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Germline genetic architecture of Alzheimer's disease (AD) indicates microglial mechanisms of disease susceptibility and outcomes. However, the mechanisms that enable microglia to mediate protective responses to AD pathology remain elusive. Adgrg1 is specifically expressed in yolk-sac-derived microglia. This study reveals the role of yolk-sac-derived microglia in AD pathology, highlighting the function of ADGRG1 in modulating microglial protective responses to amyloid deposition. Utilizing both constitutive and inducible microglial Adgrg1 knockout 5xFAD models, we demonstrate that Adgrg1 deficiency leads to increased amyloid deposition, exacerbated neuropathology, and accelerated cognitive impairment. Transcriptomic analyses reveal a distinct microglial state characterized by downregulated genes associated with homeostasis, phagocytosis, and lysosomal functions. Functional assays in mouse models and human embryonic stem cells-derived microglia support that microglial ADGRG1 is required for efficient Aß phagocytosis. Together, these results uncover a GPCR-dependent microglial response to Aß, pointing towards potential therapeutic strategies to alleviate disease progression by enhancing microglial functional competence.
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Germinal matrix hemorrhage (GMH) is a devastating neurodevelopmental condition affecting preterm infants, but why blood vessels in this brain region are vulnerable to rupture remains unknown. Here we show that microglia in prenatal mouse and human brain interact with nascent vasculature in an age-dependent manner and that ablation of these cells in mice reduces angiogenesis in the ganglionic eminences, which correspond to the human germinal matrix. Consistent with these findings, single-cell transcriptomics and flow cytometry show that distinct subsets of CD45+ cells from control preterm infants employ diverse signaling mechanisms to promote vascular network formation. In contrast, CD45+ cells from infants with GMH harbor activated neutrophils and monocytes that produce proinflammatory factors, including azurocidin 1, elastase and CXCL16, to disrupt vascular integrity and cause hemorrhage in ganglionic eminences. These results underscore the brain's innate immune cells in region-specific angiogenesis and how aberrant activation of these immune cells promotes GMH in preterm infants.
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The ability to spatially map multiple layers of the omics information over different time points allows for exploring the mechanisms driving brain development, differentiation, arealization, and alterations in disease. Herein we developed and applied spatial tri-omic sequencing technologies, DBiT ARP-seq (spatial ATAC-RNA-Protein-seq) and DBiT CTRP-seq (spatial CUT&Tag-RNA-Protein-seq) together with multiplexed immunofluorescence imaging (CODEX) to map spatial dynamic remodeling in brain development and neuroinflammation. A spatiotemporal tri-omic atlas of the mouse brain was obtained at different stages from postnatal day P0 to P21, and compared to the regions of interest in the human developing brains. Specifically, in the cortical area, we discovered temporal persistence and spatial spreading of chromatin accessibility for the layer-defining transcription factors. In corpus callosum, we observed dynamic chromatin priming of myelin genes across the subregions. Together, it suggests a role for layer specific projection neurons to coordinate axonogenesis and myelination. We further mapped the brain of a lysolecithin (LPC) neuroinflammation mouse model and observed common molecular programs in development and neuroinflammation. Microglia, exhibiting both conserved and distinct programs for inflammation and resolution, are transiently activated not only at the core of the LPC lesion, but also at distal locations presumably through neuronal circuitry. Thus, this work unveiled common and differential mechanisms in brain development and neuroinflammation, resulting in a valuable data resource to investigate brain development, function and disease.
