Distinctive pattern of Bergmann glial pathology in human hepatic encephalopathy

Alzheimer type II astrocytosis is the pathological hallmark of hepatic encephalopathy. These astrocytes undergo a characteristic morphological change and, in addition, lose immunoreactivity for glial fibrillary acidic protein (GFAP). However, a previous study in the portacaval shunted rat, a model of hepatic encephalopathy, revealed increased rather than decreased GFAP immunoreactivity in Bergmann glia, a specialized group of cerebellar astrocytes. In the present study, sections of cerebellar vermis from 15 cirrhotic patients with hepatic encephalopathy and varying degrees of Alzheimer type II astrocytosis were stained using antisera to GFAP. The Bergmann glial cells did not show altered GFAP immunoreactivity compared to controls. In addition, the degree of GFAP immunoreactivity was not correlated with the degree of Alzheimer type II change nor related to the aetiology of the liver disease. These results suggest a differential response of Bergmann glia in human hepatic encephalopathy.     

Down-regulation of astroglial proteins in the rat cerebellum after portacaval anastomosis

The effect of short-term portacaval anastomosis (PCA) on the expression of specific astroglial markers [glial fibrillary acidic protein (GFAP) and glutamine synthetase (GS)] in the rat cerebellum was examined to determine the influences of PCA on astroglial cells. The results suggest that PCA directly interferes with astroglial cytoskeleton, as indicated by the irregular distribution and reduced expression of GFAP observed after 1 month. PCA also decreased GS immunoreactivity in the Bergmann glial processes of the molecular layer, as well as in astrocytes of the granule cell layer. It might also modulate glutamatergic nervous activity as GS expression was reduced in 1 month post-PCA brains. Moreover, the GFAP and GS levels in PCA-exposed rats were lower than in control rats. This might contribute to the appearance of encephalopathy by increasing extracellular glutamate and/or ammonia concentrations. These results show that short-term PCA interferes with astroglial protein expression, with both GFAP and GS levels falling in astroglial cells.     

Reactive astroglia-neuron relationships in the human cerebellar cortex: A quantitative, morphological and immunocytochemical study in Creutzfeldt-Jakob disease

In order to investigate the role of neuron-glia interactions in the response of astroglial to a non-invasive cerebellar cortex injury, we have used two cases of the ataxic form of Creutzfeldt-Jakob disease (CJD) with distinct neuronal loss and diffuse astrogliosis. The quantitative study showed no changes in cell density of either Purkinje or Bergmann glial cells in CJ-1, whereas in the more affected CJ-2 a loss of Purkinje cells and an increase of Bergmann glial cells was found. The granular layer in both CJD cases showed a similar loss of granule cells (about 60% ) in parallel with the significant increase in GFAP+ reactive astrocytes. GFAP immunostaining revealed greater reactivity of Bergmann glia in CJ-2 than in CJ-1, as indicated by the thicker glial processes and the higher optical density. Granular layer reactive astrocytes were regularly spaced. In both CJD cases there was strict preservation of the spatial arrangement of all astroglial subtypes—Fañanas cells, Bergmann glia and granular layer astrocytes. Reactive Fañanas and Bergmann glial cells and microglia/macrophages expressed vimentin, while only a few vimentin+ reactive astrocytes were detected in the granular layer. Karyometric analysis showed that the increase in nuclear volume in reactive astrloglia was directly related with the level of glial hypertrophy. The number of nucleoli per nuclear section was constant in astroglial cells of human controls and CJD, suggesting an absence of polyploidy in reactive astroglia. Ultrastructural analysis revealed junctional complexes formed by the association of macula adherens and gap junctions. In the molecular layer numerous vacant dendritic spines were ensheathed by lamellar processes of reactive Bergmann glia. Our results suggest that quantitative (neuron/astroglia ratio) and qualitative changes in the interaction of neurons with their region-specific astroglial partners play a central role in the astroglial response pattern to the pathogenic agent of CJD.     

