Neural Development最新文献

Microglia are increasingly shown to be key players in neuron development and synapse connectivity. However, the underlying mechanisms by which microglia regulate neuron function remain poorly understood in part because such analysis is challenging in the brain where neurons and synapses are intermingled and connectivity is only beginning to be mapped. Here, we discuss the features and function of microglia in the ordered mammalian retina where the laminar organization of neurons and synapses facilitates such molecular studies. We discuss microglia origins and consider the evidence for molecularly distinct microglia subpopulations and their potential for differential roles with a particular focus on the early stages of retina development. We then review the models and methods used for the study of these cells and discuss emerging data that link retina microglia to the genesis and survival of particular retina cell subtypes. We also highlight potential roles for microglia in shaping the development and organization of the vasculature and discuss cellular and molecular mechanisms involved in this process. Such insights may help resolve the mechanisms by which retinal microglia impact visual function and help guide studies of related features in brain development and disease.

In the developing neural tube in chicken and mammals, neural stem cells proliferate and differentiate according to a stereotyped spatiotemporal pattern. Several actors have been identified in the control of this process, from tissue-scale morphogens patterning to intrinsic determinants in neural progenitor cells. In a previous study (Bonnet et al. eLife 7, 2018), we have shown that the CDC25B phosphatase promotes the transition from proliferation to differentiation by stimulating neurogenic divisions, suggesting that it acts as a maturating factor for neural progenitors. In this previous study, we set up a mathematical model linking fixed progenitor modes of division to the dynamics of progenitors and differentiated populations. Here, we extend this model over time to propose a complete dynamical picture of this process. We start from the standard paradigm that progenitors are homogeneous and can perform any type of divisions (proliferative division yielding two progenitors, asymmetric neurogenic divisions yielding one progenitor and one neuron, and terminal symmetric divisions yielding two neurons). We calibrate this model using data published by Saade et al. (Cell Reports 4, 2013) about mode of divisions and population dynamics of progenitors/neurons at different developmental stages. Next, we explore the scenarios in which the progenitor population is actually split into two different pools, one of which is composed of cells that have lost the capacity to perform proliferative divisions. The scenario in which asymmetric neurogenic division would induce such a loss of proliferative capacity appears very relevant.

Background: Purkinje cells play a central role in establishing the cerebellar circuit. Accordingly, disrupting Purkinje cell development impairs cerebellar morphogenesis and motor function. In the Car8^wdl mouse model of hereditary ataxia, severe motor deficits arise despite the cerebellum overcoming initial defects in size and morphology.

Methods: To resolve how this compensation occurs, we asked how the loss of carbonic anhydrase 8 (CAR8), a regulator of IP3R1 Ca²⁺ signaling in Purkinje cells, alters cerebellar development in Car8^wdl mice. Using a combination of histological, physiological, and behavioral analyses, we determined the extent to which the loss of CAR8 affects cerebellar anatomy, neuronal firing, and motor coordination during development.

Results: Our results reveal that granule cell proliferation is reduced in early postnatal mutants, although by the third postnatal week there is enhanced and prolonged proliferation, plus an upregulation of Sox2 expression in the inner EGL. Modified circuit patterning of Purkinje cells and Bergmann glia accompany these granule cell adjustments. We also find that although anatomy eventually normalizes, the abnormal activity of neurons and muscles persists.

Conclusions: Our data show that losing CAR8 only transiently restricts cerebellar growth, but permanently damages its function. These data support two current hypotheses about cerebellar development and disease: (1) Sox2 expression may be upregulated at sites of injury and contribute to the rescue of cerebellar structure and (2) transient delays to developmental processes may precede permanent motor dysfunction. Furthermore, we characterize waddles mutant mouse morphology and behavior during development and propose a Sox2-positive, cell-mediated role for rescue in a mouse model of human motor diseases.

