bluuebunny/crossref_metadata_2025 huggingface dataset:
Lactic acid inhibition of gap junctional intercellular communication in in vitro astrocytes as measured by fluorescence recovery after laser photobleaching
Authors: Anders Juanita J.
Published: January 2005

Lactic acid can permeate plasma membranes, causing intracellular acidosis. Gap junctions are sensitive to pHi and can be reversibly uncoupled by weak acids. In this study, dye coupling between in vitro astrocytes, presumably mediated by gap junctions, was measured in the absence and presence of lactic acid. Fluorescence recovery after laser photobleaching (gap‐FRAP analysis) was used to measure dye coupling. Astrocytes bathed in Eagle's minimum essential medium (EMEM) with lactic acid, pHo 5.5–6, showed no difference in their dye coupling (mean recovery of fluorescence 30%) when compared to control astrocytes (mean recovery of fluorescence 26%). However, 24 mM lactic acid in EMEM, pHo 4.5, decreased dye coupling (mean recovery of fluorescence 2.0%). This effect occurred within 5 min of treatment. When lactic acid‐EMEM, pH 4.5, was removed from astrocytes after 30 min and the cells were incubated in EMEM for 24 hr, decreased coupling was not reversed (mean recovery 4.0%). When lactic acid‐treated astrocytes were incubated in EMEM for 48 hr, the mean recovery of fluorescence increased to 15% (i.e., 42% of the recovery seen in controls). These observations suggest that brief exposure to high concentrations of lactic acid can have immediate and long‐lasting effects on glial gap junctional communication. Under pathological circumstances, such a sequence could be intiated, and this might impair astrocytic control of the central nervous system microenvironment mediated by spatial buffering.
Heterogeneity of potassium currents in cultured oligodendrocytes
Authors: Sontheimer H. , Kettenmann H.
Published: January 2005

In the present study we have analyzed the membrane currents of mature oligodendrocytes in cultures from dissociated fetal mouse cerebral hemispheres and explant cultures from fetal mouse spinal cord. Both types of oligodendrocytes showed large voltage‐dependent, but time‐independent inward and outward currents that were partially blocked by Ba2+. In addition, time‐ and voltage‐dependent inward and outward currents were observed in a minority of oligodendrocytes from spinal cord. All voltage‐dependent currents were completely blocked by Ba2+, and inward currents were completely blocked by Cs+, suggesting that they are mediated by K+ channels. Current‐voltage curves of mouse spinal cord oligodendrocytes varied from being linear to outwardly or inwardly rectifying. In contrast, oligodendrocytes cultured from mouse brain always showed an inward rectification of the current voltage relation and a lack of time‐dependent currents. It thus appears that mature oligodendrocytes in explant cultures of mouse spinal cord, in contrast to oligodendrocytes from dissociated brain, consist of different cell populations that are distinguished by their expression by their expression or active state of K+ channels.
Astroglial cells in vitro are heterogeneous with respect to expression of the α1‐adrenergic receptor
Authors: Lerea Leslie S. , McCarthy Ken D.
Published: January 2005

A wide variety of approaches have been used to examine the expression of neuroligand receptors by cultured astroglial cells. The results of such studies make it clear that these cells exhibit most, if not all, of the different receptors known to be associated with central neurons. However, it has been more difficult to determine if there are multiple populations of astroglia that can be distinguished on the basis of their complement of neuroligand receptors. To address this question, we established a radioligand binding assay that enabled us to visualize α1‐adrenergic receptors (α1‐ARs) on immunocytochemically defined neural cells. Saturation, time course, and competition binding experiments determined that 125I‐HEAT could be used to identify α1‐AR binding sites on immunocytochemically defined astroglial cells. Our results indicate that approximately 66% of cortical polygonal astroglia express detectable numbers of α1‐ARs. 3H‐thymidine labeling experiments indicate that both dividing and nondividing astroglia exhibit α1‐ARs. These results greatly contrast with studies on β‐adrenergic receptor expression (β‐AR), in which 100% of polygonal astroglia express β‐ARs. Process‐bearing astroglia also exhibit α1‐ARs, which is in marked contrast to the lack of β‐AR expression by these cells. The results presented here suggest that astroglia, like neurons, differ in the receptors they express and therefore may be able to selectively respond to extracellular stimulation.
Myelination in the jimpy mouse in the absence of proteolipid protein
Authors: Duncan I.D. , Hammang J.P. , Goda S. , Quarles R.H.
Published: January 2005

A point mutation in the gene for proteolipid protein (PLP) has been suggested to account for the dysmyelination seen in the jimpy mouse mutant. Despite the absence of PLP, the major integral membrane protein of central nervous system (CNS) myelin, this study shows that there are many scattered myelinated fibers present in the spinal cord of this murine mutant which are immunocytochemically positive for myelin basic protein (MBP), yet negative for PLP. This lack of PLP results in an abnormal compaction of the extracellular leaflets of the myelin sheath and the formation of an abnormal intraperiod line. These results are similar to those seen in another X‐linked myelin mutant, the myelin‐deficient rat (Duncan et al.: Proc. Natl. Acad. Sci. U.S.A., 84:6287–6291, 1987), and show that a multilamellar membrane can be formed in the absence of its major integral membrane protein.
Trauma‐induced proliferation of astrocytes in the brains of young and aged rats
Authors: Topp Kimberly S. , Faddis Brian T. , Vijayan Vijaya K.
Published: January 2005

