Figure 4.
Radial glia transform into astrocytes in the absence of signaling via erbB2, and erbB2 expression induces astrocyte to radial glia transformation. (a–c) Radial glial cells (E16) were cultured in the presence of anti-erbB2 mAb (b) or control (a) antibodies. Under control conditions, 71 ± 4% cells were radial glial cells (arrow, a); the rest were astrocytes (28 ± 3%). However, when erbB2 was blocked, only 21 ± 4% of the cells were of radial glial phenotype and 79 ± 3% of cells displayed the transformed phenotype of astrocytes (arrow, b; c). Cells were labeled with rat-401 antibodies. Data shown are mean ± SEM. (Scale bar = 30 μm.) (d) C6 cells were transfected with erbB2-GFP, GFP, or DNerbB2-GFP plasmids, treated with 50 ng/ml NRG-1 for 10 min, and immunoblotted with antiphosphotyrosine or acetylated tubulin antibodies. DNerbB2 drastically reduced NRG-1-induced tyrosine phosphorylation of ErbB receptors (185 kDa). (e–i) Radial glia from E15 mouse cortices were transfected with DNerbB2-GFP (e and f) or control GFP (g and h) plasmids. After 48 h, cells that are positive for GFP and the radial glial (RC2-blue; e and f) or astrocyte markers (anti-GFAP-red; g and h) were counted according to their phenotype. Glial cells expressing DNerbB2-GFP were mainly of astrocyte phenotype (radial glia/astrocyte ratio = 13.5 ± 2.1:85.65 ± 2; e, f, and i). Cells transfected with control plasmids were mainly of radial glial phenotype (radial glia/astrocyte ratio = 70.31 ± 3:29.7 ± 3.34; g–i). (Scale bar = 30 μm.) (j–p) Mature astrocytes from 4-week-old mouse cortices were transfected with GFP (j and k) or erbB2-GFP (l–o) plasmids. After 48 h, GFP-positive cells labeled with anti-GFAP (k and m; red) or RC2 (o; blue) antibodies were quantified. Cells expressing erbB2 were predominantly of radial glial phenotype (percentage of glial cells that are radial glia: GFP, 14 ± 4.6, erbB2-GFP, 65.7 ± 5.9), whereas cells transfected with GFP were mainly astrocytes (percentage of glial cells that are astrocytes: GFP, 85.3 ± 4.5; erbB2-GFP, 34.2 ± 6). (Scale bar = 30 μm.)