Development of astroglial elements in the suprachiasmatic nucleus of the rat: with special reference to the involvement of the optic nerve

The development of astroglial cells and the effect of the retinohypothalamic tract on it were studied by vimentin and glial fibrillary acidic protein (GFAP) immunocytochemistry in the suprachiasmatic nucleus (SCN) of the rat. At the embryonic stage, vimentin-immunoreactive (VIM-IR) radial glia, precursors of astrocytes, were dominant. However, their filaments vanished in the first few postnatal days. Instead of VIM-IR glial filaments, GFAP-immunoreactive (GFAP-IR) astrocytes appeared at E20 and grew rapidly from the P3 stage. GFAP immunoreactivity in the ventrolateral portion of the SCN (VLSCN) was measured using a computer-assisted image analyzing system. In normal rats, GFAP immunoreactivity showed a stepwise pattern with two slopes at P3-P4 and P20-P25. Bilaterally eye-enucleated rats operated on the day of birth showed lower GFAP immunoreactivity than normal rats and the GFAP immunoreactivity did not increase between P20 and P25 when GFAP-IR glial processes rapidly expand. Electron microscopic investigation at P50 (adult stage) revealed that neurons in the VLSCN had often direct apposition without astroglial processes and the frequency of this finding was significantly higher in eye-enucleated rats than in the control rats. These findings strongly suggest that the postnatal development of astroglial elements, particularly the expansion of GFAP-IR processes in the SCN, is regulated by retinohypothalamic projection.     

Heterogeneous astroglial response in the rat spinal cord to long-term portacaval shunt: an immunohistochemical study.

Glial fibrillary acidic protein (GFAP) immunoreactivity has been used to study the astroglial response in the rat spinal cord to long-term portacaval shunt (PCS). The astroglial response in PCS rats is heterogeneous. In general, astrocytes show a loss of GFAP immunoreactivity, as well as shrinking and pyknosis in their nuclei; however, while GFAP reactivity was unchanged in the periependymal region, it was strongly increased in the dorsolateral region of the spinal cord (lateral spinal nucleus, dorsal root entry zone, and the most dorsal region of the dorsal horn). Three possibilities are postulated to explain how astrocytes, in the periependymal and dorsolateral regions, can support the effects of PCS: a) astrocytes related to glutamatergic pathways ought to possess a more efficient ammonia uptake and detoxification system, b) long-term PCS can activate nociceptive pathways (substancePergic fibers), and c) astrocytes located in periependymal and dorsolateral regions can be exposed to lower concentrations of ammonia because of its diffusion into the cerebro-spinal fluid close to these regions.     

Reduced glial fibrillary acidic protein and glutamine synthetase expression in astrocytes and Bergmann glial cells in the rat cerebellum caused by delta(9)-tetrahydrocannabinol administration during development

In this study we analyzed the responses of cerebellar astroglial cells to pre- and perinatal delta(9)-tetrahydrocannabinol (THC) exposure in three postnatal ages and both sexes. To determine whether THC during development directly modifies astroglial growth, this study investigated the effects of THC on astroglial morphological changes and on the expression of specific astroglial markers (glial fibrillary acidic protein: GFAP and glutamine synthetase: GS). Our results demonstrated that the administration of THC during development has deleterious effects on astroglial maturation in the cerebellum. These results also indicate that THC might interfere with astroglial differentiation in a way dependent on sex. The effect of cannabinoids on the development of cerebellar astroglial cells (astrocytes and Bergmann glial cells) is to reduce protein synthesis, since both GFAP and GS decreased in astroglial cells, not only during THC exposure but also in adult ages. Our data suggest that pre- and perinatal THC exposure directly interferes with astroglial maturation by disrupting normal cytoskeletal formation, as indicated by the irregular disposition of GFAP and the lower GFAP expression observed at all the ages studied. THC exposure during development may also modulate glutamatergic nervous activity since GS expression is reduced in THC-exposed brains. GS expression increased progressively after THC withdrawal, but GS expression had still not reached control values two months after THC withdrawal. This indicates that glutamate uptake is lower in glial cells exposed to THC, since GS expression is lower than in older controls. Consequently, glutamatergic neurotransmission may be affected by cannabinoid exposure during gestation. Therefore, cannabinoids exert developmental toxicity, at least on astroglial cells, which could contribute to fetal brain growth retardation.     

