Glial fibrillary acidic protein immunodetection and immunoreactivity in the anterior and posterior medial amygdala of male and female rats

The medial amygdala (MeA) has receptors for gonadal hormones and modulates reproductive behaviors in rats. Adult male and female rats were used for the immunodetection, a less accurate technique, and the immunohistochemistry for the astrocytic marker glial fibrillary acidic protein (GFAP) in the anterior and posterior MeA. Both procedures were done using polyclonal anti-GFAP and were quantified by densitometry. The first technique provided no evidence for a difference between sexes in the immunocontent of GFAP in any region of the MeA (p > 0.1). Nevertheless, the measure of the intensity of GFAP immunoreactivity (GFAP-IR) showed that females had a higher GFAP-IR in the posterodorsal (p < 0.01) and in the posteroventral subregions of the MeA (p < 0.01) than males. No sex difference was found in its anterodorsal part (p > 0.1). The present results point out the differences between these two above-mentioned techniques but add a new finding to the previously described sexual dimorphism in the MeA, i.e., the GFAP-IR. Data also suggest that probably astrocytes can be affected by sex steroids in this brain area. It is likely that this regionally specific difference in the GFAP-IR may contribute to the distinct functional roles that the MeA subregions have in male and female rats.     

Expression of functional receptors and transmitter enzymes in cultured Muller cells

Glia represents the most numerous group of nervous system cells and CNS development and function depend on glial cells. We developed a purified Muller glia culture to investigate the expression of several neurotransmitter markers on these cells, such as dopaminergic, cholinergic, GABAergic and peptidergic receptors or enzymes, based on functional assays measuring second messenger levels or Western blot for specific proteins. Purified Muller cell culture was obtained from 8-day-old (E8) embryonic chick. Glial cells cultured for 15 days (E8C15) expressed D1A and D1B receptors mRNAs, but not D1D, as detected by RT-PCR. The binding of [3H]-SCH 23390 revealed an amount of expressed receptors around 40 fmol/mg protein. Dopamine (100 microM), PACAP (50 nM) and forskolin (10 microM) induced a 50-, 30- and 40-fold cAMP accumulation on glial cells, respectively, but not ip3 production. The dopamine-promoted cAMP accumulation was blocked by 2 microM SCH 23390. Carbachol stimulated a 3-fold ip3 accumulation. Western blot analysis also revealed the expression of tyrosine hydroxylase, L-dopa decarboxylase, PAC1 receptor, GAD67 and beta2-nicotinic receptor subunit by these cells. These results indicate that several components of neurotransmitter signaling and metabolism are found in cultured Muller cells.     

Glial fibrillary acidic protein (GFAP) from goldfish: its localisation in visual pathway

An intermediate filament fraction, isolated from goldfish brain, contains a prominent protein having a molecular weight of 51 kDa. In normal goldfish visual pathway, this protein is present in tectum and tract, but not in optic nerve. A polyclonal antibody raised to this protein clearly labels ependymal glial profiles in tectum and parallel processes in the tract, whereas optic nerve is unlabelled; Müller fibres in the retina are also labelled. A similar, but less prominent, pattern of staining is observed with antibodies, raised elsewhere, against glial fibrillary acidic protein from human and porcine. These results suggest that the 51 kDa protein is a GFAP, demonstrate the heterogeneity of astrocytes in goldfish visual pathway, and are consistent with the idea that GFAP is well conserved in vertebrate phylogeny.     

Ovarian cycle-related changes of glial fibrillary acidic protein (GFAP) immunoreactivity in the rat interpeduncular nucleus

The interpeduncular nucleus (IPN) of female rats was studied across the estrous cycle to observe whether the expression of the astroglial marker, glial fibrillary acidic protein (GFAP) reacts to hormonal changes in an area not belonging to the 'endocrine brain'. A marked reduction of immunoreactive GFAP was observed in estrus as compared to the immunoreactivities in met- and proestrus. This finding is consistent with earlier observations in the endocrine hypothalamus, but also proves that gonadal steroids influence astroglia in brain regions not involved in neuroendocrine regulation. Since cyclic fluctuations of synaptic numbers in the female have been described only for the endocrine hypothalamus, decrease of immunoreactive GFAP in the IPN during estrus may reflect a down-regulation of GFAP synthesis.     

