The Allen Mouse Brain Common Coordinate Framework: A 3D Reference Atlas.

Recent large-scale collaborations are generating major surveys of cell types and connections in the mouse brain, collecting large amounts of data across modalities, spatial scales, and brain areas. Successful integration of these data requires a standard 3D reference atlas. Here, we present the Allen Mouse Brain Common Coordinate Framework (CCFv3) as such a resource. We constructed an average template brain at 10 µm voxel resolution by interpolating high resolution in-plane serial two-photon tomography images with 100 µm z-sampling from 1,675 young adult C57BL/6J mice. Then, using multimodal reference data, we parcellated the entire brain directly in 3D, labeling every voxel with a brain structure spanning 43 isocortical areas and their layers, 329 subcortical gray matter structures, 81 fiber tracts, and 8 ventricular structures. CCFv3 can be used to analyze, visualize, and integrate multimodal and multiscale datasets in 3D and is openly accessible (https://atlas.brain-map.org/).

Morphological diversity of single neurons in molecularly defined cell types.

Dendritic and axonal morphology reflects the input and output of neurons and is a defining feature of neuronal types1,2, yet our knowledge of its diversity remains limited. Here, to systematically examine complete single-neuron morphologies on a brain-wide scale, we established a pipeline encompassing sparse labelling, whole-brain imaging, reconstruction, registration and analysis. We fully reconstructed 1,741 neurons from cortex, claustrum, thalamus, striatum and other brain regions in mice. We identified 11 major projection neuron types with distinct morphological features and corresponding transcriptomic identities. Extensive projectional diversity was found within each of these major types, on the basis of which some types were clustered into more refined subtypes. This diversity follows a set of generalizable principles that govern long-range axonal projections at different levels, including molecular correspondence, divergent or convergent projection, axon termination pattern, regional specificity, topography, and individual cell variability. Although clear concordance with transcriptomic profiles is evident at the level of major projection type, fine-grained morphological diversity often does not readily correlate with transcriptomic subtypes derived from unsupervised clustering, highlighting the need for single-cell cross-modality studies. Overall, our study demonstrates the crucial need for quantitative description of complete single-cell anatomy in cell-type classification, as single-cell morphological diversity reveals a plethora of ways in which different cell types and their individual members may contribute to the configuration and function of their respective circuits.

A guide to the BRAIN Initiative Cell Census Network data ecosystem.

Characterizing cellular diversity at different levels of biological organization and across data modalities is a prerequisite to understanding the function of cell types in the brain. Classification of neurons is also essential to manipulate cell types in controlled ways and to understand their variation and vulnerability in brain disorders. The BRAIN Initiative Cell Census Network (BICCN) is an integrated network of data-generating centers, data archives, and data standards developers, with the goal of systematic multimodal brain cell type profiling and characterization. Emphasis of the BICCN is on the whole mouse brain with demonstration of prototype feasibility for human and nonhuman primate (NHP) brains. Here, we provide a guide to the cellular and spatial approaches employed by the BICCN, and to accessing and using these data and extensive resources, including the BRAIN Cell Data Center (BCDC), which serves to manage and integrate data across the ecosystem. We illustrate the power of the BICCN data ecosystem through vignettes highlighting several BICCN analysis and visualization tools. Finally, we present emerging standards that have been developed or adopted toward Findable, Accessible, Interoperable, and Reusable (FAIR) neuroscience. The combined BICCN ecosystem provides a comprehensive resource for the exploration and analysis of cell types in the brain.

Hierarchical organization of cortical and thalamic connectivity.

The mammalian cortex is a laminar structure containing many areas and cell types that are densely interconnected in complex ways, and for which generalizable principles of organization remain mostly unknown. Here we describe a major expansion of the Allen Mouse Brain Connectivity Atlas resource1, involving around a thousand new tracer experiments in the cortex and its main satellite structure, the thalamus. We used Cre driver lines (mice expressing Cre recombinase) to comprehensively and selectively label brain-wide connections by layer and class of projection neuron. Through observations of axon termination patterns, we have derived a set of generalized anatomical rules to describe corticocortical, thalamocortical and corticothalamic projections. We have built a model to assign connection patterns between areas as either feedforward or feedback, and generated testable predictions of hierarchical positions for individual cortical and thalamic areas and for cortical network modules. Our results show that cell-class-specific connections are organized in a shallow hierarchy within the mouse corticothalamic network.