Background: Functioning of the adult nervous system depends on the establishment of neural circuits during embryogenesis. In vertebrates, neurons that make up motor circuits form in distinct domains along the dorsoventral axis of the neural tube. Each domain is characterized by a unique combination of transcription factors (TFs) that promote a specific fate, while repressing fates of adjacent domains. The prdm12 TF is required for the expression of eng1b and the generation of V1 interneurons in the p1 domain, but the details of its function remain unclear.

Methods: We used CRISPR/Cas9 to generate the first germline mutants for prdm12 and employed this resource, together with classical luciferase reporter assays and co-immunoprecipitation experiments, to study prdm12b function in zebrafish. We also generated germline mutants for bhlhe22 and nkx6.1 to examine how these TFs act with prdm12b to control p1 formation.

Results: We find that prdm12b mutants lack eng1b expression in the p1 domain and also possess an abnormal touch-evoked escape response. Using luciferase reporter assays, we demonstrate that Prdm12b acts as a transcriptional repressor. We also show that the Bhlhe22 TF binds via the Prdm12b zinc finger domain to form a complex. However, bhlhe22 mutants display normal eng1b expression in the p1 domain. While prdm12 has been proposed to promote p1 fates by repressing expression of the nkx6.1 TF, we do not observe an expansion of the nkx6.1 domain upon loss of prdm12b function, nor is eng1b expression restored upon simultaneous loss of prdm12b and nkx6.1.

Conclusions: We conclude that prdm12b germline mutations produce a phenotype that is indistinguishable from that of morpholino-mediated loss of prdm12 function. In terms of prdm12b function, our results indicate that Prdm12b acts as transcriptional repressor and interacts with both EHMT2/G9a and Bhlhe22. However, bhlhe22 function is not required for eng1b expression in vivo, perhaps indicating that other bhlh genes can compensate during embryogenesis. Lastly, we do not find evidence for nkx6.1 and prdm12b acting as a repressive pair in formation of the p1 domain - suggesting that prdm12b is not solely required to repress non-p1 fates, but is specifically needed to promote p1 fates.

Background: The cerebellum is a foliated posterior brain structure involved in coordination of motor movements and cognition. The cerebellum undergoes rapid growth postnataly due to Sonic Hedgehog (SHH) signaling-dependent proliferation of ATOH1+ granule cell precursors (GCPs) in the external granule cell layer (EGL), a key step for generating cerebellar foliation and the correct number of granule cells. Due to its late development, the cerebellum is particularly vulnerable to injury from preterm birth and stress around birth. We recently uncovered an intrinsic capacity of the developing cerebellum to replenish ablated GCPs via adaptive reprogramming of Nestin-expressing progenitors (NEPs). However, whether this compensation mechanism occurs in mouse mutants affecting the developing cerebellum and could lead to mis-interpretation of phenotypes was not known.

Methods: We used two different approaches to remove the main SHH signaling activator GLI2 in GCPs: 1) Our mosaic mutant analysis with spatial and temporal control of recombination (MASTR) technique to delete Gli2 in a small subset of GCPs; 2) An Atoh1-Cre transgene to delete Gli2 in most of the EGL. Genetic Inducible Fate Mapping (GIFM) and live imaging were used to analyze the behavior of NEPs after Gli2 deletion.

Results: Mosaic analysis demonstrated that SHH-GLI2 signaling is critical for generating the correct pool of granule cells by maintaining GCPs in an undifferentiated proliferative state and promoting their survival. Despite this, inactivation of GLI2 in a large proportion of GCPs in the embryo did not lead to the expected dramatic reduction in the size of the adult cerebellum. GIFM uncovered that NEPs do indeed replenish GCPs in Gli2 conditional mutants, and then expand and partially restore the production of granule cells. Furthermore, the SHH signaling-dependent NEP compensation requires Gli2, demonstrating that the activator side of the pathway is involved.

Conclusion: We demonstrate that a mouse conditional mutation that results in loss of SHH signaling in GCPs is not sufficient to induce long term severe cerebellum hypoplasia. The ability of the neonatal cerebellum to regenerate after loss of cells via a response by NEPs must therefore be considered when interpreting the phenotypes of Atoh1-Cre conditional mutants affecting GCPs.