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The nuclear genome is spatially organized into a three-dimensional (3D) architecture by physical association of large chromosomal domains with subnuclear compartments including the nuclear lamina at the radial periphery and nuclear speckles within the nucleoplasm1-5. However, how spatial genome architecture regulates human brain development has been overlooked owing to technical limitations. Here, we generate high-resolution maps of genomic interactions with the lamina and speckles in cells of the neurogenic lineage isolated from midgestational human cortex, uncovering an intimate association between subnuclear genome compartmentalization, chromatin state and transcription. During cortical neurogenesis, spatial genome organization is extensively remodeled, relocating hundreds of neuronal genes from the lamina to speckles including key neurodevelopmental genes bivalent for H3K27me3 and H3K4me3. At the lamina, bivalent genes have exceptionally low expression, and relocation to speckles enhances resolution of bivalent chromatin to H3K4me3 and increases transcription >7-fold. We further demonstrate that proximity to the nuclear periphery - not the presence of H3K27me3 - is the dominant factor in maintaining the lowly expressed, poised state of bivalent genes embedded in the lamina. In addition to uncovering a critical role of subnuclear genome compartmentalization in neurogenic transcriptional regulation, our results establish a new paradigm in which knowing the spatial location of a gene is necessary to understanding its epigenomic regulation.
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The development of the human neocortex is a highly dynamic process and involves complex cellular trajectories controlled by cell-type-specific gene regulation1. Here, we collected paired single-nucleus chromatin accessibility and transcriptome data from 38 human neocortical samples encompassing both the prefrontal cortex and primary visual cortex. These samples span five main developmental stages, ranging from the first trimester to adolescence. In parallel, we performed spatial transcriptomic analysis on a subset of the samples to illustrate spatial organization and intercellular communication. This atlas enables us to catalog cell type-, age-, and area-specific gene regulatory networks underlying neural differentiation. Moreover, combining single-cell profiling, progenitor purification, and lineage-tracing experiments, we have untangled the complex lineage relationships among progenitor subtypes during the transition from neurogenesis to gliogenesis in the human neocortex. We identified a tripotential intermediate progenitor subtype, termed Tri-IPC, responsible for the local production of GABAergic neurons, oligodendrocyte precursor cells, and astrocytes. Remarkably, most glioblastoma cells resemble Tri-IPCs at the transcriptomic level, suggesting that cancer cells hijack developmental processes to enhance growth and heterogeneity. Furthermore, by integrating our atlas data with large-scale GWAS data, we created a disease-risk map highlighting enriched ASD risk in second-trimester intratelencephalic projection neurons. Our study sheds light on the gene regulatory landscape and cellular dynamics of the developing human neocortex.
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The vasculature of the central nervous system is a 3D lattice composed of laminar vascular beds interconnected by penetrating vessels. The mechanisms controlling 3D lattice network formation remain largely unknown. Combining viral labeling, genetic marking, and single-cell profiling in the mouse retina, we discovered a perivascular neuronal subset, annotated as Fam19a4/Nts-positive retinal ganglion cells (Fam19a4/Nts-RGCs), directly contacting the vasculature with perisomatic endfeet. Developmental ablation of Fam19a4/Nts-RGCs led to disoriented growth of penetrating vessels near the ganglion cell layer (GCL), leading to a disorganized 3D vascular lattice. We identified enriched PIEZO2 expression in Fam19a4/Nts-RGCs. Piezo2 loss from all retinal neurons or Fam19a4/Nts-RGCs abolished the direct neurovascular contacts and phenocopied the Fam19a4/Nts-RGC ablation deficits. The defective vascular structure led to reduced capillary perfusion and sensitized the retina to ischemic insults. Furthermore, we uncovered a Piezo2-dependent perivascular granule cell subset for cerebellar vascular patterning, indicating neuronal Piezo2-dependent 3D vascular patterning in the brain.
