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Role of MTs in apico-basal elongation and INM, in the vertebrate neural plate. (a) Paraxial alignment of MTs causes epiblast cells to undergo apico-basal elongation, producing the high columnar shape that characterizes cells of the early bending neural plate in birds, amphibia and mammals. Images are schematic transverse sections. (b) Immunohistochemistry (green) for α-tubulin in transverse sections of the closing mouse spinal neural plate at the 21 somite stage. Apico-basally aligned MTs are visible in both the dorsal (b') and ventral (b'') regions. Blue: DAPI-stained nuclei. Scale bars: 80 μm (b); 20 μm (b', b''). (c) During INM, nuclei (blue) move basally during the G1-phase and remain at the basal neuroepithelial surface during the S-phase. During the S-to-G2 transition, dynein is activated and moves the nucleus toward the minus end of MTs at the centrosome (red), which is rooted in an apical cilium (pink). During the G2-phase, the cilium disassembles, allowing the newly-untethered centrosome to relocate to the nucleus, where it initiates mitosis. (c) Reproduced with permission from of Spear & Erickson, 2012, Developmental Biology 370: 33–41.
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Microtubules (MTs) are key cellular components, long known to participate in morphogenetic events that shape the developing embryo. However, the links between the cellular functions of MTs, their effects on cell shape and polarity, and their role in large-scale morphogenesis remain poorly understood. Here, these relationships were examined with res...
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... to play a role in mediating cell shape change during neural tube closure. Half a century ago, found that the blastopore cells of gastrulating amphibia, which develop a high columnar wedge-like shape, contain cyto- plasmic MTs aligned parallel to their long axis, suggesting that MTs might be responsible for cellular apico-basal elon- gation (Fig. 2a, b). They tentatively suggested that these principles might translate to the cells of the neural plate, which undergo similar morphological changes early in neu- rulation . Various studies subse- quently provided electron-microscopic evidence for this view, describing the growth and so-called 'paraxial' align- ment of MTs in the elongating ...
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... and S-phase occurs basally (Sauer, 1935;Langman et al. 1966). MT dis- ruption inhibits INM, producing a neural plate in which nuclei are distributed throughout the full thickness of the neuroepithelium (Messier, 1978). In the chick embryo, apical migration of G2 nuclei was found to be MT-dependent, with blockade by the MT inhibitor colcemid ( Fig. 2c; Spear & Erickson, 2012). In mouse brain slices, G2 nuclear move- ment was found to require Tpx2, a MT-nucleating/bundling protein that dissociates from the nucleus and localizes in the apical portion of the neuroepithelial cell during apical nuclear movement. In contrast, migration of nuclei basally during G1 appears to be a largely ...
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Tissue internalisation is a key morphogenetic mechanism by which embryonic tissues generate complex internal organs and a number of studies of epithelia have outlined a general view of tissue internalisation. Here we have used quantitative live imaging and mutant analysis to determine whether similar mechanisms are responsible for internalisation i...
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... The development of mammalian neural tubes requires proper coordination of cell proliferation, adhesion, migration, differentiation and apoptosis. Thus, in mice, NTDs can be caused by exposure to antimitotic agents [29], excessive cell proliferation [30], and mutation of genes encoding proteins associated with cell cycle progression or prevention of neuronal differentiation [31][32][33]. Li et al. [34] reported that miR-10a-5p inhibits chicken granulosa cell proliferation by targeting MAPRE1. ...
... NTDs are among the most serious birth defects among humans, and their incidence is as high as 1 in 1000 births [23]. Failure of morphogenetic events that occur during the neurulation process could lead to serious neurological consequences or even lethality [1,24,25], placing a significant burden on both the affected individuals and society. The etiology of NTDs is multifaceted, and the mechanism of how they develop is not fully understood. ...
