Fig. 4. Fluorescent micrographs of b-galactosidase (b-gal)-stained sections of adult GLAST::CreERT2;R26R cortices without lesion (C and C') or at 3 days postlesion (dpl) (A, D, and D') or 7 dpl (B) (lesion, stab wound; dashed line, lesion track). Sections depicted in A and B are derived from mice treated for 5 days twice daily with corn oil. Note that only small precipitates of the fluorescent tyramide complex, but no reporter⁺ cells, are detectable (compare to Fig. 1A, same magnification). (C-D') b-gal and the endothelial PECAM are not colocalized in the intact cortex (C and C') or after lesion (D and D'). Note that also in the endfeet of b-gal-expressing cells attaching to the blood vessels there is no colocalization of the reporter with PECAM (see projections at the side of C' and D'). (Scale bars: 100 mm in A and B, 50 mm in C and D, and 30 mm in C' and D'.)

Fig. 5. Histograms in A-C depict reporter⁺ (b-gal⁺) cells (A and C) and reporter^- cells (B) in GLAST::CreERT2;R26R cortices. The proportion of reporter⁺ cells among all astrocytes is shown in A in the intact cortex and at the lesion site. (B) The proportion of dividing (BrdU⁺) reporter^- astrocytes in the intact cortex and after stab wound that is very similar to the proportion of BrdU⁺ cells among the reporter⁺ astrocytes in Fig. 1C. The histogram in C depicts the percentage of increase in the number of b-gal⁺ cells in the lesioned area compared to the contralateral side. (D) Fluorescent micrograph showing b-gal and S100b coexpression 30 days after stab wound (dashed line, lesion track). (D) Fluorescent micrograph illustrating the participation of astrocytes genetically labeled before lesion in scar-forming GFAP⁺ astroglia (arrows). (Scale bars: 100 mm in C and 50 mm in D.)

Fig. 6. (A) Graph showing the number of multipotent cortical neurospheres at different passages. Note that this number is relatively constant, suggesting that neurospheres contain self-renewing stem cells. (B andC) Sorting profile of cells isolated from the lesion side of hGFAP-eGFP mice (GFP intensity is depicted on the x axis) labeled with NG2-directed (B) and PDGFRa-directed (C) antibodies detected with APC-coupled secondary antibodies (fluorescence intensity on the y axis) 3 days postlesion (dpl). Both NG2⁺ or PDGFRa⁺ single positive populations (upper left quadrant in B and C, respectively) and the double-positive populations (upper right quadrant) were sorted separately and cultured under neurosphere conditions with or without additional PDGF (see Materials and Methods). No neurospheres were observed in cultures of cells that were exclusively labeled by NG2 or PDGFRa, whereas few spheres (<0.01%) were formed by the double-positive cells. These spheres formed by the double-positive cells were, however, not multipotent and never gave rise to neurons (two experimental batches). These data are referred to together in the main text as none of these cells giving rise to multipotent neurospheres.

Fig. 7. (A and B) Fluorescent micrographs of frontal sections of the telencephalon stained for GFP (green) and GFAP (red) 3 days postinjection (3 dpi/dpl) of GFP-containing lentivirus into the subependymal zone (SEZ) and stab wound lesion in the cerebral cortex ipsilateral to the site of injection. Note that most GFP⁺ cells migrate within the rostral migratory stream (RMS) (A and B), but some also enter the white matter (WM) of the cerebral cortex. No GFP⁺ cells were detectable in the parenchyma of the cerebral cortex surrounding the injury site. (C and D) Dot blots of FACS analysis depicting forward scatter on the x axis and green fluorescence on the y axis for cells isolated from the SEZ 3 dpi (2.6% GFP⁺ cells in yellow in C) and cells isolated from the tissue surrounding the injury site (0% GFP⁺ cells in D). (Scale bars: 10 mm.)