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Cerebelo , Neurônios , Retina , Animais , Feminino , Masculino , Camundongos , Cerebelo/metabolismo , Cerebelo/irrigação sanguínea , Cerebelo/citologia , Canais Iônicos/metabolismo , Camundongos Endogâmicos C57BL , Neurônios/metabolismo , Retina/citologia , Retina/metabolismo , Células Ganglionares da Retina/metabolismo , Vasos Retinianos/metabolismoRESUMO
The temporal lobe of the human brain contains the entorhinal cortex (EC). This region of the brain is a highly interconnected integrative hub for sensory and spatial information; it also has a key role in episodic memory formation and is the main source of cortical hippocampal inputs1-4. The human EC continues to develop during childhood5, but neurogenesis and neuronal migration to the EC are widely considered to be complete by birth. Here we show that the human temporal lobe contains many young neurons migrating into the postnatal EC and adjacent regions, with a large tangential stream persisting until the age of around one year and radial dispersal continuing until around two to three years of age. By contrast, we found no equivalent postnatal migration in rhesus macaques (Macaca mulatta). Immunostaining and single-nucleus RNA sequencing of ganglionic eminence germinal zones, the EC stream and the postnatal EC revealed that most migrating cells in the EC stream are derived from the caudal ganglionic eminence and become LAMP5+RELN+ inhibitory interneurons. These late-arriving interneurons could continue to shape the processing of sensory and spatial information well into postnatal life, when children are actively interacting with their environment. The EC is one of the first regions of the brain to be affected in Alzheimer's disease, and previous work has linked cognitive decline to the loss of LAMP5+RELN+ cells6,7. Our investigation reveals that many of these cells arrive in the EC through a major postnatal migratory stream in early childhood.
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Movimento Celular , Neurônios , Lobo Temporal , Animais , Pré-Escolar , Humanos , Lactente , Córtex Entorrinal/citologia , Córtex Entorrinal/fisiologia , Eminência Ganglionar/citologia , Interneurônios/citologia , Interneurônios/fisiologia , Macaca mulatta , Neurônios/citologia , Neurônios/fisiologia , Análise da Expressão Gênica de Célula Única , Lobo Temporal/citologia , Lobo Temporal/crescimento & desenvolvimentoRESUMO
The development and diversity of neuronal subtypes in the human hypothalamus has been insufficiently characterized. To address this, we integrated transcriptomic data from 241,096 cells (126,840 newly generated) in the prenatal and adult human hypothalamus to reveal a temporal trajectory from proliferative stem cell populations to mature hypothalamic cell types. Iterative clustering of the adult neurons identified 108 robust transcriptionally distinct neuronal subtypes representing 10 hypothalamic nuclei. Pseudotime trajectories provided insights into the genes driving formation of these nuclei. Comparisons to single-cell transcriptomic data from the mouse hypothalamus suggested extensive conservation of neuronal subtypes despite certain differences in species-enriched gene expression. The uniqueness of hypothalamic neuronal lineages was examined developmentally by comparing excitatory lineages present in cortex and inhibitory lineages in ganglionic eminence, revealing both distinct and shared drivers of neuronal maturation across the human forebrain. These results provide a comprehensive transcriptomic view of human hypothalamus development through gestation and adulthood at cellular resolution.
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Hipotálamo , Neurônios , Camundongos , Animais , Humanos , Hipotálamo/metabolismo , Neurônios/metabolismo , Transcriptoma , Perfilação da Expressão Gênica , GenômicaRESUMO
We analyzed >700,000 single-nucleus RNA sequencing profiles from 106 donors during prenatal and postnatal developmental stages and identified lineage-specific programs that underlie the development of specific subtypes of excitatory cortical neurons, interneurons, glial cell types, and brain vasculature. By leveraging single-nucleus chromatin accessibility data, we delineated enhancer gene regulatory networks and transcription factors that control commitment of specific cortical lineages. By intersecting our results with genetic risk factors for human brain diseases, we identified the cortical cell types and lineages most vulnerable to genetic insults of different brain disorders, especially autism. We find that lineage-specific gene expression programs up-regulated in female cells are especially enriched for the genetic risk factors of autism. Our study captures the molecular progression of cortical lineages across human development.
