Ultrafine Diesel Exhaust Particles Induce Apoptosis of Oligodendrocytes by Increasing Intracellular Reactive Oxygen Species through NADPH Oxidase Activation.

Diesel exhaust particles (DEPs) are a main contributor to air pollution. Ultrafine DEPs can cause neurodegenerative diseases by increasing intracellular reactive oxygen species (ROS). Compared with other cells in the brain, oligodendrocytes responsible for myelination are more susceptible to oxidative stress. However, the mechanisms underlying ROS generation in oligodendrocytes and the susceptibility of oligodendrocytes to ROS by ultrafine DEPs remain unclear. Herein, we examined the effects of excessive ROS generated by NOX2, an isoform of the NADPH oxidase family, after exposure to ultrafine DEPs (200 µg/mL) on the survival of two types of oligodendrocytes-oligodendrocyte precursor cells (OPCs) and mature oligodendrocytes (mOLs)--isolated from the brain of neonatal rats. In addition, mice were exposed to ultrafine DEP suspension (20 µL, 0.4 mg/mL) via the nasal route for 1 week, after which the expression of NOX2 and cleaved caspase-3 was examined in the white matter of the cerebellum. Exposure to DEPs significantly increased NOX2 expression and ROS generation in OPCs and mOLs. OPCs and mOLs clearly exhibited viability reduction, and a significant change in p53, Bax, Bcl-2, and cleaved caspase-3 expression, after DEP exposure. In contrast, treatment with berberine (BBR), an NOX2 inhibitor, significantly mitigated these effects. In mice exposed to DEP, the presence of NOX2-positive and cleaved caspase-3-positive oligodendrocytes was demonstrated in the cerebellar white matter; NOX2 and cleaved caspase-3 expression in the cerebellum lysates was significantly increased. BBR treatment returned expression of these proteins to control levels. These results demonstrate that the susceptibility of OPCs and mOLs to ultrafine DEPs is, at least in part, caused by excessive ROS produced by NOX2 and the sequential changes in the expression of p53, Bax, Bcl-2, and cleaved caspase-3. Overall, NOX2 inhibitor enhances the survival of two types of oligodendrocytes.

Effects of diesel exhaust particles and urban particles on brain endothelial cells.

Exposure to diesel exhaust particles (DEPs) and urban particles (UPs) increases the incidence of degenerative brain diseases as well as respiratory diseases. However, there is limited evidence on the mechanism of neurotoxicity on exposure to these particles. In the present study, the damage to blood-brain barrier (BBB) function by DEP or UP exposure was evaluated in bEnd.3 cells, which are derived from the brain tissue of Balb/c mice. It was demonstrated that DEP and UP exposure may induce oxidative stress via increasing reactive oxygen species (ROS) and decreasing total antioxidant capacity (TAC) level in bEnd.3 cells. In addition, cells exposed to DEP and UP demonstrated a resistance value of about 50% each compared to the value noted prior to exposure; additionally, Claudin-5 and ZO-1 expression levels were significantly decreased compared to the corresponding levels in the control. It was inferred that DEP or UP exposure diminishes the expression of tight junction proteins in endothelial cells through ROS generation, thereby enhancing endothelial membrane permeability. This study showed that DEPs or UPs induced cell permeability and oxidative stress by increasing ROS generation in bEnd.3 cells. This suggests the possibility that exposure to DEPs or UPs may compromise the integrity of the BBB and induce adverse effects in the CNS.

High Vulnerability of Oligodendrocytes to Oxidative Stress Induced by Ultrafine Urban Particles.