Glial fiber pattern in the developing chicken cerebellum: vimentin and glial fibrillary acidic protein (GFAP) immunostaining

The possible relation between glial fibers and the formation of longitudinal granule cell migration patterns that occur in the cerebellar anlage of the chicken was investigated by immunocytochemistry of vimentin (monoclonal antibody) and glial fibrillary acidic protein (polyclonal antibody against GFAP, PGF) on fixed and unfixed brain tissues. In addition, neuronal development was studied with a monoclonal antibody for neurofilament. Vimentin was present in radial and tangential fibers in the cerebellar anlage during granule cell migration in almost all parts of the anlage. However, no specific topographic relation of vimentin and GFAP to the migration pattern of granule cells was observed. In adults, Bergmann fibers and astroglia were stained with vimentin antiserum and not with GFAP antiserum. Conclusions are that radial fibers do not determine the formation of longitudinal cytoarchitectonic patterns in the chick cerebellum and that vimentin is the main cytoskeletal component of Bergmann fibers and astroglial cells in embryonic and adult chicken cerebellum.     

Glial fibrillary acidic protein (GFAP) in rat brain tumors transplacentally induced by ethylnitrosourea (ENU)

The immunohistochemical distribution of glial fibrillary acidic protein (GFAP) in neoplastic lesions induced in the rat by ENU is reported. GFAP was present in hypertrophic reactive astrocytes, which were numerous in early neoplastic proliferations, in microtumors of the white matter, and in those collected at the periphery of large tumors. They were absent in cortical oligodendroglial foci and microtumors. No GFAP-positive cells were observed in hyperplasias of the white matter: astrocyte-like cells of large tumors were GFAP-negative. The significance of reactive astrocytes and the problem of the astrocytic component in transplacental ENU tumors are discussed.     

Glial fibrillary acidic protein in giant cell tumors of brain and other gliomas

Summary The giant cell tumor of brain (glioblastoma/sarcoma) has been considered a glioma by some and a sarcoma by others. This study shows that glial fibrillary acidic protein (GFAP), a specific marker for astrocytes, is present in the tumor cells, thus indicating that the cell of origin is the astrocyte and that the tumor should be called a giant cell glioblastoma. GFAP is present in the smaller cells of this tumor and in larger mononucleated cells, but little if any is detectable in multinucleated giant cells.In a different kind of tumor, the giant cell astrocytoma assonciated with tuberosclerosis, GFAP is restricted in most cells to a narrow peripheral zone.Immunocytochemical localization of GFAP is superior to special stains to differentiate giant cell glioblastomas from true sarcomas and giant cell bone tumors, since the latter are both negative for GFAP.Comparison of GFAP in all tumors of astrocyte origin shows that the cells that appear to contain the most GFAP include low grade well differentiated stellate cells, elongated piloid cells, and gemistocytic astrocytoma cells. Highly malignant undifferentiated cells, with less well developed processes, are less densely positive.Although there is in general an inverse relationship between GFAP content and degree of tumor malignancy, a more complex relationship exists with respect to individual cells; more GFAP is present in well differentiated cells with well-developed processes and filaments than in undifferentiated cells and large multinucleated cells. It is suggested that the pleomorphism of more malignant cells may relate to their relatively low GFAP content and perhaps to the disassembly of their glial filaments.Supported in part by the American Cancer Society, Inc., Grant No. PDT-162     

Glial fibrillary acidic protein (GFAP) in ependymal cells during development. An immunocytochemical study

