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. 2015 Apr 15;8:10. doi: 10.3389/fnmol.2015.00010

Figure 1.

Figure 1

IUE-mediated transgene delivery to astrocytes and neurons in various regions of the brain. The binary co-transfection system consists of a (A) helper plasmid carrying the piggyBac transposase enzyme and a (B) donor plasmid carrying the GCaMP (3, 6s, or 6f)-IRES-tdTomato construct flanked with piggyBac inverted terminal repeats. A subset of donor plasmids include an N-terminal Lck-tagged GCaMP for plasma membrane targeting. All transgenes are expressed under the control of CAG. (C–G) IUE of CAG-GFP-IRES-Neo transfected astrocytes and neurons in the (C,D) hippocampus, (C,G) cortex, the (E) dentate gyrus, and (F) nucleus accumbens. Fluorophore-conjugated anti-GFP antibody enhanced GFP is shown in green. In (D–F) astrocyte-labeling anti-GFAP antibody is shown in red and overlap (transfected astrocytes) in yellow. (G) GFAP overlap with GFP was sometimes difficult to detect, so we also stained astrocyte-specific Aldh1L1 which is shown in red with transfected astrocytes in yellow. (H) Hippocampal expression pattern of tdTomato expression (naïve fluorescence) and (I) co-expression (yellow) of GCaMP3 (green) and tdTomato (red) in putative thalamic neurons following IUE transfection of CAG-GCaMP3-IRES-tdTomato (naïve fluorescence). White arrows denote putative astrocytes.