NUT Expression Is of Diagnostic Utility in the Distinction of Digital Papillary Carcinoma From Poroid Hidradenoma.

ABSTRACT: The distinction between digital papillary adenocarcinoma (DPAC) and benign cutaneous adnexal tumors is clinically important and can be challenging. Poroid hidradenoma frequently occurs at acral sites and can show a number of histological features, which overlap with digital papillary adenocarcinoma. Recent work has shown that YAP1-NUTM1 fusions are frequent in poroid hidradenoma and are associated with nuclear protein in testis (NUT) expression by immunohistochemistry. We evaluated the expression of NUT-1 by immunohistochemistry in 4 cases of DPAC and 4 cases of poroid hidradenoma. Three of 4 cases of poroid hidradenoma showed strong NUT-1 expression, with no staining in any of the cases of DPAC. These results suggest that NUT-1 immunohistochemistry may be a useful additional tool in evaluating this differential diagnosis.

Highlights from the Era of Open Source Web-Based Tools.

High digital connectivity and a focus on reproducibility are contributing to an open science revolution in neuroscience. Repositories and platforms have emerged across the whole spectrum of subdisciplines, paving the way for a paradigm shift in the way we share, analyze, and reuse vast amounts of data collected across many laboratories. Here, we describe how open access web-based tools are changing the landscape and culture of neuroscience, highlighting six free resources that span subdisciplines from behavior to whole-brain mapping, circuits, neurons, and gene variants.

A comprehensive transcriptional map of primate brain development.

The transcriptional underpinnings of brain development remain poorly understood, particularly in humans and closely related non-human primates. We describe a high-resolution transcriptional atlas of rhesus monkey (Macaca mulatta) brain development that combines dense temporal sampling of prenatal and postnatal periods with fine anatomical division of cortical and subcortical regions associated with human neuropsychiatric disease. Gene expression changes more rapidly before birth, both in progenitor cells and maturing neurons. Cortical layers and areas acquire adult-like molecular profiles surprisingly late in postnatal development. Disparate cell populations exhibit distinct developmental timing of gene expression, but also unexpected synchrony of processes underlying neural circuit construction including cell projection and adhesion. Candidate risk genes for neurodevelopmental disorders including primary microcephaly, autism spectrum disorder, intellectual disability, and schizophrenia show disease-specific spatiotemporal enrichment within developing neocortex. Human developmental expression trajectories are more similar to monkey than rodent, although approximately 9% of genes show human-specific regulation with evidence for prolonged maturation or neoteny compared to monkey.

A mesoscale connectome of the mouse brain.

Comprehensive knowledge of the brain's wiring diagram is fundamental for understanding how the nervous system processes information at both local and global scales. However, with the singular exception of the C. elegans microscale connectome, there are no complete connectivity data sets in other species. Here we report a brain-wide, cellular-level, mesoscale connectome for the mouse. The Allen Mouse Brain Connectivity Atlas uses enhanced green fluorescent protein (EGFP)-expressing adeno-associated viral vectors to trace axonal projections from defined regions and cell types, and high-throughput serial two-photon tomography to image the EGFP-labelled axons throughout the brain. This systematic and standardized approach allows spatial registration of individual experiments into a common three dimensional (3D) reference space, resulting in a whole-brain connectivity matrix. A computational model yields insights into connectional strength distribution, symmetry and other network properties. Virtual tractography illustrates 3D topography among interconnected regions. Cortico-thalamic pathway analysis demonstrates segregation and integration of parallel pathways. The Allen Mouse Brain Connectivity Atlas is a freely available, foundational resource for structural and functional investigations into the neural circuits that support behavioural and cognitive processes in health and disease.

Transcriptional landscape of the prenatal human brain.

The anatomical and functional architecture of the human brain is mainly determined by prenatal transcriptional processes. We describe an anatomically comprehensive atlas of the mid-gestational human brain, including de novo reference atlases, in situ hybridization, ultra-high-resolution magnetic resonance imaging (MRI) and microarray analysis on highly discrete laser-microdissected brain regions. In developing cerebral cortex, transcriptional differences are found between different proliferative and post-mitotic layers, wherein laminar signatures reflect cellular composition and developmental processes. Cytoarchitectural differences between human and mouse have molecular correlates, including species differences in gene expression in subplate, although surprisingly we find minimal differences between the inner and outer subventricular zones even though the outer zone is expanded in humans. Both germinal and post-mitotic cortical layers exhibit fronto-temporal gradients, with particular enrichment in the frontal lobe. Finally, many neurodevelopmental disorder and human-evolution-related genes show patterned expression, potentially underlying unique features of human cortical formation. These data provide a rich, freely-accessible resource for understanding human brain development.

