bluuebunny/crossref_metadata_2025_split huggingface dataset:
Binding and action of glucagon in cultured mouse astrocytes
Authors: Cockram C. S. , Kum W. , Ho S. K. S. , Zhu S. O. , Young J. D. 
Published: December 2004

AbstractThis study investigates glucagon binding in primary cultures of differentiated mouse astrocytes and the effect of glucagon on intracellular cAMP accumulation.Binding of 125I‐glucagon (0.53 nM) to mouse astrocyte suspensions reached equilibrium after 10 min at 22°. Equilibrium binding corresponded to 46 · 15 pmol/mg protein (n = 3) representing approximately 10,000 occupied sites per cell at the tracer concentration used. Dissociation occurred with a half‐time of 2.5 min at 22°C and was not accelerated in the presence of unlabelled glucagon (1 M). Scatchard analysis suggested the presence of more than one class of binding site. The Ka for the higher affinity sites was 5.7–7.4 × 106 M−1. The Ka for the lower affinity sites was 3.6–5.3 104 M−1. The results suggest the presence of approximately 43,000 high affinity sites per cell. Binding was inhibited by unlabelled glucagon with an IC50 of 50 nM but unaffected by insulin and somatostatin. However, no 125I‐glucagon binding could be detected when intact monolayer cells attached to culture dishes were used.Glucagon stimulated cyclic‐AMP accumulation in both cell suspensions and intact monolayer cells in a dose‐dependent fashion. However high concentrations were required when compared to the receptor‐binding studies.Marked degradation of 125I‐glucagon by astrocytes during binding experiments was observed and this was inhibited by unlabelled glucagon but also by insulin and desoctapeptide insulin. © 1995 Wiley‐Liss, Inc.
Glial cell types, lineages, and response to injury in rat and fish: Implications for regeneration
Authors: Sivron Tomer , Schwartz Michal 
Published: December 2004

AbstractAxons of the mammalian central nervous system do not regenerate spontaneously after axonal injury, unlike the central nervous system axons of fish and amphibians and the peripheral nervous system of mammals, which possess a good regenerative ability (Grafstein: The Retina: A Model for Cell Biology Studies, Part II, 1986; Kiernan: Biol Rev 54:155–197, 1979; Murray: J Comp Neurol 168:175–196, 1976; Ramony Cajal: Degeneration and Regeneration of the Nervous System, 1928; Reier and Webster: J Neurocytol 3:591–618, 1974; Sperry: Physiol Zool 23:351–361, 1948). It was previously believed that intrinsic differences between the central nervous system neurons of mammals and fish account for their differences in regenerative ability. The past decade, however, has seen an accumulation of evidence, indicating that mammalian central nervous system neurons are able to regenerate injured axons, at least to some extent. This was first demonstrated by Aguayo and colleagues (David and Aguayo: Science 214:931–933, 1981; Kierstead et al: Science 246:255–257, 1989), who showed that injured mammalian central nervous system axons can grow for a considerable distance into an autograft of a peripheral nerve. It was also demonstrated that injured rabbit optic axons can regenerate into their own environment (i.e., into the distal part of the injured optic nerve), if the injured nerve is treated so as to make it conducive for growth (Lavie et al: J Comp Neurol 298:293–314, 1990; Eitan et al: Science 264:1764–1768, 1994). It is generally believed today that the glial cells surrounding the axons play a crucial role in determining the regenerative capacity of any central nervous system. In this review, we discuss various aspects of glial cells in the context of regeneration, with special emphasis on similarities and differences between the glial cells of rat and fish optic nerves, representing a nonregenerative and a regenerative system, respectively. © 1995 Wiley‐Liss, Inc.
Differential Na+ channel β1 subunit mRNA expression in stellate and flat astrocytes cultured from rat cortex and cerebellum: A combined in situ hybridization and immunocytochemistry study
Authors: Oh Youngsuk , Waxman Stephen G. 
Published: December 2004

