Initial stages of radial glia astrocytic transformation in the early postnatal anterior subventricular zone

In the early postnatal subventricular zone (SVZ), two seemingly unrelated events occur simultaneously: a massive tangential migration of neuroblasts towards the olfactory bulb, known as the rostral migratory stream (RMS), and the outward movement of radial glia (RG) undergoing astrocytic transformation. Because of the orthogonal arrangement between these two sets of cells, little, if any, relevance has been ascribed for their possible interactions. By depositing DiI at the pial surface we have studied RG transformation within the SVZ/RMS, from birth up to the end of the first postnatal week. While still within the SVZ/RMS, RG morphology changed from simple bipolar to highly complex branched profiles, attaining their highest degree of complexity at the interface of the SVZ with the overlying white matter. At this interface cell bodies of radial glia accumulate and their processes run tangentially, surrounding the SVZ/RMS. Processes of RG surrounding the SVZ/RMS could also be observed by immunostaining for vimentin, GFAP, and nestin. In contrast, in the white matter all DiI-labeled RG presented a simple bipolar profile. These results indicate that the outward radial migration of the transforming RG does not occur uniformly. Instead, the different morphologies and cell densities that RG assume when they cross the SVZ/RMS and overlying white matter imply different migratory behaviors. Finally, our data suggest that RG provide a cellular scaffold to the early postnatal SVZ/RMS, much in the same way as astrocytes in the adult RMS.     

Gap junctions are involved in cell migration in the early postnatal subventricular zone

The massive migration of neuroblasts and young neurons through the anterior extension of the postnatal subventricular zone (SVZ), known as the rostral migratory stream (RMS) is still poorly understood on its molecular basis. In this work, we investigated the involvement of gap junctional communication (GJC) in the robust centrifugal migration from SVZ/RMS explants obtained from early postnatal (P4) rats. Cells were dye‐coupled in homocellular and heterocellular pairings and expressed at least two connexins, Cx 43 and 45. Treatment with the uncoupler agent carbenoxolone (CBX, 10–100 μM) reversibly reduced outgrowth from SVZ explants, while its inactive analog, glycyrhizinic acid (GZA), had no effect. Consistent with a direct effect on cell migration, time‐lapse video microscopy show that different pharmacological uncouplers cause an abrupt and reversible arrest of cell movement in explants. Our results indicate that GJC is positively involved in the migration of neuroblasts within the SVZ/RMS. © 2009 Wiley Periodicals, Inc. Develop Neurobiol, 2009     

NPY promotes chemokinesis and neurogenesis in the rat subventricular zone

The subventricular zone (SVZ) is a major reservoir for stem cells in the adult mammalian brain. Neural stem cells supply the olfactory bulb with new interneurons and provide cells that migrate towards lesioned brain areas. Neuropeptide Y (NPY), one of the most abundant neuropeptides in the brain, was previously shown to induce neuroproliferation on mice SVZ cells. In the present study, performed in rats, we demonstrate the endogenous synthesis of NPY by cells in the SVZ that suggests that NPY could act as an autocrine/paracrine factor within the SVZ area. We observed that NPY promotes SVZ cell proliferation as previously reported in mice, but does not affect self-renewal of SVZ stem cells. Additionally, this study provides the first direct evidence of a chemokinetic activity of NPY on SVZ cells. Using pharmacological approaches, we demonstrate that both the mitogenic and chemokinetic properties of NPY involve Y1 receptor-mediated activation of the ERK1/2 MAP kinase pathway. Altogether, our data establish that NPY through Y1 receptors activation controls chemokinetic activity and, as for mice, is a major neuroproliferative regulator of rat SVZ cells.     

