bluuebunny/crossref_metadata_2025 huggingface dataset:
Sodium channel expression detected with antibody 7493 in A2B5+ and A2B5− astrocytes from rat optic nerve in vitro
Authors: Minturn J. E. , Black J. A. , Angelides K. J. , Waxman S. G.
Published: January 2005

Astrocytes cultured from neonatal rat optic nerve can be classified into two subtypes, distinguished by their morphology (stellate or fibroblast‐like) and their ability to bind monoclonal antibody A2B5. The presence of sodium channels in astrocytes cultured from rat optic nerve was demonstrated by indirect immunofluorescence with polyclonal antibody 7493, which is directed against purified rat brain sodium channel protein. Astrocytes cultured from postnatal day 7 (P7) rat optic nerves exhibited sodium channel immunostaining on both A2B5+ and A2B5− astrocytes up to 6 days in vitro (DIV). Staining was distributed throughout the cytoplasm and cell processes, with areas of greater intensity in the perinuclear region. At 6 DIV, the A2B5−/GFAP+ cells exhibited a loss of sodium channel immunostaining, while the A2B5+/GFAP+ cells continued to display 7493 immunoreactivity. This sodium channel staining pattern persisted for up to 28 DIV (the longest time point examined). Astrocyte cultures derived from P0 rat optic nerves exhibited sodium channel immunoreactivity during the first 6 DIV. The P0 astrocyte cultures, in which the vast majority of cells are A2B5−/GFAP+, displayed a similar staining pattern to those astrocytes with corresponding phenotype derived from P7 optic nerves. PO‐derived A2B5− astrocytes showed loss of 7493 immunostaining at 6 DIV, while the rare (< 1% of cells) A2B5+/GFAP+ cells continued to express sodium channels reactive to 7493. The reduction of sodium channel immunoreactivity in A2B5−, but not A2B5+, astrocytes from both P0 and P7 optic nerves after a similar latency (<6 DIV) suggests that the loss of immunostaining may result from the absence of neuronal associations in the culture environment, rather than an intrinsic biologically timed change in astrocytic expression of sodium channels.
Control of astrocyte volume by intracellular and extracellular Ca2+
Authors: Olson James E. , Fleischhacker Deborah , Murray W. Bruce , Holtzman David
Published: January 2005

Astrocytes from primary culture were exposed to conditions that affect intracellular and extracellular Ca2+ concentrations. Astrocyte cell volume was increased approximately 16% after a 30 min exposure to isoosmotic phosphate‐buffered saline (PBS) containing the Ca2+ buffer EDTA. Cell volume returned to control values within 30 min of resuspension in normal PBS. Cellular calcium content was not affected by these treatments; however, the recovery of normal cell volume following EDTA exposure was inhibited by 0.1–1.0 mM quinine HCl in a dose‐dependent fashion suggesting that a potassium channel controlled by the intracellular Ca2+ concentration is important in this volume response. Intracellular accumulation of an exogenous Ca2+ buffer, BAPTA, also produced cell swelling that persisted following resuspension in normal PBS. Lowering the extracellular Ca2+ concentration with EDTA enhanced the swelling of BAPTA‐loaded cells. These data suggest that conditions leading to a decrease in free intracellular Ca2+ concentration may influence astrocyte volume by a mechanism similar to that described in other cell types.
Volume‐sensitive release of taurine from cultured astrocytes: Properties and mechanism
Authors: Pasantes‐Morale H. , Moran J. , Schousboe A.
Published: January 2005

Release of taurine in response to cell swelling induced by hyposmolarity was observed in cultured astrocytes. Efflux of 3H‐taurine increased by 30% and 70% upon reductions in osmolarity of only 5% and 10%. Reductions in osmolarity of 20%, 30%, and 50% stimulated basal taurine release by 300%, 500%, and 1,500%, respectively. The properties of this volume‐sensitive release of taurine were examined to investigate: 1) its association with K+ and Cl− fluxes, currently activated during volume regulation; 2) its relationship with Ca2+‐dependent reactions; and 3) the mechanism of the taurine efflux process.