Neural tube defects (NTDs), which are caused by impaired embryonic neural tube closure, are one of the most serious and common birth defects. Peptidyl-prolyl cis/trans isomerase 1 (Pin1) is a prolyl isomerase that uniquely regulates cell signaling by manipulating protein conformation following phosphorylation, although its involvement in neuronal development remains unknown. In this study, we explored the involvement of Pin1 in NTDs and its potential mechanisms both in vitro and in vivo. The levels of Pin1 expression were reduced in NTD models induced by all-trans retinoic acid (Atra). Pin1 plays a significant role in regulating the apoptosis, proliferation, differentiation, and migration of neurons. Moreover, Pin1 knockdown significantly was found to exacerbate oxidative stress (OS) and endoplasmic reticulum stress (ERs) in neuronal cells. Further studies showed that the Notch1-Nrf2 signaling pathway may participate in Pin1 regulation of NTDs, as evidenced by the inhibition and overexpression of the Notch1-Nrf2 pathway. In addition, immunofluorescence (IF), co-immunoprecipitation (Co-IP), and GST pull-down experiments also showed that Pin1 interacts directly with Notch1 and Nrf2. Thus, our study suggested that the knocking down of Pin1 promotes NTD progression by inhibiting the activation of the Notch1-Nrf2 signaling pathway, and it is possible that this effect is achieved by disrupting the interaction of Pin1 with Notch1 and Nrf2, affecting their proteostasis. Our research identified that the regulation of Pin1 by retinoic acid (RA) and its involvement in the development of NTDs through the Notch1-Nrf2 axis could enhance our comprehension of the mechanism behind RA-induced brain abnormalities.
... 6 Tau protein encourages tubulin association into microtubules (the stabilizers of the cell), participating in different cellular processes such as cell division. 7 The expression of tau protein is encoded by the MAPT gene (Microtubule-Associated Protein-Tau). 8 It was revealed that ionizing radiation can promote abnormal tau phosphorylation. ...
Most studies of the biological effects of ionizing radiation have been done on a single acute dose, while clinically and environmentally exposures occur under chronic/repetitive conditions. It is important to study effects of different patterns of ionizing radiation. In this study, a rat model was used to compare the effects of repetitive and acute exposure. Groups: (I) control, (II, III) were exposed to fractionated doses (1.5 GyX4) and (2 GyX4), respectively/24h interval, and (IV, V) were exposed to 6 Gy and 8 Gy of whole-body gamma irradiation, respectively. The gene expression of MAPT and tau phos-phorylation increased in all irradiated groups but the gene expression of PKN not affected. TGFβ% increased at dose of 2 GyX4 only. In addition, the cell cycle was arrested in S phase. Micronucleus (MN) increased and cell proliferation decreased. In conclusion, the dose and pattern of ionizing radiation do not affect the MAPT and PKN gene expression, but TGF-β, p-tau, MN assay and cell proliferation are significantly affected. The dose of 2 GyX4 showed distinctive effect. Repetitive exposure may increase TGF-β%, which causes radio-resistance and, G2/M delay. Thus, the cell cycle could be regulated in a different manner according to the dose and pattern of irradiation.
... In the literature, extensive zebrafish spinal cord nomenclature, electrophysiological studies, and static neuronal mapping of both unidentified, based on morphology and growth patterns, and identified neurons exists (Bernhardt, 1994;Bernhardt et al., 1990;Bernhardt et al., 1992;Drapeau et al., 2002;Hale et al., 2001;Higashijima et al., 2004;Kuwada et al., 1990a;Kuwada et al., 1990b;Lewis and Eisen, 2003). Immediately preceding spinal cord axonal outgrowth, dynamic cell movements during neurulation and neurocoel formation occur and was therefore occasionally also observed in this study due to the developmental spatio-temporal intertwined proximity to commissural outgrowth (Buckley et al., 2013;Cearns et al., 2016;Geldmacher-Voss et al., 2003;Kimmel et al., 1994;Papan and Campos-Ortega, 1994). ...