The time‐course and magnitude of astrocyte proliferation following neural trauma was evaluated in young adult (3 months) and mid‐aged (16–19 months) male Fischer 344 rats. One to 4 days after a needle wound was made through the cortex and the hippocampus, rats received three intraperitoneal injections of 3H‐thymidine at 8 hour intervals and were sacrificed 1 hour after the last injection. For astrocyte quantification, 3H‐thymidine autoradiography was combined with immunohistochemical staining for glial fibrillary acidic protein followed by semithin sectioning. In areas of the cortex and hippocampus adjacent to the wound, astrocytes were categorized as unlabeled or labeled with silver grains over the nuclei. Labeling index and numerical density of astrocytes were determined using stereological methods. The results showed that in both young and older rats, astrocyte proliferation is an early glial response to neural trauma, occurring during the first 4 postlesion days and contributing to an increase in astrocyte population. Regional differences in labeling index and numerical density suggest heterogeneity in the proliferative capacity of astrocyte subpopulations in the rat brain. Compared with young animals, older rats demonstrated greater labeling in the cortex but not in the hippocampus. Thus, aging is associated with region‐specific increase in astrocyte reactivity to trauma possibly due to increased availability of mitogens or enhanced sensitivity of astrocytes to mitogenic signals.
Attempt to classify glial cells by means of their process specialization using the rabbit retinal Müller cell as an example of cytotopographic specialization of glial cells
Authors: Reichenbach Andreas
Published: January 2005

The rabbit retinal Müller cell is one of the most widely studied glial cell types, and it has all forms of contacts that a glial cell can express, viz. (1) to a (ventricular) fluid space, (2) to a mesenchymal borderline (basal lamina), and (3) to neuronal compartments. This cell demonstrates the local adaptation of cell processes to the microenvironment with which they are in contact. Summarizing available data on Müller cells and other glial cell types, it is concluded that the structure with which the process is in contact determines the type of glial cell process that develops. The type I process has microvilli, desmosome‐like junctions, and high Na+, K+‐ATPase activity; this type of process is in direct contact with a fluid such as cerebrospinal fluid. The type II endfoot‐bearing process contains gliofilaments and has a high K+ conductivity; this type of process is covered by a basal lamina and is in contact with mesenchyme. The type III sheath‐bearing process insulates neuronal compartments and expresses suitable membrane properties for glia‐neuronal communication. Since structurally similar processes have been shown to have similar physiological properties, a new systematic classification of glial cells is proposed, based on the presence or absence of defined types of cell processes. This approach is believed to provide new insights into the function of neuroglia in both the central and peripheral nervous systems, in vertebrates and invertebrates, and even during ontogenetic development.
Single channel potassium currents in cultured adult bovine oligodendrocytes
Authors: McLarnon James G. , Kim Seung U.
Published: January 2005

These studies have enabled the first characterization of the properties of ion channels in adult oligodendrocytes. Cell‐attached recordings from cultured adult bovine cells showed channel activity with 140 mM KCl in the patch pipette; the amplitude of the currents was increased with increasing membrane hyperpolarization. This channel, with a conductance of 29 pS, was selective for inward K+ current; little or no outward current was measured for large depolarizing voltage steps. The channel open time was strongly dependent upon membrane potential, with membrane hyperpolarization decreasing the mean open time 100‐fold over a range of 80 mV; at the resting potential of the cell the mean open time of the channel was in excess of 50 ms. Decreasing the concentration of K+ in the pipette diminished the channel conductance with no significant effect to alter the channel open time dependence on potential. The rectification and kinetic properties of the channel would be consistent with a physiological role for the channel in the regulation of external K+ near active neurons; in particular the effect of membrane depolarization to cause maintained channel open duration could be important when the driving force for inward potassium movement through oligodendrocyte membrane was low. Channels selective for outward potassium movement were seen with inside‐out excised patch recordings with symmetrical potassium concentrations across the patch; the density of these channels in the bovine membrane was low.
Sodium channels in astrocytes of rat optic nerve in situ: Immuno‐electron microscopic studies
Authors: Black J.A. , Waxman S.G. , Friedman B. , Elmer L.W. , Angelides K.J.
Published: January 2005

Immuno‐electron microscopic localization of sodium channels within astrocyte somata and processes of adult rat optic nerve was demonstrated with polyclonal antibody 7493. In immunoblots of crude glycoproteins from adult rat optic nerve, antisera 7493, which is directed against purified rat brain sodium channels, recognizes a 260 kDa protein. Antisera 7493 intensely immunostains axon membrane at nodes of Ranvier. Associated perinodal astrocyte processes are also stained with antisera 7493. In addition, astrocyte cell bodies and major processes exhibit immunoreactivity with antibody 7493. Immunostaining with antisera 7493 is heterogeneously distributed within astrocyte cytoplasm and also appears to be associated with some regions of astrocyte plasmalemma. Glial filaments are not immunostained with 7493 antisera. Astrocyte processes forming the glial limitns and surrounding blood vessels display reduced immunoreactivity to 7493 compared to longitudinally oriented or perinodal astrocyte processes. However, some focal regions of the glial limitans exhibit robust 7493 immunostaining. Oligodendrocytes do not display 7493 antisera immunoreactivity. Optic nerve sections incubated with preimmune sera or with 7493 antisera that had been previously adsorbed with purified sodium channel protein, exhibited no immunoreactivity. These results demonstrate localization of sodium channels within astrocytes in situ of rat optic nerve and extend previous electrophysiological and pharmacological findings of sodium channels in cultured astrocytes. Possible functional roles of sodium channels within astrocytes are discussed.
Phosphorylation of myelin basic protein in intact oligodendrocytes: Inhibition by galactosylsphingosine and cyclic AMP
Authors: Vartanian Timothy , Dawson Glyn , Soliven Betty , Nelson Deborah J. , Szuchet Sara
Published: January 2005