Background: During development, gustatory (taste) neurons likely undergo numerous changes in morphology and expression prior to differentiation into maturity, but little is known this process or the factors that regulate it. Neuron differentiation is likely regulated by a combination of transcription and growth factors. Embryonically, most geniculate neuron development is regulated by the growth factor brain derived neurotrophic factor (BDNF). Postnatally, however, BDNF expression becomes restricted to subpopulations of taste receptor cells with specific functions. We hypothesized that during development, the receptor for BDNF, tropomyosin kinase B receptor (TrkB), may also become developmentally restricted to a subset of taste neurons and could be one factor that is differentially expressed across taste neuron subsets.

Methods: We used transgenic mouse models to label both geniculate neurons innervating the oral cavity (Phox2b+), which are primarily taste, from those projecting to the outer ear (auricular neurons) to label TrkB expressing neurons (TrkB^GFP). We also compared neuron number, taste bud number, and taste receptor cell types in wild-type animals and conditional TrkB knockouts.

Results: Between E15.5-E17.5, TrkB receptor expression becomes restricted to half of the Phox2b + neurons. This TrkB downregulation was specific to oral cavity projecting neurons, since TrkB expression remained constant throughout development in the auricular geniculate neurons (Phox2b-). Conditional TrkB removal from oral sensory neurons (Phox2b+) reduced this population to 92% of control levels, indicating that only 8% of these neurons do not depend on TrkB for survival during development. The remaining neurons failed to innervate any remaining taste buds, 14% of which remained despite the complete loss of innervation. Finally, some types of taste receptor cells (Car4+) were more dependent on innervation than others (PLCβ2+).

Conclusions: Together, these findings indicate that TrkB expression and dependence divides gustatory neurons into three subpopulations: 1) neurons that always express TrkB and are TrkB-dependent during development (50%), 2) neurons dependent on TrkB during development but that downregulate TrkB expression between E15.5 and E17.5 (41%), and 3) neurons that never express or depend on TrkB (9%). These TrkB-independent neurons are likely non-gustatory, as they do not innervate taste buds.

Background: Mammalian motor circuits display remarkable cellular diversity with hundreds of motor neuron (MN) subtypes innervating hundreds of different muscles. Extensive research on limb muscle-innervating MNs has begun to elucidate the genetic programs that control animal locomotion. In striking contrast, the molecular mechanisms underlying the development of axial muscle-innervating MNs, which control breathing and spinal alignment, are poorly studied.

Methods: Our previous studies indicated that the function of the Collier/Olf/Ebf (COE) family of transcription factors (TFs) in axial MN development may be conserved from nematodes to simple chordates. Here, we examine the expression pattern of all four mouse COE family members (mEbf1-mEbf4) in spinal MNs and employ genetic approaches in both nematodes and mice to investigate their function in axial MN development.

Results: We report that mEbf1 and mEbf2 are expressed in distinct MN clusters (termed "columns") that innervate different axial muscles. Mouse Ebf1 is expressed in MNs of the hypaxial motor column (HMC), which is necessary for breathing, while mEbf2 is expressed in MNs of the medial motor column (MMC) that control spinal alignment. Our characterization of Ebf2 knock-out mice uncovered a requirement for Ebf2 in the differentiation program of a subset of MMC MNs and revealed for the first time molecular diversity within MMC neurons. Intriguingly, transgenic expression of mEbf1 or mEbf2 can rescue axial MN differentiation and locomotory defects in nematodes (Caenorhabditis elegans) lacking unc-3, the sole C. elegans ortholog of the COE family, suggesting functional conservation among mEbf1, mEbf2 and nematode UNC-3.

Conclusions: These findings support the hypothesis that genetic programs controlling axial MN development are deeply conserved across species, and further advance our understanding of such programs by revealing an essential role for Ebf2 in mouse axial MNs. Because human mutations in COE orthologs lead to neurodevelopmental disorders characterized by motor developmental delay, our findings may advance our understanding of these human conditions.