Sara Gil-Perotin

PDGFRα-Positive B Cells Are Neural Stem Cells in the Adult SVZ that Form Glioma-like Growths in Response to Increased PDGF Signaling

Neurons and oligodendrocytes are produced in the adult brain subventricular zone (SVZ) from neural stem cells (B cells), which express GFAP and have morphological properties of astrocytes. We report here on the identification B cells expressing the PDGFRalpha in the adult SVZ. Specifically labeled PDGFRalpha expressing B cells in vivo generate neurons and oligodendrocytes. Conditional ablation of PDGFRalpha in a subpopulation of postnatal stem cells showed that this receptor is required for oligodendrogenesis, but not neurogenesis. Infusion of PDGF alone was sufficient to arrest neuroblast production and induce SVZ B cell proliferation contributing to the generation of large hyperplasias with some features of gliomas. The work demonstrates that PDGFRalpha signaling occurs early in the adult stem cell lineage and may help regulate the balance between oligodendrocyte and neuron production. Excessive PDGF activation in the SVZ in stem cells is sufficient to induce hallmarks associated with early stages of tumor formation.

Cellular composition and cytoarchitecture of the adult human subventricular zone: A niche of neural stem cells

The lateral wall of the lateral ventricle in the human brain contains neural stem cells throughout adult life. We conducted a cytoarchitectural and ultrastructural study in complete postmortem brains (n = 7) and in postmortem (n = 42) and intraoperative tissue (n = 43) samples of the lateral walls of the human lateral ventricles. With varying thickness and cell densities, four layers were observed throughout the lateral ventricular wall: a monolayer of ependymal cells (Layer I), a hypocellular gap (Layer II), a ribbon of cells (Layer III) composed of astrocytes, and a transitional zone (Layer IV) into the brain parenchyma. Unlike rodents and nonhuman primates, adult human glial fibrillary acidic protein (GFAP)+ subventricular zone (SVZ) astrocytes are separated from the ependyma by the hypocellular gap. Some astrocytes as well as a few GFAP‐cells in Layer II in the SVZ of the anterior horn and the body of the lateral ventricle appear to proliferate based on proliferating cell nuclear antigen (PCNA) and Ki67 staining. However, compared to rodents, the adult human SVZ appears to be devoid of chain migration or large numbers of newly formed young neurons. It was only in the anterior SVZ that we found examples of elongated Tuj1+ cells with migratory morphology. We provide ultrastructural criteria to identify the different cells types in the human SVZ including three distinct types of astrocytes and a group of displaced ependymal cells between Layers II and III. Ultrastructural analysis of this layer revealed a remarkable network of astrocytic and ependymal processes. This work provides a basic description of the organization of the adult human SVZ. J. Comp. Neurol. 494:415–434, 2006.

Loss of p53 Induces Changes in the Behavior of Subventricular Zone Cells: Implication for the Genesis of Glial Tumors

The role of multipotential progenitors and neural stem cells in the adult subventricular zone (SVZ) as cell-of-origin of glioblastoma has been suggested by studies on human tumors and transgenic mice. However, it is still unknown whether glial tumors are generated by all of the heterogeneous SVZ cell types or only by specific subpopulations of cells. It has been proposed that transformation could result from lack of apoptosis and increased self-renewal, but the definition of the properties leading to neoplastic transformation of SVZ cells are still elusive. This study addresses these questions in mice carrying the deletion of p53, a tumor-suppressor gene expressed in the SVZ. We show here that, although loss of p53 by itself is not sufficient for tumor formation, it provides a proliferative advantage to the slow- and fast-proliferating subventricular zone (SVZ) populations associated with their rapid differentiation. This results in areas of increased cell density that are distributed along the walls of the lateral ventricles and often associated with increased p53-independent apoptosis. Transformation occurs when loss of p53 is associated with a mutagenic stimulus and is characterized by dramatic changes in the properties of the quiescent adult SVZ cells, including enhanced self-renewal, recruitment to the fast-proliferating compartment, and impaired differentiation. Together, these findings provide a cellular mechanism for how the slow-proliferating SVZ cells can give rise to glial tumors in the adult brain.

