Fast, high-throughput production of improved rabies viral vectors for specific, efficient and versatile transsynaptic retrograde labeling

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Abstract

To understand the function of neuronal circuits it is crucial to disentangle the connectivity patterns within the network. However, most tools currently used to explore connectivity have low throughput, low selectivity, or limited accessibility. Here, we report the development of an improved packaging system for the production of the highly neurotropic RVdG_envA-CVS-N2c rabies viral vectors, yielding titers orders of magnitude higher with no background contamination, at a fraction of the production time, while preserving the efficiency of transsynaptic labeling. Along with the production pipeline, we developed suites of 'starter' AAV and bicistronic RVdG-CVS-N2c vectors, enabling retrograde labeling from a wide range of neuronal populations, tailored for diverse experimental requirements. We demonstrate the power and flexibility of the new system by uncovering hidden local and distal inhibitory connections in the mouse hippocampal formation and by imaging the functional properties of a cortical microcircuit across weeks. Our novel production pipeline provides a convenient approach to generate new rabies vectors, while our toolkit flexibly and efficiently expands the current capacity to label, manipulate and image the neuronal activity of interconnected neuronal circuits in vitro and in vivo.

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All source data associated with the manuscript has been included in the manuscript.

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Anton Sumser

Institute of Science and Technology Austria, Klosterneuburg, Austria

Competing interests
The authors declare that no competing interests exist.

"This ORCID iD identifies the author of this article:" 0000-0002-4792-1881
Maximilian Joesch

Institute of Science and Technology Austria, Klosterneuburg, Austria

Competing interests
The authors declare that no competing interests exist.
Peter Jonas

Institute of Science and Technology Austria, Klosterneuburg, Austria

Competing interests
The authors declare that no competing interests exist.

"This ORCID iD identifies the author of this article:" 0000-0001-5001-4804
Yoav Ben-Simon

Department of Neurophysiology and Neuropharmacology, Vienna Medical University, Vienna, Austria

For correspondence
yoav.bensimon@meduniwien.ac.at

Competing interests
The authors declare that no competing interests exist.

"This ORCID iD identifies the author of this article:" 0000-0002-7075-097X

Funding

Horizon 2020 Framework Programme (692692)

Peter Jonas

Austrian Science Fund (Z 312-B27)

Peter Jonas

The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.

Ethics

Animal experimentation: Experiments on C57BL/6 wild-type and transgenic mice were performed in strict accordance with institutional, national, and European guidelines for animal experimentation and were approved by the Bundesministerium für Wissenschaft, Forschung und Wirtschaft and Bildung, Wissenschaft und Forschung, respectively, of Austria (A. Haslinger, Vienna; BMWF-66.018/0010-WF/V/3b/2015; BMBWF-66.018/0008-WF/V/3b/2018).

Copyright

This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.