Sexual dimorphism in the hamster cerebellum demonstrated by glial fibrillary acidic protein (GFAP) and vimentin immunoreactivity.

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.     

Effects of prenatal ethanol exposure on the development of Bergmann glia and astrocytes in the rat cerebellum: an immunohistochemical study.

The consequences of prenatal ethanol exposure on the postnatal development of Bergmann glia and astrocytes in the rat cerebellum were investigated by using glial fibrillary acidic protein (GFAP) immunolabeling. Pregnant rats were either fed with an ethanol containing liquid diet (6.7% v/v) or pair-fed with an isocaloric diet throughout gestation. On postnatal day (PD) 15 and 22, parasagittal sections of the cerebellar vermis from female offspring were processed for GFAP immunohistochemistry to assess the development of Bergmann glia and astrocytes in lobules I, VII, and X and astrocytes in the central core of white matter. On PD 15, compared to control animals, ethanol exposed animals had fewer GFAP positive Bergmann glial fibers per unit length of molecular layer; a significantly greater percentage of morphologically immature Bergmann fibers; a significantly greater GFAP positive astrocytic area per unit area of internal granular layer and central white matter; and the astrocytic processes were wider and more closely packed. These glial changes were associated with significantly thicker external granular layer in all 3 lobules. However, no significant differences were seen between the ethanol exposed and control animals on PD 22, indicating "catch-up growth" in the ethanol exposed animals during the third postnatal week. These results suggest that prenatal ethanol exposure causes (1) delayed maturation of Bergmann glia, which in turn contributes to the delayed migration of granule cells; and (2) alterations in the normal postnatal development of astrocytes.     

On the plasticity of the cerebellar renin-angiotensin system: localization of components and effects of mechanical perturbation

This study focuses on the renin-angiotensin system (RAS) in the cerebellar cortex and changes within this system after mechanically induced cerebellar injury. Using radioactive and non-radioactive in situ hybridization and immunocytochemistry angiotensinogen mRNA, angiotensinogen, angiotensin II and, for the first time, N-terminal angiotensin fragment (1–7) immunoreactivities, respectively, were demonstrated in the rat cerebellum. Angiotensinogen mRNA and angiotensinogen immunoreactivity (IR) were both present in glial cell populations of all layers, especially in the Purkinje and granular cell layers and within the cerebellar nuclei. Angiotensin II IR was demonstrated in glial cell population in all layers using a monoclonal angiotensin II antibody, while with a polyclonal angiotensin II antiserum (Denise) some Purkinje cell bodies were labelled. After lesioning the cerebellar cortex mechanically by an injection cannula a strong increase in angiotensinogen gene expression as well as in angiotensin II and angiotensin (1–7) immunoreactivities were observed in the glial cell populations. Furthermore, putative Bergmann glial processes, as indicated from the morphological appearance became strongly angiotensin II and angiotensinogen immunoreactive in the region close to the mechanically induced lesion. It could inter alia be demonstrated for the first time using confocal laser microscopy of ANG II IR and GFAP IR that ANG II in vivo in the intact cerebellar cortex is present in astroglial processes in the molecular layer and presumably secreted into the extracellular space in form of small spheric bodies and/or taken up by other cell types. In contrast, the N-terminal fragment angiotensin (1–7) IR was restricted to the glial cell populations and appeared only after the lesion event. Thus, it is suggested that the cerebellar RAS shows marked changes in response to mechanically induced lesions. The expression of angiotensinogen as well as the production of angiotensinogen IR and angiotensin II like IR is even after mechanical lesion restricted to astrocytes, i.e., cerebellar astrocytes and putataive Bergmann glial cells, and in case of immunoreactivities it spreads to the radially oriented Bergmann glial processes in the molecular layer.     