Extensive migration of young neurons into the infant human frontal lobe

Building the human brain As the brain develops, neurons migrate from zones of proliferation to their final locations, where they begin to build circuits. Paredes et al. have discovered that shortly after birth, a group of neurons that proliferates near the ventricles migrates in chains alongside circulatory vessels into the frontal lobes (see the Perspective by McKenzie and Fishell). Young neurons that migrate postnatally into the anterior cingulate cortex then develop features of inhibitory interneurons. The number of migratory cells decreases over the first 7 months of life, and by 2 years of age, migratory cells are not evident. Any damage during migration, such as hypoxia, may affect the childs subsequent physical and behavioral development. Science, this issue p. 81; see also p. 38 Neurons are still finding their places as inhibitory circuits are established in the developing postnatal brain. [Also see Perspective by McKenzie] INTRODUCTION Inhibitory interneurons balance the excitation and inhibition of neural networks and therefore are key to normal brain function. In the developing brain, young interneurons migrate from their sites of birth into distant locations, where they functionally integrate. Although this neuronal migration is largely complete before birth, some young inhibitory interneurons continue to travel and add to circuits in restricted regions of the juvenile and adult mammalian brain. For example, postnatally migrating inhibitory neurons travel from the walls of the lateral ventricle, along the rostral migratory stream (RMS) into the olfactory bulb. In humans, an additional ventral route branching off the RMS, the medial migratory stream (MMS), takes young neurons into the medial prefrontal cortex. It has been suggested that recruitment of neurons during postnatal life could help shape neural circuits according to experience. Specifically, inhibitory interneuron maturation during postnatal development is associated with critical periods of brain plasticity. We asked whether neuronal recruitment continues into early childhood in the frontal lobe, a region of the human brain that has greatly increased in size and complexity during evolution. RATIONALE Migrating young neurons persist for several months after birth in an extensive region of the subventricular zone (SVZ) around the anterior lateral ventricles in the human brain. Are all these young neurons migrating into the RMS and MMS, or do they have other destinations? Using high-resolution magnetic resonance imaging (MRI), histology, and time-lapse confocal microscopy, we observed the migration of many young inhibitory interneurons around the dorsal anterior walls of the lateral ventricle and into multiple cortical regions of the human frontal cortex. We determined the location and orientation of these young neurons, demonstrated their active translocation, and inferred their fates in the postnatal anterior forebrain. RESULTS A large collection of cells expressing doublecortin (DCX), a marker of young migrating neurons, traveled and integrated within the infant frontal lobe. This migratory stream, which was most prominent during the first 2 months after birth and persisted until at least 5 months, formed a caplike structure surrounding the anterior body of the lateral ventricle. We refer to this population of young neurons as the Arc. This structure could also be visualized by brain MRI. Young neurons in the Arc appeared to move long distances in distinct regions around the ventricular wall and the developing white matter. The orientation of elongated DCX+ cells suggested that migratory neurons closer to the ventricular wall dispersed tangentially. In contrast, migratory neurons within the developing white matter tended to be orientated toward the overlying cortex. These cells expressed markers of interneurons, and their entry into the anterior cingulate cortex (a major target of the Arc used for quantification) was correlated with the emergence of specific subtypes of γ-aminobutyric acid (GABA)–expressing interneurons (neuropeptide Y, somatostatin, calretinin, and calbindin). Expression of transcription factors associated with specific sites of origin suggested that these neurons arise from ventral telencephalon progenitor domains. CONCLUSION Widespread neuronal migration into the human frontal lobe continues for several months after birth. Young neurons express markers of cortical inhibitory interneurons and originate outside the cortex, likely in the ventral forebrain. The postnatal recruitment of large populations of inhibitory neurons may contribute to maturation and plasticity in the human frontal cortex. Defects in the migration of these neurons could result in circuit dysfunction associated with neurodevelopmental disorders. Widespread neuronal migration into the human frontal lobe continues during postnatal life. (A) Sagittal schematic of the newborn forebrain shows prominent collections of young migratory neurons (illustrated in green) adjacent to the lateral ventricle (LV) and in the overlying white matter. Directional axes: D, dorsal; A, anterior. (B and C) DCX+ cells coexpress GABA and GAD67, markers of inhibitory interneurons (marked by arrows). The first few months after birth, when a child begins to interact with the environment, are critical to human brain development. The human frontal lobe is important for social behavior and executive function; it has increased in size and complexity relative to other species, but the processes that have contributed to this expansion are unknown. Our studies of postmortem infant human brains revealed a collection of neurons that migrate and integrate widely into the frontal lobe during infancy. Chains of young neurons move tangentially close to the walls of the lateral ventricles and along blood vessels. These cells then individually disperse long distances to reach cortical tissue, where they differentiate and contribute to inhibitory circuits. Late-arriving interneurons could contribute to developmental plasticity, and the disruption of their postnatal migration or differentiation may underlie neurodevelopmental disorders.