... The recordings confirm previous accounts using antibody stained micrometer thick serial sections, widefield and confocal microscopy (Buckley et al., 2013;Geldmacher-Voss et al., 2003;Kimmel et al., 1994;Papan and Campos-Ortega, 1994), and demonstrate the versatility of the light-sheet microscopy, as a complement to confocal microscopy, also for studies of this kind. No cell divisions were observed during these recordings (Buckley et al., 2013); the divisions would be occurring at the apical side of the epithelium, wherefore the daughter cell would easily become located on the other side of the midline by division across the midline in a socalled C-division, for midline Crossing division (Buckley et al., 2013;Cearns et al., 2016;Kimmel et al., 1994;Papan and Campos-Ortega, 1994). The nascent neuronal cell bodies are seen moving and aligning bilaterally along the midline (Movie S1). ...
Axonal growth and guidance at the ventral floor plate is here followed $\textit{in vivo}$ in real time at high resolution by light-sheet microscopy along several hundred micrometers of the zebrafish spinal cord. The recordings show the strikingly stereotyped spatio-temporal control that governs midline crossing. Commissural axons are observed crossing the ventral floor plate midline perpendicularly at about 20 microns/h, in a manner dependent on the Robo3 receptor and with a growth rate minimum around the midline, confirming previous observations. At guidance points, commissural axons are seen to decrease their growth rate and growth cones increase in size. Commissural filopodia appear to interact with the nascent neural network, and thereby trigger immediate plastic and reversible sinusoidal-shaped bending movements of neighboring commissural shafts. Ipsilateral axons extend concurrently, but straight and without bends, at three to six times higher growth rates than commissurals, indicating they project their path on a substrate-bound surface rather than relying on diffusible guidance cues. Growing axons appeared to be under stretch, an observation that is of relevance for tension-based models of cortical morphogenesis. The \textit{in vivo} observations provide for a discussion of the current distinction between substrate-bound and diffusible guidance cues. The study applies the transparent zebrafish model that provides an experimental model system to explore further the cellular, molecular and physical mechanisms involved during axonal growth, guidance and midline crossing through a combination of $\textit{in vitro}$ and $\textit{in vivo}$ approaches.
... In vertebrates, NTDs originate from a failure in morphogenetic events that occur during the neurulation process (16). Nerve tubes and nerve sheaths are the origins of the nervous system. ...
... Nerve tubes and nerve sheaths are the origins of the nervous system. Nerve epithelial cells are NSCs that possess a variety of differentiation potentials during the development of nerve tubes (16). The normal proliferation and differentiation of NSCs is involved in the normal development of nerve tubes (17). ...
Neural tube defects (NTDs) are the most serious and common birth defects in the clinical setting. The Notch signaling pathway has been implicated in different processes of the embryonic neural stem cells (NSCs) during neural tube development. The aim of the present study was to investigate the expression pattern and function of Notch1 (N1) in all‑ trans retinoic acid (atRA)‑induced NTDs and NSC differentiation. A mouse model of brain abnormality was established by administering 28 mg/kg atRA, and then brain development was examined using hematoxylin and eosin (H&E) staining. The N1 expression pattern was detected in the brain of mice embryos via immunohistochemistry and western blotting. NSCs were extracted from the fetal brain of C57 BL/6 embryos at 18.5 days of pregnancy. N1, Nestin, neurofilament (NF), glial fibrillary acidic protein (GFAP) and galactocerebroside (GALC) were identified using immunohistochemistry. Moreover, N1, presenilin 1 (PS1), Nestin, NF, GFAP and GALC were detected via western blotting at different time points in the NSCs with control media or atRA media. H&E staining identified that the embryonic brain treated with atRA was more developed compared with the control group. N1 was downregulated in the embryonic mouse brain between days 11 and 17 in the atRA‑treated group compared with the untreated group. The distribution of N1, Nestin, NF, GFAP and GALC was positively detected using immunofluorescence staining. Western blotting results demonstrated that there were significantly, synchronous decreased expression levels of N1 and PS1, but increased expression levels of NF, GFAP and GALC in NSCs treated with atRA compared with those observed in the controls (P<0.05). The results suggested that the N1 signaling pathway inhibited brain development and NSC differentiation. Collectively, it was found that atRA promoted mouse embryo brain development and the differentiation of NSCs by inhibiting the N1 pathway.
