Substantially improved gene transfer to interneurons with second-generation 
glutamate receptor-targeted DART-AAV vectors

Günther, D.M.; Kovacs, R.; Wildner, F.; Salivara, A.; Thalheimer, F.B.; Fries, Pascal; Geiger, J.R.P.; Buchholz, C.J.

doi:10.1016/j.jneumeth.2023.109981

Item

ITEM ACTIONSEXPORT

Add to Basket

Local TagsRelease HistoryDetailsSummary

Released

Journal Article

Substantially improved gene transfer to interneurons with second-generation glutamate receptor-targeted DART-AAV vectors

MPS-Authors

Günther, D.M.
Ernst Strüngmann Institute (ESI) for Neuroscience in Cooperation with Max Planck Society, Max Planck Society;
Fries Lab, Ernst Strüngmann Institute (ESI) for Neuroscience in Cooperation with Max Planck Society, Max Planck Society;

/persons/resource/persons141609

Fries, Pascal
Ernst Strüngmann Institute (ESI) for Neuroscience in Cooperation with Max Planck Society, Max Planck Society;
Fries Lab, Ernst Strüngmann Institute (ESI) for Neuroscience in Cooperation with Max Planck Society, Max Planck Society;

External Resource

https://www.sciencedirect.com/science/article/pii/S0165027023002005
(Publisher version)

Fulltext (restricted access)

There are currently no full texts shared for your IP range.

Fulltext (public)

Guenther_2023_SubstantiallyImproved.pdf
(Publisher version), 6MB

Supplementary Material (public)

There is no public supplementary material available

Citation

Günther, D., Kovacs, R., Wildner, F., Salivara, A., Thalheimer, F., Fries, P., et al. (2023). Substantially improved gene transfer to interneurons with second-generation glutamate receptor-targeted DART-AAV vectors. Journal of Neuroscience Methods, 399: 109981. doi:10.1016/j.jneumeth.2023.109981.

Cite as: https://hdl.handle.net/21.11116/0000-000D-ECBE-4

Abstract

Background
Adeno-associated viral vectors (AAVs) are a widely used gene transfer platform in neuroscience. Although naturally AAV serotypes can have preferences for certain tissues, selectivity for particular cell types in the CNS does not exist. Towards interneuron targeting, capsid engineering of AAV2 including display of the designed ankyrin repeat protein (DARPin) 2K19 specific for the glutamate receptor subunit 4 (GluA4) at the N-terminus of the VP2 capsid protein has been established. The resulting AAV-VP2N is highly specific for interneurons, but exhibits rather moderate transduction efficiencies.

Methods
Two alternative insertion sites for 2K19 in the GH2/GH3 loop of capsid proteins VP1 (AAV-VP1L) or VP2 (AAV-VP2L) were exploited to yield second generation GluA4-AAVs. Having packaged reporter genes under ubiquitous promoters, the vectors were characterized for biochemical properties as well as gene delivery into cell lines and rat hippocampal slice cultures. Electrophysiological recordings monitored the functional properties of transduced cells.

Results
Compared to AAV-VP2N, the second-generation vectors, especially AAV-VP1L, achieved about 2-fold higher genomic titers as well as a substantially improved GluA4 binding. Improvements in gene transfer activities were 18-fold on GluA4-overexpressing A549 cells and five-fold on rat hippocampal organotypic slice cultures reaching approximately 60 % of all parvalbumin positive interneurons upon a single administration. The spiking behaviour of transduced cells was unaltered and characteristic for a heterogeneous group of interneurons.

Conclusion
The substantially improved gene transfer activity of the second generation GluA4-targeted AAV combined with low toxicity makes this vector an attractive tool for interneuron-directed gene transfer with unrestricted promotor and transgene choice.