Taurine release was unaffected by removal of Na+, Ca2+, or Cl−, by pimozide and trifluoperazine, or by agents disrupting the cytoskeleton. The K+ channel inhibitors barium, quinidine, tetraethylammonium, and gadolinium had no effect. Taurine release was reduced by furosemide, a blocker of K+/Cl− cotransport, but not by the more specific inhibitor, bumetanide. It was markedly reduced by the inhibitors of Cl− channels DIDS, SITS, and anthracene‐9‐carboxylate. Taurine efflux was pH‐dependent, being reduced at low pH values. It was decreased at 4°C but not at 14°C or 20°C. These results suggest that the volume‐sensitive release of taurine is independent of K+ fluxes but may be associated with Cl− conductances. It also seems unrelated to Ca2+‐dependent transduction mechanisms. The Na+‐dependent taurine carrier apparently is not involved in the swelling‐induced release process.
Synthesis and phosphorylation of the glial fibrillary acidic protein during brain development: A tissue slice study
Authors: Noetzel Michael J.
Published: January 2005

Brain slices were incubated with either [3H] amino acids or [32P] orthophosphate in order to characterize the synthesis and phosphorylation of the glial fibrillary acidic protein (GFAP) in the rat nervous system. The incorporation of [3H] amino acids into GFAP was found to increase significantly during early postnatal development, reaching a peak of activity on day 5 of life and then declining over the next 2 weeks. Concomitant with this peak of synthetic activity the content of GFAP in rat brain was also observed to increase dramatically. GFAP continued to accumulate in brain through postnatal day 30 despite a decrease in the synthesis of the protein. These results indicate that the increase in GFAP during the first month of life cannot be ascribed solely to the rate of GFAP synthesis. The findings are consistent with the hypothesis that during later stages of astrocytic development the accumulation of GFAP may be primarily dependent upon a low rate of protein degradation. The pattern of GFAP phosphorylation in the developing rat brain differed from that observed for the incorporation of [3H] amino acids. The peak incorporation of 32P into GFAP occurred on postnatal day 10 at a time when synthesis of the protein had declined by 43%. These findings suggest that during development phosphorylation of GFAP is mediated by factors different from those directing its synthesis. In addition, phsophorylation of GFAP did not alter its solubility in cytoskeletal preparations indicating that GFAP phosphorylation is probably not a major regulatory mechanism in disassembly of the astroglial filaments.
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

Astrocytic 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

Methotrexate (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

The 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

During 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

The 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

Unitary 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

Blood 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

An 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

Male 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

Of 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

Astrocytes 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

Glia 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

A 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

Ionic 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

During 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

The 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.
Characteristics of fish glial cells in culture: Possible implications as to their lineage
Authors: Sivron Tomer , Jeserich Gunnar , Nona Shmaiel , Schwartz Michal
Published: January 2005

Regeneration of injured central nervous system axons is largely dependent on the response of the associated nonneuronal glial cells to injury. Glial cells of the mammalian central nervous system, unlike those of fish, are apparently not conducive to axonal regeneration. While the lineage of rat glial cells is well characterized and its role in the support or inhibition of regenerative growth is beginning to be understood, little is known about fish glial cells. Accordingly, glial cells in cultures of adult goldfish brain and of newly hatched goldfish larvae were studied in an attempt to establish their lineage. The cells were identified by means of indirect immunofluorescence, using antibodies against fish astrocytes and oligodendrocytes. The cell count in the cultures increased from a small number of cells at 24 h after plating to a large number of both astrocytes and oligodendrocytes after 1 week in culture. Both of these cell types had originated from proliferating cells, as shown by their uptake of tritiated thymidine and by the inhibition of cell proliferation by 5‐fluoro‐2′‐deoxyuridine. Both astrocytes, i.e., glial fibrillary acidic protein‐positive cells, and oligodendrocytes, i.e., 6D2‐positive cells, were positively labeled also by A2B5 antibodies, which are known to label progenitors of type‐2 astrocytes and oligodendrocytes in the rat optic nerve. The results suggest that A2B5 positive progenitor cells in the goldfish central nervous system, as in the rat optic nerve, might be a common progenitor of astrocytes and oligodendrocytes. © 1992 Wiley‐Liss, Inc.
Regulation of plasminogen activators and type‐1 plasminogen activator inhibitor by cyclic AMP and phorbol ester in rat astrocytes
Authors: Tranque Pedro , Robbins Richard , Naftolin Frederick , Andrade‐Gordon Patricia
Published: January 2005