Oligodendrocytes, myelin-forming cells in the brain, are vulnerable to oxidative stress. Recent work indicates that air pollution causes demyelinating diseases such as multiple sclerosis. However, little is known about the mechanism of toxicity of ultrafine particulate matters (PMs) to oligodendrocytes. Here, we aimed to determine whether oligodendrocyte precursor cells (OPCs) and mature oligodendrocytes (mOLs) are more vulnerable to ultrafine urban PMs (uf-UPs) than other types of brain cells and damage to adult OPCs and mOLs in the mouse brain exposed to uf-UPs. For in vitro experiments, following exposure to various concentrations (2, 20, and 200 µg/mL) of uf-UPs, we measured survival rates, the amount of reactive oxygen species (ROS), and the total antioxidant capacities (TACs) of brain cells isolated from neonatal Sprague-Dawley rats. For animal experiments, after a four-week exposure to a uf-UP suspension (20 µL, 0.4 mg/mL), we enumerated the number of damaged cells and typed damaged cells in the white matter of the cerebellum of uf-UP-exposed mice. MTT assays and Hoechst staining demonstrated that OPCs and mOLs were more vulnerable to uf-UP-induced damage than astrocytes and cortical neurons at 2, 20, and 200 µg/mL of uf-UPs examined in this study (p < 0.05). Damage to OPCs and mOLs depended on uf-UP concentration. DCF assays and DHE staining indicated that the amount of ROS generated in OPCs and mOLs was significantly higher than in other brain cell types (p < 0.05). In contrast, TAC values in OPCs and mOLs were significantly lower than those of other brain cell types (p < 0.05). Fluoro-Jade B (FJB)-positive cells in the cerebellar white matter of the uf-UP-exposed group were significantly greater in number relative to the control group. Double immunofluorescence indicated that FJB-positive cells are NG2-positive adult OPCs and carbon anhydrase II-positive mOLs. Taken together, our findings suggest that oxidative stress induced by uf-UPs in the brain impairs adult OPCs and mOLs, causing demyelination and reducing the capacity for remyelination.

The vascular endothelial cell-expressed prion protein doppel promotes angiogenesis and blood-brain barrier development.

The central nervous system (CNS) contains a complex network of blood vessels that promote normal tissue development and physiology. Abnormal control of blood vessel morphogenesis and maturation is linked to the pathogenesis of various neurodevelopmental diseases. The CNS-specific genes that regulate blood vessel morphogenesis in development and disease remain largely unknown. Here, we have characterized functions for the gene encoding prion protein 2 (Prnd) in CNS blood vessel development and physiology. Prnd encodes the glycosylphosphatidylinositol (GPI)-linked protein doppel, which is expressed on the surface of angiogenic vascular endothelial cells, but is absent in quiescent endothelial cells of the adult CNS. During CNS vascular development, doppel interacts with receptor tyrosine kinases and activates cytoplasmic signaling pathways involved in endothelial cell survival, metabolism and migration. Analysis of mice genetically null for Prnd revealed impaired CNS blood vessel morphogenesis and associated endothelial cell sprouting defects. Prnd-/- mice also displayed defects in endothelial barrier integrity. Collectively, these data reveal novel mechanisms underlying doppel control of angiogenesis in the developing CNS, and may provide new insights about dysfunctional pathways that cause vascular-related CNS disorders.

Alpha B-Crystallin Overexpression Protects Oligodendrocyte Precursor Cells Against Oxidative Stress-Induced Apoptosis Through the Akt Pathway.

Alpha B-crystallin (aBC), a member of the small heat shock protein family, is expressed in mature oligodendrocytes (mOLs), but not in oligodendrocyte precursor cells (OPCs). Our previous study found that the survival rate of OPCs was lower than that of mOLs under oxidative stress, suggesting that aBC may play a protective role in mOLs. In the present study, we investigated the effects of aBC overexpression on oxidative stress-induced cell injury in OPCs and examined the underlying mechanisms. We observed that the survival rates of aBC-overexpressed OPCs were significantly higher than those of control cells under oxidative stress induced by hydrogen peroxide. Akt activities were significantly suppressed by oxidative stress in control OPCs, but not in aBC-overexpressed OPCs. The expressions of Bax and cleaved caspase-3 were decreased, whereas Bcl-2 expression was increased in aBC-overexpressed OPCs under oxidative stress. These findings suggest that low Akt activity in OPCs due to aBC deficiency may cause high susceptibility of OPCs to oxidative stress. The findings may provide new insights into the implication of OPCs in demyelinating diseases.