NG2 cells differentiate into astrocytes in cerebellar slices

NG2-glia are an abundant population of glial cells that have been considered by many to be oligodendrocyte progenitor cells (OPCs). However, growing evidence suggests that NG2-glia may also be capable of differentiating into astrocytes and neurons under certain conditions. Here, we have examined NG2-glia in cerebellar slices, using transgenic mice in which the astroglial marker glial specific protein (GFAP) drives expression of the reporter gene enhanced green fluorescent protein (EGFP). Immunolabelling for NG2 shows that NG2-glia and GFAP-EGFP astroglia are separate populations in most areas of the brain, although a substantial population of NG2-glia in the pons also express the GFAP-EGFP reporter. In the cerebellum, NG2-glia did not express EGFP, either at postnatal day (P)12 or P29–30. We developed an organotypic culture of P12 cerebellar slices that maintain cytoarchitectural integrity of Purkinje neurons and Bergmann glia. In these cultures, BrdU labelling indicates that the majority of NG2-glia enter the cell cycle within 2 days in vitro (DIV), suggesting that NG2-glia undergo a ‘reactive’ response in cerebellar cultures. After 2 DIV NG2-glia began to express the astroglial reporter EGFP and in some cases the respective GFAP protein. However, NG2-glia did not acquire phenotypic markers of neural stem cells or neurons. The results suggest that NG2-glia are not lineage restricted OPCs and are a potential source of astrocytes in the cerebellum.     

Neuronal and astroglial response to pre- and perinatal exposure to delta-9-tetra- hydrocannabinol in the rat substantia nigra

The responses of neurons and astroglial cells to pre- and perinatal exposure to Delta(9)-tetrahydrocannabinol (Delta(9)-THC) were evaluated in the substantia nigra (SN) of male and female rats, at three postnatal ages (PD21, PD30 and PD70), by immunohistochemical detection of tyrosine hydroxylase (TH) in dopaminergic neurons and of glial fibrillary acidic protein (GFAP) in astrocytes. Our results showed that the effects of pre- and perinatal exposure to Delta(9)-THC on neuronal and astroglial immunoreactivities in the SN (compacta and reticulata) varied with sex, with male rats being more susceptible than females. Prenatal exposure to Delta(9)-THC decreased TH immunoreactivity in the SN of males on PD21 when compared to both their controls and Delta(9)-THC-exposed females of the same age. Furthermore, the TH expression decreased with age in Delta(9)-THC-exposed males in the SNc pars compacta, whereas it increased in controls. On the contrary, TH expression was maintained stable in the SN pars compacta of Delta(9)-THC-exposed females from PD21. These differences in neuronal development caused by prenatal Delta(9)-THC exposure were associated with significant differences in GFAP expression by astroglial cells in both sexes. On PD21, GFAP immunoreactivity decreased in the SN in Delta(9)-THC-exposed male rats. Although GFAP expression increased in Delta(9)-THC-exposed males with age, it did not reach control levels by PD70. On the contrary, significantly increased GFAP expression in the Delta(9)-THC-exposed females on PD21 was observed, compared to their controls and also to Delta(9)-THC-exposed male rats; however, the GFAP expression shown by Delta(9)-THC-exposed females stabilized from PD21. These Delta(9)-THC-induced changes in the glial development could indicate that Delta(9)-THC accelerated the maturation of astrocytes in female rats, whereas Delta(9)-THC delayed astrocytic maturation in Delta(9)-THC-exposed males. These findings suggest that pre- and perinatal exposure to Delta(9)-THC can lead to long-term effects in both neurons and glial cells.     

Different in vitro response to rIL-1 beta of newborn and adult rat astroglia.

In recent literature, lymphokines have been reported to be able to promote both proliferation and maturation of some glial populations. In this paper, we compare the effect of rIL-1 on newborn and adult rat astroglial cells in vitro. In newborn, but not in adult astrocytes, 100 U/ml of rIL-1 beta increase [3H]thymidine incorporation with a maximal response by 3 days as compared to the control untreated culture. In contrast, rIL-1 beta induces an increase of GFAP immunoreactivity both in newborn and in adult astrocytes, as compared to the control untreated cells. These data indicate that, while both newborn and adult astroglial cells are capable of responding to rIL-1 beta, only newborn astrocytes can respond to this lymphokine with proliferation. Thus, it appears likely that different factors, other than rIL-1 beta, are needed by adult astrocytes to proliferate.     