... However, examination of the tissue architecture in zebrafish [28][29][30] and other teleosts 31,32 revealed that the neural rod is shaped by infolding of a neural plate (albeit incompletely epithelialized), which best fits the description of primary neurulation 33 . Despite this evidence, differences in tissue architecture, the multi-layered organization of the neural plate and the apparent lack of hingepoints, neural groove and NFs are difficult to reconcile with a mode of primary neurulation and have contributed to the persistent view that neural tube formation in teleosts is different than in other vertebrates [34][35][36] . ...
Primary neurulation is the process by which the neural tube, the central nervous system precursor, is formed from the neural plate. Incomplete neural tube closure occurs frequently, yet underlying causes remain poorly understood. Developmental studies in amniotes and amphibians have identified hingepoint and neural fold formation as key morphogenetic events and hallmarks of primary neurulation, the disruption of which causes neural tube defects. In contrast, the mode of neurulation in teleosts has remained highly debated. Teleosts are thought to have evolved a unique mode of neurulation, whereby the neural plate infolds in absence of hingepoints and neural folds, at least in the hindbrain/trunk where it has been studied. Using high-resolution imaging and time-lapse microscopy, we show here the presence of these morphological landmarks in the zebrafish anterior neural plate. These results reveal similarities between neurulation in teleosts and other vertebrates and hence the suitability of zebrafish to understand human neurulation.
... These processes rely on pattern determining genes to be accurately expressed in dorsal and ventral regions of neural tubes [1]. Normal neurulation is governed by a variety of cellular events, such as cell proliferation, apoptosis, cytoskeleton establishment and cell viability, etc. [2], which are precisely regulated and controlled by a variety of signaling molecules, such as FGF, Wnt, Hh and TGF-beta [3]. Neural tube defects (NTDs) are defined as improperly developed neural tubes characterized by the open neural tubes in the brain or spinal cord at birth. ...
We, in this study, studied whether or not antioxidant activities of Baicalin could reduce the incidence of neural tube defects (NTDs) in the presence of hyperglycemia. Using early chick embryos, we demonstrated that Baicalin at 6 μM dramatically reduced NTDs rate and impaired neurogenesis in E4.5-day and HH10 chick embryo neural tubes induced by high glucose (HG). Likewise, immunofluorescent staining showed that Baicalin mitigated the HG-induced regression of Pax7 expression in neural tubes of both HH10 and E4.5-day chick embryos. Additionally, PHIS3 immunofluorescent staining in neural tubes of both HH10 and E4.5-day chick embryos manifested that cell proliferation inhibited by HG was significantly reversed by the administration of Baicalin, and similar result could also be observed in neurosphere assay in vitro. c-Caspase3 or γH2AX immunofluorescent staining and quantitative PCR showed that Baicalin administration alleviated HG-induced cell apoptosis and DNA damage. Bioinformatics results indicated that retinoic acid (RA) was likely to be the signaling pathway that Baicalin targeted on, and this was confirmed by whole-mount RALDH2 in situ hybridization and quantitative PCR of HH10 chick embryos in the absence/presence of Baicalin. In addition, blocking RA with an inhibitor abolished Baicalin's protective role in HG-induced NTDs, suppression of neurogenesis and cell proliferation, and induction of apoptosis, which further verified the centrality of RA in the process of Baicalin confronting HG-induced abnormal neurodevelopment.
... Therefore, the exencephaly phenotypes in Wdr34 −/− mice is likely more due to its function in the PCP pathway. What is more, although zebrafish does not have similar neurulation process to human or mouse [39,40], it is an excellent model to study CE and the conserved PCP signaling during its neural tube development [41]. The MO knockdown of wdr34 in zebrafish showed CE defects in neural tube (Fig. 3), which is also collaborates with the function of WDR34 in PCP signaling. ...