Iron is Responsible for Production of Reactive Oxygen Species Regulating Vasopressin Expression in the Mouse Paraventricular Nucleus.

Reactive oxygen species (ROS) act as signaling molecules for maintaining homeostasis, particularly in the regulation of body-fluid balance in the paraventricular nucleus (PVN) of the hypothalamus. However, there has been little discussion regarding the source of ROS generation in this hypothalamic region. Because iron is the most abundant metal in the brain, we hypothesized that iron may act as a source of ROS, which regulate vasopressin (VP) expression. In the present study, we compared the amount of iron in the PVN to that in other forebrain regions of normal ICR mice, and examined the relationship among iron, ROS, and VP in the PVN of the iron-overloaded with iron dextran and iron-chelated mice with deferoxamine. The amount of iron in the PVN was significantly higher than in any of the forebrain regions we examined. The amount of iron in the PVN was significantly increased in iron-overloaded mice, although not in iron-chelated mice. These results suggest that the PVN exhibits high iron affinity. Both ROS production and VP expression in the PVN of iron-overloaded mice were significantly increased relative to levels observed in control mice. VP concentration in blood of iron-overloaded mice was also significantly higher than that of control mice. Interestingly, iron overload did not alter the expression of nitric oxide synthase, another modulator of VP expression. Taken together, our results suggest that high levels of iron in the PVN induce the production of ROS, which regulate VP expression, independent of nitric oxide signaling.

RhoGDI2 expression in astrocytes after an excitotoxic lesion in the mouse hippocampus.

The Rho GDP-dissociation inhibitor (RhoGDI) originally downregulates Rho family GTPases by preventing nucleotide exchange and membrane association. Although RhoGDI2 functions as a metastasis regulator, little is known in glial cells under neuropathological conditions. We monitored RhoGDI2 expression in the mouse brain after administering a kainic acid(KA)-induced excitotoxic lesion. In control, RhoGDI2 immunoreactivity (IR) was evident in the neuronal layer of the hippocampus. However, RhoGDI2 IR was increased in astrocytes markedly throughout the hippocampus at day 3 post-treatment with KA. To further investigate the molecular mechanism of RhoGDI2-induced cellular migration, primary astrocytes were transfected with the flag-tagged RhoGDI2 cDNA. Cell migration assay revealed that RhoGDI2 cDNA transfection inhibits astrocyte migration. Overexpression of RhoGDI2 leads to inhibit protein kinase B (PKB) activation and cdc42 and cAMP-responsive element-binding protein (CREB) phosphorylation. In conclusion, our results suggested for the first time that RhoGDI2 is required for PKB and CREB activation and cdc42 expression in astrocyte migration after KA-mediated excitotoxic lesion in mouse brain.

Sequential accumulation of iron in glial cells during chicken cerebellar development.

Iron is an essential, but potentially harmful, metal in the brain. In normal brain, iron has been reported to accumulate mainly in glial cells and occasionally in neurons in some particular nuclei. However, the majority of investigations have targeted the adult brain. Here, we investigated spatiotemporal localization of iron in developing and adult chicken cerebellum using iron histochemistry. Iron reactivity was not detected in the chick cerebellum until embryonic day 12. Iron accumulation was first found in mature myelinating oligodendrocytes located in the inner part of the cerebellar folium at embryonic day 14. From embryonic day 20, iron-positive mature myelinating oligodendrocytes were localized in the white matter and the granular layer. From post-hatching day 2, iron accumulation was observed in Bergmann glia in the Purkinje cell layer as well as in mature myelinating oligodendrocytes. Iron accumulation in microglia was observed in the granular and molecular layers at post-hatching month 12. Our data indicate that during cerebellar development iron is accumulated in a unique sequence according to individual requirements or microenvironmental demands.

Differential expression of αB-crystallin causes maturation-dependent susceptibility of oligodendrocytes to oxidative stress.