Human ependymal cells show positive immunostaining for glial fibrillary acidic protein (GFAP) at one stage of the fetal development. The reaction seems to coincide with maturation of the epithelial layer and development of cilia. Two types of reactive cells are present: epithelial and tanycytes. The GFAP-positive reaction in both these cells is transient, appearing at different times and with different patterns in the various regions of the ventricular system. In order to explain the presence of detectable GFAP in developing ependymal cells and its absence in mature cells, it is proposed that either the synthesis of detectable amounts of GFAP occurs only at a stage of ependymal cell maturation, or that the intermediate filaments assembled in developing ependymal cells are antigenically distinct from those of the mature cells. The present findings indicate that tanycytes are not an immature form of ependymal cells but that they develop parallel to the epithelial cells. The role of the tanycytes remains obscure, but it is suggested that they are not related to radial glia.     

Glial fibrillary acidic protein (GFAP) in oligodendroglial tumors: Gliofibrillary oligodendroglioma and transitional oligoastrocytoma as subtypes of oligodendroglioma

Summary Immuno-reactivity to glial fibrillary acidic protein (GFAP) is mainly regarded as a sign of astroglial histogenesis and/or differentiation. The presence of astrocytes in oligodendrogliomas is a well known phenomenon; in addition, GFAP-positive neoplastic oligodendrocytes have also been described but have not yet been studied systematically.Using an anti-GFAP serum in the peroxidaseantiperoxidase (PAP) technique, 50 oligodendrogliomas and 16 mixed oligodendrocytomas were investigated; they had been diagnosed by routine histological stains. In half of all oligodendrogliomas, and only in a few (12%) of the mixed oligoastrocytomas, GFAP-positive oligodendrocytes were found in some areas of the classical honey-comb texture with a prominent vascular stroma. The term gliofibrillary oligodendrocyte (gfoc) is proposed for these immuno-reactive cells. The existence of a tumour cell combining morphological characteristics of oligodendroglia with GFAP production in its cytoplasm may be considered analogous to transient GFAP expression by myelinforming glia during normal development (Choi and Kim 1984), thus suggesting the return to a foetal behaviour by some neoplastic oligodendrocytes. Three tumours of the present series consisted largely of gfocs and, therefore, may be termed gliofibrillary oligodendrogliomas. In about 32% of all oligodendrogliomas, but only once in the mixed tumour group, a gradual morphological transition from gfocs to gemistocytic astrocytes was observed. This suggested a transitional cell type or transitional oligoastrocytoma as a further subtype of oligodendroglioma, including one example in which minigemistocytes dominated over gfocs (minigemistocytoma). These oligodendroglioma subtypes with GFAP-containing cells are different from the mixed oligoastrocytoma which is a tumour of two distinct and non-transitional cell populations.Supported in part by grants from The Netherlands' Organization for the Advancement of Pure Research (ZWO) and the Foundation for Scientific Education Limburg (SWOL), The Netherlands. Drs. Herper's present address is: Dept. of Neurology, De Wever Hospital, P.O. Box 4446, NL-6401 CX Heerlen, The Netherlands.     

Specific monoclonal antibodies to glial fibrillary acidic protein (GFAP)

Anti-glial fibrillary acidic protein (GFAP) monoclonal antibody has been obtained by fusing SP2/O myeloma cells with splenic cells from mice immunized with human GFAP. It belongs to the IgG1 class and it recognizes an epitope on GFAP which is shared by each fragment of the protein. Immunohistological studies show that the epitope characterized is absolutely specific for GFAP.     

Non-glial specificities of immunocytochemistry for the glial fibrillary acidic protein (GFAP)

