Becalick, Alexander;
(2025)
High throughput molecular connectomics with barcoded rabies viruses.
Doctoral thesis (Ph.D), UCL (University College London).
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Abstract
Neural circuits are comprised of diverse neuronal cell types with distinct connectivity motifs. Knowledge of this connectivity is crucial for furthering our understanding of circuit function. However, the throughput of conventional methodologies for studying the synaptic connectivity of individual neurons is limited. This prevents the ability to practically perform comparative connectomic studies or to systematically determine wiring patterns between the many different cell types. In this thesis, I describe the application of barcoded rabies viruses for mapping these connections in the mouse brain. Methods relying on DNA sequencing of oligonucleotide ``barcodes” have previously been used to uniquely label large populations of neurons. To this end, I have generated libraries of rabies viruses containing barcode sequences with sufficient diversity to uniquely label populations of over 1000 starter cells in a single experiment. Taking advantage of the ability of rabies virus to spread transsynaptically between connected neurons, infected starter cells can then transmit their barcodes to their direct presynaptic partners. Presynaptic barcodes are then matched to their respective starter cells to reconstruct putative synaptic connections. To preserve information about the spatial location of barcoded cells and avoid cell loss resulting from tissue dissociation, I have used in situ sequencing to read out viral barcodes alongside endogenous transcripts of cell-type marker genes from sections of mouse brain. This has enabled me to relate the molecular identity of barcoded neurons with their connectivity. I then explore the feasibility of using this technique to determine the connectivity of neurons in the mouse visual cortex.
| Type: | Thesis (Doctoral) |
|---|---|
| Qualification: | Ph.D |
| Title: | High throughput molecular connectomics with barcoded rabies viruses |
| Open access status: | An open access version is available from UCL Discovery |
| Language: | English |
| Additional information: | Copyright © The Author 2025. Original content in this thesis is licensed under the terms of the Creative Commons Attribution-NonCommercial 4.0 International (CC BY-NC 4.0) Licence (https://creativecommons.org/licenses/by-nc/4.0/). Any third-party copyright material present remains the property of its respective owner(s) and is licensed under its existing terms. Access may initially be restricted at the author’s request. |
| Keywords: | Connectomics, Spatial transcriptomics, Barcoded rabies virus |
| UCL classification: | UCL UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences > Faculty of Life Sciences UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences > Faculty of Life Sciences > Div of Biosciences |
| URI: | https://discovery.ucl.ac.uk/id/eprint/10210909 |
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