We have previously shown that cyclic AMP (cAMP) inhibits the protein kinase C (PKC)‐mediated phosphorylation of myelin basic protein (MBP) in cultured oligodendrocytes (OLGs). Recently, it has been demonstrated that the long chain base sphingosine inhibits PKC by competing with PKC effectors (diacylglycerol and phorbol esters) for a binding site on the kinase (Hannun and Bell: Science 235:670–674, 1987). In this report we define further the mechanism by which cAMP inhibits MBP phosphorylation by comparing the effects of cAMP with that of galactosysphingosine (psychosin), a potential catabolite of galactocerebroside, the major OLG glycosphingolipid. We identify the consequences of psychosine treatment and PKC down‐regulation on OLG morphology and electrophysiology and discuss their relevance. Our results in intact ovine oligodendrocytes are consistent with a mechanism in which cAMP inhibits MBP phosphorylation by interfering with the release of diacylglycerol (DAG) from phosphatidylinositol. First, the effects of cAMP on MBP phosphorylation are reversed with exogenous TPA; and second, cAMP inhibits the incorporation of 1‐[14C]arachidonate into DAG and specifically inhibits the turnover (as judged by 32PO3–4 incorporation) of phosphatidylinositol. Psychosine inhibits MBP phosphorylation, and its action can be reversed by TPA suggesting a mechanism of inhibition similar to that described for other systems. In addition, psychosine has profound effects on OLG morphology; it disintegrates OLG processes while leaving the cell soma intact. Stable hyperpolarized resting potentials were obtained following psychosine treatment, but there was a 66% decrease in membrane capacitance indicating a significant decrement in membrane surface area. The morphological changes induced by psychosine are reversible and can be eliminated by removing the drug but not by the addition of TPA. Whether inhibition of PKC by psychosine plays any role in process dissolution remains an unanswered question. However, current evidence suggests that a PKC‐independent mechanism may be at play. This investigation in conjunction with our previous work emphasizes a role for the interregulation of protein kinase A (PKA) and PKC in the control of OLG somal vs. myelin components. This may have significant implications for central nervous system myelin assembly.
Phosphotyrosine antibodies specifically label ameboid microglia in vitro and ramified microglia in vivo
Authors: Lee Tillotson Meryl , Wood John G.
Published: January 2005

Using an affinity‐purified, polyclonal antibody to phosphotyrosine (Wang: Molecular and Cellular Biology 5:3640–3643, 1985) we have previously demonstrated that phosphotyrosine immunoreactivity is restricted to a population of multipolar GFAP‐negative neuroglia in adult rat brain (Tillotson and Wood: Journal of Comparative Neurology 282:133–141, 1989) and retina (Tillotson and Wood: Journal of Cell Biology 107:724a, 1988). In this study, we show that the phosphotyrosine‐immunoreactive cells are microglia. This conclusion is supported by numerous morphological and ultrastructural similarities between the phosphotyrosine‐immunoreactive cells and microglia. Furthermore, phosphotyrosine co‐localizes with the microglial‐specific B4 isolectin of Bandeiraea simplifolia‐1 lectin. Phosphotyrosine antibodies also stain ameboid microglia in primary cultures of neonatal rat brain. In addition, after 7 days in vitro, microglia are the only phosphotyrosine‐immunoreactive element in the cultures. This temporal pattern of staining in vitro mimics the developmental progression of phosphotyrosine immunoreactivity in situ, in which a variety of structures stain during postnatal neural development (Tillotson and Wood: Journal of Comparative Neurology 282:133–141, 1989), but only microglia stain in mature brain. The significance of phosphotyrosine‐containing proteins potentially expressed in a microglial‐specific manner is discussed.
Visualization of oligodendrocytes and astrocytes in the intact rat optic nerve by intracellular injection of lucifer yellow and horseradish peroxidase
Authors: Butt Arthur M. , Ransom Bruce R.
Published: January 2005

