Neural Stem Cells In Subventricular Zone Research Articles

Identification of distinct ChAT⁺ neurons and activity-dependent control of postnatal SVZ neurogenesis.

Postnatal/adult SVZ neurogenesis is believed to be primarily controlled by neural stem cell (NSC)-intrinsic mechanisms, interacting with extracellular/niche-driven cues. Although behavioral paradigms and disease states have suggested possibilities for higher-level inputs, it is currently unknown if neural activity patterns from discrete circuits can directly regulate SVZ neurogenesis. We have identified a previously undescribed population of ChAT+ neurons residing within the rodent SVZ neurogenic niche. These neurons showed morphological and functional differences from neighboring striatal counterparts, and released acetylcholine locally in activity-dependent fashion. Optogenetic inhibition and stimulation of subependymal ChAT+ neurons in vivo showed that they are necessary and sufficient to control neurogenic proliferation. Furthermore, whole-cell recordings and biochemical experiments revealed direct SVZ NSC responses to local acetylcholine release, synergizing with FGF receptor activation to increase neuroblast production. These results uncovered an unknown gateway connecting SVZ neurogenesis to neuronal activity-dependent control, and possibilities for modulating neuroregenerative capacities in health and disease.

Distinct and separable roles for EZH2 in neurogenic astroglia

The epigenetic mechanisms that enable specialized astrocytes to retain neurogenic competence throughout adult life are still poorly understood. Here we show that astrocytes that serve as neural stem cells (NSCs) in the adult mouse subventricular zone (SVZ) express the histone methyltransferase EZH2. This Polycomb repressive factor is required for neurogenesis independent of its role in SVZ NSC proliferation, as Ink4a/Arf-deficiency in Ezh2-deleted SVZ NSCs rescues cell proliferation, but neurogenesis remains defective. Olig2 is a direct target of EZH2, and repression of this bHLH transcription factor is critical for neuronal differentiation. Furthermore, Ezh2 prevents the inappropriate activation of genes associated with non-SVZ neuronal subtypes. In the human brain, SVZ cells including local astroglia also express EZH2, correlating with postnatal neurogenesis. Thus, EZH2 is an epigenetic regulator that distinguishes neurogenic SVZ astrocytes, orchestrating distinct and separable aspects of adult stem cell biology, which has important implications for regenerative medicine and oncogenesis.DOI: http://dx.doi.org/10.7554/eLife.02439.001.

GSK3β regulates oligodendrogenesis in the dorsal microdomain of the subventricular zone via Wnt-β-catenin signaling

Oligodendrocytes, the myelinating cells of the CNS, are derived postnatally from oligodendrocyte precursors (OPs) of the subventricular zone (SVZ). However, the mechanisms that regulate their generation from SVZ neural stem cells (NSC) are poorly understood. Here, we have examined the role of glycogen synthase kinase 3β (GSK3β), an effector of multiple converging signaling pathways in postnatal mice. The expression of GSK3β by rt-qPCR was most prominent in the SVZ and in the developing white matter, around the first 1–2 weeks of postnatal life, coinciding with the peak periods of OP differentiation. Intraventricular infusion of the GSK3β inhibitor ARA-014418 in mice aged postnatal day (P) 8–11 significantly increased generation of OPs in the dorsal microdomain of the SVZ, as shown by expression of cell specific markers using rt-qPCR and immunolabelling. Analysis of stage specific markers revealed that the augmentation of OPs occurred via increased specification from earlier SVZ cell types. These effects of GSK3β inhibition on the dorsal SVZ were largely attributable to stimulation of the canonical Wnt/β-catenin signaling pathway over other pathways. The results indicate GSK3β is a key endogenous factor for specifically regulating oligodendrogenesis from the dorsal SVZ microdomain under the control of Wnt-signaling.