Apoptosis-inducing factor deficiency decreases the proliferation rate and protects the subventricular zone against ionizing radiation

Cranial radiotherapy in children often leads to progressive cognitive decline. We have established a rodent model of irradiation-induced injury to the young brain. A single dose of 8 Gy was administered to the left hemisphere of postnatal day 10 (P10) mice. Harlequin (Hq) mice, carrying the hypomorphic apoptosis-inducing factor AIF^Hq mutation, express 60% less AIF at P10 and displayed significantly fewer dying cells in the subventricular zone (SVZ) 6 h after IR, compared with wild type (Wt) littermates. Irradiated cyclophilin A-deficient (CypA^−/−) mice confirmed that CypA has an essential role in AIF-induced apoptosis after IR. Hq mice displayed no reduction in SVZ size 7 days after IR, whereas 48% of the SVZ was lost in Wt mice. The proliferation rate was lower in the SVZ of Hq mice. Cultured neural precursor cells from the SVZ of Hq mice displayed a slower proliferation rate and were more resistant to IR. IR preferentially kills proliferating cells, and the slower proliferation rate in the SVZ of Hq mice may, at least partly, explain the protective effect of the Hq mutation. Together, these results indicate that targeting AIF may provide a fruitful strategy for protection of normal brain tissue against the detrimental side effects of IR.     

Persistent migration of neuroblasts from the subventricular zone to the injured striatum mediated by osteopontin following intracerebral hemorrhage

We investigated intracerebral hemorrhage (ICH)-induced lateral migration of neuroblasts and the mechanism underlying this migration. ICH model was induced by collagenase injection into the striatum of adult wild-type and osteopontin (OPN) knockout mice. In the wild-type mice, the lateral migration of neuroblasts from the ipsislateral subventricular zone (SVZ) towards the hematoma started at day 3 and continued up to day 28 after ICH. In addition to migrating towards the hematoma, neuroblasts also migrated to the area of ipsilateral striatum remote to the hematoma. The migrating neuroblasts were closely associated with activated astrocytes and blood vessels in the injured striatum. Following ICH, the expression of OPN was up-regulated in the ipsilateral striatum from day 1 to day 28. In vitro , OPN treatment did not affect the proliferation of neural progenitors, but enhanced the trans-well and radial migration of neural progenitors. In vivo , OPN deficiency did not affect the proliferation of neural progenitors in the SVZ. However, following ICH a significant decrease in lateral neuroblast migration was observed in the OPN knockout mice compared with the wild-type mice. These results suggest that increased OPN expression in the injured striatum plays a significant role in the lateral migration of neuroblasts following ICH.     

Architecture and cell types of the adult subventricular zone: In search of the stem cells

Neural stem cells are maintained in the subventricular zone (SVZ) of the adult mammalian brain. Here, we review the cellular organization of this germinal layer and propose lineage relationships of the three main cell types found in this area. The majority of cells in the adult SVZ are migrating neuroblasts (type A cells) that continue to proliferate. These cells form an extensive network of tangentially oriented pathways throughout the lateral wall of the lateral ventricle. Type A cells move long distances through this network at high speeds by means of chain migration. Cells in the SVZ network enter the rostral migratory stream (RMS) and migrate anteriorly into the olfactory bulb, where they differentiate into interneurons. The chains of type A cells are ensheathed by slowly proliferating astrocytes (type B cells), the second most common cell type in this germinal layer. The most actively proliferating cells in the SVZ, type C, form small clusters dispersed throughout the network. These foci of proliferating type C cells are in close proximity to chains of type A cells. We discuss possible lineage relationships among these cells and hypothesize which are the neural stem cells in the adult SVZ. In addition, we suggest that interactions between type A, B, and C cells may regulate proliferation and initial differentiation within this germinal layer. © 1998 John Wiley & Sons, Inc. J Neurobiol 36: 234–248, 1998     

Opposing effects of retinoid signaling on astrogliogenesis in embryonic day 13 and 17 cortical progenitor cells

