Figure 2. NG2 mice for visualizing pericytes in vivo.

Images taken from adult brains of three different mouse lines: NG2-DsRed (left column); NG2-Cre crossed with Ai14 tdTomato reporter mice (middle column); and NG2-CreER™ crossed with Ai14 and induced with tamoxifen in adulthood (right column). Tamoxifen was administered intraperitoneally at a dose of 100 mg/kg dissolved in corn oil:ethanol (9:1), every 24 hours for 5 consecutive days. Mice were sacrificed for histology within 2 weeks after the last injection of tamoxifen. Brain sections (50 μm thick) from all three lines were stained with anti-CD31 antibody to label the endothelium and imaged with wide field fluorescence (A, D, G), and confocal microscopy (B, B’, E, E’, H, H’). Confocal images (maximal projections of 20 μm thick stacks) are shown in the red channel alone to highlight fluorescent protein expression (B, E and H), and with CD31 to show perivascular location of pericytes (B’, E’ and H’). Using confocal microscopy of histological slices, ovaloid pericyte cell bodies (arrowheads) are visible in all lines. Fine pericyte processes that radiate along the microvasculature, however, are most easily visualized with an NG2-CreER™ × Ai14 cross (arrow). In vivo images (maximal projections of 50 μm stacks taken 25-75 μm below the pial surface) were collected through a PoRTs window using TPLSM (C, F and I) [116]. The vascular serum (green) is labeled with intravenous FITC-dextran (2 MDa) [116]. Pericytes are difficult to discern in the NG2-Cre × Ai14 line due to excessive background neuronal labeling. In comparison, NG2-CreER™ × Ai14 and NG2-DsRed animals exhibit very sparse labeling of pyramidal neurons, and possible interneurons. Neurons appear distributed independent of cortical layer, and their number increases with time after tamoxifen induction. All animals label oligodendrocyte precursors and vascular smooth muscle cells.