Project description:Dendritic and axonal morphology reflects the input and output of neurons and is a defining feature of neuronal types, yet our knowledge of the diversity of neuronal morphology, in particular distal axon projection patterns, remains limited. To systematically obtain complete single neuron morphology on a brain-wide scale, we established a platform with five major components: sparse labeling, whole-brain imaging, reconstruction, registration, and classification. We fully reconstructed 1,741 neurons from cortex, claustrum, thalamus, striatum and other brain regions. We identify 11 major projection neuron types with distinct morphological features and corresponding transcriptomic identities. We further reveal extensive morphological diversity within each of these major types, some of which cluster into more refined morphological subtypes. We analyze this diversity at different levels following a set of generalizable organizational rules governing long-range axonal projections, including molecular correspondence, divergent or convergent projection, axon termination pattern, regional specificity, topography, and individual cell variability. We illustrate how these rules manifest in different projection neuron types. Although clear concordance with transcriptomic profiles is evident at major projection type level, fine-grained morphological diversity often does not readily correlate with transcriptomic subtypes derived from unsupervised clustering, highlighting the need for single-cell level cross-modality studies. Overall, our study provides a systematic demonstration of the crucial need for quantitative description of complete single cell anatomy in cell type classification, as the cell type-specific morphological diversity reveals a plethora of ways different cell types and individual neurons may contribute to the function of their respective circuits.
Project description:To validate different projection targets of already molecularly-defined olfactory bulb projection neurons we used viral targeting specifically into anterior or posterior cortical areas, Fluorescence Activated Nuclei Sorting (FANS) to enrich for olfactory bulb projection neurons, and single-nuclei RNA sequencing (sn-RNA seq) To isolate GFP-labelled nuclei, 1 individual replicate of AON or PCx-injected mice was used. Ipsilateral and controlateral sides were minced separately and placed into two different tubes. The minced tissue was gently homogenized in Nuclei PURE Lysis Buffer and 10% Triton X-100 using an ice-cold dounce and pestle, and filtered two times through a 40 μm cell strainer on ice. After centrifuging at 500 rpm for 5 min at 4 °C, the supernatant was aspirated and gently resuspended in 500 μl of cold buffer (1x of cold Hanks' Balanced Salt Solution HBSS, 1% nuclease-free BSA, RNasin Plus and 1/2000 DRAQ5). Our study identifies molecularly distinct subtypes of mitral cells projecting to anterior or posterior olfactory cortices.
Project description:We characterize the transcriptomic, morphological and functional features of 472 high-quality RGCs using Patch-seq, providing functional and morphological annotation of many transcriptomic defined cell types of previously established RGC atlas.
Project description:Droplet-based single cell RNA sequencing (scRNA-seq) to classify molecularly distinct neuronal and non-neuronal cell types in the mouse ventral posterior hypothalamus. Cluster analysis of >16,000 single cells revealed 20 neuronal and 18 non-neuronal cell populations, defined by suites of discriminatory markers. We validated differentially expressed genes in a selection of neuronal populations through fluorescence in situ hybridization (FISH). Focusing on the mammillary nuclei, we discovered transcriptionally-distinct clusters that broadly align with neuroanatomical compartments. This single cell transcriptomic analysis of cell types in the VPH provides a resource for interrogating the circuit-level mechanisms underlying the diverse functions of VPH circuits in health and disease.
Project description:The striatum is the interface between dopamine reward signals and cortico-basal ganglia circuits that mediate diverse behavioral functions. Medium spiny neurons (MSNs) constitute the vast majority of striatal neurons and are traditionally classified as direct- or indirect-pathway neurons. However, that traditional model does not explain the anatomical and functional diversity of MSNs. Here, we defined molecularly distinct MSN types in the primate striatum, including (1) dorsal striatum MSN types associated with striosome and matrix compartments, (2) ventral striatum types associated with the nucleus accumbens shell and olfactory tubercle, and (3) an MSN-like type restricted to mu-opioid receptor rich islands in the ventral striatum. These results lay the foundation for achieving cell type-specific transgenesis in the primate striatum and provide a blueprint for investigating circuit-specific processing.