The morphology of glial cells in the intact rat optic nerve, a central nervous system (CNS) white matter tract, was analysed by filling over 500 macroglial cells intracellularly with horseradish peroxidase (HRP) or Lucifer yellow (LY). Two main cell types were distinguished: fibrous astrocytes and cells presumed to be oligodendrocytes. Intracellularly stained astrocytes were highly complex, with 50–60 long branching processes which passed radially from the cell body and terminated in end‐feet at the pial surface or on blood vessels; some processes ended freely in the nerve parenchyma. Astrocytes filled with LY were usually dye‐coupled to other astrocytes after the first week of life. Filled oligodendrocytes had a unique appearance that unmistakably distinguished them from astrocytes and were occasionally dye‐coupled to nearby oligodendrocytes. These cells had 20–30 longitudinally oriented processes 150–200 μm long, which passed exclusively along the long axis of the nerve parallel to axons; the longitudinal processes were connected to the cell body by thin branches 15–30 μm long. The longitudinal processes probably represent the tongue processes of the internodal myelin sheaths, and thus each oligodendrocyte appears to myelinate 20–30 axons with sheaths that are 150–200 μm in length.
Human chromogranin A‐like immunoreactivity in the bergmann glia of the rat brain
Authors: McAuliffe W. Geoffrey , Hess Arthur
Published: January 2005

A mouse monoclonal antibody to human chromogranin A (LK2H10) selectively stains the Bergmann glia (Golgi epithelial cells) of the rat cerebellum, while rabbit antibodies to bovine or porcine chromogranin A do not. Chromogranin A immunostaining in the cell bodies and processes reveals the characteristic candelabra morphology seen in Golgi preparations of these cells. Immunostaining adjacent sections with antibodies to glial fibrillary acidic protein shows that a relatively small number of the glial cell processes in the molecular layer are chromogranin A positive. Recent reports indicate that the metabolites of chromogranin A play a regulatory role in endocrine and neuroendocrine systems but the function of chromogranins in neurons and glia is unknown.
Transection of the rat olfactory nerve increases glial fibrillary acidic protein immunoreactivity from the olfactory bulb to the piriform cortex
Authors: Anders Juanita J. , Johnson Julie A.
Published: January 2005

Astrocytic glial fibrillary acidic protein (GFAP) immunoreactivity in response to retrograde changes of motoneurons after axotomy has been the subject of a number of reports. In contrast, this study examined the astrocytic GFAP immunoreactivity in response to axotomy in a sensory system, the adult rat olfactory system. The purpose of this study was to determine, by immunolabeling GFAP, the extent and transience of astrocytic reactivity in the olfactory system. Unilateral transection of the olfactory nerve fascicles was performed intracranially at the level of the cribriform plate. Rats were allowed to survive from 24 hours to 1 month after axotomy. GFAP immunolabeling was examined throughout the rat olfactory system using the peroxidase–anti‐peroxidase method. After axotomy, a transient increase occurred in the astrocytic GFAP immunore‐activity in the ipsilateral olfactory system. The greatest enhancement of GFAP immuno‐reactivity in the olfactory system occurred at 48 hours post‐axotomy. Increased GFAP immunoreactivity occurred not only along the axons and synaptic endings of the injured primary olfactory neurons, but also along the dendrites, cell bodies, axons, and synaptic endings of the secondary sensory neurons. The increased GFAP immunoreactivity was specifically associated with the anatomical distribution pathways of the primary and secondary olfactory neurons. Increased GFAP immunoreactivity was not altered until 14 days post‐axotomy. At 1 month post‐axotomy, GFAP immunoreactivity returned to control levels. The time course and transience of increased GFAP immunoreativity closely correlates with the time course of rat primary olfactory neuronal degeneration and regeneration after axotomy. The increase in astrocytic GFAP immunoreactivity along the secondary olfactory neurons suggests the existence of a transsynaptic effect that is directly related to the state of degeneration and regeneration of the primary olfactory neurons.
Expression of glial fibrillary acidic protein (GFAP) in goldfish optic nerve following injury
Authors: Stafford C. A. , Shehab S. A. S. , Nona S. N. , Cronly‐Dillon J. R.
Published: January 2005

By using an antibody to goldfish glial fibrillary acidic protein (GFAP), the reaction of goldfish optic nerve to injury has been studied by immunoblotting and immunohistochemical methods. Goldfish optic nerve, which normally lacks GFAP immunoreactivity (Nona et al.: Glia, 2:189–200, 1989), expresses GFAP following injury. This immunoreactivity, which is observed as early as 10 days after crush and which is still evident at 30 days after crush, all but disappears by 150 days after crush. Since it is well established that functional restoration of synaptic connections and the recovery of vision takes place in goldfish following optic nerve injury, our results indicate that reactive astrocytes do not represent an impediment to regeneration in goldfish visual system.
A functional parameter to study heterogeneity of glial cells in rat brain slices: Cyclic guanosine monophosphate production in atrial natriuretic factor (ANF)‐responsive cells
Authors: De Vente J. , Manshanden C. G. , Sikking R. A. , Ramaekers F. C. S. , Steinbusch H. W. M.
Published: January 2005

Stimulation of guanylate cyclase in vitro by atrial natriuretic factor (ANF) or sodium nitroprusside was studied in rat brain tissue slices biochemically as well as by means of cyclic guanosine monophosphate (cGMP) immunocytochemistry. The ANF‐responsive, cGMP‐producing cells were studied in the olfactory bulb, the septal area, the hippocampus, the medial amygdala, and the medial preoptic area. These cells, having the ANF‐stimulated particulate guanylate cyclase, were characterized as astroglial cells on the basis of their glial fibrillary acidic protein (GFAP) immunostaining, although not all astroglial cells in these areas could be identified as cGMP‐immunoreactive cells. Sodium nitroprusside‐stimulated soluble guanylate cyclase activity was demonstrated in neuronal cell bodies and varicose fibers and was associated with blood vessel walls.

