Heterogeneity of cycling glial progenitors in the adult mammalian cortex and white matter.

The heterogeneity and differentiation potential of mitotically active cells in the adult brain were studied by labeling adult rats with BrdU, and isolating an enriched population of cycling cells from neocortex and from subcortical white matter. The majority of this population isolated from either brain region labeled with O4, an early oligodendrocyte marker. In tissue culture, these O4(+) progenitors acquired galactocerebroside, a glycolipid of mature oligodendrocytes, but not GFAP, an intermediate filament of astrocytes. A minority population expressed the intermediate filament protein, vimentin, but not O4. This population expressed GFAP after several days in culture. A third population of cycling cells, expressing the gangliosides labeled with the A2B5 antibody, represented a minority population in subcortical white matter, but one of the major cycling populations in cortex, with substantial overlap with O4. Small populations of cycling NG2(+) cells also were observed. Thus, the cycling cells in the adult brain are heterogeneous, and the majority appear to belong to glial lineages.     

Heterogeneity of cycling glial progenitors in the adult mammalian cortex and white matter

The heterogeneity and differentiation potential of mitotically active cells in the adult brain were studied by labeling adult rats with BrdU, and isolating an enriched population of cycling cells from neocortex and from subcortical white matter. The majority of this population isolated from either brain region labeled with O4, an early oligodendrocyte marker. In tissue culture, these O4⁺ progenitors acquired galactocerebroside, a glycolipid of mature oligodendrocytes, but not GFAP, an intermediate filament of astrocytes. A minority population expressed the intermediate filament protein, vimentin, but not O4. This population expressed GFAP after several days in culture. A third population of cycling cells, expressing the gangliosides labeled with the A2B5 antibody, represented a minority population in subcortical white matter, but one of the major cycling populations in cortex, with substantial overlap with O4. Small populations of cycling NG2⁺ cells also were observed. Thus, the cycling cells in the adult brain are heterogeneous, and the majority appear to belong to glial lineages. © 2001 John Wiley & Sons, Inc. J Neurobiol 48: 75–86, 2001     

High-affinity glycine and glutamate transport in pig forebrain white and gray matter: a quantitative study

High-affinity uptake of glycine and glutamate modulates glutamatergic neurotransmission in gray matter. N-Methyl-D-aspartate (NMDA) receptors were recently described on white matter oligodendrocytes, therefore uptake of glutamate and glycine in white matter may also modulate NMDA receptor function. We found that glycine uptake in white structures of pig forebrain (corpus callosum, fimbria, subcortical pyramidal tracts, and occipital subcortical white matter) was similar to that in gray structures (frontal and temporal cortices and hippocampus), and that it was sensitive to sarcosine, a GLYT1 inhibitor (IC(50) 15 microM). Glutamate uptake in white matter was approximately 10% of that in gray; it was sensitive to dihydrokainate, an EAAT2 inhibitor. The levels of glycine and its precursor serine were similar in white and gray matter: approximately 2 and 1 nmol/mg tissue, respectively. The white matter level of glutamate was approximately 7.6 nmol/mg tissue, or approximately 74% of gray matter levels. The activity of serine hydroxymethyl transferase, which converts serine into glycine, was similar in white and gray matter (11-18 pmol/(mg tissue)min), whereas the white matter activity of phosphate-activated glutaminase, which converts glutamine into glutamate, was approximately 100 pmol/(mg tissue)min, or approximately 34% of gray matter activity. The white matter activity of glutamine synthetase, the glial enzyme that converts glutamate into glutamine, was 20-40 nmol/(mg tissue)min in neocortex and 5-6 nmol/(mg tissue)min in white matter. The data show that forebrain white matter is equipped to regulate extracellular levels of glycine and glutamate, functions that may modulate white matter NMDA receptor function.     