AbstractAstrocytes have been shown to express voltage‐sensitive Na+ channels, but the molecular structure of these channels is not yet known. Recent studies have demonstrated the expression of rat brain voltage‐sensitive Na+ channel mRNAs in astrocytes. In this study, we used a combined non‐radioactive in situ hybridization immunocytochemistry method to investigate the expression of voltage‐sensitive Na+ channel ß1 subunit (Naß1) mRNA in definitively identified, GFAP‐positive astrocytes cultured from two different regions of the rat brain, cerebrum and cerebellum. In general, two morphologically distinct types of GFAP‐positive astrocytes were observed in culture: flat, fibroblast‐like and stellate, process‐bearing. We observed a differential expression ofNaß1 mRNA in GFAP‐positive astrocytes: (1) stellate astrocytes expressed Naß1 mRNA, although the level of Naß1 mRNA expression was variable, and (2) flat astrocytes generally did not express Naß1 mRNA. Moreover, Bergmann‐like cells from cerebellum did not express Naß mRNA, while the granule cells associated with Bergmann‐like cell expressed Naß mRNA. These observations indicate that Naß mRNA is differentially expressed in rat astrocytes with various morphologies in vitro. © 1995 Wiley‐Liss, Inc.
Indicator expression directed by regulatory sequences of the glial fibrillary acidic protein (GFAP) gene: In vivo comparison of distinct GFAP‐lacZ transgenes
Authors: Johnson William B. , Ruppe Mary D. , Rockenstein Edward M. , Price Jennifer , Sarthy Vijay P. , Verderber Lisa C. , Mucke Lennart 
Published: December 2004

AbstractAn increase in the expression of the glial fibrillary acidic protein (GFAP) gene by astrocytes appears to constitute a crucial component of the brain's response to injury because it is seen in many different species and features prominently in diverse neurological diseases. Previously, we have used a modified GFAP gene (C‐339) to target the expression of β‐galactosidase (β‐gal) to astrocytes in transgenic mice (Mucke et al.; New Biol 3:465–474 1991). To determine to what extent the in vivoThroughout this text, “in vitro” refers to cells grown in culture in culture and “in vivo” to living animals.
expression of GFAP‐driven fusion genes is influenced by intragenic GFAP sequences, the E. coli lacZ reporter gene was either placed downstream of approximately 2 kb ofmurine GFAP 5′ flanking region (C‐259) or ligated into exon 1 of the entire murine GFAP gene (C‐445). Transgenic mice expressing C‐259 versus C‐445 showed similar levels and distributions of β‐gal activity in their brains. Exclusion of intragenic GFAP sequences from the GFAP‐lacZ fusion gene did not diminish injury‐induced upmodulation ofastroglial β‐gal expression or increase β‐gal expression in non‐astrocytic brain cells. These results demonstrate that 2 kb ofmurine GFAP 5′ flanking region is sufficient to restrict transgene expression primarily to astrocytes and to mediate injury‐responsiveness in vivo. This sequence therefore constitutes a critical target for mediators of reactive astrocytosis. While acute penetrating brain injuries induced focal increases in β‐gal expression around the lesion sites in C‐259, C‐445, and C‐339 transgenic mice, infection of C‐339 transgenic mice with scrapie led to a widespread upmodulation of astroglial β‐gal expression. Hence, GFAP‐lacZ transgenic mice can be used to monitor differential patterns of astroglial activation in vivo. These and related models should facilitate the assessment of strategies aimed at the in vivo manipulation of GFAP expression and astroglial activation. © 1995 Wiley‐Liss, Inc.
The receptor for tumor necrosis factor on murine astrocytes: Characterization, intracellular degradation, and regulation by cytokines and theiler's murine encephalomyelitis virus
Authors: Aránguez Isabel , Torres Concepción , Rubio Nazario 
Published: December 2004

AbstractPrevious reports have shown that tumor necrosis factor (TNF) exerts a role on the physiology of astrocytes under inflammatory situations. The signalling for biological effects of this and other cytokines are usually exerted through cell surface receptors. In this study, we have demonstrated the presence of a surface TNF α receptor type I in murine astrocytes of both SJL/J and BALB/c origin, using 125I‐labelled recombi‐nant mouse TNFα. A linear Scatchard plot indicates the presence of only one type of receptor with a MW of 58 kDa (Type I TNF receptor) that binds the ligand with a Kd of 1 10−9 M. There are 3,000 copies of this receptor on untreated astrocytes. The results also indicate that receptor‐bound TNF is rapidly internalized at 37°C and degraded intracellularly to a principal molecular species which elutes from HPLC reverse‐phase columns at 38% acetonitrile rather than at 60% as native TNFα does. The binding is up‐regulated by increasing the number of receptors (but not its affinity) by treatments with Theiler's murine encephalomyelitis virus (TMEV), Con A and inflammatory cyto‐kines such as IL‐1, IL‐6 and INF‐β. It is not influnced by vaccinia virus, IL‐2, or LPS. This receptor may contribute to the initiation of perpetuation of the immune response which mediates the demyelinating inflammation induced by Theiler's virus. © 1995 Wiley‐Liss, Inc.
Disruption of the blood‐brain interface in neonatal rat neocortex induces a transient expression of metallothionein in reactive astrocytes
Authors: Penkowa Milena , Moos Torben 
Published: December 2004