Upon maturation, a significant decrease in cGMP production was found after stimulation by 100 nM ANF‐(103–126) in the olfactory bulb, the medial amygdala, and the hippocampus, but not in the septal area; no change was found in these areas in cGMP content after stimulation of cGMP production by 10 μM sodium nitroprusside. Via cGMP immunocytochemistry, no qualitative differences were seen in the ANF‐responsive, cGMP‐producing cells upon maturation.
Region‐ and sex‐related differences in maturation of astrocytes in dissociated cell cultures of embryonic rat brain
Authors: Beyer C. , Epp B. , Fassberg J. , Reisert I. , Pilgrim C.
Published: January 2005

Previous studies using dissociated cell cultures of fetal rat brain have revealed considerable regional diversity as well as sex steroid‐independent sex differences in developmental schedules of dopaminergic neurons. Because these phenomena might be related to glial heterogeneity, cultures of dissociated male and female diencephalon, mesencephalon, and rhombencephalon of gestational day 14 rats were investigated with respect to the development of astrocytic markers. Cultures were incubated for 3–8 days in vitro (DIV) in serum‐supplemented or serum‐free medium. Vimentin and glial fibrillary acidic protein (GFAP) were quantified by counting of immunolabeled cells and immunoblotting. Vimentin and GFAP content rose from DIV 3 to 6 in all cultures. Regional variation of vimentin content was low, but large differences occurred in amounts of GFAP. GFAP reached high levels in rhombencephalon, especially when supplemented with serum, but remained very low or not detectable in mesencephalon. Simultaneous immunostaining for both cytoskeletal proteins revealed the presence of large numbers of vimentin single‐labeled and small numbers of vimentin/GFAP double‐labeled cells. Numbers of cells expressing GFAP showed similar regional variations as GFAP contents in both serum‐free and serum‐supplemented medium. They rose steeply from DIV 3 to 8 in rhomb‐ and diencephalon but not in mesencephalon. Transiently, female diencephalic cultures contained slightly more GFAP‐immunoreactive cells than male cultures. The results thus demonstrate considerable regional heterogeneity of astrocytic maturation. However, neither the regional nor the sex differences show a consistent correlation with previous data on development of dopaminergic and other monoaminergic neurons in vitro. It seems likely that the dependence of neurons on glial environment for realization of an inherent developmental program varies among neuronal phenotypes.
Intracellular pH shifts capable of uncoupling cultured oligodendrocytes are seen only in low HCO3− solution
Authors: Kettenmann H. , Ransom B. R. , Schlue W.‐R.
Published: January 2005

Electrical coupling between cultured mouse oligodendrocytes was transiently blocked when pHi was decreased below about 6.5 using the NH4+ prepulse method. This uncoupling could, however, only be achieved if the dominant pHi regulating mechanism in these cells, the Na+/HCO3− cotransporter, was blocked by lowering bath [HCO3−]. Under this condition, an NH4+ prepulse caused pHi to decrease toward the passive distribution for H+ (i.e., about pH 6.2). In the presence of normal bath [HCO3−] an NH4+ prepulse did not decrease pHi below 6.5 even when the second pHi regulating mechanism, the Na+/H+ exchanger, was blocked by amiloride, and consequently oligodendrocytes could not be uncoupled. Increasing CO2, which uncouples glial cells in situ (Connors et al: J. Neurosci. 4:1324–1330, 1984), did not uncouple cultured oligodendrocytes in the presence of normal bath [HCO3−], but did cause uncoupling in low [HCO3−] solution. These results indicate that electrical coupling between cultured oligodendrocytes is sensitive to pHi; in normal bath [HCO3−], however, the pHi regulation of these cells is so effective that standard techniques for intracellular accidification are unable to lower pHi to levels which cause the closure of oligodendrocyte gap junctions.
Schwann cells depleted of galactocerebroside express myelin‐associated glycoprotein and initiate but do not continue the process of myelination
Authors: Owens Geoffrey C. , Bunge Richard P.
Published: January 2005

Two peripheral myelin components, galactocerebroside (GalC) and myelin‐associated glycoprotein (MAG), are known to be expressed early in Schwann cell differentiation, prior to the formation of definitive myelin segments containing compacted membrane. To discern the relative roles of these myelin components, cultures of Schwann cells and dorsal root ganglion neurons were treated with antigalactocerebroside mAbs in order to remove GalC from the Schwann cell surface (Ranscht et al., 1987). In the continuous presence of anti‐GalC antibodies and in a medium containing serum plus ascorbic acid, Schwann cells assemble a basal lamina and progress to the one:one stage of Schwann cell:axon interaction but do not differentiate further. Immunostaining with anti‐MAG antibodies revealed that GalC‐depleted Schwann cells expressed high levels of MAG. Double staining with anti‐MAG and anti‐Po antibodies showed that there was essentially no Po immunoreactivity in the same cells. In those Schwann cells that had attained a one:one association with large‐diameter axons, the inner‐axon‐related cytoplasmic process had passed under the outer mesaxon but had not completed a full turn around the axon. The expression of MAG on the single cytoplasmic process apposed to the axon in Schwann cells depleted of GalC further implicates MAG in the initial envelopment of the axon during myelination.
Neuronal‐glial interactions: Complexity of neurite outgrowth correlates with substrated adhesivity of serotonergic neurons
Authors: Lieth Erich , McClay David R. , Lauder Jean M.
Published: January 2005

