[Abstract Title]. - Society for Neuroscience

Time: Sunday, November 16, 2008, 1:00 pm - 5:00 pm
Program#/Poster#: 242.11/I6
Topic: B.11.b. Cell biology and signalling
Support: NINDS
NIMH
Title: Astrocyte-specific transgene expression using adeno-associated virus
Authors: *J. DONG, A. MUNGENAST, H. TAKANO, E. STAMBROOK, H. ZHOU, P.
HAYDON;
Univ. Pennsylvania Sch. Med., Philadelphia, PA
Abstract: Recombinant adeno-associated virus (rAAV) is a highly effective vehicle for gene
delivery into CNS. Towards the goal of manipulating gene expression in astrocyte specifically,
we examined the transduction pattern and cellular tropism of AAV2 vector sequences
pseudotyped with capsid sequences from serotypes 1, 2, 5, 7, 8, and 9. Quantification of the
EGFP expression 7 days after in vivo injection showed that recombinant AAV2/1, AAV2/2,
AAV2/7, AAV2/8 and AAV2/9 displayed predominantly neuronal tropism in both cortex and
hippocampus. Comparatively, AAV2/5 was demonstrated to show significantly more astrocyte
transduction. To further investigate whether the astrocyte specific GFAP promoter can enhance
astrocyte specificity of the AAV2/5 pseudotype, we generated rAAV vectors with transgenes
driven by the human GFAP promoter (AAV-Gfa2-eGFP), a downsized GFAP promoter (AAV-
Gfa104-eGFP and AAV-Gfa104-Venus-DAGK). AAV2/5 containing the GFAP promoter
exhibited efficient transduction both in vitro and in vivo and the transduction was restricted to
astrocytes. This specificity was verified by colabeling using anti-GFAP and anti- glutamine
synthetase antibodies, well-known markers for astrocytes, and a lack of co-labeling with other
cell-specific antibodies including anti-NeuN, anti-NG2, anti-Oligodendrocyte and anti-Iba1. We
also demonstrated the possibility of repeated imaging on the same astrocyte transduced by
AAV2/5 in living mice using 2-photon microscopy, which allows us to study astrocyte dynamics
and function in vivo over time. Our study suggests that specifically targeting astrocytes via the
combination of the AAV2/5 pseudotype and the GFAP promoter will become a powerful tool to
investigate the role of astrocytes in vivo.
Disclosures: J. Dong , None; A. Mungenast, None; H. Takano, None; E. Stambrook,
None; H. Zhou, None; P. Haydon, None.
Poster
242. Glial Neuronal Interactions: Glutamate and Calcium