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Development of an in vivo adeno-associated virus-mediated siRNA approach to knockdown tyrosine hydroxylase in the lateral retrochiasmatic area of the ovine brain

Abstract : We developed a new technique of gene knockdown (KD) in a specific brain area of the ewe using an adeno-associated virus (AAV)-mediated short interfering RNA (siRNA) method to elucidate the importance of key factors of seasonal reproduction. Two 19-nucleotide sequences (TH1 or TH2) were chosen from the tyrosine hydroxylase (TH) gene. TH1, TH2 or a random sequence (TH3) was incorporated into an eGFP expressing AAV vector. Firstly, 5 mu l of AAV-TH1 or AAV-TH2 solutions (8-9 x 10(11) Vg/ml) were stereotaxically injected into one A 15 nucleus while the other received a control treatment. Ewes were killed after 15 or 75 days. The number of TH neurons was 49% and 36% lower on the AAV-TH1 treated side than on the control side 15 and 75 days post-injection, respectively. AAV-TH2 did not induce a significant variation in TH cell population. Finally, in order to increase the KD, two groups of ewes received 10 mu l of AAV-TH1 either in a bolus injection or in two 5 mu l inoculations carried out 2 weeks apart. Only ewes receiving a bolus injection showed a larger KD reaching 66% 2 months after inoculation. This method proved effective in reducing TH expression and will be further developed to understand cellular mechanisms driving seasonal functions. (c) 2008 Elsevier B.V. All rights reserved.
Mots-clés : REPRODUCTION SAISONNEE
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https://hal.inrae.fr/hal-02662545
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Submitted on : Sunday, May 31, 2020 - 12:32:04 AM
Last modification on : Tuesday, November 2, 2021 - 9:18:01 AM

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Laurence Dufourny, Martine Migaud, Jean-Claude Thiéry, Benoit Malpaux. Development of an in vivo adeno-associated virus-mediated siRNA approach to knockdown tyrosine hydroxylase in the lateral retrochiasmatic area of the ovine brain. Journal of Neuroscience Methods, Elsevier, 2008, 170 (1), pp.56-66. ⟨10.1016/j.jneumeth.2007.12.018⟩. ⟨hal-02662545⟩

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