diverse as neurons in their functional characteristics.
Regulation of carbachol‐ and histamine‐induced inositol phospholipid hydrolysis in a human oligodendroglioma
Authors: Post Ginell Ristic , Dawson Glyn 
Published: January 2005

AbstractA stable cell line derived from a human oligodendroglioma (HOG) was used to study the regulation of muscarinic‐ and histamine receptor‐mediated phosphoinositide hydrolysis. Both carbachol and histamine increased inositol monophosphate (InsP) accumulation in a dose‐and time‐dependent manner in the presence of lithium and the effect of simultaneous addition of carbachol and histamine was additive, implying independent signal transduction pathways. Homologous desensitization of muscarinic, but not histamine receptors, could be demonstrated although neither receptor type appeared to be heterologously desensitized. [3H]InsP accumulation in HOG cells was also stimulated by fluoride, suggesting guanosine triphosphate (GTP)‐binding protein involvement, but phosphoinositide (PtdIns) hydrolysis was not sensitive to pertussis toxin. Phorbol ester‐activation of protein kinase C (PKC) inhibited both muscarinic and histamine receptor‐stimulated InsP release but did not attenuate either the fluoride‐induced release of InsP nor β‐adrenergic receptor‐mediated stimulation of adenylate cyclase activity. Taken together, we conclude that muscarinic and histamine receptors are differentially regulated through both PKC‐dependent and ‐independent mechanisms, and that feedback inhibition of PtdIns turnover occurs proximal to the GTP binding proteins.
Identification of RBK1 potassium channels in C6 astrocytoma cells
Authors: Wang Sho‐Ya , Castle Neil A. , Wang Ging Kuo 
Published: January 2005

AbstractIonic currents in C6 astrocytoma cells were studied using the patch clamp technique under the whole cell configuration. A delayed rectifier K+ current with an amplitude of ∼ 1 nA at +50 mV was observed in 86% (92/107) of the cells examined. This K+ current resembled the delayed rectifier present in type‐1 and type‐2 astrocytes in vitro and could be inhibited by a variety of K+ channel blockers, including TEA (IC50: 0.5 mM), 4‐aminopyridine (IC50: 0.2 mM), MCD peptide (IC50 : 52 nM), dendrotoxin I (IC50: 9 nM), and charybdotoxin (74% inhibition at 50 nM). Northern blot analysis, cloning of cDNA and subsequent sequencing showed that the C6 cell delayed rectifier K+ channel is equivalent to the RBK1 K+ channel derived from a rat brain cDNA library. The level of RBK1 transcripts in C6 cells was comparable to that reported in rat brain. The C6 delayed rectifier K+ channel is probably a homomeric RBK1 K+ channel judging from its pharmacological properties which are similar to the RBK1 channel expressed in Xenopus oocytes. Some C6 cells also expressed a transiently activated outward K+ current (IA). This current was found in less than 50% of the cells and in general contributed no more than 8% of the total outward current. No voltage‐dependent inward Na+ or Ca2+ currents or inwardly rectifying K+ currents were observed in over 100 C6 cells examined. The present results show that the dominant voltage gated ionic current in C6 cells is the RBK1 delayed rectifier K+ channel. Since C6 cells express RBK1 mRNA at a level comparable to its rat brain counterpart, these cells may be a valuable system to study the RBK1 channel and its gene expression.
Autocrine inhibition of mitotic activity in cultured oligodendrocyte‐type‐2 astrocyte (O‐2A) precursor cells
Authors: Louis Jean‐Claude , Muir David , Varon Silvio 
Published: January 2005

AbstractDuring development, oligodendrocytes are generated from a bipotential glial stem cell, the oligodendrocyte‐type‐2 astrocyte precursor (O‐2A). O‐2A cells are under the mitogenic influence of the platelet‐derived growth factor (PDGF) released from type‐1 astrocytes. In vitro experiments have shown that O‐2A cells stimulated by PDGF are limited to a set number of divisions and then differentiate to oligodendrocytes by becoming unresponsive to the growth factor. In the healthy adult central nervous system, oligodendrocyte proliferation remains generally quiescent and is possibly under negative growth control. The view that O‐2A lineage cells are capable of negatively regulating their own proliferation is supported by the demonstration that conditioned medium obtained from O‐2A cultures inhibits their DNA synthesis. In addition to O‐2A cells, the newly established CG4 cell line, a derivative of O‐2A cells, was found to inhibit O‐2A lineage cell proliferation. The antiproliferative activity was present in the media conditioned by CG4 cells that were expanded as undifferentiated O‐2A precursors, as well as by CG4 cells induced to differentiate to nonproliferating oligodendrocytes. Moreover, the inhibitory activity was produced by CG4 cells (source cells) propagated by various mitogens. The inhibition of mitotic activity was nearly complete, dose‐dependent, fully reversible, and exhibited when CG4 cells (test cells) were stimulated to divide by various mitogens, such as PDGF, basic fibroblast growth factor, or medium conditioned by the neuronal B104 cell line. The inhibition of proliferation was accompanied by the conversion of the phenotype of CG4 cells, from A2B5+/O4− precursors to A2B5−/O4+ prooligodendrocytes. This change of antigenic phenotype, like the inhibition of proliferation, was reversible. These results suggest that O‐2A cells secrete an autocrine inhibitory factor capable of reversibly withdrawing them from the cell cycle, which suggests that the commitment to the oligodendrocyte lineage might proceed through more than one discrete stage. © 1992 Wiley‐Liss, Inc.
A histochemical study of the activity of nonspecific cholinesterase in C6 glioma cells in culture
Authors: Dubový P. , Mareš V. , Lisá V. 
Published: January 2005

AbstractThe activity of nonspecific cholinesterase (nChE) was revealed histochemically in a small number of C6 glioma cells under conditions of spontaneous growth in culture. The nChE‐positive cells (nChE+) first appeared in 4‐day‐old cultures. They occurred solitarily or in small groups with a cluster‐ or chain‐like arrangement. The incidence of nChE+ cells increased with in vitro age and/or higher population densities of cultures but it never exceeded 3% of the whole population. These results indicate the necessity of histochemical as well as biochemical data on nChE activity in glioma cells both in situ and in culture for the discovery of the heterogeneous distribution of this enzyme. db‐cAMP induced a decrease in the proportion of nChE+ cells to total number of cells. These changes suggest that the original C6 glioma cell seed stock contains a small subclone of less differentiated cells exhibiting nChE activity. The greater number of nChE+ cells in older cultures is probably due to recovery and multiplication of initially scarce subclone cells. C6 glioma cells in culture form a suitable model for investigation of changes in nChE activity of glial cells provided that a histochemical method of detection is applied. © 1992 Wiley‐Liss, Inc.