Glial cell lineage in vivo in the mouse cerebellum

Glial cells of the cerebellum originate from cells of the ventricular germinative layer, but their lineage has not been fully elucidated. For studying the glial cell lineage in vivo by retrovirus-mediated gene transfer, we introduced a marker retrovirus into the ventricular germinative layer of embryonic day 13 mice. In the resulting adult cerebella, virus-labeled glial cells were grouped in discrete clusters, and statistical analysis showed that these clusters represented clones in high probability. Of 71 of the virus-labeled glial clusters, 33 clusters were composed of astrocytes/Bergmann glia, 10 were composed of only white matter astrocytes, and 24 were composed of only oligodendrocytes. No glial clusters contained virus-labeled neurons. These results suggest that astrocytes/Bergmann glia, white matter astrocytes and oligodendrocytes immediately arise from separate glial precursors: these three glial lineages may diverge in the course of cerebellar development.     

Glial progenitors of the neonatal subventricular zone differentiate asynchronously, leading to spatial dispersion of glial clones and to the persistence of immature glia in the adult mammalian CNS

The subventricular zone (SVZ) of the developing mammalian forebrain gives rise to astrocytes and oligodendrocytes in the neocortex and white matter, and neurons in the olfactory bulb in perinatal life. We have examined the developmental fates and spatial distributions of the descendants of single SVZ cells by infecting them in vivo at postnatal day 0-1 (P0-1) with a retroviral "library". In most cases, individual SVZ cells gave rise to either oligodendrocytes or astrocytes, but some generated both types of glia. Members of glial clones can disperse widely through the gray and white matter. Progenitors continued to divide after stopping migration, generating clusters of related cells. However, the progeny of a single SVZ cell does not differentiate synchronously: individual clones contained both mature and less mature glia after short or long intervals. For example, progenitors that settled in the white matter generated three types of clonal oligodendrocyte clusters: those composed of only myelinating oligodendrocytes, of both myelinating oligodendrocytes and non-myelinating oligodendrocytes, or of only non-myelinating cells of the oligodendrocyte lineage. Thus, some progenitors do not fully differentiate, but remain immature and may continue to cycle well into adult life.     

Monoclonal antibody analysis of MHC expression in human brain biopsies: tissue ranging from "histologically normal" to that showing different levels of glial tumor involvement

The expression of class I and class II MHC products in human brain was studied. Radioimmunoassay confirmed weak expression of HLA-A,B,C and beta 2-microglobulin (beta 2-m) in brain extract. Quantitative inhibition assay showed brain had 1/70 as much activity as spleen, per microgram of extract protein. Immunoblot assay confirmed that HLA chains and beta 2-m were present in the brain extract. Class II was not detected. Microscopic analysis was performed on eight brain biopsies. The histologic appearance ranged from "apparently normal," to the presence of reactive astrocytes, to the presence of glial tumor. In every case, HLA-A,B,C and beta 2-m activity was concentrated at blood vessel walls. Small and medium-sized vessels were uniformly stained. Cell body staining was not seen in neurons, glia, oligodendrocytes, microglia, reactive astrocytes, or the majority of glial tumor cells. Class II activity was seen in occasional cell bodies in both grey matter and white matter in the microscopic assays. These cells had the morphologic appearance of microglia or reactive astrocytes. Occasional blood vessels also showed class II activity. Unlike the class I activity, the class II blood vessel stain was often discontinuous. More class II+ cell bodies were seen in tumor-associated tissue.     

猫腰髓白质年龄相关的形态学变化

以青年成年猫(1-3龄,2-2．5 kg)和老年猫(12龄,3-3．5kg)L6段脊髓白质为研究对象,用 神经丝蛋白(NF)免疫染色显示神经纤维,用改良的Holzer结晶紫染色显示所有胶质细胞并用成年动物Golgi 法显示其形态,用胶质纤维酸性蛋白(GFAP)免疫染色显示星形胶质细胞。光镜下对青年猫与老年猫腰髓白质 中神经纤维和胶质细胞进行形态学观察和定量研究。与青年猫相比,老年猫腰髓白质中的神经纤维密度显著下 降(P相似文献   

A comparative investigation of neuroglia in representative vertebrates: a silver carbonate study

