Cell-type-specific and projection-specific brain-wide reconstruction of single neurons

We developed a dual-adeno-associated-virus expression system that enables strong and sparse labeling of individual neurons with cell-type and projection specificity. We demonstrated its utility for whole-brain reconstruction of midbrain dopamine neurons and striatum-projecting cortical neurons. We further extended the labeling method for rapid reconstruction in cleared thick brain sections and simultaneous dual-color labeling. This labeling system may facilitate the process of generating mesoscale single-neuron projectomes of mammalian brains.An AAV-based platform achieves sparse yet bright labeling of neurons with cell-type specificity. This technology will facilitate the reconstruction of neurons in the mouse brain.

[1]  Kristina J. Nielsen,et al.  Targeting Single Neuronal Networks for Gene Expression and Cell Labeling In Vivo , 2010, Neuron.

[2]  P. Osten,et al.  Mapping brain circuitry with a light microscope , 2013, Nature Methods.

[3]  Jing Yuan,et al.  TDat: An Efficient Platform for Processing Petabyte-Scale Whole-Brain Volumetric Images , 2017, Front. Neural Circuits.

[4]  Shaoqun Zeng,et al.  Embedding and Chemical Reactivation of Green Fluorescent Protein in the Whole Mouse Brain for Optical Micro-Imaging , 2017, Front. Neurosci..

[5]  C. S. Raymond,et al.  High-Efficiency FLP and ΦC31 Site-Specific Recombination in Mammalian Cells , 2007, PloS one.

[6]  Swapnil Bhatia,et al.  A single-layer platform for Boolean logic and arithmetic through DNA excision in mammalian cells , 2017, Nature Biotechnology.

[7]  J. Tepper,et al.  Heterogeneity and Diversity of Striatal GABAergic Interneurons , 2010, Front. Neuroanat..

[8]  Kenneth D Harris,et al.  A genuine layer 4 in motor cortex with prototypical synaptic circuit connectivity , 2014, eLife.

[9]  N. Plesnila,et al.  Shrinkage-mediated imaging of entire organs and organisms using uDISCO , 2016, Nature Methods.

[10]  W Ding,et al.  Automatic macroscopic density artefact removal in a Nissl‐stained microscopic atlas of whole mouse brain , 2013, Journal of microscopy.

[11]  D. G. Gibson,et al.  Enzymatic assembly of DNA molecules up to several hundred kilobases , 2009, Nature Methods.

[12]  Andreas T Schaefer,et al.  Transfection via whole-cell recording in vivo: bridging single-cell physiology, genetics and connectomics , 2011, Nature Neuroscience.

[13]  K. Svoboda,et al.  Genetic Dissection of Neural Circuits: A Decade of Progress , 2018, Neuron.

[14]  M. Nakayama,et al.  Recombinase-mediated cassette exchange (RMCE) and BAC engineering via VCre/VloxP and SCre/SloxP systems. , 2011, BioTechniques.

[15]  Karel Svoboda,et al.  The Past, Present, and Future of Single Neuron Reconstruction , 2011, Neuroinformatics.

[16]  Marten Postma,et al.  mScarlet: a bright monomeric red fluorescent protein for cellular imaging , 2016, Nature Methods.

[17]  Edward M. Callaway,et al.  Genetic Dissection of Neural Circuits: A Decade of Progress. , 2018, Neuron.

[18]  T. Schlake,et al.  Use of mutated FLP recognition target (FRT) sites for the exchange of expression cassettes at defined chromosomal loci. , 1994, Biochemistry.

[19]  W. Denk,et al.  Targeted patch-clamp recordings and single-cell electroporation of unlabeled neurons in vivo , 2008, Nature Methods.

[20]  Y. Yoshimura,et al.  NMDAR-Regulated Dynamics of Layer 4 Neuronal Dendrites during Thalamocortical Reorganization in Neonates , 2014, Neuron.

[21]  Concha Bielza,et al.  New insights into the classification and nomenclature of cortical GABAergic interneurons , 2013, Nature Reviews Neuroscience.

[22]  Shaoqun Zeng,et al.  High-throughput dual-colour precision imaging for brain-wide connectome with cytoarchitectonic landmarks at the cellular level , 2016, Nature Communications.

[23]  D. Duan,et al.  Recombinant adeno-associated viral vector production and purification. , 2012, Methods in molecular biology.

[24]  E. Suzuki,et al.  VCre/VloxP and SCre/SloxP: new site-specific recombination systems for genome engineering , 2011, Nucleic acids research.

[25]  Cheuk Y. Tang,et al.  Mapping of Brain Activity by Automated Volume Analysis of Immediate Early Genes , 2016, Cell.

[26]  L. Looger,et al.  A Designer AAV Variant Permits Efficient Retrograde Access to Projection Neurons , 2016, Neuron.

[27]  Lief E. Fenno,et al.  Targeting cells with single vectors using multiple-feature Boolean logic , 2014, Nature Methods.

[28]  B. Roska,et al.  Virus stamping for targeted single-cell infection in vitro and in vivo , 2017, Nature Biotechnology.

[29]  Ian R. Wickersham,et al.  Cortical representations of olfactory input by trans-synaptic tracing , 2011, Nature.

[30]  Shaoqun Zeng,et al.  Rapid imaging of large tissues using high-resolution stage-scanning microscopy. , 2015, Biomedical optics express.

[31]  J. DeFelipe,et al.  The pyramidal neuron of the cerebral cortex: Morphological and chemical characteristics of the synaptic inputs , 1992, Progress in Neurobiology.

[32]  G. Shepherd.  Corticostriatal connectivity and its role in disease , 2013, Nature Reviews Neuroscience.

[33]  Minmin Luo,et al.  Whole-brain mapping of the direct inputs and axonal projections of POMC and AgRP neurons , 2015, Front. Neuroanat..

[34]  Jeff W. Lichtman,et al.  Clarifying Tissue Clearing , 2015, Cell.

[35]  S. Itohara,et al.  Supernova: A Versatile Vector System for Single-Cell Labeling and Gene Function Studies in vivo , 2016, Scientific Reports.