Differentiation Of Neural Precursors Research Articles

Zebrafish PEX1 Is Required for the Generation of GABAergic Neuron in p3 Domain.

Maintenance of neural progenitors requires Notch signaling in vertebrate development. Previous study has shown that Jagged2-mediated Notch signaling maintains proliferating neural progenitors in the ventral spinal cord. However, components for Jagged-mediated signaling remain poorly defined during late neurogenesis. Here we performed yeast-two hybrid screening by using the intracellular domain (ICD) of zebrafish Jagged2, and investigated a possible role of PEX1 as a component of Notch signaling for the cell-fate decision and the differentiation of neural precursors in p3 domain. Western blotting showed that zebrafish PEX1 might interacts with the ICD of zebrafish Jagged2 physically. PEX1 morpholino-injected embryos showed the increased number of GABAergic KA" neurons as well as the ectopic expression of secondary motor neurons in the p3 domain. The increased number of KA" neurons was also observed in the zebrafish embryos with PEX1 mutation induced by CRISPR/Cas9. These phenotypes resemble with that of Jagged2 morphant. Our observations imply that a critical role of PEX1 in the cell-fate decision of proliferating neural precursors in the p3 domain during the continuing growth and development of the vertebrate nervous system.

SOX10 Loss Sensitizes Melanoma Cells to Cytokine-Mediated Inflammatory Cell Death.

These data support a role for SOX10 expression altering the response to T cell-mediated cell death and contribute to a broader understanding of the interaction between immune cells and melanoma cells.

CHANGES IN THE NEUROGENIC NICHE OF THE RAT HIPPOCAMPUS UNDER HYPOXIC EXPOSURE

One of the most vulnerable brain structures to hypoxia is the hippocampus. Maintenance of the neurogenic niche cell pool in the subgranular zone of the hippocampus (SGZ) is provided by adaptive mechanisms. Among them are changes in the functional activity of mitochondrial respiratory chain complexes and the reaction of astroglia, which provides metabolic support for neurons. In order to study the dynamics of adaptive changes in neurons and glia in the dentate gyrus of the hippocampus under hypoxic conditions on a model of intermitten hypobaric hypoxia (5000 m, equivalent to 10.5% O2), with a single (60 min) and multiple (8 and 20 episodes) exposure in low-resistant rats, immunomorphological methods revealed the features of localization and content in the neurons of complex IV mitochondrial respiratory chain (MTCO1), astrocyte marker proteins glutamine synthetase (GS) and GFAP, and doublecortin (DCX) in immature neurons. With a single hypoxia, the content of MTCO1 in neurons significantly increased, and after eight exposures, the amount of glutamine synthetase (GS) in astrocytes of the dentate gyrus of the hippocampus increased. Changes in the content of GS were most pronounced in the processes of astrocytes, which indicates a redistribution of GS. The number of DCX+ neurons in the SGZ significantly decreased after 20 episodes of hypoxia. At the same time, DCX+ cells of glial morphology were found in the polymorphic layer, and staining for GFAP showed an increase in the number of astrocytes. This may be due to a shift in the direction of cell differentiation in the neurogenic niche. Thus, in hypoxia, at the initial stage, a functional restructuring of the respiratory chain of neurons of the granular layer occurs. Subsequently, it is noted by the activation of astrocytes that modulate glutamate metabolism. The presence of a relationship between the dynamics of adaptive reactions of energy exchange in neurons and glia and changes in neurogenesis during 20 episodes of hypoxia suggests that during long-term hypoxia, the differentiation of neural precursors of SGZ in the direction of astroglia occurs, however, this issue requires further study in order to more accurately determine the nature of DCX+ cells.