AbstractExposure of the adult rat brain parenchyma to zinc induces an increase in the intracerebral expression of the metal‐binding protein, metallothionein, which is normally confined to astrocytes, ependymal cells, choroid plexus epithelial cells, and brain endothelial cells. Metallothionein is expressed only in diminutive amounts in astrocytes of the neonatal rat brain, which could imply that neonatal rats are devoid of the capacity to detoxify free metals released from a brain wound. In order to examine the influence of a brain injury on the expression of metallothionein in the neonatal brain, PO rats were subjected to a localized freeze lesion of the neocortex of the right temporal cortex. This lesion results in a disrupted blood‐brain interface, leading to extravasation of plasma proteins. From 16 h, reactive astrocytosis, defined as an increase in the number and size of cells expressing GFAP and vimentin, was observed surrounding the neocortical lesion site. Astrocytes and pial cells situated adjacent to the area of injury also became positively stained for metallothionein. At 3–6 days post‐lesion, the highest level of reactive astrocytes expressing metallothionein was observed. Neo‐Timm staining revealed that histochemically reactive zinc had disappeared from the lesion site. Extracellular albumin and metallolhionein‐positive astrocytes were absent approximately 2 weeks after the lesion, whereas reactive astrocytosis was still observed. These results show that a lesion of the neonatal rat brain induces a transient expression of metallothionein in reactive astrocytes, probably as a response to metals released from the site of the brain injury. © 1995 Wiley‐Liss, Inc.
The human blood‐brain barrier glucose transporter (GLUT1) is a glucose transporter of gray matter astrocytes
Authors: Morgello Susan , Uson Ronald R. , Schwartz Elissa J. , Haber Richard S. 
Published: December 2004

AbstractHuman and monkey brain sections were examined by immunohistochemical light and electron microscopy to determine the distribution of GLUT1, a glucose transporter isoform associated with erythrocytes and endothelial cells of the human blood‐brain barrier. Protein immunoblotting of fractionated human brain membranes was performed to determine the distribution of molecular forms of the transporter. GLUT1 staining was abundant in erythrocytes and cerebral endothelium of gray and white matter but was also present diffusely in gray matter neuropil when viewed by light microscopy. Immunoelectron microscopy confirmed the gray matter and vascular localization of GLUT1, with specific GLUT1 staining seen in erythrocytes, gray and white matter endothelial cells, astrocyte foot processes surrounding gray matter blood vessels, and in astrocyte processes adjacent to synaptic contacts. No astrocytic staining was identified in white matter. Astrocyte GLUT1 staining was identified only in mature gray matter regions; undifferentiated regions of preterm (22–23 weeks gestation) cortex had GLUT1 staining only in blood vessels and erythrocytes, as did germinal matrix. Immunoblots of adult human frontal cortex revealed that two forms of GLUT1 (45 and 52 kDa) were present in unfractionated brain homogenates. Immunoblots of vessel‐depleted frontal lobe revealed only the 45 kDa form in gray matter fractions, and depleted in membranes prepared from white matter regions. We conclude that the GLUT1 isoform of glucose transporter is present both in endothelium of the blood‐brain barrier and in astrocytes surrounding gray matter blood vessels and synapses. Furthermore, the form present in astrocytes is likely to have a lower molecular weight than the form found in cerebral endothelium. The GLUT1 transporter may play an important role not only in astrocyte metabolism, but also in astrocyte‐associated pathways supporting neuronal energy metabolism. © 1995 Wiley‐Liss, Inc.
Hypoxia‐induced vascular endothelial growth factor expression in normal rat astrocyte cultures
Authors: Ijichi Akihiro , Sakuma Shirou , Tofilon Philip J. 
Published: December 2004

