Fig. 1.
Cell death of CGCs (neuronal–astrocytic cultures, 9 DIV) induced by coculture with activated microglia or by direct exposure to proinflammatory cytokines or NOC-18 (NO donor).A, In the control culture of CGCs, note the phase-bright normal cell bodies and dense neuritic network. B, Coculture of CGCs (9 DIV, 0.25 × 106cells/cm2) with LPS/IFN-γ-activated microglia (0.2 × 106cells/cm2) for 24 hr induced cell death of all neurons. Note shrunken cell bodies and nuclei and loss of all neurites (phase-contrast photographs). C, Addition of LPS (4 ng/ml) and IFN-γ (100 U/ml) simultaneously for 48 hr to the CGCs, cultured in the presence of glial cells (9 DIV, nontreated with ara-C), caused severe neuronal cell death, but dead cells were not phagocytosed. D, Morphological analysis of nuclear chromatin in CGC culture (neuronal–astrocytic, 9 DIV) exposed to 500 μm NOC-18 for 4 hr using DNA-binding fluorochrome Hoechst 33342 (fluorescence microscope). Viable cells showed round nuclei with weak fluorescence, but some nuclei had strongly condensed chromatin (bright fluorescence), predominantly without fragmentation. However, a few nuclei with fragmented chromatin were also present. Particular cell or nuclear types are indicated by the following abbreviations (placed to the right of the cell): n, healthy neurons; a, astrocyte; am, activated microglia; dn, dead neurons; cc, condensed chromatin; fn, fragmented chromatin;ncc, non-condensed chromatin. Scale bar (shown inC): A–C, 40 μm;D, 20 μm.