All-trans retinoic acid (RA) is a differentiation factor in many tissues. However, its role in astrogliogenesis has not been extensively studied. Here, we investigated the effect of RA on the regulation of astrogliogenesis at different cortical developmental stages. We prepared rat cortical progenitor cells from embryonic day (E) 13 and E17, which correspond to the beginning of neurogenic and astrogliogenic periods, respectively. Surprisingly, RA promoted astrogliogenesis at E17 but inhibited astrogliogenesis induced by ciliary neurotrophic factor (CNTF) at E13. The inhibitory effect of RA on astrogliogenesis at E13 was not due to premature commitment of progenitors to a neuronal or oligodendroglial lineage. Rather, RA retained more progenitors in a proliferative state. Furthermore, RA inhibition of astrogliogenesis at E13 was independent of STAT3 signaling and required the function of the α and β isoforms of the RA receptors (RAR). Moreover, the differential response of E13 and E17 progenitors to RA was due to differences in the intrinsic properties of these cells that are preserved in vitro . The inhibitory effect of RA on cytokine-induced astrogliogenesis at E13 may contribute to silencing of any potential precocious astrogliogenesis during the neurogenic period.     

Notch1 signaling modulates neuronal progenitor activity in the subventricular zone in response to aging and focal ischemia

Neurogenesis diminishes with aging and ischemia‐induced neurogenesis also occurs, but reduced in aged brain. Currently, the cellular and molecular pathways mediating these effects remain largely unknown. Our previous study has shown that Notch1 signaling regulates neurogenesis in subventricular zone (SVZ) of young adult brain after focal ischemia, but whether a similar effect occurs in aged normal and ischemic animals is unknown. Here, we used normal and ischemic aged rat brains to investigate whether Notch1 signaling was involved in the reduction of neurogenesis in response to aging and modulates neurogenesis in aged brains after focal ischemia. By Western blot, we found that Notch1 and Jagged1 expression in the SVZ of aged brain was significantly reduced compared with young adult brain. Consistently, the activated form of Notch1 (Notch intracellular domain; NICD) expression was also declined. Immunohistochemistry confirmed that expression and activation of Notch1 signaling in the SVZ of aged brain were reduced. Double or triple immunostaining showed that that Notch1 was mainly expressed in doublecortin (DCX)‐positive cells, whereas Jagged1 was predominantly expressed in astroglial cells in the SVZ of normal aged rat brain. In addition, disruption or activation of Notch1 signaling altered the number of proliferating cells labeled by bromodeoxyuridine (BrdU) and DCX in the SVZ of aged brain. Moreover, ischemia‐induced cell proliferation in the SVZ of aged brain was enhanced by activating the Notch1 pathway and was suppressed by inhibiting the Notch1 signaling. Reduced infarct volume and improved motor deficits were also observed in Notch1 activator–treated aged ischemic rats. Our data suggest that Notch1 signaling modulates the SVZ neurogenesis in aged brain in normal and ischemic conditions.     

Identification and characterization of neuroblasts in the subventricular zone and rostral migratory stream of the adult human brain

It is of great interest to identify new neurons in the adult human brain, but the persistence of neurogenesis in the subventricular zone (SVZ) and the existence of the rostral migratory stream (RMS)-like pathway in the adult human forebrain remain highly controversial. In the present study, we have described the general configuration of the RMS in adult monkey, fetal human and adult human brains. We provide evidence that neuroblasts exist continuously in the anterior ventral SVZ and RMS of the adult human brain. The neuroblasts appear singly or in pairs without forming chains; they exhibit migratory morphologies and co-express the immature neuronal markers doublecortin, polysialylated neural cell adhesion molecule and βIII-tubulin. Few of these neuroblasts appear to be actively proliferating in the anterior ventral SVZ but none in the RMS, indicating that neuroblasts distributed along the RMS are most likely derived from the ventral SVZ. Interestingly, no neuroblasts are found in the adult human olfactory bulb. Taken together, our data suggest that the SVZ maintains the ability to produce neuroblasts in the adult human brain.     

