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Leaky expression of channelrhodopsin-2 (ChR2) in Ai32 mouse lines

Optogenetics enables the selective activation of genetically-targeted neuronal populations using light-sensitive ion channels. Genetic strategies using Cre-dependent mouse strains, especially the Ai32 line expressing Channelrhodopsin (ChR2)-EYFP fusion protein, have been a popular means to drive ops...

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Autores principales: Prabhakar, Arthi, Vujovic, Dragan, Cui, Lian, Olson, William, Luo, Wenqin
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Public Library of Science 2019
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6435231/
https://www.ncbi.nlm.nih.gov/pubmed/30913225
http://dx.doi.org/10.1371/journal.pone.0213326
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author Prabhakar, Arthi
Vujovic, Dragan
Cui, Lian
Olson, William
Luo, Wenqin
author_facet Prabhakar, Arthi
Vujovic, Dragan
Cui, Lian
Olson, William
Luo, Wenqin
author_sort Prabhakar, Arthi
collection PubMed
description Optogenetics enables the selective activation of genetically-targeted neuronal populations using light-sensitive ion channels. Genetic strategies using Cre-dependent mouse strains, especially the Ai32 line expressing Channelrhodopsin (ChR2)-EYFP fusion protein, have been a popular means to drive opsin expression in a cell-type specific manner. Here we report a low level of leaky ‘off-target’ (Cre-independent) ChR2-EYFP expression in Ai32/Ai32 homozygous mice throughout the nervous system. This leaky off-target expression was characterized in multiple prevalent nervous system regions using anti-EYFP immunostaining. Expression of full-length ChR2-EYFP protein was confirmed using immunoprecipitation followed by Western blotting. Notably, light stimulation of these ChR2-EYFP expressing neurons in the spinal cord dorsal horn did not induce detectable photocurrents in juvenile 4-week old mice. Given the wide use of the Ai32 line by many labs, our results suggest researchers should be vigilant of possible off-target ChR2-EYFP expression in their region of interest, especially when generating Ai32/Ai32 homozygotes to drive high levels of ChR2-EYFP expression in adult mice.
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spelling pubmed-64352312019-04-08 Leaky expression of channelrhodopsin-2 (ChR2) in Ai32 mouse lines Prabhakar, Arthi Vujovic, Dragan Cui, Lian Olson, William Luo, Wenqin PLoS One Research Article Optogenetics enables the selective activation of genetically-targeted neuronal populations using light-sensitive ion channels. Genetic strategies using Cre-dependent mouse strains, especially the Ai32 line expressing Channelrhodopsin (ChR2)-EYFP fusion protein, have been a popular means to drive opsin expression in a cell-type specific manner. Here we report a low level of leaky ‘off-target’ (Cre-independent) ChR2-EYFP expression in Ai32/Ai32 homozygous mice throughout the nervous system. This leaky off-target expression was characterized in multiple prevalent nervous system regions using anti-EYFP immunostaining. Expression of full-length ChR2-EYFP protein was confirmed using immunoprecipitation followed by Western blotting. Notably, light stimulation of these ChR2-EYFP expressing neurons in the spinal cord dorsal horn did not induce detectable photocurrents in juvenile 4-week old mice. Given the wide use of the Ai32 line by many labs, our results suggest researchers should be vigilant of possible off-target ChR2-EYFP expression in their region of interest, especially when generating Ai32/Ai32 homozygotes to drive high levels of ChR2-EYFP expression in adult mice. Public Library of Science 2019-03-26 /pmc/articles/PMC6435231/ /pubmed/30913225 http://dx.doi.org/10.1371/journal.pone.0213326 Text en © 2019 Prabhakar et al http://creativecommons.org/licenses/by/4.0/ This is an open access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/4.0/) , which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.
spellingShingle Research Article
Prabhakar, Arthi
Vujovic, Dragan
Cui, Lian
Olson, William
Luo, Wenqin
Leaky expression of channelrhodopsin-2 (ChR2) in Ai32 mouse lines
title Leaky expression of channelrhodopsin-2 (ChR2) in Ai32 mouse lines
title_full Leaky expression of channelrhodopsin-2 (ChR2) in Ai32 mouse lines
title_fullStr Leaky expression of channelrhodopsin-2 (ChR2) in Ai32 mouse lines
title_full_unstemmed Leaky expression of channelrhodopsin-2 (ChR2) in Ai32 mouse lines
title_short Leaky expression of channelrhodopsin-2 (ChR2) in Ai32 mouse lines
title_sort leaky expression of channelrhodopsin-2 (chr2) in ai32 mouse lines
topic Research Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6435231/
https://www.ncbi.nlm.nih.gov/pubmed/30913225
http://dx.doi.org/10.1371/journal.pone.0213326
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