Adult subventricular zone neural stem cells as a potential source of dopaminergic replacement neurons

Clinical trials engrafting human fetal ventral mesencephalic tissue have demonstrated, in principle, that cell replacement therapy provides substantial long-lasting improvement of motor impairments generated by Parkinson's Disease (PD). The use of fetal tissue is not practical for widespread clinical implementation of this therapy, but stem cells are a promising alternative source for obtaining replacement cells. The ideal stem cell source has yet to be established and, in this review, we discuss the potential of neural stem cells in the adult subventricular zone (SVZ) as an autologous source of replacement cells. We identify three key challenges for further developing this potential source of replacement cells: (1) improving survival of transplanted cells, (2) suppressing glial progenitor proliferation and survival, and (3) developing methods to efficiently produce dopaminergic neurons. Subventricular neural stem cells naturally produce a dopaminergic interneuron phenotype that has an apparent lack of vulnerability to PD-mediated degeneration. We also discuss whether olfactory bulb dopaminergic neurons derived from adult SVZ neural stem cells are a suitable source for cell replacement strategies.

Titration of GLI3 Repressor Activity by Sonic Hedgehog Signaling Is Critical for Maintaining Multiple Adult Neural Stem Cell and Astrocyte Functions

Sonic hedgehog (SHH), a key regulator of embryonic neurogenesis, signals directly to neural stem cells (NSCs) in the subventricular zone (SVZ) and to astrocytes in the adult mouse forebrain. The specific mechanism by which the GLI2 and GLI3 transcriptional activators (GLI2(A) and GLI3(A)) and repressors (GLI2(R) and GLI3(R)) carry out SHH signaling has not been addressed. We found that the majority of slow-cycling NSCs express Gli2 and Gli3, whereas Gli1 is restricted ventrally and all three genes are downregulated when NSCs transition into proliferating progenitors. Surprisingly, whereas conditional ablation of Smo in postnatal glial fibrillary acidic protein-expressing cells results in cell-autonomous loss of NSCs and a progressive reduction in SVZ proliferation, without an increase in glial cell production, removal of Gli2 or Gli3 does not alter adult SVZ neurogenesis. Significantly, removing Gli3 in Smo conditional mutants largely rescues neurogenesis and, conversely, expression of a constitutive GLI3(R) in the absence of normal Gli2 and Gli3 abrogates neurogenesis. Thus unattenuated GLI3(R) is a primary inhibitor of adult SVZ NSC function. Ablation of Gli2 and Gli3 revealed a minor role for GLI2(R) and little requirement for GLI(A) function in stimulating SVZ neurogenesis. Moreover, we found that similar rules of GLI activity apply to SHH signaling in regulating SVZ-derived olfactory bulb interneurons and maintaining cortical astrocyte function. Namely, fewer superficial olfactory bulb interneurons are generated in the absence of Gli2 and Gli3, whereas astrocyte partial gliosis results from an increase in GLI3(R). Thus precise titration of GLI(R) levels by SHH is critical to multiple functions of adult NSCs and astrocytes.

CHD7 and retinoic acid signaling cooperate to regulate neural stem cell and inner ear development in mouse models of CHARGE syndrome

CHARGE syndrome is a multiple congenital anomaly disorder that leads to life-threatening birth defects, such as choanal atresia and cardiac malformations as well as multiple sensory impairments, that affect hearing, vision, olfaction and balance. CHARGE is caused by heterozygous mutations in CHD7, which encodes an ATP-dependent chromatin remodeling enzyme. Identification of the mechanisms underlying neurological and sensory defects in CHARGE is a first step toward developing treatments for CHARGE individuals. Here, we used mouse models of Chd7 deficiency to explore the function of CHD7 in the development of the subventricular zone (SVZ) neural stem cell niche and inner ear, structures that are important for olfactory bulb neurogenesis and hearing and balance, respectively. We found that loss of Chd7 results in cell-autonomous proliferative, neurogenic and self-renewal defects in the perinatal and mature mouse SVZ stem cell niche. Modulation of retinoic acid (RA) signaling prevented in vivo inner ear and in vitro neural stem cell defects caused by Chd7 deficiency. Our findings demonstrate critical, cooperative roles for RA and CHD7 in SVZ neural stem cell function and inner ear development, suggesting that altered RA signaling may be an effective method for treating Chd7 deficiency.