A decrease in the addition of new cells in the nucleus accumbens and prefrontal cortex between puberty and adulthood in male rats

Adolescence involves shifts in social behaviors, behavioral flexibility, and adaptive risk‐taking that coincide with structural remodeling of the brain. We previously showed that new cells are added to brain regions associated with sexual behaviors, suggesting that cytogenesis may be a mechanism for acquiring adult‐typical behaviors during adolescence. Whether pubertal cell addition occurs in brain regions associated with behavioral flexibility or motivation and whether these patterns differ between pubertal and adult animals had not been determined. Therefore, we assessed patterns of cell proliferation or survival in the prefrontal cortex and nucleus accumbens. Pubertal and adult male rats were given injections of bromo‐deoxyuridine (BrdU). To assess cell proliferation, half of the animals from each group were sacrificed 24 h following the last injection. The remaining animals were sacrificed at Day 30 following the last injection to evaluate cell survival. Adult animals had significantly lower densities of BrdU‐immunoreactive (ir) cells in the prefrontal cortex, irrespective of post‐BrdU survival time, whereas in the nucleus accumbens, adult animals had a lower density of BrdU‐ir cells at the short survival time; however, the density of BrdU‐ir cells was equivalent in pubertal and adult animals at the longer survival time. These data provide evidence that cell addition during puberty may contribute to the remodeling of brain regions associated with behavioral flexibility and motivation, and this cell addition continues into adulthood, albeit at lower levels. Higher levels of cell proliferation or survival in younger animals may reflect a higher level of plasticity, possibly contributing to the dynamic remodeling of the pubertal brain. © 2013 Wiley Periodicals, Inc. Develop Neurobiol 74: 633–642, 2014     

Aging results in copper accumulations in glial fibrillary acidic protein‐positive cells in the subventricular zone

Analysis of rodent brains with X‐ray fluorescence (XRF) microscopy combined with immunohistochemistry allowed us to demonstrate that local Cu concentrations are thousands of times higher in the glia of the subventricular zone (SVZ) than in other cells. Using XRF microscopy with subcellular resolution and intracellular X‐ray absorption spectroscopy we determined the copper (I) oxidation state and the sulfur ligand environment. Cu K‐edge X‐ray absorption near edge spectroscopy is consistent with Cu being bound as a multimetallic Cu‐S cluster similar to one present in Cu‐metallothionein. Analysis of age‐related changes show that Cu content in astrocytes of the SVZ increases fourfold from 3 weeks to 9 months, while Cu concentration in other brain areas remain essentially constant. This increase in Cu correlates with a decrease in adult neurogenesis assessed using the Ki67 marker (both, however, can be age‐related effects). We demonstrate that the Cu distribution and age‐related concentration changes in the brain are highly cell specific.     

Species‐specific injury‐induced cell proliferation in the hippocampus and subventricular zone of food‐storing and nonstoring wild birds

Cells are continuously born and incorporated into the adult hippocampus (HP). Adult neurogenesis might act to increase the total number of cells or replace dead cells. Thus, neurogenesis might be a primary factor in augmenting, maintaining, or even recovering functions. In zebra finches, HP injury increases cell proliferation in the HP and stem cell rich subventricular zone (SVZ). It is unknown what effect injury has on a species dependent upon the HP for survival in the wild. In food‐storing birds, recovery of caches is seasonal, necessary for survival, dependent upon the HP and is concomitant with a peak in HP neurogenesis. During the fall, food‐storing black‐capped chickadees (BCCs) and nonstoring dark‐eyed juncos (DEJs) were captured and given a unilateral penetrating lesion to the HP one day later. On day 3, birds were injected with the mitotic marker 5‐bromo‐2′‐deoxyuridine (BrdU) and perfused on day 10. If unlesioned, more BrdU‐labeled cells were observed in the HP and SVZ of BCCs compared to DEJs, indicating higher innate cell proliferation or incorporation in BCCs. If lesioned, BrdU‐labeled cells increased in the injured HP of both species; however, lesions caused larger increases in DEJs. DEJs also showed increases in BrdU‐labeled cells in the SVZ and contralateral HP. BCCs showed no such increases on day 10. Thus, during the fall food‐storing season, storers showed suppressed injury‐induced cell proliferation and/or reduced survival rates of these new cells compared to nonstorers. These species differences may provide a useful model for isolating factors involved in cellular responses following injury. © 2009 Wiley Periodicals, Inc. Develop Neurobiol, 2010     