Two plasminogen activators (PAs): tissue‐type plasminogen activator (t‐PA) and urokinase‐type plasminogen activator (u‐PA), as well as the type‐1 plasminogen activator inhibitor (PAI‐1) are synthesized and secreted by rat astrocytes. Preliminary studies suggest that PA activity plays a role in astrocyte development and differentiation. We have examined the regulation of the PA system by the cAMP‐dependent protein kinase (PKA) and protein kinase C (PKC) in purified rat astrocyte cultures. PKA activity was increased by exposing cultured astrocytes to forskolin or dibutyryl cyclic AMP, whereas PKC activity was stimulated with phorbol‐12‐myristate 13‐acetate (PMA). Activation of both second‐messenger pathways produced a time‐ and dose‐dependent increase in the total PA activity. However, based on SDS‐PAGE/zymography we found that forskolin increased t‐PA activity and reduced u‐PA activity, whereas PMA treatment caused a significant increase in u‐PA activity without altering t‐PA activity. Reverse zymography analysis revealed that astrocyte PAI‐1 activity is decreased by forskolin and increased by PMA. Together, these results demonstrate that the components of the PA system in rat astrocytes are independently and reciprocally regulated by PKA and PKC. Our findings raise the possibility that the plasminogen activator system could be involved in some of the actions of growth factors and/or neuromodulators that modulate PKC or PKA in astrocytes. © 1992 Wiley‐Liss, Inc.
Altered antigen expression of microglia in the aged rodent CNS
Authors: Perry V. H. , Matyszak M. K. , Fearn S.
Published: January 2005

Microglia, the resident macrophages of the central nervous system, are characterised by a highly specialized morphology and unusual antigenic phenotype. Microglia appear to be downregulated by their microenvironment when compared to other tissue macrophages. We have studied the microglia in brains of healthy, aged rats with a panel of monoclonal antibodies. We have found that microglia in the brains of these aged rats express antigens that are downregulated or absent from microglia of juvenile rats. The stimuli which give rise to this upregulated phenotype are not known. Age related changes in the phenotype of microglia should be taken into account when considering the possible role of microglia in neuropathological conditions.
Polymorphonuclear leukocytes in brain parenchyma after injury and their interaction with purified astrocytes in culture
Authors: Moreno‐Flores M. Teresa , Bovolenta Paola , Nieto‐Sampedro Manuel
Published: January 2005

At a time after brain injury when removal of debris and secondary cell death were prevalent, many polymorphonuclear neutrophils were observed in injured tissue. Because neural damage could be mediated by activated neutrophils, we tested in vitro the effect of these leukocytes and other blood, components on central nervous system cells. At concentrations similar to those present in blood, polymorphonuclear leukocytes resulted in astrocyte detachment from the substrate and aggregation. These neutrophil concentrations affected both epithelioid and stellate HNK1/A2B5‐negative (type 1) astrocytes but not hippocampal neurons. Substrate detachment was partially prevented by corticosterone, but not by protease inhibitors or free‐radical scavengers.

Co‐cultures of purified cortical astrocytes with neutrophils (1/20 cell ratio) contained at the beginning of the experiment ε93‐98% astrocytes with type 1 markers. After 6‐8 days co‐culture, many stellate cells insensitive to neutrophils seemed to migrate out of the aggregates. About 70% of these resistant cells had immunological markers typical of type 2 astrocytes. The possible relevance of these findings to reactive astrogliosis and secondary neuronal death is discussed.
Development of cell‐cell coupling among cells of the oligodendrocyte lineage
Authors: von Blankenfeld G. , Ransom B. R. , Kettenmann H.
Published: January 2005

The development of functional gap‐junctional communication was studied in cells of the oligodendrocyte lineage. The presence of cell‐cell coupling was determined by the passage of current between cells using a double whole‐cell patch‐clamp system or by injecting the low molecular weight dye Lucifer yellow into individual cells via a patch pipette and observing the diffusion of the dye into adjacent cells. The developmental stage of the cells under study was determined using antibodies to specific surface markers (04,01, and 010) that characterize cells of sequential maturity along the oligodendrocyte lineage (Kuhlmann‐Krieg et al., 1988; Sommer and Schachner, 1981; 1982). Both stages of precursor cells of this lineage, 04+ and 04−, almost never showed dye or electrical coupling, even though they were in close physical contact with other cells. The O1‐positive oligodendrocytes with simple morphology were also noncoupled, with only few exceptions. In contrast, more than 40 percent of more mature, O10‐positive oligoden‐drocytes showed cell‐cell coupling detectable with both dye and current injection. Thus, the formation of gap junctions between cells of the oligodendrocyte lineage does not occur with, but some time after, the commitment of the cell to becoming an oligodendrocyte.
Effects of interleukin‐1β and tumor necrosis factor‐α on the expression of glial fibrillary acidic protein and transferrin in cultured astrocytes
Authors: Oh Young J. , Markelonis George J. , Oh Tae H.
Published: January 2005