Multimodal imaging of subventricular zone neural stem/progenitor cells in the cuprizone mouse model reveals increased neurogenic potential for the olfactory bulb pathway, but no contribution to remyelination of the corpus callosum

Multiple sclerosis is a devastating demyelinating disease of the central nervous system (CNS) in which endogenous remyelination, and thus recovery, often fails. Although the cuprizone mouse model allowed elucidation of many molecular factors governing remyelination, currently very little is known about the spatial origin of the oligodendrocyte progenitor cells that initiate remyelination in this model. Therefore, we here investigated in this model whether subventricular zone (SVZ) neural stem/progenitor cells (NSPCs) contribute to remyelination of the splenium following cuprizone-induced demyelination. Experimentally, from the day of in situ NSPC labeling, C57BL/6J mice were fed a 0.2% cuprizone diet during a 4-week period and then left to recover on a normal diet for 8weeks. Two in situ labeling strategies were employed: (i) NSPCs were labeled by intraventricular injection of micron-sized iron oxide particles and then followed up longitudinally by means of magnetic resonance imaging (MRI), and (ii) SVZ NSPCs were transduced with a lentiviral vector encoding the eGFP and Luciferase reporter proteins for longitudinal monitoring by means of in vivo bioluminescence imaging (BLI). In contrast to preceding suggestions, no migration of SVZ NSPC towards the demyelinated splenium was observed using both MRI and BLI, and further validated by histological analysis, thereby demonstrating that SVZ NSPCs are unable to contribute directly to remyelination of the splenium in the cuprizone model. Interestingly, using longitudinal BLI analysis and confirmed by histological analysis, an increased migration of SVZ NSPC-derived neuroblasts towards the olfactory bulb was observed following cuprizone treatment, indicative for a potential link between CNS inflammation and increased neurogenesis.

Egr-1 is a Critical Regulator of EGF-Receptor-Mediated Expansion of Subventricular Zone Neural Stem Cells and Progenitors During Recovery from Hypoxia–Hypoglycemia

We recently established that the EGF-R (epidermal growth factor receptor) (EGF-R) is an essential regulator of the reactive expansion of SVZ (subventricular zone) NPs (neural precursors) that occurs during recovery from hypoxic-ischemic brain injury. The purpose of the current studies was to identify the conditions and the transcription factor (s) responsible for inducing the EGF-R. Here, we show that the increase in EGF-R expression and the more rapid division of the NPs can be recapitulated in in vitro by exposing SVZ NPs to hypoxia and hypoglycemia simultaneously, but not separately. The EGF-R promoter has binding sites for multiple transcription factors that includes the zinc finger transcription factor, Egr-1. We show that Egr-1 expression increases in NPs, but not astrocytes, following hypoxia and hypoglycemia where it accumulates in the nucleus. To determine whether Egr-1 is necessary for EGF-R expression, we used SiRNAs (small interfering RNA) specific for Egr-1 to decrease Egr-1 expression. Knocking-down Egr-1 decreased basal levels of EGF-R and it abolished the stress-induced increase in EGF-R expression. By contrast, HIF-1 accumulation did not contribute to EGF-R expression and FGF-2 only modestly induced EGF-R. These studies establish a new role for Egr-1 in regulating the expression of the mitogenic EGF-R. They also provide new information into mechanisms that promote NP expansion and provide insights into strategies for amplifying the numbers of stem cells for CNS (central nervous system) regeneration.

Integration of Genome-wide Approaches Identifies lncRNAs of Adult Neural Stem Cells and Their Progeny In Vivo

Long noncoding RNAs (lncRNAs) have been described in cell lines and various whole tissues, but lncRNA analysis of development invivo is limited. Here, we comprehensively analyze lncRNA expression for the adult mouse subventricular zone neuralstem cell lineage. We utilize complementary genome-wide techniques including RNA-seq, RNA CaptureSeq, and ChIP-seq to associate specific lncRNAs with neural cell types, developmental processes, and human disease states. By integrating data from chromatin state maps, custom microarrays, and FACS purification of the subventricular zone lineage, we stringently identify lncRNAs with potential roles in adult neurogenesis. shRNA-mediated knockdown of two such lncRNAs, Six3os and Dlx1as, indicate roles for lncRNAs in the glial-neuronal lineage specification of multipotent adult stem cells. Our data and workflow thus provide a uniquely coherent invivo lncRNA analysis and form the foundation of a user-friendly online resource for the study of lncRNAs in development and disease.