Summary In an extensive immunocytochemistry study for glial fibrillary acidic protein (GFAP) of human neuropathological biopsy or autopsy tissue specimens examined for diagnostic or research purposes, rare non-glial specificities of the GFAP immunostain were observed: Schwann cells of some small nerves in salivary gland, renal capsule, and in epidural fat adjacent to a metastatic carcinoma, Schwann and satellite cells in a spinal ganglion invaded by tumor, chondrocytes of epiglottic cartilage, few cells of a malignant pleomorphic adenoma of salivary gland, most cells of a recurrent papillary meningioma with areas similar to the hemangiopericytic variant, and many cells of a renal carcinoma metastatic to brain; the primary renal tumor had been operated 4 years earlier and focally contained some GFAP-positive cells. To ascertain the specificity of such unexpected immunoreactivities for GFAP and to exclude possible crossreactivities with other intermediate filament (IF) proteins, a panel of different antibodies was used for immunocytochemistry with the peroxidase-antiperoxidase (polyclonal antisera) or labeled biotin-avidin (monoclonal antibodies) techniques: two monoclonal and four polyclonal anti-GFAP, three monoclonal and one polyclonal anti-cytokeratins (CK), and two monoclonal anti-vimentin (VIM) antibodies. Triple expression of GFAP, VIM and CK was found in the papillary meningioma (in patterns suggesting frequent co-localization), in the metastatic carcinoma (in patterns suggesting little co-localization), and in the pleomorphic adenoma (only few GFAP-positive cells). Co-expression of GFAP and VIM was seen in epiglottic chondrocytes and reactive astroglia; another metastatic carcinoma was labeled only for CKs. In the light of previous reports on non-glial specificities of the GFAP immunostain, and of the consistency of our immunostaining results obtained by all monospecific anti-GFAP antibodies as well as the lack of immunocytochemically evident crossreactivity with other IF proteins, authentic GFAP production by some rare non-glial tissues and tumors is suggested.Dedicated to Prof. F. Seitelberger on the occasion of his seventieth birthdaySupported by a project fund of the Austrian Ministry for Science and Research     

Production of glial fibrillary acidic protein (GFAP) by neoplastic cells: Adaptation to the microenvironment

Summary In 80 specimens of human glioma the production of glial fibrillary acidic protein (GFAP) by tumour cells invading meninges or connective tissue was studied immuno-cytochemically by the PAP technique. In 38 of 55 cases of astrocytoma, glioblastoma, gliosarcoma, and oligoastrocytoma, GFAP immuno-reactivity was greater in the invading cells as compared with the main part of the neoplasm. Fifty-eight percent of the astroglial tumours invading the leptomeninges, all astroglial tumours invading connective tissue and all gliosarcomas showed enhanced GFAP immuno-reactivity of tumour cells getting in contact with collagenous tissue, whereas meningeal infiltrates of 25 non-astroglial tumours (oligodendroglioma, ependymoma, medulloblastoma) remained GFAP-negative like the main part of the respective tumours. In the majority of astroglial tumours an increase of GFAP immunoreactivity was found also in perivascular cells of the main part of the tumour.It is concluded that glioma cells are capable of adapting their cytoskeleton to their micro-environment. Contact with dense collagenous tissue appears as an important factor able to induce an increased production of GFAP by adjacent glial cells.Supported in part by grants from The Netherlands Organisation for the Advancement of Pure Research (ZWO) and the Foundation for Scientific Education Limburg (SWOL), The Netherlands     

Neuronal modulation of Schwann cell glial fibrillary acidic protein (GFAP)

Adult rat sciatic nerves contain cytoskeletal peptides that resemble CNS glial fibrillary acidic protein (GFAP) in immunoreactivity and molecular weight. Immunohistological examination of teased nerve fascicles indicated that these peptides are expressed selectively by Schwann cells related to small axons. Radiolabelled mouse and rat CNS GFAP cDNA probes hybridized with a single, 2.7 kb RNA band in Northern blots prepared from total RNA from both rat sciatic nerve and rat brain. Sciatic nerve GFAP mRNA was detectable by this means in adult, 2 month, or 21 day postnatal rats, but not in 3,6, or 10 day postnatal rats. Sciatic nerve transection caused a marked reduction in the level of GFAP mRNA in the axotomized distal stump. We conclude that Schwann cell synthesis of GFAP is developmentally regulated and that Schwann cells, unlike astroglia, require continued trophic input from small axons in order to express GFAP.     

