Astroglial differentiation of cortical precursor cells triggered by activation of the cAMP-dependent signaling pathway.

In the developing brain, differentiation of neural precursors into neurons or glial cells occurs in response to neurotrophic factors acting on the cell surface. Intracellular signaling mechanisms that relay information to initiate differentiative responses of neural precursor cells are poorly understood. To investigate whether stimulation of the cAMP-dependent signaling pathway participates in differentiative responses of cells in the developing CNS, we performed experiments using both conditionally immortalized neural precursor cells (RC2.E10 cells) and primary cultures of cells from developing rat cortex. Initially, we determined that RC2.E10 cells retain phenotypic features of neural precursors after inactivation of the immortalizing oncogene, a temperature-sensitive mutant of the simian virus 40 large-T antigen (SV40T). We found that, once SV40T is inactivated, RC2.E10 cells cease to divide and die. However, RC2. E10 cells can proliferate in the presence of basic fibroblast growth factor. In addition, they express nestin, a marker of neural precursor cells. Both RC2.E10 cells and primary cortical precursor cells undergo astroglial differentiation in response to cAMP stimulation by treatment with 8-bromo-cAMP. In both cases, cAMP-induced astrocyte differentiation is characterized by morphological changes, stimulation of glial fibrillary acidic protein expression, downregulation of nestin expression, and decreased proliferation. No increases in the expression of neuronal or oligodendrocytic markers were observed. Our results support the notion that the developing CNS contains neural precursor cells with the capacity of undergoing astrocyte differentiation in response to increased intracellular cAMP concentrations.

The effects of diethyldithiocarbamate (DDC) on the astrocytic cytoskeleton.

The dithiocarbamates are a group of compounds that are used extensively in industry, agriculture and medicine. Exposure to these compounds has caused deleterious effects to both the central and peripheral nervous systems. Cultured rat hippocampal astroglia treated with 35 micrograms/ml diethyldithiocarbamate (DDC) in media were studied for alterations to the cytoskeleton. Examination by both immunohistochemistry and scanning electron microscopy revealed disruption of the cytoskeletal elements. This occurred in a progressive time-dependent manner. Electrophoretic patterns demonstrated two cytoskeletal protein alterations. The microtubular protein, beta-tubulin, appeared to have an altered mobility while the major intermediate filament protein, glial fibrillary acidic protein (GFAP), was decreased. The cytoskeleton appears to be an important cellular target for injury by DDC exposure. This study has demonstrated that DDC induces alterations in the architecture of the cytoskeleton of astroglia and suggests that these changes involve microtubular and intermediate filament proteins.