Neonatal Subventricular Zone Electroporation

Neural stem cells (NSCs) line the postnatal lateral ventricles and give rise to multiple cell types which include neurons, astrocytes, and ependymal cells. Understanding the molecular pathways responsible for NSC self-renewal, commitment, and differentiation is critical for harnessing their unique potential to repair the brain and better understand central nervous system disorders. Previous methods for the manipulation of mammalian systems required the time consuming and expensive endeavor of genetic engineering at the whole animal level. Thus, the vast majority of studies have explored the functions of NSC molecules in vitro or in invertebrates. Here, we demonstrate the simple and rapid technique to manipulate neonatal NPCs that is referred to as neonatal subventricular zone (SVZ) electroporation. Similar techniques were developed a decade ago to study embryonic NSCs and have aided studies on cortical development. More recently this was applied to study the postnatal rodent forebrain. This technique results in robust labeling of SVZ NSCs and their progeny. Thus, postnatal SVZ electroporation provides a cost and time effective alternative for mammalian NSC genetic engineering.

Neonatal Subventricular Zone Electroporation

Neural stem cells (NSCs) line the postnatal lateral ventricles and give rise to multiple cell types which include neurons, astrocytes, and ependymal cells1. Understanding the molecular pathways responsible for NSC self-renewal, commitment, and differentiation is critical for harnessing their unique potential to repair the brain and better understand central nervous system disorders. Previous methods for the manipulation of mammalian systems required the time consuming and expensive endeavor of genetic engineering at the whole animal level2. Thus, the vast majority of studies have explored the functions of NSC molecules in vitro or in invertebrates. Here, we demonstrate the simple and rapid technique to manipulate neonatal NPCs that is referred to as neonatal subventricular zone (SVZ) electroporation. Similar techniques were developed a decade ago to study embryonic NSCs and have aided studies on cortical development3,4 . More recently this was applied to study the postnatal rodent forebrain5-7. This technique results in robust labeling of SVZ NSCs and their progeny. Thus, postnatal SVZ electroporation provides a cost and time effective alternative for mammalian NSC genetic engineering.

Chromatin-based epigenetics of adult subventricular zone neural stem cells.

In specific regions of the adult mammalian brain, neural stem cells (NSCs) generate new neurons throughout life. Emerging evidence indicate that chromatin-based transcriptional regulation is a key epigenetic mechanism for the life-long function of adult NSCs. In the adult mouse brain, NSCs in the subventricular zone (SVZ) retain the ability to produce both neurons and glia for the life of the animal. In this review, we discuss the origin and function of SVZ NSCs as they relate to key epigenetic concepts of development and potential underlying mechanism of chromatin-based transcriptional regulation. A central point of discussion is how SVZ NSCs – which possess many characteristics of mature, non-neurogenic astrocytes – maintain a “youthful” ability to produce both neuronal and glial lineages. In addition to reviewing data regarding the function of chromatin-modifying factors in SVZ neurogenesis, we incorporate our growing understanding that long non-coding RNAs serve as an important element to chromatin-based transcriptional regulation, including that of SVZ NSCs. Discoveries regarding the epigenetic mechanisms of adult SVZ NSCs may provide key insights into fundamental principles of adult stem cell biology as well as the more complex and dynamic developmental environment of the embryonic brain.

Polymeric Nanoparticles to Control the Differentiation of Neural Stem Cells in the Subventricular Zone of the Brain

Herein, we report the use of retinoic acid-loaded polymeric nanoparticles as a potent tool to induce the neuronal differentiation of subventricular zone neural stem cells. The intracellular delivery of retinoic acid by the nanoparticles activated nuclear retinoic acid receptors, decreased stemness, and increased proneurogenic gene expression. Importantly, this work reports for the first time a nanoparticle formulation able to modulate in vivo the subventricular zone neurogenic niche. The work further compares the dynamics of initial stages of differentiation between SVZ cells treated with retinoic acid-loaded polymeric nanoparticles and solubilized retinoic acid. The nanoparticle formulation developed here may ultimately offer new perspectives to treat neurodegenerative diseases.