The phylogenetic development of neuroglia (astrocytes, oligodendrocytes) was investigated in homologous cortical and subcortical forebrain regions of selected vertebrates. Microglia were not considered in the current study. Four to seven brains from each species were used. Scharenberg's modification for astroglia of del Rio Hortega's silver carbonate technique was used. The analysis of neuroglia cells was based on (1) the characteristic cellular morphology found in each species, (2) a comparison of the selected regions in each animal, (3) the interrelationships of astrocytes and their relations to neurons, blood vessels, and oligodendrocytes. The predominant type of neuroglia found in the fish, frog, and lizard was the ependymal cell; however, non-ependymal glial cells were also present. The bird represented a transitional phylogenetic stage from a predominance of ependymal glial to a predominance of non-ependymal glia. A progressive increase in the morphological relationships of glial cell bodies and processes to neurons was found with ascension of the phylogenetic scale from fish through primate. Interrelations were observed between adjacent astrocytic processes and cell bodies, and between astrocytes and oligodendrocytes. The processes of adjacent glial cells also appeared to show an increase in thickness at the point of approximation. A variety of astrocytes were observed ranging from small, round-oval shaped cells to large polygonal or stellate forms. Variations in the number of astrocytic processes, their thickness, and degree of secondary branching were described, and their possible functional significance was discussed.     

Recognition of Bergmann glial and ependymal cells in the mouse nervous system by monoclonal antibody

A monoclonal antibody designated anti-Cl was obtained from a hybridoma clone isolated from a fusion of NS1 myeloma with spleen cells from BALB/c mice injected with homogenate of white matter from bovine corpus callosum. In the adult mouse neuroectoderm, C1 antigen is detectable by indirect immunohistology in the processes of Bergmann glial cells (also called Golgi epithelial cells) in the cerebellum and of Muller cells in the retina, whereas other astrocytes that express glial fibrillary acidic protein in these brain areas are negative for C1. In addition, C1 antigen is expressed in most, if not all, ependymal cells and in large blood vessels, but not capillaries. In the developing, early postnatal cerebellum, C1 antigen is not confined to Bergmann glial and ependymal cells but is additionally present in astrocytes of presumptive white matter and Purkinje cell layer. In the embryonic neuroectoderm, C1 antigen is already expressed at day 10, the earliest stage tested so far. The antigen is distinguished in radially oriented structures in telencephalon, pons, pituitary anlage, and retina. Ventricular cells are not labeled by C1 antibody at this stage. C1 antigen is not detectable in astrocytes of adult or nearly adult cerebella from the neurological mutant mice staggerer, reeler, and weaver, but is present in ependymal cells and large blood vessels. C1 antigen is expressed not only in the intact animal but also in cultured cerebellar astrocytes and fibroblastlike cells. It is localized intracellularly.     

White matter injury following systemic endotoxemia or asphyxia in the fetal sheep

White matter injury is the most frequently observed brain lesion in preterm infants. The etiology remains unclear, however, both cerebral hypoperfusion and intrauterine infections have been suggested as risk factors. We compared the neuropathological outcome, including the effect on oligodendrocytes, astrocytes, and microglia, following either systemic asphyxia or endotoxemia in fetal sheep at midgestation. Fetal sheep were subjected to either 25 minutes of umbilical cord occlusion or systemic endotoxemia by administration of Escherichia coli lipopolysaccharide (LPS O111:B4, 100 ng/kg, IV). Periventricular white matter lesions were observed in 2 of 6 asphyxiated fetuses, whereas the remaining animals showed diffuse injury throughout the subcortical white matter and neuronal necrosis in subcortical regions, including the striatum and hippocampus. LPS-treatment resulted in focal inflammatory infiltrates and cystic lesions in periventricular white matter in 2 of 5 animals, but with no neuron specific injury. Both experimental paradigms resulted in microglia activation in the white matter, damaged astrocytes, and loss of oligodendrocytes. These results show that the white matter at midgestation is sensitive to injury following both systemic asphyxia and endotoxemia. Asphyxia induced lesions in both white and subcortical grey matter in association with microglia activation, and endotoxemia resulted in selective white matter damage and inflammation.     