Ultrastructure of the subventricular zone in Macaca fascicularis and evidence of a mouse‐like migratory stream

Recent publications have shown that the lateral wall of the lateral ventricles in the Macaca fascicularis brain, in particular the subventricular zone (SVZ), contains neural stem cells throughout adulthood that migrate through a migratory pathway (RMS) to the olfactory bulb (OB). To date, a detailed and systematic cytoarchitectural and ultrastructural study of the monkey SVZ and RMS has not been done. We found that the organization of the SVZ was similar to that of humans, with the ependymal layer surrounding the lateral ventricles, a hypocellular GAP layer formed by astrocytic and ependymal expansions, and the astrocyte ribbon, composed of astrocytic bodies. We found no cells corresponding to the type C proliferating precursor of the rodent brain. Instead, proliferating cells, expressed as Ki‐67 immunoreactivity, were predominantly young neurons concentrated in the anterior regions, and occasional astrocytes of the ribbon. We observed displaced ependymal cells of still unknown significance. New neurons tended to organize in chain‐like structures, which were surrounded by astrocytes. This pattern was highly reminiscent of that observed in rodent RMS, but not in humans. These chains spread from the frontal SVZ along a GAP‐like layer, uniquely composed of astrocytic expansions, to the olfactory bulb (OB). The number of neuronal chains and the number of chain‐forming cells decreased gradually upon reaching the OB. The purpose of this work is to provide a reference for future studies in the field of adult neurogenesis that may lead to an understanding of the fate and functionality of newborn neurons in primates, and ultimately in humans. J. Comp. Neurol. 514:533–554, 2009.

Defective Postnatal Neurogenesis and Disorganization of the Rostral Migratory Stream in Absence of the Vax1 Homeobox Gene

The subventricular zone (SVZ) is one of the sources of adult neural stem cells (ANSCs) in the mouse brain. Precursor cells proliferate in the SVZ and migrate through the rostral migratory stream (RMS) to the olfactory bulb (OB), where they differentiate into granule and periglomerular cells. Few transcription factors are known to be responsible for regulating NSC proliferation, migration, and differentiation processes; even fewer have been found to be responsible for the organization of the SVZ and RMS. For this reason, we studied the ventral anterior homeobox (Vax1) gene in NSC proliferation and in SVZ organization. We found that Vax1 is strongly expressed in the SVZ and in the RMS and that, in the absence of Vax1, embryonic precursor cells proliferate 100 times more than wild-type controls, in vitro. The SVZ of Vax1-/- brains is hyperplastic and mostly disorganized, and the RMS is missing, causing a failure of precursor cell migration to the OBs, which as a result are severely hypoplastic. Moreover, we found that Vax1 is essential for the correct differentiation of ependyma and astrocytes. Together, these data indicate that Vax1 is a potent regulator of SVZ organization and NSC proliferation, with important consequences on postnatal neurogenesis.

The LIM Homeodomain Factor Lhx2 Is Required for Hypothalamic Tanycyte Specification and Differentiation

Hypothalamic tanycytes, a radial glial-like ependymal cell population that expresses numerous genes selectively enriched in embryonic hypothalamic progenitors and adult neural stem cells, have recently been observed to serve as a source of adult-born neurons in the mammalian brain. The genetic mechanisms that regulate the specification and maintenance of tanycyte identity are unknown, but are critical for understanding how these cells can act as adult neural progenitor cells. We observe that LIM (Lin-11, Isl-1, Mec-3)-homeodomain gene Lhx2 is selectively expressed in hypothalamic progenitor cells and tanycytes. To test the function of Lhx2 in tanycyte development, we used an intersectional genetic strategy to conditionally delete Lhx2 in posteroventral hypothalamic neuroepithelium, both embryonically and postnatally. We observed that tanycyte development was severely disrupted when Lhx2 function was ablated during embryonic development. Lhx2-deficient tanycytes lost expression of tanycyte-specific genes, such as Rax, while also displaying ectopic expression of genes specific to cuboid ependymal cells, such as Rarres2. Ultrastructural analysis revealed that mutant tanycytes exhibited a hybrid identity, retaining radial morphology while becoming multiciliated. In contrast, postnatal loss of function of Lhx2 resulted only in loss of expression of tanycyte-specific genes. Using chromatin immunoprecipitation, we further showed that Lhx2 directly regulated expression of Rax, an essential homeodomain factor for tanycyte development. This study identifies Lhx2 as a key intrinsic regulator of tanycyte differentiation, sustaining Rax-dependent activation of tanycyte-specific genes while also inhibiting expression of ependymal cell-specific genes. These findings provide key insights into the transcriptional regulatory network specifying this still poorly characterized cell type.