Oligodendrocyte precursor cells (OPCs) are most susceptible to oxidative stress in the brain. However, the cause of differences in susceptibility to oxidative stress between OPCs and mature oligodendrocytes (mOLs) remains unclear. Recently, we identified in vivo that αB-crystallin (aBC) is expressed in mOLs but not in OPCs. Therefore, we examined in the present study whether aBC expression could affect cell survival under oxidative stress induced by hydrogen peroxide using primary cultures of OPCs and mOLs from neonatal rat brains. Expression of aBC was greater in mOLs than in OPCs, and the survival rate of mOLs was significantly higher than that of OPCs under oxidative stress. Suppression of aBC by siRNA transfection resulted in a decrease in the survival rate of mOLs under oxidative stress. These data suggest that higher susceptibility of OPCs than mOLs to oxidative stress is due, at least in part, to low levels of aBC expression.

Electrophysiological and Histologic Evaluation of the Time Course of Retinal Degeneration in the rd10 Mouse Model of Retinitis Pigmentosa.

Among several animal models of retinitis pigmentosa (RP), the more recently developed rd10 mouse with later onset and slower rate of retinal degeneration than rd1 mouse is a more suitable model for testing therapeutic modalities. We therefore investigated the time course of retinal degeneration in rd10 mice before adopting this model in our interventional studies. Electroretinogram (ERG) recordings were carried out in postnatal weeks (PW) 3~5 rd10 (n=23) and wild-type (wt) mice (n=26). We compared the amplitude and implicit time of the b-wave of ERG records from wt and rd10 mice. Our results showed that b-wave amplitudes in rd10 mice were significantly lower and the implicit time of b-waves in rd10 mice were also significantly slower than that in wt mice (20~160 µV vs. 350~480 µV; 55~75 ms vs. 100~150 ms: p<0.001) through PW3 to PW5. The most drastic changes in ERG amplitudes and latencies were observed during PW3 to PW4. In multichannel recording of rd10 retina in PW2 to PW4.5, we found no significant difference in mean spike frequency, but the frequency of power spectral peak of local field potential at PW3 and PW3.5 is significantly different among other age groups (p<0.05). Histologic examination of rd10 retinae showed significant decrease in thickness of the outer nuclear layer at PW3. TUNEL positive cells were most frequently observed at PW3. From these data, we confirm that in the rd10 mouse, the most precipitous retinal degeneration occurs between PW3~PW4 and that photoreceptor degeneration is complete by PW5.

Expression of LC3 and Beclin 1 in the spinal dorsal horn following spinal nerve ligation-induced neuropathic pain.

Impaired spinal GABAergic inhibitory function is known to be pivotal in neuropathic pain (NPP). At present, data concerning time-dependent alterations in cell type and cell death in the spinal dorsal horn are highly controversial, likely related to the experimental NPP model used. In this study, we examined the expression of autophagy using a L5 spinal nerve ligation (SNL)-induced neuropathic pain rat model. Following ligation of the spinal nerve, neuropathic pain behavior, such as mechanical allodynia, was induced rapidly and maintained for 14 days. After testing for mechanical allodynia, we assessed the changes in expression of LC3 and Beclin 1 in the spinal cord following SNL. Immunohistochemical analysis showed that the levels of LC3 and Beclin 1 protein in the ipsilateral L5 spinal dorsal horn were significantly elevated on day 14 following SNL. Double immunohistochemical analysis further confirmed increases in LC3 and Beclin 1 in mostly neurons and a few astrocytes following SNL. LC3 and Beclin 1 expressions were upregulated in GABAergic interneurons of spinal dorsal horn after SNL, while the loss of GABAergic interneurons did not change significantly. Our results suggest that autophagic disruption in GABAergic interneurons and astrocytes following peripheral nerve injury might be involved in the induction and maintenance of neuropathic pain.

Caffeine-induced endothelial cell death and the inhibition of angiogenesis.