Recent evidence suggests that interleukin (IL)‐1 and tumor necrosis factor (TNF) may play a role in astrogliosis following injury to the CNS. The short‐term biochemical effects of these immune‐related cytokines were determined on cultured rat polygonal and process‐bearing astrocytes. Both IL‐1 and TNF stimulated the rate of thymidine incorporation in polygonal astrocytes up to 137% and 215%, respectively, over the level observed in untreated controls. By contrast, thymidine incorporation was relatively unaffected by these cytokines in process‐bearing astrocytes. The cytokines did not significantly affect the level of glial fibrillary acidic protein (GFAP) within polygonal astrocytes, even though they appeared to downregulate the expression of GFAP mRNA by as much as 62%. Both cytokines increased the intracellular expression of transferrin (Tf) within some polygonal astrocytes. In untreated control cultures, fewer than 2% of polygonal astrocytes were immunoreactive for Tf. By contrast, approximately 30% of polygonal astrocytes treated with IL‐1 or TNF‐α became strongly immunoreactive for Tf. Neither IL‐2 nor a number of other known growth factors appeared to alter the level of immunoreactive Tf in these cells. Process‐bearing astrocytes were negative for Tf, regardless of the treatment used. Northern blot analysis demonstrated that the level of Tf mRNA in cultures of polygonal astrocytes increased 148% above the level observed in untreated controls following treatment with either IL‐1 or TNF, whereas no change was observed following treatment with IL‐2. These results suggest that increased levels of particular cytokines known to be present in injured CNS can produce pronounced biochemical alterations within a subtype of cultured astrocytes. © 1993 Wiley‐Liss, Inc.
N‐ and R‐cadherin expression in the optic nerve of the chicken embryo
Authors: Redies Christoph , Takeichi Masatoshi
Published: January 2005

Cadherins are a family of molecules mediating Ca2+‐dependent cell‐cell adhesion in various tissues. N ‐ and R‐cadherin are expressed in the chick embryonic CNS and differ in their expression pattern during development. Here we focus on the differential expression of N ‐ and R‐cadherin in the early optic nerve. N‐cadherin is expressed by the retinal neurites growing through the optic nerve. R‐cadherin is expressed by the early optic nerve glia, which derives from the optic stalk neuroepithelium and corresponds to an immature form of the type‐1 astrocyte described in rat optic nerve. The close contact between the plasma membranes of the retinal neurites and the optic nerve glia is believed to be important in guiding retinal axons through the optic nerve. Using neuroblastoma cell lines transfected with R‐cadherin, we demonstrate that the N‐cadherin‐positive retinal axons can use R‐cadherin as a substrate for axon elongation. These results suggest that the R‐cadherin expressed by the early optic nerve glia might provide a molecular substrate for the growth of N‐cadherin‐positive retinal axons through the optic nerve. © 1993 Wiley‐Liss, Inc.
Development of an astrocytic response to lesions of the spinal cord in the north american opossum: An immunohistochemical study using anti‐glial fibrillary acidic protein
Authors: Ghooray Ganesh T. , Martin George F.
Published: January 2005

We have shown previously that rubral axons grow around a lesion of their pathway in developing opossums and that a critical period exists for that plasticity. The critical period begins when rubral axons first reach the level of the lesion and ends sometime between postnatal days (PD) 26 and 30. The aim of the present study was to examine the development of an astrocytic response to lesioning the spinal cord to determine if there is a temporal correlation between the development of such a response and the end of the critical period. The astrocytic response was examined immunohistochemically, 2 and 4 weeks after hemisecting the thoracic spinal cord, using an antibody to glial fibrillary acidic protein (GFAP). A response was first seen at PD21 in the 2‐week series. The response was relatively mild, however, and limited to the white matter. When the lesion was made at PD26. the response was still restricted to the white matter, but hypertrophied astrocytes were found at the gray/white matter junction and cystic cavities were present. When the lesion was made at PD41, the response had spread to the gray matter and it occupied a larger area rostral and caudal to the lesion than at earlier ages. The animals allowed to survive 4 weeks after lesioning were subjected to a second operation 4‐5 days before sacrifice so that Fast Blue could be injected bilaterally two to three segments caudal to the lesion. When the hemisection was made at PD15, a response was present in the ventral and ventrolateral funiculi, but not in that part of the lateral funiculus that contains rubrospinal axons. As expected from previous studies, rubral neurons were labeled contralateral to the lesion, suggesting that their axons had grown around it. When the lesion was made at PD21, the glial response was still limited to the white matter, but it extended into that part of the lateral funiculus that contains rubral axons. In spite of the presence of a glial response, rubral neurons were still labeled contralateral to the lesion. When the lesion was made at PD26, hypertrophied astrocytes were present at the gray/white matter junction and small cavities were noted at the lesion site. In such cases, there was no evidence for rubrospinal plasticity. An astrocytic response was not observed in the gray matter of the dorsal horn, an area used by rubral axons to grow around a lesion of their pathway, until well after the end of the critical period. We conclude that the initial development of a glial scar in the white matter after lesioning does not determine the end of the critical period for rubrospinal plasticity. Loss of rubrospinal plasticity correlates most closely with the appearance of hypertrophied astrocytes at the gray/white matter junction and the formation of cystic cavities.
Voltage‐dependent sodium and potassium currents in cultured trout astrocytes
Authors: Glassmeier G. , Jeserich G. , Krüppel T.
Published: January 2005