Expression of ezrin in subventricular zone neural stem cells and their progeny in adult and developing mice

Ezrin is a member of the ezrin-radixin-moesin (ERM) family of proteins, which link the cytoskeleton and cell membrane. ERM proteins are involved in pivotal cellular functions including cell-matrix recognition, cell-cell communication, and cell motility. Several recent studies have shown that ERM proteins are expressed in specific cell types of the adult rostral migratory stream (RMS). In this study, we found that ERM proteins are expressed highly in the early postnatal RMS and the ventricular zone of embryonic cerebral cortex, suggesting that these proteins may be expressed by neural progenitors. Furthermore, whereas ezrin previously was found to be expressed exclusively by astrocytes of the adult RMS, we found that ezrin-expressing cells also expressed the markers for indicating neuroblasts in vivo and in vitro, and that ezrin expression by neuroblasts decreases progressively as neuroblasts migrate. Using in vitro differentiation of adult neural stem cells, we found that ezrin is expressed by neural stem cells and their progeny (neuroblasts and astrocytes), but not by oligodendrocytic progeny. Collectively our findings demonstrate that adult neural stem cells and neuroblasts express ezrin and that ezrin may be involved in intracellular actin remodeling.

Multifaces of neuropeptide Y in the brain – Neuroprotection, neurogenesis and neuroinflammation

Neuropeptide Y (NPY) has been implicated in the modulation of important features of neuronal physiology, including calcium homeostasis, neurotransmitter release and excitability. Moreover, NPY has been involved as an important modulator of hippocampal and thalamic circuits, receiving particular attention as an endogenous antiepileptic peptide and as a potential master regulator of feeding behavior. NPY not only inhibits excessive glutamate release (decreasing circuitry hyperexcitability) but also protects neurons from excitotoxic cell death. Furthermore, NPY has been involved in the modulation of the dynamics of dentate gyrus and subventricular zone neural stem cell niches. In both regions, NPY is part of the chemical resource of the neurogenic niche and acts through NPY Y1 receptors to promote neuronal differentiation. Interestingly, NPY is also considered a neuroimmune messenger. In this review, we highlight recent evidences concerning paracrine/autocrine actions of NPY involved in neuroprotection, neurogenesis and neuroinflammation. In summary, the three faces of NPY, discussed in the present review, may contribute to better understand the dynamics and cell fate decision in the brain parenchyma and in restricted areas of neurogenic niches, in health and disease.

Spatiotemporal Changes to the Subventricular Zone Stem Cell Pool through Aging

Through adulthood, the rodent subventricular zone (SVZ) stem cell niche generates new olfactory bulb interneurons. We had previously reported that the number of new neurons produced in the SVZ declines through aging; however, age-related changes attributable specifically to the SVZ neural stem cell (NSC) population have not been fully characterized. Here, we conducted a spatiotemporal evaluation of adult SVZ NSCs. We assessed ventricle-contacting NSCs, which together with ependymal cells form regenerative units (pinwheels) along the lateral wall of the lateral ventricle. Based on their apical GFAP-expressing process, individual NSCs were identified across the ventricle surface using serial reconstruction of the SVZ. We observed an 86% decline in total NSCs/mm² of intact ependyma in 2-year old versus 3-month-old mice, with fewer NSC processes within each aged pinwheel. This resulted in an associated 78% decline in total pinwheel units/mm². Regional analysis along the lateral ventricle surface revealed that the age-dependent decline of NSCs and pinwheels is spatially uniform and ultimately maintains the conserved ratio of olfactory bulb interneuron subtypes generated in young mice. However, the overall neurogenic output of the aged SVZ is reduced. Surprisingly, we found no significant change in the number of actively proliferating NSCs per mm² of ventricle surface. Instead, our data reveal that, although the total NSC number, pinwheel units and NSCs per pinwheel decline with age, the percentage of actively, mitotic NSCs increases, indicating that age-related declines in SVZ-mediated olfactory bulb neurogenesis occur downstream of NSC proliferation.

Thyroid Hormone Signaling Acts as a Neurogenic Switch by Repressing Sox2 in the Adult Neural Stem Cell Niche

The subventricular zone (SVZ) neural stem cell niche contains mixed populations of stem cells, transit-amplifying cells, and migrating neuroblasts. Deciphering how endogenous signals, such as hormones, affect the balance between these cell types is essential for understanding the physiology of niche plasticity and homeostasis. We show that Thyroid Hormone (T(3)) and its receptor, TRα1, are directly involved in maintaining this balance. TRα1 is expressed in amplifying and migrating cells. In vivo gain- and loss-of-function experiments demonstrate first, that T(3)/TRα1 directly repress Sox2 expression, and second, that TRα1 overexpression in the niche favors the appearance of DCX+ migrating neuroblasts. Lack of TRα increases numbers of SOX2+ cells in the SVZ. Hypothyroidism increases proportions of cells in interphase. Thus, in the adult SVZ, T(3)/TRα1 together favor neural stem cell commitment and progression toward a migrating neuroblast phenotype; this transition correlates with T(3)/TRα1-dependent transcriptional repression of Sox2.