Numerous studies have shown that adenosine or adenosine agonists can stimulate angiogenesis. However, the effect of caffeine (a known adenosine receptor antagonist) on angiogenesis has not been previously studied. Accordingly, this study was undertaken to examine the effect of caffeine on angiogenesis and to clarify the mechanism involved. Chick chorioallantoic membrane assays were used to investigate the effect of caffeine on angiogenesis and proliferation assays using human umbilical vein endothelial cells (HUVECs), were used to study its effects on specific aspects of angiogenesis. The expressions of caspase-3 and Bcl-2 were examined by western blotting, immunofluorescence staining was used to identify HUVEC morphological changes, and fluorescence activated cell sorting (FACS) and DAPI staining were used to detect HUVEC apoptosis. Caffeine was found to inhibit blood vessel formation dose-dependently and to inhibit the proliferation of HUVECs time- and dose-dependently. FACS analysis and DAPI staining showed that inhibitory effect of caffeine on HUVEC proliferation was the result of apoptosis and the up-regulation of thrombospondin-1 (TSP-1). Furthermore, TSP-1 levels were down-regulated by NECA but were unaffected by CGS21680, indicating that caffeine regulated TSP-1 expression via adenosine A2B receptor. In addition, caffeine up-regulated caspase-3 and down-regulated Bcl-2 at the protein level. These results suggest that the inhibitory effect of caffeine on angiogenesis is associated, at least in part, with its induction of endothelial cell apoptosis, probably mediated by a caspase-3 dependent mechanism.

Expression of CD200 in alternative activation of microglia following an excitotoxic lesion in the mouse hippocampus.

CD200 is a glycoprotein that is expressed on the surfaces of neurons and other cells. It interacts with its receptor, CD200R, which is expressed on cells of the myeloid lineage, including microglia. The interaction of CD200 with its receptor plays a significant role in maintaining microglia in a quiescent state; thus, a decrease in CD200 expression in the brain is associated with evidence of microglial activation. However, their roles in pathological progression remain unclear. We examined the expression of CD200 in kainic acid (KA)-induced neurodegeneration of the mouse hippocampus. Our quantitative analysis revealed that CD200 was constitutively expressed in the normal brain and transiently upregulated by KA treatment. At the cellular level, CD200 was expressed in neurons in control, and was upregulated primarily in the microglia of KA-treated mouse hippocampi. We examined the contribution of CD200 to both the classical and alternative activation of microglia in vitro using an adult microglia culture, which was exposed to interleukin-4 (IL-4) with and without lipopolysaccharide (LPS). CD200 expression was increased after exposure to IL-4, but not to LPS. These in vivo experiments demonstrated that CD200 was transiently expressed in microglia in a process mediated by the inflammatory response. Based on CD200R expression in microglia, it suggests that microglia is maintained in an activated state with autocrine signaling by interactions between microglial CD200 and its CD200R. Moreover, we suggest that CD200 may be expressed in the alternative activation of microglia and play a beneficial role in neuroinflammation.

alpha B-crystallin is expressed in myelinating oligodendrocytes of the developing and adult avian retina.

It is well known that the expression of α B-crystallin (aBC) is increased in neurons and glia under pathologic conditions. However, the expression of aBC during the normal development of the central nervous system has not been reported. This study aimed to clarify the cell type in the chick retina in which aBC is expressed and timing of aBC expression in this cell type during development. Double immunofluorescence with cell-specific markers demonstrated that aBC was selectively expressed in oligodendrocytes (OLs) in the embryonic day 20 (E20) chick retina. A small number of aBC-expressing OLs first appeared in the nerve fiber layer of the central and peripheral retina at E16. Faint aBC expression was also observed in myelin sheaths near cell bodies in the central retina. The number of aBC-expressing OLs and intensity of aBC expression in myelin sheaths were increased in the periphery as well as in the center of the E19 retina. aBC signals in the post-hatching day 120 retina were observed in the entire nerve fiber layer. The spatiotemporal expression pattern of aBC was identical to that of myelin basic protein. These data indicate that aBC-expressing OLs are myelinating OLs among OL-lineage cells. Besides, intrayolk injection of tocopherol, an antioxidant, provoked a decrease in the levels of aBC expression in myelinating OLs. These data suggest that aBC expression in myelinating OLs responds to the change of physiological oxidative stress.

