Biomedical subjects

A Represa

Publications and source records attributed to A Represa.

At least 37 records · Page 2Linked to original sources

PubMed1998

Ultrastructural morphology of neuronal death following reversible focal ischemia in the rat.

Electron microscopy and terminal deoxynucleotidyl transferase (TdT) mediated dUTP-biotin nick end-labelling (TUNEL) were used to illustrate the different stages and subcellular alterations of cell degeneration that occur in the striatum of the rat after transient focal ischemia. Degenerating neurons exhibited different morphological types: apoptosis Type 1 (aggregation of dense masses of chromatin beneath the 'intact' nuclear membrane) and Type 2 (high cytoplasmic vacuolization), and necrosis. These profiles were localized in different part of the striatum. Type 1 was found in the head of the caudate putamen, Type 2 in the middle part of the striatum and necrosis in the striatal core. These ultrastructural results demonstrated that apoptosis occurs in neurons following focal ischemia in the striatal penumbra. In contrast, necrosis can be observed in the ischemic core, the region maximally affected by the ischemia. Finally, the presence of astrocytes throughout both the penumbra and ischemic core displaying numerous cytoplasmic vacuoles suggested an activation of glial cells.

Journal Article↗

PubMed1998

MAP2d mRNA is expressed in identified neuronal populations in the developing and adult rat brain and its subcellular distribution differs from that of MAP2b in hippocampal neurones.

The brain microtubule-associated protein MAP2 family is composed of high-molecular-weight (MAP2a and MAP2b) and low-molecular-weight (MAP2c and MAP2d) isoforms. The common C-terminal region of HMW MAP2 and MAP2c contains three repeated microtubule-binding domains while MAP2d comprises four repeats. MAP2c mRNA is known to be expressed at high levels in the immature brain. We show that in the brains of rat pups, MAP2c mRNAs are indeed expressed at high levels compared with MAP2d. However, in adult rat brains, MAP2d mRNA levels are higher than MAP2c. In order to identify the neural cells expressing MAP2d, we used in situ hybridization. In vivo, we show that MAP2d mRNA is expressed in well-identified neuronal populations of the brain. In primary cultures of hippocampal neurones, double-labelling experiments confirm that MAP2d is clearly expressed in neurones. We also evaluated in this study the subcellular distribution of the MAP2d mRNAs in cultured hippocampal neurones and we report that in contrast with MAP2b mRNAs, mostly localized in dendrites, MAP2d mRNAs are essentially located in neuronal cell bodies.

Animals↗

PubMed1998

Long-lasting enhanced expression in the rat hippocampus of NMDAR1 splice variants in a kainate model of epilepsy.

Chronic epilepsy is associated with increased excitability which may result from abnormal glutamatergic synaptic transmission involving altered properties of N-methyl-D-aspartate (NMDA) receptors. To date two gene families encoding NMDA receptor subunits have been cloned, NR1 and NR2. Eight NR1 mRNAs are generated by alternative splicing of exons 5, 21 and 22; the NR1-1 to NR1-4 C-terminal variants exist in the a or b version depending on the presence or absence of the domain encoded by exon 5. Epilepsy was induced in rats by unilateral intra-amygdalar injection of kainate and animals were killed from 6 h to 4 months following the injection. Increased NR1 mRNA levels were observed during status epilepticus (6-24 h after the injection), both psilateral and contralateral, while a second wave of NMDAR1 mRNA increase occurred in chronic epileptic animals, between 21 days and 4 months following kainate injection. Our data show: (i) a permanent increase of the NR1-2a and NR1-2b mRNA species (containing exon 22) in all hippocampal fields, both ipsilateral and contralateral, and (ii) an increase of the NR1-3 (a and b) mRNAs (containing exon 21) in the ipsilateral CA1, and NR1-3a mRNA in the ipsilateral dentate gyrus. No long-term changes were observed for the NR1-1 and NR14 splice variants. In the ipsilateral CA3 area a globally decreased mRNA expression was associated with neuronal loss. A possible contribution to the maintenance of the epileptic state by an increased expression of NMDA receptors is discussed.