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Encefalopatias , Córtex Cerebral , Neurônios , Feminino , Humanos , Recém-Nascido , Gravidez , Encefalopatias/genética , Córtex Cerebral/crescimento & desenvolvimento , Redes Reguladoras de Genes , Interneurônios/metabolismo , Neurônios/metabolismo , Análise de Célula Única , Masculino , Fatores de RiscoRESUMO
Radial glial (RG) development is essential for cerebral cortex growth and organization. In humans, the outer radial glia (oRG) subtype is expanded and gives rise to diverse neurons and glia. However, the mechanisms regulating oRG differentiation are unclear. oRG cells express leukemia-inhibitory factor (LIF) receptors during neurogenesis, and consistent with a role in stem cell self-renewal, LIF perturbation impacts oRG proliferation in cortical tissue and organoids. Surprisingly, LIF treatment also increases the production of inhibitory interneurons (INs) in cortical cultures. Comparative transcriptomic analysis identifies that the enhanced IN population resembles INs produced in the caudal ganglionic eminence. To evaluate whether INs could arise from oRGs, we isolated primary oRG cells and cultured them with LIF. We observed the production of INs from oRG cells and an increase in IN abundance following LIF treatment. Our observations suggest that LIF signaling regulates the capacity of oRG cells to generate INs.
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Células Ependimogliais , Neurogênese , Humanos , Diferenciação Celular/fisiologia , Neurogênese/fisiologia , Córtex Cerebral , Interneurônios/fisiologiaRESUMO
The molecular mechanisms and evolutionary changes accompanying synapse development are still poorly understood1,2. Here we generate a cross-species proteomic map of synapse development in the human, macaque and mouse neocortex. By tracking the changes of more than 1,000 postsynaptic density (PSD) proteins from midgestation to young adulthood, we find that PSD maturation in humans separates into three major phases that are dominated by distinct pathways. Cross-species comparisons reveal that human PSDs mature about two to three times slower than those of other species and contain higher levels of Rho guanine nucleotide exchange factors (RhoGEFs) in the perinatal period. Enhancement of RhoGEF signalling in human neurons delays morphological maturation of dendritic spines and functional maturation of synapses, potentially contributing to the neotenic traits of human brain development. In addition, PSD proteins can be divided into four modules that exert stage- and cell-type-specific functions, possibly explaining their differential associations with cognitive functions and diseases. Our proteomic map of synapse development provides a blueprint for studying the molecular basis and evolutionary changes of synapse maturation.
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Proteômica , Sinapses , Adolescente , Animais , Criança , Pré-Escolar , Humanos , Lactente , Recém-Nascido , Camundongos , Adulto Jovem , Cognição/fisiologia , Espinhas Dendríticas , Idade Gestacional , Macaca , Neurônios/metabolismo , Densidade Pós-Sináptica/metabolismo , Fatores de Troca de Nucleotídeo Guanina Rho/metabolismo , Transdução de Sinais , Especificidade da Espécie , Sinapses/metabolismo , Sinapses/fisiologiaRESUMO
Chronic neurodegeneration and acute injuries lead to neuron losses via diverse processes. We compared retinal ganglion cell (RGC) responses between chronic glaucomatous conditions and the acute injury model. Among major RGC subclasses, αRGCs and intrinsically photosensitive RGCs (ipRGCs) preferentially survive glaucomatous conditions, similar to findings in the retina subject to axotomy. Focusing on an αRGC intrinsic factor, Osteopontin (secreted phosphoprotein 1 [Spp1]), we found an ectopic neuronal expression of Osteopontin (Spp1) in other RGCs subject to glaucomatous conditions. This contrasted with the Spp1 downregulation subject to axotomy. αRGC-specific Spp1 elimination led to significant αRGC loss, diminishing their resiliency. Spp1 overexpression led to robust neuroprotection of susceptible RGC subclasses under glaucomatous conditions. In contrast, Spp1 overexpression did not significantly protect RGCs subject to axotomy. Additionally, SPP1 marked adult human RGC subsets with large somata and SPP1 expression in the aqueous humor correlated with glaucoma severity. Our study reveals Spp1's role in mediating neuronal resiliency in glaucoma.
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Glaucoma , Doenças do Nervo Óptico , Humanos , Células Ganglionares da Retina/metabolismo , Osteopontina , Nervo Óptico/metabolismo , Doenças do Nervo Óptico/metabolismoRESUMO
Radial glial cells, the stem cells of the cerebral cortex, extend a long basal fiber that ends in basal endfeet. A new study in PLOS Biology found that non-muscle myosins control basal endfoot integrity to regulate interneuron organization.