Changes in Glial Fibrillary Acidic Protein Immunoreactivity in Response to Experimental Hepatic Encephalopathy in the Rat Hippocampus

The response of astroglial cells in the hippocampus to long-term portacaval anastomosis (PCA), an experimental model of hepatic encephalopathy, was studied in adult male rats and compared with controls. Six months after PCA, the rat hippocampi were processed for glial fibrillary acidic protein (GFAP). GFAP-immunopositive astroglial profiles were observed in all hippocampal layers in PCA rats, but GFAP distribution differed in PCA rats and controls. In PCA rats, cell bodies and cell processes immunostained with GFAP were observed mainly in the CA1-CA3 layers in relation to pyramidal neurons, whereas the number and length of the astroglial processes decreased in the rest of the hippocampal regions. In addition, decreased GFAP immunoreactivity in the perivascular processes was observed in PCA rats compared with controls. These results show that long-term PCA elicited different responses in GFAP expression in different hippocampal regions, which might suggest a regional variation in glial sensitivity.     

Acidic calponin immunoreactivity in postnatal rat brain and cultures: subcellular localization in growth cones, under the plasma membrane and along actin and glial filaments.

Acidic calponin, an F-actin-binding protein, is particularly enriched in brain, where calponin protein and mRNA are mainly expressed by neurons. The presence of calponin immunoreactivity in cultured astroglial cells has been reported, but the presence of acidic calponin in astrocytes in vivo appears equivocal. For the present study, we raised a specific polyclonal antibody against the 16-residue synthetic peptide covering the sequence E311-Q326 (EYPDEYPREYQYGDDQ) situated at the carboxy-terminal end of rat acidic calponin, and we investigated the cellular and subcellular localization of the protein in the developing central nervous system. Our results show that acidic calponin is particularly enriched in: (i) growth cones and submembranous fields of maturing cerebellar and cortical cells, where it codistributes with microfilaments and (ii) glial cells in vivo, including radial glia, glia limitans, Bergmann glia and mature astrocytes, and ex vivo, where acidic calponin strongly colocalizes with intermediate glial fibrillary acidic protein (GFAP) and vimentin filaments. Finally, up to four acidic calponin subtypes with different isoelectric point (pI) values were identified by two-dimensional gel electrophoresis of cerebellar and hippocampal extracts. The more acidic isoforms were developmentally regulated. As only one single mRNA for acidic calponin has been identified, these isoforms must reflect postsynthesis changes probably related to the particular functions of acidic calponin in maturing cells. Although brain acidic calponin's exact role remains uncertain, the present data suggest that it is involved in neuronal and glial plasticity.     

Dynamic transformation of Bergmann glial fibers proceeds in correlation with dendritic outgrowth and synapse formation of cerebellar Purkinje cells

Bergmann glia (BG) are unipolar cerebellar astrocytes, whose radial (or Bergmann) fibers associate with developing granule cells and mature Purkinje cells (PCs). In the present study, we investigated the morphodifferentiation of BG by immunohistochemistry for glutamate transporter GLAST and electron microscopy. GLAST was expressed widely in cerebellar radial glia/astrocytes during fetal and neonatal periods and became concentrated in BG postnatally. During the second postnatal week when PC dendrites grow actively, GLAST immunostaining revealed dynamic cytologic changes in Bergmann fibers in a deep-to-superficial gradient; Bergmann fibers traversing the external granular layer were stained as rod-like fibers, whereas in the molecular layer, the rod-like pattern was gradually replaced with a reticular meshwork. At postnatal day 10, the superficial rod-like domain was composed of glial fibrillary acidic protein (GFAP)-positive/GLAST-positive straight fibers, forming cytoplasmic swellings and short filopodia. Along this domain, the tip of growing PC dendrites ascended vertically and entered the base of the external granular layer. The deeper reticular domain of Bergmann fibers was characterized by active expansion of GFAP-negative/GLAST-positive lamellate processes, which surrounded PC synapses almost completely. Therefore, the transformation of Bergmann fibers proceeds in correlation with dendritic differentiation of PCs. The intimate PC-BG relationships during cerebellar development raise the possibility that a preexisting glial shaft could serve as a structural substrate that directs dendritic outgrowth toward the pial surface, whereas the successive formation of a reticular glial meshwork should lead to structural maturation of newly formed PC synapses.     