Animals↗

PubMed1997

Differential expression of fibronectin, tenascin-C and NCAMs in cultured hippocampal astrocytes activated by kainate, bacterial lipopolysaccharide or basic fibroblast growth factor.

Different reports demonstrated that reactive glial cells express increased amounts of adhesion and matrix molecules. Despite a wealth of information on the expression of these molecules during development and after lesion, very little is known of how this expression is regulated. In the present report we used Western blots and immunocytochemistry to investigate the expression of neural cell adhesion molecule (NCAM), fibronectin and tenascin-C in cultured astrocytes from rat hippocampus. The effects of three different extracellular signals were analyzed: the glutamatergic receptor agonist kainic acid, the basic fibroblast growth factor (bFGF) and the bacterial lipopolysaccharide. Each treatment had a specific pattern of glial activation and differentially modified the expression of these proteins. Treatment of astrocytes with kainic acid resulted in an increase of tenascin-C, a decrease of fibronectin and a shift of NCAMs isoforms: NCAM 140 and PSA-NCAM (polysialic acid-rich NCAMs) were increased while NCAM 120 was decreased, bFGF increased fibronectin, tenascin-C and NCAM 120, while decreasing PSA-NCAM. Finally, the treatment of astrocytes with lipopolysaccharide induced a significant increase of fibronectin, tenascin-C and NCAM 120 but did not modify the expression of NCAM 140 and PSA-NCAM. These data suggest different mechanisms for modulation of cell surface interactions. They suggest that glial activation by bFGF and lipopolysaccharide are associated with an increase of the adhesive properties, while kainate action is rather associated with a decrease of the adhesiveness of astrocytes.

Animals↗

PubMed1997

Effects of tenascin-C in cultured hippocampal astrocytes: NCAM and fibronectin immunoreactivity changes.

Tenascin-C is an extracellular matrix glycoprotein with trophic and repulsive properties on neuronal cells, involved in migratory processes of immature neurons. Previous reports demonstrated that this molecule is produced and secreted by astrocytes, in vitro after activation by bFGF or in vivo after CNS lesion. In injured brain the expression of tenascin-C has been correlated with the glial reaction since it was observed in regions suffering a dramatic glial proliferation and hypertrophy. In this report we show that the treatment of cultured hippocampal astrocytes with tenascin-C results in an increased fibronectin and NCAM immunoreactivities. In addition, treated astrocytes form longer extensions than control ones. The number of cells as well as the levels of GFAP mRNA and protein immunoreactivity are not modified after tenascin-C treatment. The present changes may, therefore, be related to the modification of the adhesive properties of astrocytes to the substrate. These observations are compatible with the hypothesis that tenascin-C may contribute to the glial scarring process.

Animals↗

PubMed1997

NCAM is essential for axonal growth and fasciculation in the hippocampus.

The neural cell adhesion molecule (NCAM), probably the best characterized and most abundant cell adhesion molecule on neurons, is thought to be a major regulator of axonal growth and pathfinding. Here we present a detailed analysis of these processes in mice deficient for all NCAM isoforms, generated by gene targeting. The hippocampal mossy fiber tract shows prominent expression of polysialylated NCAM and the generation of new axonal projections throughout life. Focusing on this important intrahippocampal connection, we demonstrate that in the absence of NCAM, fasciculation and pathfinding of these axons are strongly affected. In addition we show alterations in the distribution of mossy fiber terminals. The phenotype is more severe in adult than in young animals, suggesting an essential role for NCAM in the maintenance of plasticity in the mature nervous system.

Aging↗

PubMed1997

FGF-2 induces nerve growth factor expression in cultured rat hippocampal neurons.