Abstract 3167: The scaffolding protein DLG5 is necessary to maintain Sonic Hedgehog signaling in glioblastoma stem cells and promote tumor growth

Abstract Proteins complexes that maintain the apico-basal polarity of epithelial cells and organize cell-signaling domains are necessary for normal tissue organization and therefore regarded as tumor suppressors. Accordingly, the scaffolding proteins that form the Scribble polarity complex (SCRIB, LLGL1, and DLG members) are usually downregulated in cancer, in particular in highly aggressive or invasive tumors. Because gliomas arise from non-epithelial precursors that are likely motile before transformation, the expression and functions of these scaffolding proteins in malignant brain tumors has remained almost entirely unexplored. Surprisingly, we have found that one member of the DLG family, DLG5, is highly upregulated in malignant gliomas compared to other solid tumors. Indeed, DLG5 was the predominant DLG member found in all molecular subtypes of glioblastoma (GBM). Knockdown of DLG5 reduced the viability, invasiveness, and self-renewal of GBM stem cells belonging to the proneural and mesenchymal molecular subtypes. As a result, intracranial tumors defficient in DLG5 were smaller and less proliferative than controls, resulting in extended animal survival. At the molecular level, DLG5 knockdown decreased the expression of stemness-related genes (SOX2, NANOG, POU5F1) but increased the expression of LLGL1, a component of the Scribble complex that is associated with neural precursor differentiation. These changes were accompanied by downregulation of Sonic Hedgehog (SHh) signaling, both in vitro and in vivo, which drives GBM cell stemness and proliferation. Moreover, DLG5-deficient GBM stem cells became insensitive to exogenous SHh and were unable to upregulate Gli1 or increase cell invasion in presence of added SHh, compared to control cells. We further confirmed that DLG5 downregulation increased the ubiquitination of Gli1-containing complexes as well as Gli1 proteasomal degradation, providing a suitable mechanism for the loss of SHh signaling and tumor stemness. These results describe for the first time the expression of a DLG family member in malignant gliomas and reveal a novel, polarity-independent, pro-tumoral function of DLG5, which differs from its proposed tumor-suppressor role in other solid tumors. Citation Format: Somanath Kundu, Mohan S. Nandhu, Sharon L. Longo, John A. Longo, Shawn Rai, Lawrence S. Chin, Timothy E. Richardson, Mariano S. Viapiano. The scaffolding protein DLG5 is necessary to maintain Sonic Hedgehog signaling in glioblastoma stem cells and promote tumor growth [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 3167.

Deficiency of N-glycanase 1 perturbs neurogenesis and cerebral development modeled by human organoids

Mutations in N-glycanase 1 (NGLY1), which deglycosylates misfolded glycoproteins for degradation, can cause NGLY1 deficiency in patients and their abnormal fetal development in multiple organs, including microcephaly and other neurological disorders. Using cerebral organoids (COs) developed from human embryonic stem cells (hESCs) and induced pluripotent stem cells (hiPSCs), we investigate how NGLY1 dysfunction disturbs early brain development. While NGLY1 loss had limited impact on the undifferentiated cells, COs developed from NGLY1-deficient hESCs showed defective formation of SATB2-positive upper-layer neurons, and attenuation of STAT3 and HES1 signaling critical for sustaining radial glia. Bulk and single-cell transcriptomic analysis revealed premature neuronal differentiation accompanied by downregulation of secreted and transcription factors, including TTR, IGFBP2, and ID4 in NGLY1-deficient COs. NGLY1 malfunction also dysregulated ID4 and enhanced neuronal differentiation in CO transplants developed in vivo. NGLY1-deficient CO cells were more vulnerable to multiple stressors; treating the deficient cells with recombinant TTR reduced their susceptibility to stress from proteasome inactivation, likely through LRP2-mediated activation of MAPK signaling. Expressing NGLY1 led to IGFBP2 and ID4 upregulation in CO cells developed from NGLY1-deficiency patient’s hiPSCs. In addition, treatment with recombinant IGFBP2 enhanced ID4 expression, STAT3 signaling, and proliferation of NGLY1-deficient CO cells. Overall, our discoveries suggest that dysregulation of stress responses and neural precursor differentiation underlies the brain abnormalities observed in NGLY1-deficient individuals.