To study the interactions between neurons of known transmitter phenotype and non‐neuronal cells of glial or fibroblastic origin, serotonergic (5‐HT) neurons were tested for their strength of adhesion and neurite outgrowth patterns on substrates of astrocytes or fibroblasts using a cell adhesion assay for transmitter‐identified neurons, and morphometry of immunocytochemically stained neurons in dissociated cell cultures. Both the strength of adhesion and the rate and complexity of neurite outgrowth by 5‐HT neurons were significantly greater on substrates of astrocytes compared to fibroblasts. These results provide evidence that 5‐HT neurons can interact selectively with glia via cell surface determinants, and that this process may be important for the development of complex (dendrite‐like) neuritic arbors. The methods developed in this study will be useful for future studies of interactions between transmitter‐identified neurons and glial cells during ontogeny of the embryonic brain.
Growth of adult rat retinal ganglion cell neurites on astrocytes
Authors: Baehr M. , Bunge R. P.
Published: January 2005

Astrocytes, as well as Schwann cells (SC), can provide suitable substrata for embryonic neurites during development, but their abilities to support adult regenerating neurites have not been directly compared. The aim of the present study was to determine the ability of astrocytes to promote adult rat retinal ganglion cell (RGC) regeneration in vitro and to compare this to previously determined growth on the surface of Schwann cells. We prepared Type I astrocytes (Raff et al: J. Neurosci. 3:1289–1300, 1983) from perinatal rats. These were subcultured and maintained in either a serum‐free medium for at least 2 weeks (stellate astrocytes with little immunoreactivity for laminin) or in serum containing medium for 7 to 10 days (flat and polygonal astrocytes with immunoreactivity for laminin). Stellate astrocytes might therefore represent mature astrocytes in vivo (Ard and Bunge: J. Neurosci. 8:2844–2858, 1988), while flat astrocytes might resemble immature brain astrocytes (Liesi et al: J. Cell Biol. 96:920–924, 1983). Adult RGC survival and axonal regrowth on these glia populations was compared to that observed on different SC populations, as previously reported (Baehr and Bunge: Exp. Neurol. 106:27–40, 1989). Both astrocyte populations (either flat or stellate astrocytes) did not enhance RGC survival. Stellate astrocytes were less effective in supporting RGC axon regeneration than flat astrocytes. When these date were compared to RGC survival and axon growth on SC (Baehr and Bunge: Exp. Neurol. 106:27–40, 1989) only “ctivated” mature SC populations were superior to astrocytes in enhancing RGC survival and neurite regrowth. These results suggest 1) that astrocytes and “immature” SC are similar in their ability to support RGC survival; 2) “activated” mature SC populations are significantly better than astrocytes and “immature” SC in enhancing RGC survival and neurite growth; 3) stellate (“mature”) astrocytes, although permissive for regrowing axons, are not a favorable substrate for regenerating adult RGC neurites, nor do they effectively support RGC survival.
Dark‐rearing retards the maturation of astrocytes in restricted layers of cat visual cortex
Authors: Müller Christian M.
Published: January 2005

The cat visual cortex develops its mature appearance, i.e., its circuitry and neuronal morphology, during a limited period of postnatal development under the influence of visual experience. The critical period for cortical plasticity, which normally extends from the third to seventh postnatal week, can be prolonged by raising animals in total darkness. The prolongation of the critical period by dark‐rearing is restricted to the cortical layers except layer IV. Besides the influence of afferent activity on the physiology of cortical cells and on the interconnectivity of thalamo‐cortical afferents, visual experience has also been shown to affect the development of glial cells. The present study investigates the effects of dark‐rearing on astroglial characteristics as determined by immunostaining for glial fibrillary acidic protein (GFAP) and the S‐100 protein. The data reveal a retardation of astrocytic maturation in dark‐reared animals, shown by a reduced presence of GFAP immunoreactivity compared to light‐experienced animals. The density of astrocytic cell bodies positive for S‐100 is unaffected by dark‐rearing, suggesting that astroglial proliferation does not rely on afferent activity. However, punctate S‐100 staining in the neuropil, which has been shown to reflect astrocytic processes, was also reduced in certain cortical layers in dark‐reared animals. The effects of dark‐rearing on the expression of GFAP and S‐100 were restricted to the cortical layers except layer IV, i.e., those layers that reveal a prolongation of the critical period for cortical plasticity following dark‐rearing. It is concluded that astrocytic maturation in the visual cortex is influenced by neuronal activity. The coincidence of the location of retarded astrocytic maturation and the prolongation of the critical period for cortical plasticity is discussed in the light of the recent evidence that immature astrocytes may be involved in activity‐dependent plasticity of the visual cortex.
Simulation of aspects of ischemia in cell culture: Changes in lactate compartmentation
Authors: Walz Wolfgang , Mukerji Srimathie
Published: January 2005