Neural tube defects (NTDs) are debilitating human congenital abnormalities due to failure of neural tube closure. Sonic Hedgehog (SHH) signaling is required for dorsal–ventral patterning of the neural tube. The loss of activation in SHH signaling normally causes holoprosencephaly while the loss of inhibition causes exencephaly due to failure in neural tube closure. WDR34 is a dynein intermedia chain component which is required for SHH activation. However, Wdr34 knockout mouse exhibit exencephaly. Here we screened mutations in WDR34 gene in 100 anencephaly patients of Chinese Han population. Compared to 1000 Genome Project data, two potentially disease causing missense mutations of WDR34 gene (c.1177G>A; p.G393S and c.1310A>G; p.Y437C) were identified in anencephaly patients. These two mutations did not affect the protein expression level of WDR34. Luciferase reporter and endogenous target gene expression level showed that both mutations are lose-of-function mutations in SHH signaling. Surprisingly, WDR34 could promote planar cell polarity (PCP) signaling and the G393S lost this promoting effect on PCP signaling. Morpholino knockdown of wdr34 in zebrafish caused severe convergent extension defects and pericardial abnormalities. The G393S mutant has less rescuing effects than both WT and Y437C WDR34 in zebrafish. Our results suggested that mutation in WDR34 could contribute to human NTDs by affecting both SHH and PCP signaling.
... In invertebrates, PAR-and CRB-complex proteins directly interact to determine the apicobasal axis and the position and stability of cell-cell adherens junctions [29][30][31][32][33][34][35][36]. PAR and CRB complex components are evolutionarily conserved and similarly regulate polarity, integrity, and morphogenesis of vertebrate epithelia, including the neural tube neuroepithelium [37][38][39]. In the mouse, Crb2 (one of the three vertebrate Crb genes) is required for maintenance of the apical polarity complex [40][41][42], and in zebrafish, the two crb2 genes have been implicated in retinal organisation. ...
In the spinal cord, the central canal forms through a poorly understood process termed dorsal collapse that involves attrition and remodelling of pseudostratified ventricular layer (VL) cells. Here, we use mouse and chick models to show that dorsal ventricular layer (dVL) cells adjacent to dorsal midline Nestin⁽⁺⁾ radial glia (dmNes⁺RG) down-regulate apical polarity proteins, including Crumbs2 (CRB2) and delaminate in a stepwise manner; live imaging shows that as one cell delaminates, the next cell ratchets up, the dmNes⁺RG endfoot ratchets down, and the process repeats. We show that dmNes⁺RG secrete a factor that promotes loss of cell polarity and delamination. This activity is mimicked by a secreted variant of Crumbs2 (CRB2S) which is specifically expressed by dmNes⁺RG. In cultured MDCK cells, CRB2S associates with apical membranes and decreases cell cohesion. Analysis of Crb2F/F/Nestin-Cre+/− mice, and targeted reduction of Crb2/CRB2S in slice cultures reveal essential roles for transmembrane CRB2 (CRB2TM) and CRB2S on VL cells and dmNes⁺RG, respectively. We propose a model in which a CRB2S–CRB2TM interaction promotes the progressive attrition of the dVL without loss of overall VL integrity. This novel mechanism may operate more widely to promote orderly progenitor delamination.
... by tissue folding and fusion ( Araya et al., 2016;Cearns et al., 2016;Lowery and Sive, 2004). 515 ...
Circulation of the cerebrospinal fluid (CSF) contributes to body axis formation and brain development. Here, we investigated the unexplained origins of the CSF flow bidirectionality in the central canal of the spinal cord of 30 hpf zebrafish embryos and its impact on development. Experiments combined with modeling and simulations demonstrate that the CSF flow is generated locally by caudally-polarized motile cilia along the ventral wall of the central canal. The closed geometry of the canal imposes the average flow rate to be null, explaining the reported bidirectionality. We also demonstrate that at this early stage, motile cilia ensure the proper formation of the central canal. Furthermore, we demonstrate that the bidirectional flow accelerates the transport of particles in the CSF via a coupled convective-diffusive transport process. Our study demonstrates that cilia activity combined with muscle contractions sustain the long-range transport of extracellular lipidic particles, enabling embryonic growth.