Deficiency of nde1 in zebrafish induces brain inflammatory responses and autism-like behavior

SummaryThe cytoskeletal protein NDE1 plays an important role in chromosome segregation, neural precursor differentiation, and neuronal migration. Clinical studies have shown that NDE1 deficiency is associated with several neuropsychiatric disorders including autism. Here, we generated nde1 homologous deficiency zebrafish (nde1−/−) to elucidate the cellular molecular mechanisms behind it. nde1−/− exhibit increased neurological apoptotic responses at early infancy, enlarged ventricles, and shrank valvula cerebelli in adult brain tissue. Behavioral analysis revealed that nde1−/− displayed autism-like behavior traits such as increased locomotor activity and repetitive stereotype behaviors and impaired social and kin recognition behaviors. Furthermore, nde1 mRNA injection rescued apoptosis in early development, and minocycline treatment rescued impaired social behavior and overactive motor activity by inhibiting inflammatory cytokines. In this study, we revealed that nde1 homozygous deletion leads to abnormal neurological development with autism-related behavioral phenotypes and that inflammatory responses in the brain are an important molecular basis behind it.

A Functional Signature in the Developing Cerebellum: Evidence From a Preclinical Model of Autism.

Autism spectrum disorders (ASD) are pervasive neurodevelopmental conditions detected during childhood when delayed language onset and social deficits are observed. Children diagnosed with ASD frequently display sensorimotor deficits associated with the cerebellum, suggesting a dysfunction of synaptic circuits. Astroglia are part of the tripartite synapses and postmortem studies reported an increased expression of the glial fibrillary acidic protein (GFAP) in the cerebellum of ASD patients. Astroglia respond to neuronal activity with calcium transients that propagate to neighboring cells, resulting in a functional response known as a calcium wave. This form of intercellular signaling is implicated in proliferation, migration, and differentiation of neural precursors. Prenatal exposure to valproate (VPA) is a preclinical model of ASD in which premature migration and excess of apoptosis occur in the internal granular layer (IGL) of the cerebellum during the early postnatal period. In this study we tested calcium wave propagation in the IGL of mice prenatally exposed to VPA. Sensorimotor deficits were observed and IGL depolarization evoked a calcium wave with astrocyte recruitment. The calcium wave propagation, initial cell recruitment, and mean amplitude of the calcium transients increased significantly in VPA-exposed mice compared to the control group. Astrocyte recruitment was significantly increased in the VPA model, but the mean amplitude of the calcium transients was unchanged. Western blot and histological studies revealed an increased expression of GFAP, higher astroglial density and augmented morphological complexity. We conclude that the functional signature of the IGL is remarkably augmented in the preclinical model of autism.

The Effects of the Olig Family on the Regulation of Spinal Cord Development and Regeneration.

Neurons and glial cells in the central nervous system (CNS) are generated from neuroepithelial cells in the ventricular zone that surrounds the embryonic neural tube. The proliferation and distinct differentiation of neural precursors occurs at certain stages and are regulated by a series of transcription factors leading to the generation of neuronal and glial cell subtypes. In this manuscript, we review the effects of the Olig family, namely, members Olig1, Olig2 and Olig3, on the distinct differentiation of glial and neuronal cells in the developing spinal cord and injured neural tissue.

Disrupted Trigeminal Ganglion Development in a Mouse Model of Familial Dysautonomia