Aspects of hyperglycemic ischemia were simulated in cultures of astrocytes and of neurons by high glucose and dinitrophenol exposure. Lactate release increased almost sevenfold and it was found that astrocytes were responsible for 92% of the release. There was no significant increase in internal lactate content. Experiments involving loading of astrocytes with lactate at different external pH values showed that lactate accumulation was increased by an increased inward proton gradient. This inward transport of lactate probably consists of two transport components, a passive diffusion of its neutral form and transport via a recently described monocarboxylic acid carrier. It was found that lactate did not get trapped in astrocytes, despite the fact that loading of astrocytes with lactic acid by exposure to 30 mM lactic acid increased the membrane input resistance dramatically. We conclude that lactate is released as lactic acid from astrocytes and equilibrates quickly with all CNS compartments. Thus we argue against a role of lactate accumulation in cytotoxic swelling.
Astrocytes: A possible primary site for colchicine‐mediated neurotoxicity in the rat striatum
Authors: Pitts Olin M. , Cohen Martin R. , McAmis William , Gutman Robert L.
Published: January 2005

Ultrastructural changes in rat astrocytes including endfoot swelling and rupture of plasma membranes were seen as early as 1 h after injection of 6.0 μg colchicine (CO) into the striatum. Significant change in neuronal morphology was not seen by 24 h. Astrocytes may therefore be primary targets for CO‐mediated damage, which then may have secondary neurotoxic consequences.
Ontogeny of radial and other astroglial cells in murine cerebral cortex
Authors: Misson Jean‐Paul , Takahashi Takao , Caviness Verne S.
Published: January 2005

Three cell forms of astroglial lineage populate the prenatal and early postnatal murine cerebral wall. In the present review we consider the ontogeny of these cell forms with respect to histogenetic events of the perinatal period.

Classic bipolar radial glial cells predominate prior to E17. The bipolar coexist with monopolar radial forms in the perinatal period. Both bipolar and monopolar radial forms coexist with multipolar astrocytes in the course of the first postnatal week and are ultimately succeeded by the multipolar cells. The shift from bipolar to monopolar radial forms is initially coincident with translocation of somata of bipolar cells from the ventricular zone to the upper intermediate zone and cortical strata. Arborization appears to occur both at the growing tips and along the shaft of the processes of both bipolar and monopolar radial cell types. As arborization continues, the processes of the monopolar radial cells come to resemble those of the multipolar astrocytes. Eventually the radial cells are fully transformed into the multipolar astrocytic forms. During this period of transition, radial processes in the cortex appear to be degenerating, suggesting that regressive processes contribute to the cytologic transformation.

This sequence of transformations begins late in the period of neuronal migration and continues through the early stages of growth and differentiation in the murine cerebral cortex. The signals that induce these changes may arise from differentiating neurons within the cortex. These transformations occur at a time when radial glial fibers are no longer required as guides for neuronal migration, and the glial population assumes new roles related to the development and operation of cortical neuronal circuits.
Muscle‐derived factors induce proliferation and astrocytic phenotypic expression in C‐6 glial cells
Authors: Brodie Chaya , Vernadakis Antonia
Published: January 2005

Previous studies from our laboratory have shown that C‐6 glial cells in culture express astrocytic phentoypes with increasing cell passage. Early passage glial cells have been shown to respond to various factors by expressing either astrocytic or oligodendrocytic phenotype. In view of the numerous reports on the trophic effects of muscle‐derived factors on spinal cord neurons, we examined the effect of muscle‐dervied factors on C‐6 glial cells from both early and late passages. Soluble factors were obtained from either leg (LME) or breast (BME) muscle from 15‐day‐old chick embryos by homogenization followed by high velocity centrifugation. Both LME‐ and BME‐derived factors triggered the proliferation and differentiation of early passage glial cells as assessed by cell number and glutamine synthetase activity, respectively. Late passage glial cells responded to muscle‐derved factors with enhanced proliferation and only with a slight increase in glutamine synthetase activity. These findings indicate that glial cells of both early and late passages respond to signals produced by muscle cells.
Gomori‐positive astrocytes: Biological properties and implications for neurologic and neuroendocrine disorders
Authors: Schipper Hyman M.
Published: January 2005

Granule laden astrocytes exhibiting an affinity for chrome alum hematoxylin and aldehyde fuchsin (Gomori stains) have been described in the periventricular brain of all terrestrial vertebrate species examined to date including humans. The astrocytic inclusions are rich in sulfhydryl groups, emit an orange‐red autofluorescence, and stain intensely with diaminobenzidine, a marker of endogenous peroxidase activity. The distinct autofluorescence pattern and the absence of neutral lipid, acid phosphatase, and β‐glucuronidase activity exclude lipofuscin or lysosomes as components of these astrocytic granules. The emission of orange‐red autofluorescence and the nonenzymatic nature of the peroxidase activity implicate the presence of porphyrins and metalloporphyrins such as heme as major constituents of these cytoplasmic gliosomes. The role of Gomori‐positive astrocytes under normal and pathologic conditions is incompletely understood. In vivo, numbers of astrocytic granules increase as a function of advancing age, in response to chronic estrogen stimulation, and following X‐irradiation. In vitro, these cells accumulate with increasing time in culture and following exposure to the sulfhydryl agent, cysteamine. Gomori‐positive astrocytes may supply heme to neurons for the synthesis of cytochromes, catalases, and other heme enzymes. They may play a role in photostimulation of sexual cyclicity, the promotion of neuritic development, the degradation of toxic lipoperoxides, and the metabolism of various neurotransmitters. Conversely, these cells may contribute to the pathogenesis of several neurologic and neuroendocrine disorders. Examples of the latter include (a) augmentation of goldthioglucose neurotoxicity, (b) induction of hypothalamic anovulation and reproductive failure, (c) exacerbation of porphyric encephalopathy, and (d) potentiation of parkinsonism and other free radical‐related neurodegenerations.
Glial fibrillary acidic protein in the fish optic nerve
Authors: Blaugrund Eran , Cohen Ilana , Shani Yael , Schwartz Michal
Published: January 2005