Upregulation of peroxiredeoxin III in the hippocampus of acute immobilization stress model rats and the Foxo3a-dependent expression in PC12 cells.

Stress induces structural plasticity in neurons of the adult central nervous system (CNS) and alters the levels of cellular production of reactive oxygen species (ROS), and these changes might involve modifications of the antioxidant defense system. This study investigated whether acute stress altered the expression pattern of peroxiredoxin (Prx) III, which is an antioxidant enzyme that controls cytokine-induced peroxide levels. Prx III immunoreactivity was upregulated in the pyramidal neurons of the hippocampus and in the motor neurons of the spinal cord in an acute immobilization stress (AIS) model. In addition, we tested whether the transcription factor Foxo3a was necessary for the expression of Prx III. The depletion of Foxo3a led to a marked reduction of Prx III and a compensatory enhancement of mitochondrial superoxide dismutase (Mn-SOD) in PC12 cells. The results of this study suggest that Foxo3a mediates the neuronal levels of expression of Prx III and the levels of expression of Mn-SOD in mitochondria. These mechanisms may play an important role in neuroprotection against oxidative stress. Furthermore, Prx III upregulation might be an useful approach for the management of stress.

Characterization of the antigenic phenotype of αB-crystallin-expressing peripapillary glial cells in the developing chick retina.

Radial glia are transdifferentiated into astrocytes within the developing brain and spinal cord. The neural retina contains Müller cells, which are retinal radial glia. Some of the cells that surround the optic nerve head among Müller cells in the chicken retina are called peripapillary glial cells (PPGCs). PPGCs express different molecules compared to typical Müller cells. However, an antigenic PPGC phenotype has not yet been clearly established. In this study, we classified the antigenic PPGC phenotypes and identified the differentiation stages of these cells. At embryonic day (E)8, αB-crystallin-positive PPGCs had a bipolar shape with long processes that traversed entire layers of the retina. Pax2 and vimentin were expressed in αB-crystallin-positive PPGCs. Glial fibrillary acidic protein (GFAP) immunoreactivity was not observed in PPGCs. At E18, αB-crystallin immunoreactivity disappeared from the vitread processes of PPGCs. However, the PPGC cell bodies and ventricular processes contained αB-crystallin protein, and the PPGCs retained the same Pax2-positive/vimentin-positive/GFAP-negative profile as that seen at E8. At post-hatch day 120, αB-crystallin and Pax2 immunoreactivity was not observed, but vimentin and GFAP expression was clearly observed in the presumptive location of the PPGCs. Furthermore, these two proteins overlapped within that location. Considering that vimentin expression is prolonged until the post-hatching period in chicken brain, these findings suggest that Pax2-negative/vimentin-positive/GFAP-positive PPGCs are phenotypically identical to mature astrocytes in this avian species.

Expression of αB-crystallin in the peripapillary glial cells of the developing chick retina.

Peripapillary glial cells (PPGCs) are a peculiar macroglia in avian species, located in the central retina adjacent to the optic nerve head. PPGCs have a similar shape and orientation to Müller cells, which traverse the entire layer of the retina; however, there are differences in protein expression between the two cell types. In the present study, we first demonstrated that PPGCs expressed αB-crystallin, which is not expressed in Müller cells, during retinal development. αB-crystallin was first faintly expressed in PPGCs of the E5 retina, adjacent to the optic nerve head. Further, αB-crystallin was exclusively expressed in PPGCs up to E14. The shape of these cells was bipolar with vitread and ventricular processes. The vitread processes of αB-crystallin+ PPGCs became finer at E18. Double labeling analysis clearly demonstrated that only vimentin+ or GFAP+ astrocytes were located in the optic nerve head and were demarcated from the retina by αB-crystallin+ PPGCs. Furthermore, we determined that αB-crystallin+ PPGCs, with a number of processes, completely wrapped the optic nerve head and were densely located in the junction of the optic nerve head and the retina in a whole mount preparation and in vertical-sectioned retinae. The results of present study, together with reports that retinal astrocytes migrate from the optic nerve head, suggest that PPGCs prevent astrocytes from migrating into the retina in avian species.