Neurons in the trigeminal ganglion (TG) relay somatosensory information from the face and oral cavity to the central nervous system. Arising from the differentiation of both neural crest cell and ectodermal placode cell precursors during embryogenesis, neurons in the TG ultimately encode one sensory modality (i.e., touch, pain, temperature). TG neuron identity is discernable by particular molecular markers, including the mutually exclusive expression of the neurotrophin receptors TrkA, TrkB, or TrkC, which are required for long-term neuronal function and survival. While little is known about the mechanisms that establish and maintain distinct neuronal subpopulations in the TG, disease studies offer some insight. Familial Dysautonomia (FD) is a hereditary neuropathy caused by a mutation in Elongator complex protein 1 (ELP1), which leads to reduced ELP1 protein levels in sensory and autonomic neurons. FD patients have clinical deficits that point to abnormal TG function, including impaired sensation of facial pain and temperature. Moreover, FD patients have smaller trigeminal nerves compared to healthy controls, in the absence of overt degeneration. Given these findings, we hypothesize that ELP1 is necessary for the development and survival of TG sensory neurons. Our initial results uncovered cytoplasmic Elp1 expression in both placode cell- and neural crest cell-derived neurons of the developing mouse TG starting from embryonic day (E)10.5. Using a conditional knockout mouse model in which Elp1 is deleted from Wnt1-positive neural crest cells and derivatives, including sensory neurons, we next examined the effect of Elp1 loss on TG development. Our preliminary data reveal Elp1 depletion from neural crest cells leads to abnormal TG morphology and fewer trigeminal nerve branches as early as embryonic day (E)11, a phenotype that persists throughout development and suggests defects in both neural crest- and placode cell-derived neurons in the TG. TrkA-expressing neurons, which typically function in pain perception, are especially vulnerable to Elp1 loss during this period of TG formation. By E12.5, we observe decreased TrkA expression in distal trigeminal nerve endings and fewer TrkA-positive neuron cell bodies in the TG. Analyses from TUNEL staining reveal increased cell death in the TG at E12.5, suggesting TrkA-expressing neurons, progenitor cells, and/or supporting cells may undergo aberrant apoptosis in Elp1 conditional knockout TG. Together, our results confirm a role for Elp1 in the development and survival of TG sensory neurons. These findings help explain the loss of facial sensation in FD and may guide future therapeutic interventions to increase quality of life for FD patients.

Immune-tolerance to human iPS-derived neural progenitors xenografted into the immature cerebellum is overridden by species-specific differences in differentiation timing

We xeno-transplanted human neural precursor cells derived from induced pluripotent stem cells into the cerebellum and brainstem of mice and rats during prenatal development or the first postnatal week. The transplants survived and started to differentiate up to 1 month after birth when they were rejected by both species. Extended survival and differentiation of the same cells were obtained only when they were transplanted in NOD-SCID mice. Transplants of human neural precursor cells mixed with the same cells after partial in vitro differentiation or with a cellular extract obtained from adult rat cerebellum increased survival of the xeno-graft beyond one month. These findings are consistent with the hypothesis that the slower pace of differentiation of human neural precursors compared to that of rodents restricts induction of immune-tolerance to human antigens expressed before completion of maturation of the immune system. With further maturation the transplanted neural precursors expressed more mature antigens before the graft were rejected. Supplementation of the immature cells suspensions with more mature antigens may help to induce immune-tolerance for those antigens expressed only later by the engrafted cells.

Transcription Factor-Based Fate Specification and Forward Programming for Neural Regeneration.

Traditionally, in vitro generation of donor cells for brain repair has been dominated by the application of extrinsic growth factors and morphogens. Recent advances in cell engineering strategies such as reprogramming of somatic cells into induced pluripotent stem cells and direct cell fate conversion have impressively demonstrated the feasibility to manipulate cell identities by the overexpression of cell fate-determining transcription factors. These strategies are now increasingly implemented for transcription factor-guided differentiation of neural precursors and forward programming of pluripotent stem cells toward specific neural subtypes. This review covers major achievements, pros and cons, as well as future prospects of transcription factor-based cell fate specification and the applicability of these approaches for the generation of donor cells for brain repair.

Chemotherapy-Induced Neuropathy and Drug Discovery Platform Using Human Sensory Neurons Converted Directly from Adult Peripheral Blood.

