Figure 5.

Activation of Fas triggers PCD of motoneurons even in the presence of optimal trophic support. Different agents known to activate Fas receptor by clustering were tested for their ability to trigger motoneuron cell death. (A) Purified E12.5 mouse motoneurons were cultured in the presence of BDNF (1 ng/ml), CT-1 (10 ng/ml), and GDNF (0.1 ng/ml) (+NTFs) for 1 d. Subsequently, the indicated concentrations of anti-Fas antibody were added in the continued presence of NTFs, and motoneuron survival was counted 2 d later, at 3 DIV. Counts were expressed relative to the value for NTFs at 3 DIV. Fas antibody induced a dose-dependent loss of 45% of the motoneurons. (B) An analogous experiment using purified motoneurons from E14 rat, cultured in the continued presence of BDNF (1 ng/ml). After 1 DIV, tagged soluble FasL was added at the indicated concentrations in the presence of 1 μg/ml of enhancer antibody, which had no effect when tested alone. Survival was counted 2 d later. (C) Comparison of the fractions of E14 rat motoneurons lost after 2 d of incubation with 10 ng/ml sFasL, in the presence of indicated neurotrophic factors used at the same concentrations as for mouse motoneurons in A. The number of motoneurons lost was expressed as a percentage of the total number of motoneurons present in the same conditions but without sFasL. All histograms are representative of at least three independent experiments. Error bars represent the mean ± range of duplicate wells.