Exposure to N-Ethyl-N-Nitrosourea in Adult Mice Alters Structural and Functional Integrity of Neurogenic Sites

Background Previous studies have shown that prenatal exposure to the mutagen N-ethyl-N-nitrosourea (ENU), a N-nitroso compound (NOC) found in the environment, disrupts developmental neurogenesis and alters memory formation. Previously, we showed that postnatal ENU treatment induced lasting deficits in proliferation of neural progenitors in the subventricular zone (SVZ), the main neurogenic region in the adult mouse brain. The present study is aimed to examine, in mice exposed to ENU, both the structural features of adult neurogenic sites, incorporating the dentate gyrus (DG), and the behavioral performance in tasks sensitive to manipulations of adult neurogenesis. Methodology/Principal Findings 2-month old mice received 5 doses of ENU and were sacrificed 45 days after treatment. Then, an ultrastructural analysis of the SVZ and DG was performed to determine cellular composition in these regions, confirming a significant alteration. After bromodeoxyuridine injections, an S-phase exogenous marker, the immunohistochemical analysis revealed a deficit in proliferation and a decreased recruitment of newly generated cells in neurogenic areas of ENU-treated animals. Behavioral effects were also detected after ENU-exposure, observing impairment in odor discrimination task (habituation-dishabituation test) and a deficit in spatial memory (Barnes maze performance), two functions primarily related to the SVZ and the DG regions, respectively. Conclusions/Significance The results demonstrate that postnatal exposure to ENU produces severe disruption of adult neurogenesis in the SVZ and DG, as well as strong behavioral impairments. These findings highlight the potential risk of environmental NOC-exposure for the development of neural and behavioral deficits.

Roles of p53 and p27 Kip1 in the regulation of neurogenesis in the murine adult subventricular zone

The tumor suppressor protein p53 (Trp53) and the cell cycle inhibitor p27 Kip1 (Cdknb1) have both been implicated in regulating proliferation of adult subventricular zone (aSVZ) cells. We previously reported that genetic ablation of Trp53 (Trp53 −/−) or Cdknb1 (p27 Kip1−/−) increased proliferation of cells in the aSVZ, but differentially affected the number of adult born neuroblasts. We therefore hypothesized that these molecules might play non‐redundant roles. To test this hypothesis we generated mice lacking both genes (Trp53 −/−;p27 Kip1−/−) and analysed the consequences on aSVZ cells and adult neuroblasts. Proliferation and self‐renewal of cultured aSVZ cells were increased in the double mutants compared with control, but the mice did not develop spontaneous brain tumors. In contrast, the number of adult‐born neuroblasts in the double mutants was similar to wild‐type animals and suggested a complementation of the p27 Kip1−/− phenotype due to loss of Trp53. Cellular differences detected in the aSVZ correlated with cellular changes in the olfactory bulb and behavioral data on novel odor recognition. The exploration time for new odors was reduced in p27 Kip1−/− mice, increased in Trp53 −/−mice and normalized in the double Trp53−/−;p27 Kip1−/− mutants. At the molecular level, Trp53 −/−aSVZ cells were characterized by higher levels of NeuroD and Math3 and by the ability to generate neurons more readily. In contrast, p27 Kip1−/− cells generated fewer neurons, due to enhanced proteasomal degradation of pro‐neural transcription factors. Together, these results suggest that p27 Kip1 and p53 function non‐redundantly to modulate proliferation and self‐renewal of aSVZ cells and antagonistically in regulating adult neurogenesis.

Identification and characterization of neural progenitor cells in the adult mammalian brain.

Adult neurogenesis has been questioned for many years. In the early 1900s, a dogma was established that denied new neuron formation in the adult brain. In the last century, however, new discoveries have demonstrated the real existence of proliferation in the adult brain, and in the last decade, these studies led to the identification of neural stem cells in mammals. Adult neural stem cells are undifferentiated cells that are present in the adult brain and are capable of dividing and differentiating into glia and new neurons. Newly formed neurons terminally differentiate into mature neurons in the olfactory bulb and the dentate gyrus of the hippocampus. Since then, a number of new research lines have emerged whose common objective is the phenotypical and molecular characterization of brain stem cells. As a result, new therapies are successfully being applied to animal models for certain neurodegenerative diseases or stroke. This work is being or will be extended to the adult human brain, and so it provides purpose and hope to all previous studies in this field. We are still far from clinical therapies because the mechanisms and functions of these cells are not completely understood, but we appear to be moving in the right direction.