PARP‐1 deletion promotes subventricular zone neural stem cells toward a glial fate

Identification of critical factors involved in oligodendroglial fate specification from endogenous neural stem cells is relevant to the development of therapeutic interventions that aim to promote remyelination. Here we report a novel role of the DNA repair protein poly-ADP-ribose polymerase-1 (PARP-1) in regulating the neural stem cell profile in the postnatal mouse forebrain subventricular zone (SVZ). We observed increased expression of Sox2 and Sox10 in the SVZ of postnatal day 11 (P11) PARP-1 knockout mice. This increase corresponded to increased Olig2 expression in Sox2-positive cells of the PARP-1 knockout mouse SVZ and decreased Map2abc expression compared with Sox2/Olig2 and Sox2/Map2abc expression in wild-type mice. We noted enhanced expression of proliferating oligodendrocyte progenitor cells (OPCs) at the expense of proliferating neuroblasts in the SVZ of PARP-1 knockout mice, by using Olig1/Ki67/DCX, NG2/Ki67/DCX, and PDGFR/BrdU/TuJ1 immunofluorescence labeling. In addition, the percentage of BrdU/Olig2 double-labeled cells increased in the SVZ and corpus callosum of PARP-1 knockout mice compared with wild-type mice. We also observed a decrease in DCX-positive cells without a decrease in the overall SVZ area in PARP-1 knockout mice, further indicating a switch from neuroblast to OPC fate. PARP-1 knockout mice displayed thinning of MBP expression in the corpus callosum and external capsule, suggesting that the enhanced OPC proliferation in the SVZ might compensate for deficiency in myelination. Together, our results show that PARP-1 deletion promotes SVZ neural stem cells toward a glial fate and suggest that future studies target PARP-1 as a potential therapeutic strategy for demyelinating diseases.

Olig1 and Olig2 promotes oligodendrocyte differentiation of neural stem cells in adult mice injured by EAE

Investigating neural stem cell plasticity in the hippo-campal niche, we demonstrate that retroviral forced expression of Mash1 (Mammalian Achaete-Scute Homolog 1), Olig1(Oligodendrocyte transcription factor 1), and Olig2 (Oligodendrocyte transcription factor 2) genes, transcription factors involved in enhanced oligodendrogenesis, can contribute to directing the differentiation of adult subventricular zone neural stem cells to functional oligodendrocytes. We found that Mash1, Olig1 and Olig2 all induced oligodendrocyte differentiation. However, Olig1 and Olig2 induction resulted in an elevated number of generated oligoden-drocytes without a significant production of other cell lineages, unlike Mash1. These newly differentiated cells are also capable of migration and possible myelination, showing that targeting oligodendrocyte production and possible remyelination is a viable therapeutic strategy for restoration of neuronal function.

The path from the choroid plexus to the subventricular zone: go with the flow!

In adult mammals, under physiological conditions, neurogenesis, the process of generating new functional neurons from precursor cells, occurs mainly in two brain areas: the subgranular zone in the dentate gyrus of the hippocampus, and the subventricular zone (SVZ) lining the walls of the brain lateral ventricles. Taking into account the location of the SVZ and the cytoarchitecture of this periventricular neural progenitor cell niche, namely the fact that the slow dividing primary progenitor cells (type B cells) of the SVZ extend an apical primary cilium toward the brain ventricular space which is filled with cerebrospinal fluid (CSF), it becomes likely that the composition of the CSF can modulate both self-renewal, proliferation and differentiation of SVZ neural stem cells. The major site of CSF synthesis is the choroid plexus (CP); quite surprisingly, however, it is still largely unknown the contribution of molecules specifically secreted by the adult CP as modulators of the SVZ adult neurogenesis. This is even more relevant in light of recent evidence showing the ability of the CP to adapt its transcriptome and secretome to various physiologic and pathologic stimuli. By giving particular emphasizes to growth factors and axonal guidance molecules we will illustrate how CP-born molecules might play an important role in the SVZ niche cell population dynamics.