AbstractVascular endothelial growth factor (VEGF) is an endothelial cell‐specific mitogen, which also enhances vascular permeability. Because this angiogenic factor has been suggested to play a role in brain tumor biology, we have begun to investigate the regulation of VEGF expression in cultures of rat type I astrocytes. In this report, we have focused on the influence of hypoxia on VEGF expression. Under standard in vitro conditions (21% O2) VEGF expression in astrocytes is barely detectable by northern analysis. However, after exposure to 0.2% O2 for as little as 3 h VEGF mRNA levels are markedly increased reaching a maximum by approximately 8 h of exposure. Treatment of astrocytes with CoCl2 or desferrioxamine results in a similar induction of VEGF, suggesting that the oxygen sensor regulating VEGF expression in astrocytes is a heme‐containing molecule. Although acute treatment with TPA (6 h) induces VEGF expression, chronic exposure to TPA (24 h) to deplete PKC activity does not reduce the hypoxia‐induced VEGF expression. These data indicate that VEGF induction in astrocytes can proceed through PKC‐dependent and ‐independent pathways. Furthermore, chronic exposure to TPA or treatment with herbimycin A results in the enhancement of the hypoxia‐mediated increase in VEGF mRNA levels. These results suggest that PKC and herbimycin‐sensitive tyrosine kinase may serve as negative regulators of the hypoxia‐activated signal transduction pathway that leads to the induction of VEGF expression. However, treatment of astrocytes with the nonspecific kinase inhibitors H7 and H8 reduced the level of VEGF induction by hypoxia, indicating that some type of kinase activity is required in this signaling pathway. © 1995 Wiley‐Liss, Inc.
Interferon‐γ and lipopolysaccharide reduce cAMP responses in cultured glial cells: Reversal by a type IV phosphodiesterase inhibitor
Authors: Patrizio Mario , Costa Tommaso , Levi Giulio 
Published: December 2004

AbstractThe aim of the present study was to determine whether two classical macrophage activators, bacterial lipopolysaccharide (LPS) and interferon‐γ (IFN‐γ) could affect the accumulation of the second messenger cAMP in cultured rat microglia and astrocytes. Purified microglia and astrocyte secondary cultures obtained from the neonatal rat were grown for 3 days in basal medium Eagle (BME) + 10% fetal calf serum (FCS). Exposure of microglia to LPS resulted into a dose‐ and time‐dependent decrease in the accumulation of cAMP induced by receptor‐mediated (isoproterenol or prostaglandin E2) or direct (forskolin) activation of adenylate cyclase. The inhibitory effect of LPS was rapid (a 10 min preincubation was sufficient to approach a maximal effect), occurred at low doses (IC50 = 1.2 ng/ml), and was not abrogated by pertussis toxin. A selective inhibitor of type IV phosphodiesterase (rolipram, 100 nM) prevented the effect of LPS on cAMP accumulation, while inhibitors of other forms of phosphodiesterase were unable to do so. IFN‐γ (100 u/ml) also caused a depression of the evoked cAMP accumulation in microglia after a 10 min preincubation, and its effect was prevented by rolipram, as in the case of LPS. Astrocytes differed from microglia in that LPS (1–100 ng/ml) did not inhibit the accumulation of cAMP induced by either isoproterenol or forskolin; on the other hand, IFN‐γ did have an inhibitory effect (though less pronounced than in microglia) that could be prevented by rolipram. Our observations indicate that two potent activators of microglia acting at different receptors, LPS and IFN‐γ, can diminish the accumulation of cAMP through a common mechanism, the stimulation of a specific form of cAMP phosphodiesterase. The fact that IFN‐γ, but not LPS, was effective in astrocytes suggests that LPS receptors are scarcely, if at all, expressed in these cells, or that they are differently coupled to second messengers. Selective inhibitors of type IV phosphodiesterase might prevent some of the obnoxious actions of LPS or IFN‐γ in the living organism. © 1995 Wiley‐Liss, Inc.
Multiply myelinated axons in the optic nerve of mice deficient for the myelin‐associated glycoprotein
Authors: Bartsch Udo , Montag Dirk , Bartsch Susanne , Schachner Melitta 
Published: December 2004

AbstractWe recently reported that some retinal ganglion cell axons in mice deficient for the myelin‐associated glycoprotein are concentrically surrounded by more than one myelin sheath. In the present study, we demonstrate that myelin sheaths displaced from the axon reveal a normal ultrastructure of compact myelin, with the only exception that multiple myelination of axons frequently correlates with the presence of unfused regions of major dense lines. Supernumerary sheaths terminated on other sheaths or on astrocyte cell surfaces in a pattern closely resembling the morphology of a true paranode. The thickness of compact myelin of multiply myelinated axons was significantly increased when compared with axons of similar caliber surrounded by a single myelin sheath. Our observations demonstrate that maintenance of compact myelin and paranodal regions is not dependent on direct axonal contact and that the presence of more than one concentric myelin sheath around an axon results in dysregulation of the axon‐to‐fiber ratio. © 1995 Wiley‐Liss, Inc.
Isoform‐specific expression of sodium channels in astrocytes in vitro: Immunocytochemical observations
Authors: Black J. A. , Westenbroek R. , Minturn J. E. , Ransom B. R. , Catterall W. A. , Waxman S. G. 
Published: December 2004

