Fig. 7.

Effect of co-exposure to A1254 and PKC inhibitor, bis, on dbcAMP-induced GFAP immunoreactivity and morphological change in C6 cells. C6 cells were subjected to different treatments, fixed and then immunocytochemistry (A) and phase-contrast (B) analysis were performed as detailed in Section 2.4. Cells were treated with bis (0.125 μM) or co-exposed to bis (0.125 μM) and A1254 (3 or 9 μM) in presence of dbcAMP (1 mM) in serum-deprived medium containing 0.1% (v/v) DMSO. After 24 h incubation, cells were subjected to immunofluorescence analysis to reveal GFAP immunoreactivity (green). Immunocytochemistry of cells treated with 0.1% (v/v) DMSO (vehicle) (A–C); cells exposed to bis (0.125 μM) (D–F). Astrocytic differentiation induced by treatment with dbcAMP (1 mM) in presence of bis (0.125 μM) (G–I). dbcAMP-differentiated cells co-exposed to: bis (0.125 μM) and 3 μM A1254 (J–L): bis (0.125 μM) and 9 μM A1254 (M–O). GFAP immunostaining (A, D, G, J and M). DAPI-nuclear stain (blue) of the same field (B, E, H, K, and N). Merge for composite images (C, F, I, L and O). Scale bar = 50 μm. (B) Phase-contrast micrographs of C6 cells cultured in serum-free DMEM in presence (B) or absence (A) of bis (0.125 μM). dbcAMP-differentiating cells were treated with bis (0.125 μM) alone (C) or in combination with A1254 3 μM (D) or 9 μM (E). All the treatments were performed for 24 h under serum-free conditions in presence of 0.1% (v/v) DMSO used as vehicle for A1254 and bis. Scale bar = 25 μm. (For interpretation of color in Fig. 7, the reader is referred to the web version of this article.)