Traumatic dental injuries presenting to a paediatric emergency department in a tertiary children's hospital, Adelaide, Australia.

BACKGROUND/AIM: There is a scarcity of data regarding paediatric traumatic dental injuries (TDI) in tertiary teaching hospitals. Therefore, the aim of this study was to review the TDI presenting to the Women's and Children's Hospital in Adelaide (Australia) on an emergency basis and to identify the characteristics of the presenting patient cohort, their accident, injuries and management. METHODS: Medical health records of 337 paediatric patients attending the Paediatric Emergency Department (PED) for the management of TDI over 18 months were prospectively reviewed. RESULTS: TDI were more frequent in children under 5 years of age (56.1%) with a predominance of injuries sustained by males (63.8%). The accident characteristics included weekend occurrence (35.6%), the most common aetiology was falls (64.4%) and many incidents occurred at home (48.5%). Overall, 654 teeth were injured with the majority affecting deciduous teeth (58.4%) and the maxillary central incisors (69.9%). The most frequent injury was lateral luxation (27.5%). The majority of patients were referred to the Paediatric Dentistry Department (60.8%). However, almost half of presenting patients did not require further management locally and were subsequently discharged to their dental practitioners (39.2%). Most patients receiving treatment were managed under general anaesthetic (36.9%), and there was often a delay of 3-12 hours before treatment commenced (49.1%). Similarly, more severe injuries in the permanent dentition (avulsion, extrusion, root fracture, intrusion, alveolar fracture) were more frequently managed between 3 and 12 hours following the accident. CONCLUSION: The patient, accident, injury and management characteristics are comparable to what has previously been reported in other studies in paediatric populations. Injuries affecting the permanent dentition are more likely to be managed within 3 and 12 hours in an outpatient setting, whereas injuries affecting the deciduous dentition had a delay in management between 12 and 24 hours under general anaesthetic.

An anatomically comprehensive atlas of the adult human brain transcriptome.

Neuroanatomically precise, genome-wide maps of transcript distributions are critical resources to complement genomic sequence data and to correlate functional and genetic brain architecture. Here we describe the generation and analysis of a transcriptional atlas of the adult human brain, comprising extensive histological analysis and comprehensive microarray profiling of ∼900 neuroanatomically precise subdivisions in two individuals. Transcriptional regulation varies enormously by anatomical location, with different regions and their constituent cell types displaying robust molecular signatures that are highly conserved between individuals. Analysis of differential gene expression and gene co-expression relationships demonstrates that brain-wide variation strongly reflects the distributions of major cell classes such as neurons, oligodendrocytes, astrocytes and microglia. Local neighbourhood relationships between fine anatomical subdivisions are associated with discrete neuronal subtypes and genes involved with synaptic transmission. The neocortex displays a relatively homogeneous transcriptional pattern, but with distinct features associated selectively with primary sensorimotor cortices and with enriched frontal lobe expression. Notably, the spatial topography of the neocortex is strongly reflected in its molecular topography-the closer two cortical regions, the more similar their transcriptomes. This freely accessible online data resource forms a high-resolution transcriptional baseline for neurogenetic studies of normal and abnormal human brain function.

A genome-wide RNAi screen identifies multiple RSK-dependent regulators of cell migration.

To define the functional pathways regulating epithelial cell migration, we performed a genome-wide RNAi screen using 55,000 pooled lentiviral shRNAs targeting ∼11,000 genes, selecting for transduced cells with increased motility. A stringent validation protocol generated a set of 31 genes representing diverse pathways whose knockdown dramatically enhances cellular migration. Some of these pathways share features of epithelial-to-mesenchymal transition (EMT), and together they implicate key regulators of transcription, cellular signaling, and metabolism, as well as novel modulators of cellular trafficking, such as DLG5. In delineating downstream pathways mediating these migration phenotypes, we observed universal activation of ERKs and a profound dependence on their RSK effectors. Pharmacological inhibition of RSK dramatically suppresses epithelial cell migration induced by knockdown of all 31 genes, suggesting that convergence of diverse migratory pathways on this kinase may provide a therapeutic opportunity in disorders of cell migration, including cancer metastasis.