Distribution of glial fibrillary acidic protein-immunopositive structures in the brain of the domestic chicken (Gallus domesticus)

The present study is the first comprehensive mapping of glial fibrillary acidic protein (GFAP)-immunopositive structures in the avian brain. Two main types of GFAP-immunopositive elements were observed: (1) nonbranching fibers, occasionally twisted or varicose, and (2) star-shaped cells. Long immunostained fibers orignate from the lateral ventricle to form three bundles. Fibers of the dorsal group, emanating from the dorsal/lateral corner of the ventricle, course in lateral, anterior, and ventral directions forming a semidome, which separates the outer pallial (lateral cortical) regions from the underlying striatal mass. The middle group of fibers is directed anteriorly and laterally corresponding to the laminae frontales superior and suprema. The ventral fiber bundle is conical and traverses the lobus parolfactorius, crossing also the lamina medullaris dorsalis (the latter consisting mainly of star-shaped cells). The hippocampus, septum, and hypothalamus also contain straight radial fibers. In some areas, given their variable orientation, the fibers cannot be regarded as merely persisting radial glia. In the telencephalon, the nuclei basalis, accumbens, ectostriatum, paleostriatum primitivum, and the ventral paleostriatum are particularly rich in GFAP-positive cells, whereas the neostriatum, hyperstriatum, and paleostriatum augmentatum are almost devoid of GFAP labelling. Certain nuclei of the thalamus and the lower brainstem are conspicuous by their low levels of GFAP immunoreactivity. The Bergmann glia were GFAP-immunonegative.     

Glial fibrillary acidic protein (GFAP) in oligodendrogliomas: a reflection of transient GFAP expression by immature oligodendroglia

Fourteen pure oligodendrogliomas were studied by light- and electronmicroscopy and immunohistochemistry to examine glial fibrillary acidic protein (GFAP) positivity in the tumors. To compare the immunohistochemical staining patterns of neoplastic oligodendroglia and immature oligodendroglia, myelination glia in the white matter of eight normal brains from children under 6 months of age were studied. The tumors possessed light microscopic and ultrastructural features characteristic of oligodendrogliomas. Microtubules were found in the cytoplasm of nine tumors on electronmicroscopy. In one, intermediate filaments and microtubules were observed in occasional tumor cells with polygonal crystalline structures in the cytoplasm. Using the peroxidase-antiperoxidase technique, all specimens were stained for GFAP, vimentin, S-100 and neuron-specific enolase (NSE). In nine tumors, variable numbers of cells with an oligodendroglial morphology reacted positively for GFAP. All tumors were positive for S-100 and negative for vimentin and NSE. The myelination glia in the eight normal brains stained positively for GFAP but not for vimentin. Vimentin is expressed by developing, reactive and neoplastic astrocytes. Thus, GFAP positivity combined with vimentin negativity in both neoplastic and immature oligodendroglia suggests that GFAP positivity in oligodendrogliomas may reflect the transient expression of this intermediate filament by immature oligodendroglia.     

Glial fibrillary acidic protein (GFAP): the major protein of glial intermediate filaments in differentiated astrocytes

The glial fibrillary acidic protein (GFA protein or GFAP) is the major protein constituent of glial intermediate filaments in differentiated fibrous and protoplasmic astrocytes of the central nervous system. Proteins having similar molecular weights, isoelectric points, and immunoreactivity with GFAP have been found in cells of neural crest and ectodermal origin. A putative function ascribed to glial filaments is its role as a component of the cytoskeleton in defining and maintaining the shape of the astrocyte. Since 1980, over 350 reports have utilized antisera to GFAP for immunochemical and immunocytochemical studies.     