Basic fibroblast growth factor (FGF-2) is expressed in the hippocampus and has been demonstrated to promote neurotrophic effects on hippocampal neurons in vitro. We show that these neurons, even at the embryonic stage, express the mRNAs encoding the FGF receptors, bek and flg. We have characterized the effects of FGF-2 on the expression of nerve growth factor (NGF) using the reverse transcription-coupled polymerase chain reaction, in situ hybridization and immunocytochemistry. In hippocampal neurons grown in the absence of serum, FGF-2 exposure induces an important elevation of NGF mRNA expression followed by a marked increase in NGF immunoreactivity. Combining in situ hybridization with an NGF probe and microtubule-associated protein-2 (MAP2) immunocytochemistry we show that the induction of NGF mRNA by FGF-2 is localized in MAP2-immunoreactive neurons. These results suggest roles for FGF-2 in the development of hippocampal neurons and in the maintenance of connections in the central nervous system, particularly the septo-hippocampal pathway, via the regulation of an important neurotrophin.

Animals↗

PubMed1997

Molecular and cellular cascades in seizure-induced neosynapse formation.

Limbic seizures induce in vulnerable structures, such as the hippocampal complex, morphologic changes that may contribute to the development of epilepsy. They include neuronal cell death, reactive gliosis, and neosynapse formation. In temporal lobe epilepsy patients, cell death develops in Ammon's horn and the hilus. This cell death involves both necrotic and apoptotic mechanisms and is likely responsible for the initiation of the glial reaction that consists of astroglial and macrophage proliferation and hypertrophy. Reactive astrocytes acquire the phenotypic properties of type 2 astrocytes and express trophic factors (e.g., bFGF), cell adhesion molecules (e.g., NCAM), and substrate molecules (e.g., tenascin-C). Seizures induce in the hippocampus a synaptic remodeling of mossy fibers. Mossy fiber collaterals innervate granule cell dendrites, creating recurrent excitatory circuits. We suggest that collateral branches of MF originate under the influence of trophic factors and as a consequence of an overproduction of tubulin polymers. In fact, seizures induce a transient increased expression of tubulin and microtubule-associated proteins in granule cells and mossy fibers. Navigation of mossy fiber growth cones may be facilitated by the interaction with astrocytes, which would exert this effect by producing and excreting cell adhesion and substrate molecules. In light of the results discussed here, one can suggest that in the adult brain, activated astrocytes could contribute to the process of axonal outgrowth and synaptogenesis.

Animals↗

PubMed1996

Entactin immunoreactivity in immature and adult rat brain.

Entactin (nidogen) is a glycoprotein of 150 kDa mainly found in the basement membranes of peripheral tissues where it is co-localized and forms a very tight complex with the outgrowth-promoting molecule laminin. In the present report we tested by immunoblotting the specificity of polyclonal antibodies to laminin and entactin isolated from Engelbreth-Holm-Swarm (EHS) mouse sarcoma and investigated laminin and entactin immunoreactivities in the hippocampus of newborn, adult control and kainate-injured rats. The three polyclonal antibodies to laminin (two of them commercial) used in the present study stained somas of neurons, blood vessels and reactive glial cells, in agreement with previous reports. Nevertheless, all of them cross-reacted with entactin. The anti-entactin serum, which specifically recognized entactin protein, but not laminin or fibronectin, stained mainly the walls of blood vessels in rat brain slices. We observed a stronger entactin expression in immature than in adult brain, and a dramatic increase of vascular staining in kainate-injured hippocampus, suggesting a contribution of entactin to both development and reactive angiogenesis.

Animals↗

PubMed1996

The HIV-1 envelope protein gp120 induces neuronal apoptosis in hippocampal slices.

The HIV-1 envelope protein gp120 produces neuronal cell damage in primary cultures of a variety of cell types including hippocampal and retinal ganglion cell neurons. The properties of primary cell cultures are, however, often markedly different from those of cells living in their normal environment. We now report that gp120 induces widespread chromatin condensation and lesions in pyramidal granular neurones and in interneurones of rat hippocampal organotypic slice cultures. This damage is clearly of an apoptotic (programmed cell death) type. The use of an in vitro organized structure will enable the molecular and cellular mechanism of action of gp120 to be examined in conditions which are particularly suitable and relevant to the in vivo situation.

Animals↗

PubMed1996

Reactive glial cells express a vitronectin-like protein in the hippocampus of epileptic rats.