Chemotherapy‐induced peripheral neuropathy (PN) is a disorder damaging the peripheral nervous system (PNS) and represents one of the most common side effects of chemotherapy, negatively impacting the quality of life of patients to the extent of withdrawing life‐saving chemotherapy dose or duration. Unfortunately, the pathophysiological effects of PN are poorly understood, in part due to the lack of availability of large numbers of human sensory neurons (SNs) for study. Previous reports have demonstrated that human SNs can be directly converted from primitive CD34+ hematopoietic cells, but was limited to a small‐scale product of SNs and derived exclusively from less abundant allogenic sources of cord or drug mobilized peripheral blood (PB). To address this shortcoming, we have developed and report detailed procedures toward the generation of human SN directly converted from conventionally drawn PB of adults that can be used in a high‐content screening platform for discovery‐based studies of chemotherapy agents on neuronal biology. In the absence of mobilization drugs, cryogenically preserved adult human PB could be induced to (i)SN via development through expandable neural precursor differentiation. iSNs could be transferable to high‐throughput procedures suitable for high‐content screening applicable to neuropathy for example, alterations in neurite morphology in response to chemotherapeutics. Our study provides the first reported platform using adult PB‐derived iSNs to study peripheral nervous system‐related neuropathies as well as target and drug screening potential for the ability to prevent, block, or repair chemotherapy‐induced PN damage. stem cells translational medicine 2019;8:1180–1191

Mitochondrial Complex I Function Is Essential for Neural Stem/Progenitor Cells Proliferation and Differentiation.

Neurogenesis in developing and adult mammalian brain is a tightly regulated process that relies on neural stem cell (NSC) activity. There is increasing evidence that mitochondrial metabolism affects NSC homeostasis and differentiation but the precise role of mitochondrial function in the neurogenic process requires further investigation. Here, we have analyzed how mitochondrial complex I (MCI) dysfunction affects NSC viability, proliferation and differentiation, as well as survival of the neural progeny. We have generated a conditional knockout model (hGFAP-NDUFS2 mice) in which expression of the NDUFS2 protein, essential for MCI function, is suppressed in cells expressing the Cre recombinase under the human glial fibrillary acidic protein promoter, active in mouse radial glial cells (RGCs) and in neural stem cells (NSCs) that reside in adult neurogenic niches. In this model we observed that survival of central NSC population does not appear to be severely affected by MCI dysfunction. However, perinatal brain development was markedly inhibited and Ndufs2 knockout mice died before the tenth postnatal day. In addition, in vitro studies of subventricular zone NSCs showed that active neural progenitors require a functional MCI to produce ATP and to proliferate. In vitro differentiation of neural precursors into neurons and oligodendrocytes was also profoundly affected. These data indicate the need of a correct MCI function and oxidative phosphorylation for glia-like NSC proliferation, differentiation and subsequent oligodendrocyte or neuronal maturation.

Derivation of novel naive-like porcine embryonic stem cells by a reprogramming factor-assisted strategy.

The establishment of ungulate embryonic stem cells (ESCs) has been notoriously difficult via a conventional approach. We combined a traditional ESC culture method with reprogramming factors to assist the establishment of porcine naive-like ESCs (nESCs). Pig embryonic fibroblasts were transfected with a tetracycline-inducible vector carrying 4 classic mouse reprogramming factors, followed by somatic cell nuclear transfer and culturing to the blastocyst stage. Then, the inner cell mass was isolated and seeded in culture medium. The naive-like ESCs had characteristic verys similar to those of mouse ESCs and showed no signs of altered morphology or differentiation, even after 130 passages. They depended on leukemia inhibitory factor signals for maintenance of pluripotency, and the female cell lines had low expression of the X-inactive specific transcript gene and no histone H3 lysine 27 trimethylation spot. Notably, the ESCs differentiated into 3 germ layers in vitro and could be induced to undergo directional neural and kidney precursor differentiation under defined conditions, and the ESCs could keep proliferating after doxycycline was removed. nESCs can be established, and the well-characterized ESC lines will be useful for the research of transgenic pig models for human disease.-Zhang, M., Wang, C., Jiang, H., Liu, M., Yang, N., Zhao, L., Hou, D., Jin, Y., Chen, Q., Chen, Y., Wang, J., Dai, Y., Li, R. Derivation of novel naive-like porcine embryonic stem cells by a reprogramming factor-assisted strategy.