Neurogenesis and aging: FGF-2 and HB-EGF restore neurogenesis in hippocampus and subventricular zone of aged mice

Neurogenesis, which may contribute to the ability of the adult brain to function normally and adapt to disease, nevertheless declines with advancing age. Adult neurogenesis can be enhanced by administration of growth factors, but whether the aged brain remains responsive to these factors is unknown. We compared the effects of intracerebroventricular fibroblast growth factor (FGF)-2 and heparin-binding epidermal growth factor-like growth factor (HB-EGF) on neurogenesis in the hippocampal dentate subgranular zone (SGZ) and the subventricular zone (SVZ) of young adult (3-month) and aged (20-month) mice. Neurogenesis, measured by labelling with bromodeoxyuridine (BrdU) and by expression of doublecortin, was reduced by approximately 90% in SGZ and by approximately 50% in SVZ of aged mice. HB-EGF increased BrdU labelling in SGZ at 3 months by approximately 60% and at 20 months by approximately 450%, which increased the number of BrdU-labelled cells in SGZ of aged mice to approximately 25% of that in young adults. FGF-2 also stimulated BrdU labelling in SGZ, by approximately 25% at 3 months and by approximately 250% at 20 months, increasing the number of newborn neurones in older mice to approximately 20% of that in younger mice. In SVZ, HB-EGF and FGF-2 increased BrdU incorporation by approximately 140% at 3 months and approximately 170% at 20 months, so the number of BrdU-labelled cells was comparable in untreated 3-month-old and growth factor-treated 20-month-old mice. These results demonstrate that the aged brain retains the capacity to respond to exogenous growth factors with increased neurogenesis, which may have implications for the therapeutic potential of neurogenesis enhancement in age-associated neurological disorders.     

Ependymal/subependymal zone cells of postnatal and adult songbird brain generate both neurons and nonneuronal siblings in vitro and in vivo

The songbird forebrain continues to generate neurons in adulthood, from precursor cells located in the ependymal/subependymal zone (SZ) over the mediocaudal neostriatum. Precursor mitosis is followed by migration of neuronal daughter cells into the underlying forebrain, along radial fibers derived from the SZ. To define the ontogeny of both the new neurons and their radial guide cells, we employed retroviral insertion of the lacZ gene into neostriatal SZ precursor cells derived from postnatal and adult songbirds. We found that single SZ cells generate both neurons and substrate glia in vitro, and in an analogous fashion, both neurons and radial cells in vivo. This suggests that newly generated neurons and radial cells of the adult avian brain derive from a common pluripotential progenitor. © 1996 John Wiley & Sons, Inc.     

Hu protein as an early marker of neuronal phenotypic differentiation by subependymal zone cells of the adult songbird forebrain

The avian forebrain exhibits neurogenesis in adulthood, with neuronal production from ependymal/subependymal zone (SZ) precursor cells. To follow the commitment of newborn cells to neuronal lineage, we used their expression of the Hu family of neuronal RNA-binding proteins to identify them before their migration from the SZ. Adult canaries were injected with [³H]thymidine as a marker of DNA replication, sacrificed after varying intervals, stained for Hu, and autoradiographed. We found that Hu was not expressed by premitotic precursor cells, but rather appeared within hours in their neuronal progeny, which did not embark on parenchymal migration until 4 to 7 days later. Hu was expressed by all neurons, but not glia, both in vivo and in vitro, as determined by ultrastructural analysis as well as co-localization of Hu and cell-type selective antigens. In addition, co-staining for Hu and N-cadherin, whose expression is down-regulated on neuronal emigration from the SZ, revealed their initial co-expression by neuronal daughter cells still within the SZ. These results suggest that Hu expression may be used as a very early indicator of neuronal differentiation by SZ cells. Furthermore, the data indicate that in the adult avian brain, neuronal phenotype is established within hours of precursor mitosis, even though the neuronal daughter cells do not initiate parenchymal migration for at least 4 days thereafter, following their down-regulation of N-cadherin. © 1995 John Wiley & Sons, Inc.