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Córtex Cerebral , Células Ependimogliais , Interneurônios , Células-Tronco , MiosinasRESUMO
Mutations in the human progranulin (GRN) gene are a leading cause of frontotemporal lobar degeneration (FTLD). While previous studies implicate aberrant microglial activation as a disease-driving factor in neurodegeneration in the thalamocortical circuit in Grn-/- mice, the exact mechanism for neurodegeneration in FTLD-GRN remains unclear. By performing comparative single-cell transcriptomics in the thalamus and frontal cortex of Grn-/- mice and patients with FTLD-GRN, we have uncovered a highly conserved astroglial pathology characterized by upregulation of gap junction protein GJA1, water channel AQP4, and lipid-binding protein APOE, and downregulation of glutamate transporter SLC1A2 that promoted profound synaptic degeneration across the two species. This astroglial toxicity could be recapitulated in mouse astrocyte-neuron cocultures and by transplanting induced pluripotent stem cell-derived astrocytes to cortical organoids, where progranulin-deficient astrocytes promoted synaptic degeneration, neuronal stress, and TDP-43 proteinopathy. Together, these results reveal a previously unappreciated astroglial pathology as a potential key mechanism in neurodegeneration in FTLD-GRN.
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Demência Frontotemporal , Degeneração Lobar Frontotemporal , Humanos , Animais , Camundongos , Progranulinas/genética , Demência Frontotemporal/genética , Astrócitos/metabolismo , Peptídeos e Proteínas de Sinalização Intercelular/genética , Mutação , Degeneração Lobar Frontotemporal/genética , Degeneração Lobar Frontotemporal/metabolismo , Degeneração Lobar Frontotemporal/patologiaRESUMO
Apical-basal progenitor cell polarity establishes key features of the radial and laminar architecture of the developing human cortex. The unique diversity of cortical stem cell populations and an expansion of progenitor population size in the human cortex have been mirrored by an increase in the complexity of cellular processes that regulate stem cell morphology and behaviour, including their polarity. The study of human cells in primary tissue samples and human stem cell-derived model systems (such as cortical organoids) has provided insight into these processes, revealing that protein complexes regulate progenitor polarity by controlling cell membrane adherence within appropriate cortical niches and are themselves regulated by cytoskeletal proteins, signalling molecules and receptors, and cellular organelles. Studies exploring how cortical stem cell polarity is established and maintained are key for understanding the features of human brain development and have implications for neurological dysfunction.
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Polaridade Celular , Córtex Cerebral , Humanos , Células-Tronco/fisiologia , Organoides , Membrana CelularRESUMO
Interactions between angiogenesis and neurogenesis regulate embryonic brain development. However, a comprehensive understanding of the stages of vascular cell maturation is lacking, especially in the prenatal human brain. Using fluorescence-activated cell sorting, single-cell transcriptomics, and histological and ultrastructural analyses, we show that an ensemble of endothelial and mural cell subtypes tile the brain vasculature during the second trimester. These vascular cells follow distinct developmental trajectories and utilize diverse signaling mechanisms, including collagen, laminin, and midkine, to facilitate cell-cell communication and maturation. Interestingly, our results reveal that tip cells, a subtype of endothelial cells, are highly enriched near the ventricular zone, the site of active neurogenesis. Consistent with these observations, prenatal vascular cells transplanted into cortical organoids exhibit restricted lineage potential that favors tip cells, promotes neurogenesis, and reduces cellular stress. Together, our results uncover important mechanisms into vascular maturation during this critical period of human brain development.
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Células Endoteliais , Neovascularização Fisiológica , Encéfalo , Colágeno , Humanos , Laminina , Midkina , Neovascularização Patológica/patologia , Neovascularização Fisiológica/fisiologia , PericitosRESUMO
Self-organizing three-dimensional cellular models derived from human pluripotent stem cells or primary tissue have great potential to provide insights into how the human nervous system develops, what makes it unique and how disorders of the nervous system arise, progress and could be treated. Here, to facilitate progress and improve communication with the scientific community and the public, we clarify and provide a basic framework for the nomenclature of human multicellular models of nervous system development and disease, including organoids, assembloids and transplants.