DNA and RNA and the cytoarchitecture of human frontal cortex

DNA and RNA were studied in the layers of human prefrontal cortex by quantitative microchemical analyses on microtome-prepared serial frozen sections. Eleven cortical specimens from six autopsy brains were assayed. The mean total number of cells per mm³ of fresh cortex was estimated from DNA values. The number in layer I (61,000) was falsely high because of the inclusion of cells from the pia mater. From a plateau of 82,000-86,000 in layers II, IIIa and IIIb, the number of cells rose to 91,000 and 113,000 in layers IIIc and IV, respectively. The mean number was slightly lower in layer V, then gradually rose through layer VI to 127,000 cells in white matter. Per unit dry weight, DNA and cells varied much less than per unit volume; values averaged 14 per cent higher in layers II and IV than in neighbouring layers and in white matter were 20 per cent lower than in layer I. Intracortical patterns of RNA and RNA/cell reflected chiefly the distribution of neuronal cell bodies. Per unit fresh volume, RNA roughly paralleled DNA in layers I-V; through layer VI and into white matter RNA declined as DNA rose, reflecting the decline in neurons and increasing predominance of glial cells of lower RNA content. Per unit dry weight, RNA rose 50 per cent from layer I to layer II; a plateau of high values extended through layers II-V, then RNA declined rapidly through layer VI to a level in white matter that was 28 per cent of the value in layer II. Mean RNA/cell in cortex was 9-8 pg, with a maximum in layer IIIb (11.4 pg); in subcortical white matter it was 5.3 pg.     

Endothelin-converting enzyme in human tissues

Our aim was to determine whether the expression of endothelin-converting enzyme in human tissues would correlate with the distribution of its substrate, big endothelin-1, and its product, the mature peptide. Site-directed antisera raised against the conserved C-terminus of the mammalian enzyme were used to measure the immunoreactive enzyme in microsomal fractions prepared from tissue homogenates and to localize staining to the endothelial cells lining large conduit and smaller resistance vessels within cardiac, adrenal, respiratory and brain tissue. The activity of endothelin-converting enzyme was measured and characterized in isolated endothelial cells. This pattern of staining in the vascular endothelium paralleled that of mature endothelin and big endothelin-1, and these peptides were detectable by radioimmunoassay in all tissues examined. Immunoreactive endothelin-converting enzyme localized to other cell types, including bronchial epithelial cells, and to fibres within the glial limitans, neuronal processes and cell bodies of the cerebral cortex. Although perivascular astrocytes in the subcortical white matter displayed intense endothelin-converting enzyme-like immunoreactivity, endothelin staining was not detected. The results suggest that endothelin-converting enzyme has a ubiquitous distribution within the human vascular endothelium and is positioned to catalyse the conversion of big endothelin-1 to the biologically active endothelin-1, which on release may contribute to the maintenance of basal tone in humans. Endothelin-converting enzyme localized to epithelial cells in peripheral tissues or astrocytes within the brain may be upregulated in pathophysiological conditions in which endothelin levels are increased and could represent a further target for therapeutic intervention by enzyme inhibitors. © 1998 Chapman & Hall     

Glial Cell Composition and Ultrastructure of the Caudal Spinal Cord of Young and Adult Tuataras,Sphenodon punctatus

Research on the caudal spinal cord (SC) of three young and three adult tuataras (Sphenodon punctatus) has revealed that most of the glial cells were well differentiated as astrocytes and oligodendrocytes. The former type was not completely differentiated 3 months post-hatching, but was the main glial cell type at the age of 1 yr and in adulthood. Smaller numbers of oligodendrocytes were found in the white matter of adult animals than fibrous astrocytes. It was concluded that during growth, there is a progressive decrease in the dark, basophilic and electron-dense glioblasts, most of which develop as astrocytes.     