AbstractThe expression of sodium channel α‐subunit isoforms in astrocytes cultured from P‐0 rat spinal cord and P‐7 rat optic nerve was examined utilizing immunocytochemical methods with antibodies generated against conserved and isoform‐specific amino acid sequences of the rat brain sodium channel. In spinal cord cultures at 5 days in vitro (DIV), both stellate and flat astrocytes were immunostained with antibody SP20, which recognizes a conserved sequence common to sodium channel types I, II/IIA, and III. Antibody SP11‐I, which is directed against a subtype‐specific sequence in sodium channel I, did not yield detectable staining in spinal cord astrocytes. Antibody SP11‐II, which is directed against a subtype‐specific sequence in sodium channel II, immunostained both stellate and flat spinal cord astrocytes, although with less intensity than SP20. Antibody SP32‐III, which is directed against a subtype‐sequence in sodium channel III, immunostained stellate but not flat spinal cord astrocytes. SP20, SP11‐II, and SP32‐III staining persisted in stellate spinal cord astrocytes through 14–21 DIV, while SP20 and SP11‐II immunostaining in flat spinal cord astrocytes was attenuated with time in culture. In optic nerve cultures at 5 DIV, SP20 staining was present in both stellate and flat astrocytes, but at reduced levels compared to spinal cord astrocytes. With increased time in culture, SP20 staining was maintained in stellate optic nerve astrocytes but was gradually lost in flat optic nerve astrocytes. Stellate optic nerve astrocytes exhibited low levels of staining with SP11‐I, SP11‐II, and SP32‐III. Flat optic nerve astrocytes lacked or displayed very low SP11‐II staining, and SP11‐I and SP32‐III staining was not detectable. These observations demonstrate that cultured astrocytes are immunoreactive to antibodies generated against conserved and isotype‐specific peptide sequences of rat brain sodium channels, and further suggest that there are different patterns of sodium channel expression between flat vs. stellate astrocytes and in astrocytes derived from different regions of the CNS. © 1995 Wiley‐Liss, Inc.
Intracellular acidification and Ca2+ transients in cultured rat cerebellar astrocytes evoked by glutamate agonists and noradrenaline
Authors: Brune Thomas , Deitmer Joachim W. 
Published: December 2004

AbstractThe effect of different neurotransmitters on the intracellular pH (pHi) and intracellular calcium (Ca2+i) was studied in cultured astrocytes from neonatal rat cerebellum, using the fluorescent dyes 2,7′‐bis(carboxyethyl)‐5,6‐carboxy‐fluorescein (BCECF) and Fura‐2. Application of glutamate or kainate (100 μM) in a HEPES‐buffered, CO2/HCO3−‐free saline induced a decrease in pHi and an increase in Ca2+i. Amplitude and time course of the pHi and Ca2+i transients were different. Glutamate and kainate evoked a mean acidification of 0.22 ± 0.05 (n = 29) and 0.20 ± 0.04 (n = 12) pH units, respectively. The changes in pHi and Ca2+i induced by kainate, but not by glutamate, were inhibited by 6‐cyano‐7‐dinitroquinozalin‐2,3‐dion (CNQX; 50 μM). In order to elucidate the mechanism of the agonist‐induced acidification, whether the pHi changes were secondary to the Ca2+ rises was tested. In the absence of extracellular Ca2+, the kainate‐induced Ca2+i transient was suppressed, while the intracellular acidification was only reduced by 13%. Removal of extracellular Ca2+ reduced the glutamate‐induced pHi change by 8%, while the second component of the Ca2+i transient was abolished. Application of trans‐(±)‐1‐amino‐(1S,3R)‐cyclopentadicarboxylic acid (t‐ACPD, 100 μM), a metabotropic glutamate receptor agonist, and of noradrenaline (20 μM) evoked a Ca2+i increase, but no change of pHi. D‐aspartate, which has a low affinity to glutamate receptors, but is known to be transported by the glutamate uptake system in some astrocytes, evoked an intracellular acidification, similar to that induced by glutamate, but no Ca2+i transient. The results suggest that the kainate‐induced acidification is only partly due to the concomitant Ca2+i rise, while the glutamate/aspartate‐induced acidification is mainly due to the activation of the glutamate uptake system. © 1995 Wiley‐Liss, Inc.