Role of astroglial cell clones in the survival and differentiation of cerebellar embryonic neurons

To investigate the role of astrocytes in the survival and differentiation of cerebellar neurons during development, we have used astroglial cell clones, derived from 8-day postnatal cerebellar explants and which might be the in vitro equivalents of the 3 main types of cerebellar astrocytes, the Golgi epithelial cells and their Bergmann processes, the velate protoplasmic and the fibrous astrocytes (F. Alliot and B. Pessac, Brain Res., 306 (1984) 283-291). Nearly all single cells, dissociated from 15-day embryonic mouse cerebella and seeded at low density, adhered to layers of each of the cerebellar astroglial cell clones as well as to other glial lines or artificial substrates. However, the cerebellar embryonic neurons survived well only on monolayers of either the 'Golgi-Bergmann'-like or the 'velate protoplasmic'-like clones. On these layers, 60-80% of the neurons were still present after 5 days of co-culture, while only less than 5% survived on the other types of substrates. The differentiation pattern of the neurons surviving on the 'Golgi-Bergmann' and the 'velate protoplasmic' astroglial clones was studied with markers of postmitotic granule cells, the major neuronal population in adult cerebellum. The velate protoplasmic-like clone was the only one able to support the coordinate acquisition by most surviving neurons of the phenotypic characteristics of granule cells, i.e. a distinct morphology, a specific epitope binding the monoclonal antibody 7-8 D2 and immunoreactivity to glutamate. These data show a broad heterogeneity in the capacity of astroglial cell clones to support embryonic cerebellar neurons. In addition, they indicate that neuronal survival per se is not sufficient for the acquisition of a differentiated neuronal phenotype.     

Neuron–Glia Interaction in the Suprachiasmatic Nucleus: A Double Labeling Light and Electron Microscopic Immunocytochemical Study in the Rat

The morphological interactions between astroglial and neuronal elements were elucidated in the rat suprachiasmatic nucleus (SCN) by light and electron microscopic immunocytochemistry using antibodies against glial fibrillary acidic protein (GFAP), vasoactive intestinal peptide (VIP) and arginine-vasopressin (AVP). Throughout the SCN, particularly in its ventral portion, GFAP-like-immunoreactive (GFAP-LI) astroglial elements were found. These astrocytes displaying GFAP-like immunoreactivity occasionally contained fairly well-developed organelles. Some of these astrocytes were found as satellite cells in close contact with non-immunoreactive neuronal perikarya and processes. Around the neurons, GFAP-LI astroglial processes were also observed to cover some portions of presynaptic and postsynaptic elements. In addition, these astroglial elements were seen between two neuronal somata and pericytes of blood capillaries as glial endfeet. By double labeling immunoelectron microscopy using antibodies against GFAP/VIP and GFAP/AVP, some portions of VIP-like-immunoreactive or AVP-like-immunoreactive neuronal somata and processes were found to be engulfed by GFAP-LI astroglial processes. The possible functional roles of the morphological interactions between astroglial and neuronal elements are discussed.     

NG2 proteoglycan-expressing cells of the adult rat brain: possible involvement in the formation of glial scar astrocytes following stab wound

Stab wound lesion to the adult central nervous system induces strong proliferative response that is followed by the formation of a dense astroglial scar. In order to determine the origin of those astrocytes composing the glial scar, the cell proliferation marker bromodeoxyuridine (BrdU) was administered to lesioned rats that were fixed 3 h or 6 days later. At 3 h after the BrdU administration, labeled nuclei were frequently associated with either NG2(+) cells or microglia/macrophages, but rarely with astrocytes expressing glial fibrillary acidic protein (GFAP). Six days later, by contrast, numerous BrdU-labeled nuclei were associated with astrocytes located along the lesion borders. After the injection of a viral vector of the green fluorescent protein (GFP) into the lesional cavity, GFP was preferentially detected within NG2- or GFAP-labeled cells when lesioned animals were fixed 1 or 6 days after the injections, respectively. The combined detection of glial markers within cells present in the lesioned area indicated that, although they rarely express GFAP, the marker of mature astrocytes, NG2(+) cells located along the lesion borders frequently express nestin and vimentin, i.e., two markers of immature astrocytes. Lastly, chronic treatment of lesioned rats with dexamethasone was found to inhibit the proliferation of NG2(+) cells present within the lesioned area and to subsequently alter the formation of a dense astroglial scar. Taken together, these data strongly suggest that following a surgical lesion, at least a portion of the astrocytes that constitute the glial scar are issued from resident NG2(+) cells.     