Cultured astrocytes derived from corpus callosum or cortical grey matter show distinct glutamate handling properties

While the astrocytic control of extracellular glutamate concentration at synaptic contacts is well characterized, little is known regarding the clearance of glutamate along axon tracts, even though local excitotoxic damage has been reported. Therefore, we have compared glutamate handling in astrocyte cultures derived from white matter (corpus callosum) and grey matter tissues (cortical structures). These populations of astrocytes showed clearly distinct phenotypes, adopting stellate or protoplasmic morphologies respectively. In addition, white matter astrocytes showed high densities of the intermediate filament proteins glial fibrillary acidic protein, vimentin and nestin. The glutamate–aspartate transporter and glutamate transporter‐1, as well as glutamine synthetase, were found to be expressed at higher levels in white matter compared with grey matter astrocytes. Consistent with this aspartate uptake capacity was three to fourfold higher in white matter cells, and the use of specific inhibitors revealed a substantial activity of glutamate transporter‐1, contrasting with grey matter cells where this transporter appeared poorly functional. In addition, expression of type 5 metabotropic glutamate receptors was considerably higher in white matter astrocytes where the agonist (S)‐3,5‐dihydroxyphenylglycine triggered a large release of intracellular calcium. Differences in these astrocyte cultures were also observed when exposed to experimental conditions that trigger glial activation. This study highlights typical features of cultured astrocytes derived from white matter tissues, which appear constitutively adapted to handle excitotoxic insults. Moreover, the expression and activity of the astroglial components involved in the control of glutamatergic transmission are reinforced when these cells are maintained under conditions mimicking a gliotic environment.     

Immunohistochemistry of Atrial Natriuretic Peptide in Brain Infarction

Atrial natriuretic peptide (ANP) was originally isolated from cardiac atria, and has potent natriuretic, diuretic, and vasorelaxant properties. It has been localized in neurons and astrocytes in the cerebral cortex and the white matter. We hypothesize that glial ANP may contribute to the regulation of cerebral blood flow in brain infarction. In order to elucidate this possible role, the immunohistochemistry of ANP was studied in cases of brain infarction and in other cases of brain trauma for comparison. A statistically significant increase in the number of ANP-immunoreactive glial cells (mainly astrocytes) was observed in the white matter surrounding the brain infarction compared with the intact area. No statistically significant increase in ANP-immunoreactive glial cell number was observed in the cerebral white matter from brain haemorrhage, contusion and control cases. Our results indicate that glial ANP may increase in number in brain infarction, and that it may be involved in the regulation of the cerebral blood flow in the infarcted area.     

Interaction of astrochondrin with extracellular matrix components and its involvement in astrocyte process formation and cerebellar granule cell migration

We have recently characterized a chondroitin sulfate proteoglycan from the murine central nervous system which is expressed by astrocytes in vitro and carries the L2/HNK-1 and L5 carbohydrate structures. In the present study, we provide evidence that its three core proteins of different size are similar in their proteolytic peptide maps and thus designate this group of structurally related molecules astrochondrin. During development, astrochondrin and the L5 carbohydrate were hardly detectable in the brain of 14-d-old mouse embryos by Western blot analysis. Expression of astrochondrin and the L5 epitope was highest at postnatal day 8, the peak of cerebellar granule cell migration and Bergmann glial process formation, and decreased to weakly detectable levels in the adult. Immunocytochemical localization of astrochondrin in the cerebellar cortex of 6-d-old mice showed association of immunoreactivity with the cell surface of astrocytes, including Bergmann glial processes and astrocytes in the internal granular layer or prospective white matter. Endfeet of astrocytes contacting the basal lamina of endothelial and meningeal cells and contact sites between Bergmann glial processes and granule cells also showed detectable levels of astrochondrin. Furthermore, granule cell axons in the molecular layer were astrochondrin immunoreactive. In the adult, astrochondrin immunoreactivity was weakly present in the internal granular layer and white matter. Both Fab fragments of polyclonal antibodies to astrochondrin and monovalent fragments of the L5 monoclonal antibody reduced the formation of processes of mature GFAP- positive astrocytes on laminin and collagen type IV, but not on fibronectin as substrata. Interestingly, the initial attachment of astrocytic cell bodies was not disturbed by these antibodies. Antibodies to astrochondrin also reduced the migration of granule cells in the early postnatal mouse cerebellar cortex. In a solid phase radioligand binding assay, astrochondrin was shown to bind to the extracellular matrix components laminin and collagen type IV, being enhanced in the presence of Ca2+, but not to fibronectin, J1/tenascin or other neural recognition molecules. Furthermore, astrochondrin interacted with collagen types III and V, less strongly with collagen types I, II, and IX, but not with collagen type VI. The interaction of astrochondrin with collagen types III and V was saturable and susceptible to increasing ionic strength, and could be competed by chondroitin sulfate, heparin, and dextran sulfate, but not by hyaluronic acid, glucose-6-phosphate, or neuraminic acid.(ABSTRACT TRUNCATED AT 400 WORDS)     