Temporal correlation of elevated PRMT1 gene expression with mushroom body neurogenesis during bumblebee brain development

Neural development depends on the controlled proliferation and differentiation of neural precursors. In holometabolous insects, these processes must be coordinated during larval and pupal development. Recently, protein arginine methylation has come into focus as an important mechanism of controlling neural stem cell proliferation and differentiation in mammals. Whether a similar mechanism is at work in insects is unknown. We investigated this possibility by determining the expression pattern of three protein arginine methyltransferase mRNAs (PRMT1, 4 and 5) in the developing brain of bumblebees by in situ hybridisation. We detected expression in neural precursors and neurons in functionally important brain areas throughout development. We found markedly higher expression of PRMT1, but not PRMT4 and PRMT5, in regions of mushroom bodies containing dividing cells during pupal stages at the time of active neurogenesis within this brain area. At later stages of development, PRMT1 expression levels were found to be uniform and did not correlate with actively dividing cells. Our study suggests a role for PRMT1 in regulating neural precursor divisions in the mushroom bodies of bumblebees during the period of neurogenesis.

Canonical Wnt signaling regulates patterning, differentiation and nucleogenesis in mouse hypothalamus and prethalamus

The hypothalamus is a small, but anatomically and functionally complex region of the brain whose development is poorly understood. In this study, we have explored its development by studying the canonical Wnt signaling pathway, generating gain and loss of function mutations of beta-catenin (Ctnnb1) in both hypothalamic and prethalamic neuroepithelium. Deletion of Ctnnb1 resulted in an anteriorized and hypoplastic hypothalamus. Posterior structures were lost or reduced, and anterior structures were expanded. In contrast, overexpression of a constitutively active mutant form of Ctnnb1 resulted in severe hyperplasia of prethalamus and hypothalamus, and expanded expression of a subset of posterior and premamillary hypothalamic markers. Moderate defects in differentiation of Arx-positive GABAergic neural precursors were observed in both prethalamus and hypothalamus of Ctnnb1 loss of function mutants, while in gain of function mutants, their differentiation was completely suppressed, although markers of prethalamic progenitors were preserved. Multiple other region-specific markers, including several specific posterior hypothalamic structures, were also suppressed in Ctnnb1 gain of function mutations. Severe, region-specific defects in hypothalamic nucleogenesis were also observed in both gain and loss of function mutations of Ctnnb1. Finally, both gain and loss of function of Ctnnb1 also produced severe, non-cell autonomous disruptions of pituitary development. These findings demonstrate a central and multifaceted role for canonical Wnt signaling in regulating growth, patterning, differentiation and nucleogenesis in multiple diencephalic regions.

Enhancement of neuronal differentiation by using small molecules modulating Nodal/Smad, Wnt/β-catenin, and FGF signaling

Pluripotent embryonic stem cells are one of the best modalities for the disease treatment due to their potential for self-renewal and differentiation into various cell types. Induction of stem cell differentiation into specific cell lineages has been investigated for decades, especially in vitro neuronal differentiation of embryonic stem cells. However, in vitro differentiation methods do not yield sufficient amounts of neurons for use in the therapeutic treatment of neurological disorders. Here, we provide an improved neuronal differentiation method based on a combination of small regulatory molecules for specific signaling pathways (FGF4 for FGF signaling, SB431542 for Nodal/Smad signaling, and XAV939 and BIO for Wnt signaling) in N2B27 media. We found that FGF4 was required for neural induction, SB431542 accelerated neural precursor differentiation, and treatment with XAV939 and BIO at different periods enhanced neuronal differentiation. These optimized neuronal differentiation conditions may allow a greater neuron cell yield within a shorter time than current methods and be the basis for treatment of neurological dysfunction using stem cells.

Kir6.2 Deficiency Promotes Mesencephalic Neural Precursor Cell Differentiation via Regulating miR-133b/GDNF in a Parkinson's Disease Mouse Model.