Astroglial alterations in rat hippocampus during chronic lead exposure.

The present study was performed in order to follow the response of astroglial cells in the rat hippocampus to chronic low-level lead exposure. The experiments combined immunohistochemistry using anti-glial fibrillary acidic protein (GFAP) antibody and conventional transmission electron microscopy (EM). Chronic administration with drinking water [1 g% w/v (subclinical dose) of lead acetate dissolved in distilled water] was started through the mother's milk when pups were 7 days old. Following weaning, experimental offspring were treated for 3 months with the same concentration of adulterated water. The group of intoxicated animals and their controls were sacrificed by perfusion-fixation at 30, 60, and 90 days of exposure. After 60 days of lead treatment, staining of GFAP-positive cells demonstrated an astroglial transformation from the quiescent to the reactive state, characterized by an increase in GFAP. In control rats no changes in GFAP immunostaining were observed. The intensity of the astroglial response was enhanced after 90 days of lead intoxication, showing an increment of GFAP immunoreactivity. Quantification of these changes was made by computerized image analysis, confirming that the sectional areas of the astroglia in lead-exposed animals were larger than those in controls. These results are consistent with the ultrastructural alterations. Simultaneously with the increment in gliofilaments, intranuclear inclusions were seen in some astrocytes. The mechanisms by which lead affects astrocytes are unknown. Probably the astroglial changes induced by lead intoxication produce microenvironmental modifications that may disturb the neuronal function.     

Upregulation of α‐synuclein expression in the rat cerebellum in experimental hepatic encephalopathy

I. Suárez, G. Bodega and B. Fernández (2010) Neuropathology and Applied Neurobiology 36, 422–435
Upregulation of α‐synuclein expression in the rat cerebellum in experimental hepatic encephalopathy Aims: The overexpression of α‐synuclein has been associated with neurodegenerative diseases, especially when the protein aggregates to form insoluble structures. The present study examined the effect of chronic hyperammonaemia on α‐synuclein expression in the rat cerebellum following portacaval anastomosis (PCA). Methods: Immunohistochemical and western blot determinations were performed 1 month and 6 months after the PCA procedure. Results: A time‐dependent increase in α‐synuclein expression was seen in the cerebellar grey matter compared with the controls. At 1 month post PCA, α‐synuclein‐immunopositive material was observed in the molecular layer, while the Purkinje cells showed weak α‐synuclein expression, and α‐synuclein aggregates were observed throughout the granular layer. At 6 months post PCA, α‐synuclein expression was significantly increased compared with the controls. α‐synuclein‐immunostained astroglial cells were also found; the Bergmann glial cells showed α‐synuclein‐positive processes in the molecular layer of PCA‐exposed rats, and in the granular layer, perivascular astrocytes showed intense α‐synuclein immunoreactivity, as indicated by colocalization of α‐synuclein with glial fibrillary acidic protein (GFAP). In addition, ubiquitin‐immunoreactive inclusions were present in PCA‐exposed rats, although they did not colocalize with α‐synuclein. Western blotting performed at 6 months post PCA showed a reduction in the level of soluble α‐synuclein compared with 1 month post PCA and the controls; this reduction was concomitant with an increase in the insoluble form of α‐synuclein. Conclusions: Although the precise mechanism by which α‐synuclein aggregates in PCA‐treated rats remains unknown, the present data suggest an important role for this protein in the onset and progression of hepatic encephalopathy, probably via its expression in astroglial cells.