Acute lipopolysaccharide-mediated injury in neonatal white matter glia: role of TNF-alpha, IL-1beta, and calcium

Bacterial infection is implicated in the selective CNS white matter injury associated with cerebral palsy, a common birth disorder. Exposure to the bacterial endotoxin LPS produced death of white matter glial cells in isolated neonatal rat optic nerve (RON) (a model white matter tract), over a 180-min time course. A delayed intracellular Ca(2+) concentration ([Ca(2+)](i)) rise preceded cell death and both events were prevented by removing extracellular Ca(2+). The cytokines TNF-alpha or IL-1beta, but not IL-6, mimicked the cytotoxic effect of LPS, whereas blocking either TNF-alpha with a neutralizing Ab or IL-1 with recombinant antagonist prevented LPS cytotoxicity. Ultrastructural examination showed wide-scale oligodendroglial cell death in LPS-treated rat optic nerves, with preservation of astrocytes and axons. Fluorescently conjugated LPS revealed LPS binding on microglia and astrocytes in neonatal white and gray matter. Astrocyte binding predominated, and was particularly intense around blood vessels. LPS can therefore bind directly to developing white matter astrocytes and microglia to evoke rapid cell death in neighboring oligodendroglia via a calcium- and cytokine-mediated pathway. In addition to direct toxicity, LPS increased the degree of acute cell death evoked by ischemia in a calcium-dependent manner.     

Astrocytic calcium/zinc binding protein S100A6 over expression in Alzheimer's disease and in PS1/APP transgenic mice models

Astrocytes recruitment and activation are a hallmark of many neurodegenerative diseases including Alzheimer's disease (AD). We have previously observed an overexpression for S100A6 protein, a Ca(2+)/Zn(2+) binding protein presenting more affinity for zinc than for calcium, in amyotrophic lateral sclerosis (ALS). Here we demonstrated in AD patients but also in two different AD mouse models, that astrocytic S100A6 protein was homogeneously up-regulated within the white matter. However, within the grey matter, almost all S100A6 immunoreactivity was concentrated in astrocytes surrounding the Abeta amyloid deposits of senile plaques. These S100A6 neocortex labelled astrocytes were also positive for the glial fibrillary acidic protein (GFAP) and S100B protein. Contrasting with S100A6, the distribution for S100B and GFA astrocytic labelled cells was not restricted to the Abeta amyloid deposit in grey matter, but widely distributed throughout the neocortex. Coupling the knowledge that biometals such as zinc are highly concentrated in the amyloid deposits in AD and S100A6 having a high affinity for Zn(2+) may suggest that S100A6 plays a role in AD neuropathology.     

Astrocyte Atrophy and Immune Dysfunction in Self-Harming Macaques

Background

Self-injurious behavior (SIB) is a complex condition that exhibits a spectrum of abnormal neuropsychological and locomotor behaviors. Mechanisms for neuropathogenesis could include irregular immune activation, host soluble factors, and astrocyte dysfunction.

Methods

We examined the role of astrocytes as modulators of immune function in macaques with SIB. We measured changes in astrocyte morphology and function. Paraffin sections of frontal cortices from rhesus macaques identified with SIB were stained for glial fibrillary acidic protein (GFAP) and Toll-like receptor 2 (TLR2). Morphologic features of astrocytes were determined using computer-assisted camera lucida.

Results

There was atrophy of white matter astrocyte cell bodies, decreased arbor length in both white and gray matter astrocytes, and decreased bifurcations and tips on astrocytes in animals with SIB. This was combined with a five-fold increase in the proportion of astrocytes immunopositive for TLR2.

Conclusions

These results provide direct evidence that SIB induces immune activation of astrocytes concomitant with quantifiably different morphology.