The loss of dopaminergic (DA) neurons in the substantia nigra (SN) is a major feature in the pathology of Parkinson's disease (PD). Using neural stem or progenitor cells (NSC/NPCs), the prospect of replacing the missing or damaged DA neurons is very attractive for PD therapy. However, little is known about the endogenous mechanisms and molecular pathways regulating the NSC/NPC proliferation and differentiation in the development of PD. Herein, using Kir6.2 knockout (Kir6.2-/-) mice, we observed that genetic deficiency of Kir6.2 exacerbated the loss of SN DA neurons relatively early in a chronic MPTP/probenecid (MPTP/p) injection course, but rescued the damage of neurons 7days after the last MPTP/p injection. Meanwhile, we found that Kir6.2 knockout predominantly increased the differentiation of nuclear receptor-related 1 (Nurr1+) precursors to DA neurons, indicating that Kir6.2 deficiency could activate an endogenous self-repair process. Furthermore, we demonstrated in vivo and in vitro that lack of Kir6.2 promoted neuronal differentiation via inhibiting the downregulation of glia cell line-derived neurotrophic factor (GDNF), which negatively related to the level of microRNA-133b. Notably, we revealed that Gdnf is a target gene of miR-133b and transfection of miR-133b could attenuate the enhancement of neural precursor differentiation induced by Kir6.2 deficiency. Collectively, we clarify for the first time that Kir6.2/K-ATP channel functions as a novel endogenous negative regulator of NPC differentiation, and provide a promising neuroprotective target for PD therapeutics.

Larval neurogenesis in the copepod Tigriopus californicus (Tetraconata, Multicrustacea).

Arthropod early neurogenesis shows distinct patterns that have been interpreted in an evolutionary framework. For instance, crustaceans and Hexapoda form the taxon Tetraconata and share the differentiation of specific neural precursors, the neuroblasts, a character which sets them apart from Chelicerata and Myriapoda. Neuroblasts are relatively large stem cells that generate ganglion mother cells by asymmetric divisions. Ganglion mother cells typically divide once to give rise to neurons and glia cells. In hexapods, neuroblasts segregate from the neuroectoderm before they begin their characteristic proliferative activity. In the crustaceans studied so far, neuroblasts remain in the neuroectoderm. Yet, detailed studies on early neurogenesis of crustaceans at the cellular level are largely restricted to some malacostracan and branchiopod species. Crustaceans are very diverse and likely paraphyletic with respect to hexapods. Hence, knowledge about neural differentiation in other crustacean taxa might contribute to the understanding of evolution of neurogenesis in Tetraconata. Here, we describe the early neurogenesis during naupliar development of the copepod Tigriopus californicus. We show that neuroblasts are present that generate ganglion mother cells, which in turn divide to give rise to neurons of the ventral nerve cord. These two neural precursor cell types and their specific arrangement correspond to what has been found in other crustaceans. One obvious difference concerns the relative size of the neuroblasts, which are not much larger than their progeny. Our results complement the picture of neural differentiation in crustaceans and suggest that superficially located neuroblasts are likely the ancestral condition in Tetraconata.

GDNF/GFRα1 Complex Abrogates Self-Renewing Activity of Cortical Neural Precursors Inducing Their Differentiation.

SummaryThe balance between factors leading to proliferation and differentiation of cortical neural precursors (CNPs) determines the correct cortical development. In this work, we show that GDNF and its receptor GFRα1 are expressed in the neocortex during the period of cortical neurogenesis. We show that the GDNF/GFRα1 complex inhibits the self-renewal capacity of mouse CNP cells induced by fibroblast growth factor 2 (FGF2), promoting neuronal differentiation. While GDNF leads to decreased proliferation of cultured cortical precursor cells, ablation of GFRα1 in glutamatergic cortical precursors enhances its proliferation. We show that GDNF treatment of CNPs promoted morphological differentiation even in the presence of the self-renewal-promoting factor, FGF2. Analysis of GFRα1-deficient mice shows an increase in the number of cycling cells during cortical development and a reduction in dendrite development of cortical GFRα1-expressing neurons. Together, these results indicate that GDNF/GFRα1 signaling plays an essential role in regulating the proliferative condition and the differentiation of cortical progenitors.