Voltage‐gated ionic currents were recorded from cultured trout astrocytes with the whole‐cell variation of the patch‐clamp technique. In a subpopulation of astrocytes depolarizations above −40 m V activated a fast transient inward current that was identified as a sodium current by ion substitution experiments, its current reversal potential, and its TTX‐sensitivity. Regarding threshold of activation, peak current voltage, and amplitude this current closely resembled those previously described for mammalian astrocytes. Voltage‐dependence of inactivation and kinetics, however, markedly differed from the “glial‐like” sodium current occurring in mammalian hippocampal or optic nerve astrocytes, since the sodium current of trout astrocytes exhibited a faster time course of activation and decay and a more depolarized steady‐state inactivation curve with midpoints close to −60 mV. During a period of 2 weeks in culture the biophysical properties of the sodium current did not change significantly, albeit a continuous decrease in current density was observed. At depolarizing voltage steps positive to −40 mV, additionally voltage‐gated potassium outward currents were evoked, which could be separated into a steady‐state current with delayed rectifier properties and an inactivating component resembling the A‐type current. Moreover, in a subpopulation of astrocytes an inward potassium current was elicited at hyperpolarizing potentials, which exhibited biophysical features consistent with the potassium inward rectifier of mammalian astrocytes. © 1994 Wiley‐Liss, Inc.
Psychiatric morbidity among residents in a home for the elderly in spain: Prevalence of disorder and validity of screening
Authors: Lobo Antonio , Ventura Tirso , Marco Carlos
Published: January 2005

All residents in a home for elderly válidos (able) in Spain were interviewed and 101 completed the study. Non‐medical interviewers adminstered the GHQ‐28 items and either Zung's Self‐Rating Depression Scale (SDS) (N=69) or the Geriatric Depression Rating Scale (GDS) (N=31). The residents were also examined blind by psychiatrists standardized in the use of the Clinical Interview Scale (CIS), who also administered the ‘Mini‐Examen Cognoscitivo’, the Spanish version of the Mini‐Mental State Examination (MMSE), and diagnosed the identified ‘cases’ according to ICD‐9 criteria. Twenty‐one residents (20.8%) were considered to be ‘cases’ on the basis of the CIS; 14 (13.9%) were diagnosed as depressed and only four (4%) were diagnosed as being demented, although the mean age of the sample was 81.9 years (SD=5.6). Most ‘cases’; were of mild intensity and the differences with ‘non‐cases’; seemed quantitative rather than qualitative. The GHQ‐28 correctly identified 81.5% of ‘cases’; with a cutting score of 5/6 (sensitivity 70%, specificity 84.4%) and 77.4% of ‘cases’; with a cutting score of 4/5 (sensitivity 85%, specificity 75.3%), suggesting a discriminative power almost as good as in adult individuals. In the pilot study, the GDS performed better than the SDS as a screening instrument. The addition of the ‘Mini‐Examen’ would have improved the sensitivity of both the GHQ‐28 items and the SDS.
Cognitive effects of ect in the elderly: Preliminary findings
Authors: Russ Mark J. , Ackerman Sigurd H. , Burton Leslie , Shindledecker Richard D. , Goldberg Elkhonon
Published: January 2005

Clinical recovery, cognitive function, and ward behavior indicative of organic brain syndrome were assessed in 12 elderly, depressed patients before, and approximately one week after, treatment with electroconvulsive therapy (ECT). There was a statistically significant improvement in depression, and ward behavior, whereas cognitive function was unchanged. These findings support and extend prior studies which indicate that ECT in the treatment of depression in the elderly is not associated with functionally significant decrements in cognitive performance.