Glial fibrillary acidic protein (GFAP)-positive radial-like cells are present in the vicinity of proliferative progenitors in the nucleus tractus solitarius of adult rat

The dorsal vagal complex (DVC), an integrative center of autonomic functions located dorsally in the caudal brainstem, comprises the nucleus tractus solitarius (NTS), the area postrema (AP), and the dorsal motor nucleus of the vagus nerve (DMNX). Recently, this area of the brainstem was shown to retain, during adulthood, the expression of developmental markers, which is consistent with several forms of morphological and functional plasticity. These data led us to attempt to determine the structural organization and phenotypical characteristics of the astroglial compartment in the adult DVC. We report a strikingly high density of glial fibrillary acidic protein (GFAP) immunoreactive cells in the NTS and the DMNX compared to other brainstem structures. Furthermore, we observed a subpopulation of atypical GFAP+ cells in the NTS. These cells expressed vimentin and nestin and displayed unbranched processes that radiate rostrocaudally from cuboid cell bodies located in the 4th ventricle wall. Interestingly, these radiating cells were found in close association with neural progenitors whose proliferation was stimulated by intracerebroventricular injection of epidermal growth factor/basic fibroblast growth factor or lesion of the vagus nerve. Newly born neurons in the NTS identified by doublecortin (DCX) immunolabeling were also preferentially found in the vicinity of the radiating cells. Altogether, these results indicate that the adult NTS retains, during adulthood, astroglial cells that display morphological and phenotypical features seen during development. The overlap in the distribution of proliferative neural progenitors, newborn neurons, and radiating GFAP-positive cells suggest a possible role of the glial compartment of the NTS in functional plasticity in this structure.     

Glial fibrillary acidic protein and S-100 protein in human hepatic encephalopathy: Immunocytochemical demonstration of dissociation of two glia-associated proteins

Summary Immunocytochemical staining patterns for glial fibrillary acidic protein (GFAP) and S-100 protein (S100P) were compared in cerebral cortex, basal ganglia and white matter of eight cases with hepatic encephalopathy (HE), including four cases of Wilson's disease and four of liver cirrhosis, and of eight age-matched controls, using the peroxidase-antiperoxidase method on adjacent paraffin sections. The majority of Alzheimer type II glia (Alzg II) showed prominent immunoreactivity for S100P but not for GFAP, resembling normal astrocytes of protoplasmic type; Alzg II might be interpreted as being peculiar types of reactive astrocytes retaining characteristics of protoplasmic astrocytes. A small number of Alzg II cells showed slight perinuclear immunoreactivity for GFAP; some lacked both markers. This suggests a spectrum of metabolic changes in these two proteins in Alzg II. GFAP-positive Alzg II cells were restricted to basal ganglia and white matter adjacent to grey matter, indicating that expression of GFAP in Alzg II might be modulated by local factors. Alzheimer type I cells and Opalski cells in Wilson's disease were immunoreactive for both proteins, confirming their astroglial origin and different character from that of Alzg II. In morphometric comparison, the proportion of GFAP-positive glial cells decreased in the cortex (P<0.001) but not significantly in the white matter (0.05<P<0.1), confirming earlier data that the prominent reduction of GFAP in HE brains is restricted to the grey matter. In the putamen, the proportion of GFAP-positive glia decreased in acquired HE (0.01<P<0.05) but not in Wilson's disease, probably resulting from prominent fibrous glial poliferation related to severe parenchymal damage in Wilson's disease. In contrast, glial cells immunoreactive for S100P did not significantly change (P>0.1) in any of the areas studied, indicating an intact glial metabolism of S100P in HE. This clear dissociation between GFAP and S100P defines Alzg II as a peculiar glial reaction with a rather selective deficit of GFAP metabolism (gliofibrillary dystrophy).     

Glial fibrillary acidic (GFA) protein in Müller glia. Immunofluorescence study of the goldfish retina

Glial fibrillary acidic protein, the subunit of intermediate filaments specific for astrocytes, was localized by immunofluorescence in the Müller glia of goldfish retina. Based on previous studies reporting the localization in Müller glia of carbonic anhydrase C, an oligodendrocyte marker, we suggest that the main type of neuroglia in the retina combines properties which in the brain are specific for astrocytes and oligodendrocytes.