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Consenso , Sistema Nervoso , Organoides , Terminologia como Assunto , Humanos , Modelos Biológicos , Sistema Nervoso/citologia , Sistema Nervoso/patologia , Organoides/citologia , Organoides/patologia , Células-Tronco Pluripotentes/citologiaRESUMO
Zika virus (ZIKV) is a flavivirus transmitted via mosquitoes and sex to cause congenital neurodevelopmental defects, including microcephaly. Inherited forms of microcephaly (MCPH) are associated with disrupted centrosome organization. Similarly, we found that ZIKV infection disrupted centrosome organization. ZIKV infection disrupted the organization of centrosomal proteins including CEP63, a MCPH-associated protein. The ZIKV nonstructural protein NS3 bound CEP63, and expression of NS3 was sufficient to alter centrosome architecture and CEP63 localization. Loss of CEP63 suppressed ZIKV-induced centrosome disorganization, indicating that ZIKV requires CEP63 to disrupt centrosome organization. ZIKV infection or CEP63 loss decreased the centrosomal localization and stability of TANK-binding kinase 1 (TBK1), a regulator of the innate immune response. ZIKV infection also increased the centrosomal accumulation of the CEP63 interactor DTX4, a ubiquitin ligase that degrades TBK1. Therefore, we propose that ZIKV disrupts CEP63 function to increase centrosomal DTX4 localization and destabilization of TBK1, thereby tempering the innate immune response.
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Microcefalia , Infecção por Zika virus , Zika virus , Animais , Proteínas de Ciclo Celular/genética , Proteínas de Ciclo Celular/metabolismo , Centrossomo/metabolismo , Humanos , Imunidade Inata , Microcefalia/metabolismo , Zika virus/fisiologiaRESUMO
The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) readily infects a variety of cell types impacting the function of vital organ systems, with particularly severe impact on respiratory function. Neurological symptoms, which range in severity, accompany as many as one-third of COVID-19 cases, indicating a potential vulnerability of neural cell types. To assess whether human cortical cells can be directly infected by SARS-CoV-2, we utilized stem-cell-derived cortical organoids as well as primary human cortical tissue, both from developmental and adult stages. We find significant and predominant infection in cortical astrocytes in both primary tissue and organoid cultures, with minimal infection of other cortical populations. Infected and bystander astrocytes have a corresponding increase in inflammatory gene expression, reactivity characteristics, increased cytokine and growth factor signaling, and cellular stress. Although human cortical cells, particularly astrocytes, have no observable ACE2 expression, we find high levels of coronavirus coreceptors in infected astrocytes, including CD147 and DPP4. Decreasing coreceptor abundance and activity reduces overall infection rate, and increasing expression is sufficient to promote infection. Thus, we find tropism of SARS-CoV-2 for human astrocytes resulting in inflammatory gliosis-type injury that is dependent on coronavirus coreceptors.
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Astrócitos , Córtex Cerebral , SARS-CoV-2 , Tropismo Viral , Enzima de Conversão de Angiotensina 2/metabolismo , Astrócitos/enzimologia , Astrócitos/virologia , Córtex Cerebral/virologia , Humanos , Organoides/virologia , Cultura Primária de Células , SARS-CoV-2/fisiologiaRESUMO
Organoids are 3D cell culture systems derived from human pluripotent stem cells that contain tissue resident cell types and reflect features of early tissue organization. Neural organoids are a particularly innovative scientific advance given the lack of accessibility of developing human brain tissue and intractability of neurological diseases. Neural organoids have become an invaluable approach to model features of human brain development that are not well reflected in animal models. Organoids also hold promise for the study of atypical cellular, molecular, and genetic features that underscore neurological diseases. Additionally, organoids may provide a platform for testing therapeutics in human cells and are a potential source for cell replacement approaches to brain injury or disease. Despite the promising features of organoids, their broad utility is tempered by a variety of limitations yet to be overcome, including lack of high-fidelity cell types, limited maturation, atypical physiology, and lack of arealization, features that may limit their reliability for certain applications.