Cell-type-based model explaining coexpression patterns of genes in the brain.

Spatial patterns of gene expression in the vertebrate brain are not independent, as pairs of genes can exhibit complex patterns of coexpression. Two genes may be similarly expressed in one region, but differentially expressed in other regions. These correlations have been studied quantitatively, particularly for the Allen Atlas of the adult mouse brain, but their biological meaning remains obscure. We propose a simple model of the coexpression patterns in terms of spatial distributions of underlying cell types and establish its plausibility using independently measured cell-type-specific transcriptomes. The model allows us to predict the spatial distribution of cell types in the mouse brain.

Exploration and visualization of connectivity in the adult mouse brain.

The Allen Mouse Brain Connectivity Atlas is a mesoscale whole brain axonal projection atlas of the C57Bl/6J mouse brain. All data were aligned to a common template in 3D space to generate a comprehensive and quantitative database of inter-areal and cell-type-specific projections. A suite of computational tools were developed to search and visualize the projection labeling experiments, available at http://connectivity.brain-map.org. We present three use cases illustrating how these publicly-available tools can be used to perform analyses of long range brain region connectivity. The use cases make extensive use of advanced visualization tools integrated with the atlas including projection density histograms, 3D computed anterograde and retrograde projection paths, and multi-specimen projection composites. These tools offer convenient access to detailed axonal projection information in the adult mouse brain and the ability to perform data analysis and visualization of projection fields and neuroanatomy in an integrated manner.

Neuroinformatics of the Allen Mouse Brain Connectivity Atlas.

The Allen Mouse Brain Connectivity Atlas is a mesoscale whole brain axonal projection atlas of the C57Bl/6J mouse brain. Anatomical trajectories throughout the brain were mapped into a common 3D space using a standardized platform to generate a comprehensive and quantitative database of inter-areal and cell-type-specific projections. This connectivity atlas has several desirable features, including brain-wide coverage, validated and versatile experimental techniques, a single standardized data format, a quantifiable and integrated neuroinformatics resource, and an open-access public online database (http://connectivity.brain-map.org/). Meaningful informatics data quantification and comparison is key to effective use and interpretation of connectome data. This relies on successful definition of a high fidelity atlas template and framework, mapping precision of raw data sets into the 3D reference framework, accurate signal detection and quantitative connection strength algorithms, and effective presentation in an integrated online application. Here we describe key informatics pipeline steps in the creation of the Allen Mouse Brain Connectivity Atlas and include basic application use cases.

Allen Brain Atlas: an integrated spatio-temporal portal for exploring the central nervous system.

The Allen Brain Atlas (http://www.brain-map.org) provides a unique online public resource integrating extensive gene expression data, connectivity data and neuroanatomical information with powerful search and viewing tools for the adult and developing brain in mouse, human and non-human primate. Here, we review the resources available at the Allen Brain Atlas, describing each product and data type [such as in situ hybridization (ISH) and supporting histology, microarray, RNA sequencing, reference atlases, projection mapping and magnetic resonance imaging]. In addition, standardized and unique features in the web applications are described that enable users to search and mine the various data sets. Features include both simple and sophisticated methods for gene searches, colorimetric and fluorescent ISH image viewers, graphical displays of ISH, microarray and RNA sequencing data, Brain Explorer software for 3D navigation of anatomy and gene expression, and an interactive reference atlas viewer. In addition, cross data set searches enable users to query multiple Allen Brain Atlas data sets simultaneously. All of the Allen Brain Atlas resources can be accessed through the Allen Brain Atlas data portal.

Data-driven fine-grained region discovery in the mouse brain with transformers.

Technologies such as spatial transcriptomics offer unique opportunities to define the spatial organization of the mouse brain. We developed an unsupervised training scheme and novel transformer-based deep learning architecture to detect spatial domains across the whole mouse brain using spatial transcriptomics data. Our model learns local representations of molecular and cellular statistical patterns which can be clustered to identify spatial domains within the brain from coarse to fine-grained. Discovered domains are spatially regular, even with several hundreds of spatial clusters. They are also consistent with existing anatomical ontologies such as the Allen Mouse Brain Common Coordinate Framework version 3 (CCFv3) and can be visually interpreted at the cell type or transcript level. We demonstrate our method can be used to identify previously uncatalogued subregions, such as in the midbrain, where we uncover gradients of inhibitory neuron complexity and abundance. Notably, these subregions cannot be discovered using other methods. We apply our method to a separate multi-animal whole-brain spatial transcriptomic dataset and show that our method can also robustly integrate spatial domains across animals.