Injection of kainic acid into the amygdala induces in addition to a local cell loss a seizure related distal damage of the hippocampal complex, in particular in the CA3 field and hilus. This neuronal lesion is associated with hypertrophy and proliferation of astroglial cells which start around 3 days after kainate and peaks within 20 days of kainate. We now report that reactive astrocytes are labelled with antibodies against vitronectin in the CA3 field and hilus. In the present study we also exclude that the presence of vitronectin into the brain is due to an extravasation from serum throughout a blood brain barrier leakage. The present results constitute the first demonstration for a glial expression of vitronectin in vivo. Vitronectin is an extracellular matrix glycoprotein involved in axonal growth. The glial expression of vitronectin may therefore contribute to the synaptic remodeling of mossy fibers induced in the hippocampus by such treatment.

Amygdala↗

PubMed1996

Transient increase of tenascin-C in immature hippocampus: astroglial and neuronal expression.

In the present report we describe the anatomical localization of cells expressing tenascin-C, an extracellular matrix glycoprotein, in the hippocampal complex of developing rats. We report a development-dependent down regulation of both tenascin-C protein and mRNA. The highest levels of expression of tenascin-C was observed in rat pups from embryonic day 18 to postnatal day 7. Double labelling experiments performed with a tenascin-C antibody or tenascin-C probes combined with specific markers of astrocytes (GFAP) or neurons (MAP2 and Tau) allowed us to demonstrate that tenascin-C is expressed by both immature astrocytes and neurons in immature hippocampus. The temporal and topographic distribution of cells expressing tenascin-C (in the hilus and the stratum oriens of CA3) correlate with the localization and period of migration and maturation of post-mitotic cells. In view of these data we discuss the hypothesis that tenascin-C, as a mediator of neuron-glia interactions, may contribute to the development of hippocampal cells.

Animals↗

PubMed1996

Seizures induce tenascin-C mRNA expression in neurons.

Tenascin-C, an extracellular matrix glycoprotein that exhibits both growth-promoting and growth-inhibiting properties, is produced in the CNS mainly by astrocytes. In the present study we show that kainate-induced seizures result in an increased expression of tenascin-C in rat brain. Tenascin-C mRNA was increased mainly in the granule cell layer of the hippocampal complex, but tenascin-C mRNA expression was also observed in the pyriform cortex and amygdalo-cortical nucleus. Double labelling experiments using tenascin-C probes and MAP2 (a neuronal microtubule associated protein) antibodies revealed many neurons in these layers that express tenascin-C mRNA. These results support our previous findings of an increased tenascin-C immunoreactivity associated with the axons of granule cells. Tenascin-C expression is rapidly induced by seizures (6 h), preceding any lesion and glial reaction. In this pathological condition tenascin-C appears to be produced by both glia and neurons. The functional repercussions on the scarring and remodelling processes are also discussed.

Animals↗

PubMed1996

Apoptotic features of selective neuronal death in ischemia, epilepsy and gp 120 toxicity.

The occurrence of physiological cell death has been known for decades, but interest in the subject was renewed in 1972 when Kerr, Wyllie and Currie described in detail the ultrastructural changes characteristic of dying cells and coined the term apoptosis to describe the process. Cells display a wide variety of morphological changes when dying during development or following a toxic insult. A binary classification scheme suggests that physiologically appropriate death is due to apoptosis and that pathological mechanisms involve necrosis. However, recent studies indicate a potential involvement of apoptotic cell death in ischemia, status epilepticus and HIV-1 infection.

Animals↗

PubMed1996

Apoptosis and necrosis after reversible focal ischemia: an in situ DNA fragmentation analysis.

