Development of an in vivo adeno-associated virus-mediated siRNA approach to knockdown tyrosine hydroxylase in the lateral retrochiasmatic area of the ovine brain - INRAE - Institut national de recherche pour l’agriculture, l’alimentation et l’environnement Accéder directement au contenu
Article Dans Une Revue Journal of Neuroscience Methods Année : 2008

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

Résumé

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.

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Dates et versions

hal-02662545 , version 1 (31-05-2020)

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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, 2008, 170 (1), pp.56-66. ⟨10.1016/j.jneumeth.2007.12.018⟩. ⟨hal-02662545⟩
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