2. Academic Validation
  3. High-Complexity Barcoded Rabies Virus for Scalable Circuit Mapping Using Single-Cell and Single-Nucleus Sequencing

High-Complexity Barcoded Rabies Virus for Scalable Circuit Mapping Using Single-Cell and Single-Nucleus Sequencing

  • bioRxiv. 2024 Dec 11:2024.10.01.616167. doi: 10.1101/2024.10.01.616167.
David Shin 1 2,  Madeleine E Urbanek 1 2,  H Hanh Larson 2,  Anthony J Moussa 3,  Kevin Y Lee 2,  Donovan L Baker 2,  Elio Standen-Bloom 2,  Sangeetha Ramachandran 1 2,  Derek Bogdanoff 4,  Cathryn R Cadwell 2 5 6 7 8,  Tomasz J Nowakowski 2 7 9 10 11
Affiliations

Affiliations

  • 1 Biomedical Sciences Graduate Program, University of California, San Francisco, CA, USA.
  • 2 Department of Neurological Surgery, University of California, San Francisco, CA, USA.
  • 3 Medical Scientist Training Program, University of California, San Francisco, CA, USA.
  • 4 Tetrad Graduate Program, University of California, San Francisco, CA, USA.
  • 5 Department of Pathology, University of California, San Francisco, CA, USA.
  • 6 Weill Neurohub, University of California, San Francisco, CA, USA.
  • 7 Weill Institute for Neurosciences, University of California, San Francisco, CA, USA.
  • 8 Kavli Institute for Fundamental Neuroscience, University of California, San Francisco, CA, USA.
  • 9 Department of Anatomy, University of California, San Francisco, CA, USA.
  • 10 Department of Psychiatry and Behavioral Sciences, University of California, San Francisco, CA, USA.
  • 11 Eli and Edythe Broad Center for Regeneration Medicine and Stem Cell Research, University of California, San Francisco, CA, USA.
PMID: 39713304 DOI: 10.1101/2024.10.01.616167
Abstract

Single cell genomics has revolutionized our understanding of neuronal cell types. However, scalable technologies for probing single-cell connectivity are lacking, and we are just beginning to understand how molecularly defined cell types are organized into functional circuits. Here, we describe a protocol to generate high-complexity barcoded rabies virus (RV) for scalable circuit mapping from tens of thousands of individual starter cells in parallel. In addition, we introduce a strategy for targeting RV-encoded barcode transcripts to the nucleus so that they can be read out using single-nucleus RNA Sequencing (snRNA-seq). We apply this tool in organotypic slice cultures of the developing human cerebral cortex, which reveals the emergence of cell type-specific circuit motifs in midgestation. By leveraging the power and throughput of single cell genomics for mapping synaptic connectivity, we chart a path forward for scalable circuit mapping of molecularly-defined cell types in healthy and disease states.

Keywords

Barcoded rabies virus; barcode complexity; barcode diversity; brain development; cortex; human; single-cell RNA sequencing; single-nucleus RNA sequencing; subplate.

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