The ANTsX Ecosystem for Mapping the Mouse Brain.

Precision mapping techniques coupled with high resolution image acquisition of the mouse brain permit the study of the spatial organization of gene expression and their mutual interaction for a comprehensive view of salient structural/functional relationships. Such research is facilitated by standardized anatomical coordinate systems, such as the well-known Allen Common Coordinate Framework (AllenCCFv3), and the ability to spatially map to such standardized spaces. The Advanced Normalization Tools Ecosystem is a comprehensive open-source software toolkit for generalized quantitative imaging with applicability to multiple organ systems, modalities, and animal species. Herein, we illustrate the utility of ANTsX for generating precision spatial mappings of the mouse brain and potential subsequent quantitation. We describe ANTsX-based workflows for mapping domain-specific image data to AllenCCFv3 accounting for common artefacts and other confounds. Novel contributions include ANTsX functionality for velocity flow-based mapping spanning the spatiotemporal domain of a longitudinal trajectory which we apply to the Developmental Common Coordinate Framework. Additionally, we present an automated structural morphological pipeline for determining volumetric and cortical thickness measurements analogous to the well-utilized ANTsX pipeline for human neuroanatomical structural morphology which illustrates a general open-source framework for tailored brain parcellations.

Cross-modality mapping using image varifolds to align tissue-scale atlases to molecular-scale measures with application to 2D brain sections.

This paper explicates a solution to building correspondences between molecular-scale transcriptomics and tissue-scale atlases. This problem arises in atlas construction and cross-specimen/technology alignment where specimens per emerging technology remain sparse and conventional image representations cannot efficiently model the high dimensions from subcellular detection of thousands of genes. We address these challenges by representing spatial transcriptomics data as generalized functions encoding position and high-dimensional feature (gene, cell type) identity. We map onto low-dimensional atlas ontologies by modeling regions as homogeneous random fields with unknown transcriptomic feature distribution. We solve simultaneously for the minimizing geodesic diffeomorphism of coordinates through LDDMM and for these latent feature densities. We map tissue-scale mouse brain atlases to gene-based and cell-based transcriptomics data from MERFISH and BARseq technologies and to histopathology and cross-species atlases to illustrate integration of diverse molecular and cellular datasets into a single coordinate system as a means of comparison and further atlas construction.

The transcriptomic and spatial organization of telencephalic GABAergic neuronal types.

The telencephalon of the mammalian brain comprises multiple regions and circuit pathways that play adaptive and integrative roles in a variety of brain functions. There is a wide array of GABAergic neurons in the telencephalon; they play a multitude of circuit functions, and dysfunction of these neurons has been implicated in diverse brain disorders. In this study, we conducted a systematic and in-depth analysis of the transcriptomic and spatial organization of GABAergic neuronal types in all regions of the mouse telencephalon and their developmental origins. This was accomplished by utilizing 611,423 single-cell transcriptomes from the comprehensive and high-resolution transcriptomic and spatial cell type atlas for the adult whole mouse brain we have generated, supplemented with an additional single-cell RNA-sequencing dataset containing 99,438 high-quality single-cell transcriptomes collected from the pre- and postnatal developing mouse brain. We present a hierarchically organized adult telencephalic GABAergic neuronal cell type taxonomy of 7 classes, 52 subclasses, 284 supertypes, and 1,051 clusters, as well as a corresponding developmental taxonomy of 450 clusters across different ages. Detailed charting efforts reveal extraordinary complexity where relationships among cell types reflect both spatial locations and developmental origins. Transcriptomically and developmentally related cell types can often be found in distant and diverse brain regions indicating that long-distance migration and dispersion is a common characteristic of nearly all classes of telencephalic GABAergic neurons. Additionally, we find various spatial dimensions of both discrete and continuous variations among related cell types that are correlated with gene expression gradients. Lastly, we find that cortical, striatal and some pallidal GABAergic neurons undergo extensive postnatal diversification, whereas septal and most pallidal GABAergic neuronal types emerge simultaneously during the embryonic stage with limited postnatal diversification. Overall, the telencephalic GABAergic cell type taxonomy can serve as a foundational reference for molecular, structural and functional studies of cell types and circuits by the entire community.