Apoptosis is one of the two forms of cell death and occurs under a variety of physiological and pathological conditions. Cells undergoing apoptotic cell death reveal a characteristic sequence of cytological alternations including membrane blebbing and nuclear and cytoplasmic condensation. Early activation of an endonuclease has been previously demonstrated after a transient focal ischemia in the rat brain Charriaut-Marlangue C, Margaill I, Plotkine M, Ben-Ari Y (1995) Early endonuclease activation following reversible focal ischemia. J Cereb Blood Flow Metab 15:385-388). We now show that a significant number of striatal and cortical neurons, exhibited chromatin condensation, nucleus segmentation, and apoptotic bodies increasing with recirculation time, as demonstrated by in situ labeling of DNA breaks in cryostat sections. Apoptotic nuclei were also detected in the horizontal limb diagonal band, accumbens nucleus and islands of Calleja. Several necrotic neurons, in which random DNA fragmentation occurs, were also shown at 6 h recirculation, in the ischemic core. Further investigation with hematoxylin/eosin staining revealed that apoptotic nuclei were present in cells with a large and swelled cytoplasm and in cells with an apparently well-preserved cytoplasm. These two types of cell death were reminiscent of those described in developmental cell death. Our data suggested that apoptosis may contribute to the expansion of the ischemic lesion.

Animals↗

PubMed1996

Acidic calponin cloned from neural cells is differentially expressed during rat brain development.

Calponin is an actin-, tropomyosin- and Ca2+ calmodulin-binding protein that inhibits in vitro the actomyosin MgATPase. Basic and acidic variants of calponin have been described to date. Although the cerebral expression of calponin remained controversial for some time, transcripts encoding acidic calponin in the adult rat brain and in cultured cerebellar cells have been reported. In the present work, we report the expression of acidic calponin mRNAs and the isolation of cDNAs encoding the full-length acidic calponin in cultured neuronal and glial cells and in adult rat brain. Sequence analysis reveals that acidic calponin in the brain is identical to that previously described in rat aortic vascular smooth muscle. In situ hybridization shows that calponin is highly expressed during ontogenesis in granule cells of the dentate gyrus of the hippocampus, in all layers of the olfactory bulb and in cerebellar granule neurons of the external and internal layers. In the adult rat brain, calponin expression decreased in these fields, but increased in choroid plexus cells. Bergmann glial cells were also labelled by a calponin probe. The reverse transcription-coupled polymerase chain reaction confirms that calponin mRNA levels are highest in the early stages of hippocampal development and that expression levels are low in adult hippocampi. The developmental expression pattern of brain acidic calponin suggests that calponin could be involved in contractile activity associated with neural cell proliferation or neuronal migration.

Acids↗

PubMed1996

MAP2d promotes bundling and stabilization of both microtubules and microfilaments.

Two low molecular weight MAP2 variants have been described, MAP2c and MAP2d. These variants are produced from a single gene by alternative splicing, and in their C-terminal regions contain, respectively, 3 and 4 tandem repeats, some of which are known to be involved in binding to microtubules. Substantial differences in the developmental expression pattern of MAP2c and MAP2d suggest they have different functions in neural cells. In order to investigate the respective roles of these MAP2 variants, we have analyzed the effects of MAP2c and MAP2d expression on microtubule and microfilament organization in transiently transfected cells. Our results show that both variants stabilize microtubules, but only MAP2d stabilizes microfilaments.

Actin Cytoskeleton↗

PubMed1996

Tenascin-C mRNA and tenascin-C protein immunoreactivity increase in astrocytes after activation by bFGF.

Tenascin-C is an extracellular matrix glycoprotein with trophic and repulsive properties, involved in migratory processes in CNS. Previous reports demonstrated that this molecule is produced and secreted by astrocytes. Preliminary data on fibroblasts and astrocytes have suggested that bFGF may modulate tenascin-C expression. bFGF is a mitogenic growth factor, involved in cell differentiation and neovascularization. In the present study, we examined whether bFGF modulates the expression of tenascin-C in hippocampal astrocytes from newborn rats. Our results suggest that bFGF increases the production of tenascin-C by cultured hippocampal astrocytes. We found that both tenascin-C mRNA and protein immunoreactivity were increased after bFGF treatment. Our results also demonstrated that tenascin-C polypeptides were secreted into the extracellular medium. In agreement with previous studies, we suggest that secreted tenascin-C is mainly the high molecular weight form. In addition, our results suggest that a cleavage of the high molecular weight form. In addition, our results suggest that a cleavage of the high molecular weight form may occur in the extracellular medium causing production of proteolytic fragments, that may modify the biological properties of tenascin-C. The present results may be relevant to the understanding of lesion scarring and regeneration process.

Animals↗