Spatiotemporal gradient of astrocyte development in the chick optic tectum: evidence for multiple origins and migratory paths of astrocytes.

Astrocytes have been considered to be transformed from radial glial cells that appear at early stage of development and play a scaffold-role for neuronal cell migration. Recent studies indicate that neuroepithelial cells in the spinal cord also give rise to astrocytes. However, the mode of astroglial generation and migration in the ventricular neuroepithelium remains poorly understood. In this study, we have utilized immunohistochemical and retroviral lineage tracing methods to characterize the developmental profiles of astrocytes in the chick optic tectum, which develops from both the neural tube and invasion of optic tract. Chick vimentin and glial fibrillary acidic protein (GFAP) were found as single bands at molecular weights consistent with those reported for mammalian species. Differential developmental trends were observed for both proteins with relative vimentin levels decreasing and GFAP levels increasing with embryonic age. We observed two streams of tectal GFAP-labeled astrocytes originated from the tectal ventricle (intrinsic origin) and the optic tract (extrinsic origin). The extrinsic astrocytes arose from the ventral neuroepithelium of the third ventricle, dispersed bilaterally to the optic tract, and subsequently to the outer layer of optic tectum, indicating migration of astrocytes along retinal ganglion cell axons. On the other hand, the intrinsic astrocytes from the tectal ventricular neuroepithelium appeared first in the ventral part of the optic tectum, and then in the lateral and dorsal tectum. The intrinsic tectal astrocytes closely associated with fascicles of vimentin-labeled radial glial cells, indicating a presumptive radial migration of astrocytes. These results demonstrated that the optic tectum contains heterogeneous populations of astrocytes developed from the different origins and routes of migration.

Induction of transcription factor A-myb expression in reactive astrocytes following an excitotoxic lesion in the mouse hippocampus.

In the present study, we examined patterns of A-myb expression in the kainic acid (KA)-treated mouse hippocampus. Western blot analysis revealed that A-myb expression was dramatically increased in brain 3 days after KA treatment, and was sustained for more than 7 days. A-myb immunoreactivity was restricted to hippocampal neurons in control mice. Three days after KA treatment, strong A-myb immunoreactivity was observed in reactive astrocytes throughout the CA3 region. Thereafter, A-myb immunoreactive astrocytes gradually concentrated around the CA3 region in parallel with selective neuronal loss, and only a few A-myb immunoreactive astrocytes persisted in the CA3 region 14 days after KA treatment. These findings suggest that the A-myb plays a role in the reactive gliosis signaling pathway in KA-induced excitotoxic lesions.

The multiple dorsoventral origins and migratory pathway of tectal oligodendrocytes in the developing chick.

Oligodendrocytes have been considered to originate in a restricted ventricular zone of the ventral neural tube and to migrate and mature in their final targets. However, recent studies indicate that oligodendrocytes arise from multiple distinct dorsoventral origins. In this study, we investigate oligodendrocyte lineage cells in the embryonic optic tectum of chick, which develops from the dorsal region of the neural tube and invasion of optic tract. Oligodendrocyte precursor cells (OPCs) first appeared bilaterally on either side of the floor plate at E5. With further development, OPCs increased and spread laterally and dorsally to populate the optic tectum. At E7, OPCs appeared in another site along the ventral midline of the third ventricle, just dorsal to the optic chiasm. To examine the migration routes of these ventrally derived OPCs, we used DiI tracing in the organic culture and retinal denervation. Our results reveal that OPCs dispersed bilaterally along the optic tract and then migrated to the optic tectum in the stratum opticum (SO). In addition to these extrinsic OPCs, OPCs intrinsic to the tectal ventricle zone were identified at E14 using a combination of immunohistochemistry and retroviral mediated lineage tracing studies. These data support stage-specific dorsoventral origins and distribution of oligodendrocytes populating the optic tectum.