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Neuronal tropism of transgene expression in rat PVN mediated by an AAV2 vector containing the CBA promoter
Author(s) -
Gao Yongxin,
Li Hongwei,
Sumners Colin
Publication year - 2006
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.20.4.a408-a
Subject(s) - green fluorescent protein , transgene , biology , microbiology and biotechnology , neun , microinjection , viral vector , recombinant dna , immunology , immunohistochemistry , gene , genetics
In previous studies we have utilized an adenoviral vector system to over express macrophage migration inhibitory factor (MIF) within rat paraventricular nucleus (PVN), in order to determined the regulatory effects of this protein on blood pressure and drinking behavior. However, the expression of MIF produced by the Ad system is transient, lasting for only 7–14 days. To obtain longer term neuronal over expression of MIF in the PVN, we plan to construct a MIF recombinant adeno‐associated virus 2 (AAV2) virus. Recombinant AAV2 containing a cytomegalovirus/chicken β‐actin (CBA) promoter has been shown to efficiently transduce a number of brain structures. Here the objective was to determine whether this AAV2‐CBA vector will be able to target primarily neurons in PVN, and result in sustained long‐term expression of transgenes. One μl of AAV2‐GFP containing the CBA promoter to control transgene (GFP) expression (2.0x10 10 or 1.0 x 10 11 particles), was infused stereotaxically into the PVN, the nucleus of the solitary tract (NTS) or the hippocampus of adult Sprague Dawley male rats. Rats were sacrificed and the brain analyzed for GFP expression on days 7, 14, 60 post‐injection. Localization of the injection site and neuron‐specific transduction was confirmed by visualization of GFP and immunostaining with antibodies against the neuron‐specific protein NeuN. Microinjection of both doses of AAV2‐GFP into SD rat PVN resulted in sustained expression of GFP from days 7 to 60. The expression was selective for neurons, as judged by the exclusive neuronal localization of GFP. Similar results were obtained from injections into the nucleus of the solitary tract (NTS) or hippocampus. These results suggest that this AAV2‐CBA construct is a very efficient and suitable vector for targeting neurons within rat PVN. This work was supported by NIH grant HL‐076803.