The intermediate filament glial fibrillary acidic protein (GFAP) is the predominant cytoskeletal protein of mature glial cells in the mammalian nervous system. The nervous systems of lower vertebrates, such as fish, have been examined for the presence of GFAP and several investigators have shown that goldfish (Carassius auratus) brain contains GFAP‐positive astrocytes. The same studies have demonstrated that, in contrast to the brain, the optic nerve of goldfish did not show any GFAP immunoreactivity, suggesting that this intermediate filament protein is not expressed in fish optic nerve astrocytes. The present study shows, however, that the monoclonal antibodies to porcine GFAP react with the optic nerve of carp (Cyprinus carpio), another member of the goldfish family. These antibodies to porcine GFAP cross react with rat brain and carp optic nerve, yielding a band of approximately 52 kDa in both species. Northern blot analysis using mouse GFAP DNA probe revealed that carp optic nerve RNA contains two transcripts, of 2.3 and 2.1 kb, which hybridize with the mouse GFAP probe. Injury to the carp optic nerve was followed by a decrease of GFAP immunoreactivity from neural tissue and a strong expression around blood vessels and connective tissues. On the basis of these observations and within the limitation of the techniques it is reasonable to conclude that the carp optic nerve expresses GFAP immunoreactivity and that the pattern of expression of this intermediate filament protein is altered after injury. Such an alteration might be relevant to the process of regeneration.
Axon‐glia interactions in the crayfish: Glial cell oxygen consumption is tightly coupled to axon metabolism
Authors: Hargittai Pa'l T. , Lieberman Edward M.
Published: January 2005

Oxygen consumption (Q̇) of single isolated axons and their associated glial cell sheath was investigated under a variety of conditions to determine the contribution of each cell type to whole tissue Q̇. It was found that the Q̇ of the sheath, in the absence of a functional axon, represented approximately 30% of the total tissue Q̇. When the axon was injected with carboxyatractyloside, an inhibitor of mitochondrial oxidative phosphorylation that is membrane impermeant, electrophysiological properties of the axon were not affected and glial sheath respiratory activity was stimulated by 1.7 to 2.7 times the untreated control level.

These results suggest that glial cell metabolic activity is regulated by the metabolic activity of the axon. Depending on the experimental conditions the glial sheath accounts for 30% to nearly 100% of the Q̇ of axon‐glial cell tissue. On the basis of these and morphometric measurements we estimated that in a normally functioning axon‐glial cell system the glial sheath accounts for 90% of the tissue Q̇.
Cerebral type 2 astroglia are heterogeneous with respect to their ability to respond to neuroligands linked to calcium mobilization
Authors: Dave Vijendra , Gordon Gerald W. , McCarthy Ken D.
Published: January 2005

Very little information is available concerning the pharmacology of type 2 astroglia. During the past decade it has become apparent that two distinct lineages of astroglial cells can be defined in vitro. These two lineages are commonly referred to as type 1 and type 2 and are distinguished from each other on the basis of their morphological features and antigenic phenotypes. In contrast to type 1 astroglia, very little is known about the pharmacology of type 2 astroglia. The lack of information concerning the responsiveness of these cells stems primarily from difficulties encountered in isolating large numbers of type 2 astroglia free of other cell types. In the present study video‐ and photometer‐based imaging systems were used to monitor the influence of a series of neuroligands on the intracellular calcium levels of individual cerebral type 2 astroglia in order to assess their expression of calcium‐mobilizing receptors. The responses of 85 immunocytochemically identified cerebral type 2 astroglia to bradykinin (BK), norepinephrine (NE), histamine (HIST), carbachol (CARB), 2‐methyl‐thio ATP (2MT‐ATP), glutamate (GLUT), and serotonin (5‐HT) were analyzed. Approximately 50% of cerebral type 2 astroglia responded to BK, NE, HIST, CARB, and 2MT‐ATP whereas only 16% and 9% of the cells responded to GLUT and 5‐HT, respectively. The number of neuroligands that increased calcium in individual cells ranged from 0 to 6. These responses are quite similar to those previously demonstrated in cultured cerebral type 1 astroglia. No pattern of receptor co‐expression was observed for the different neuroligands tested. While variable, distinct differences were observed in the time course of the calcium responses elicited by different neuroligands. The present study demonstrates that cultured cerebral type 2 astroglia are heterogenous when judged by their ability to respond to neuroligands with a rise in intracellular calcium.