Interactions between GSK3β and caspase signalling pathways during NGF deprivation induced cell death

Withdrawal of NGF (NGF-W) in PC12 cells leads to caspase and GSK3 beta activation which results in cell death. Our recent findings suggest that inhibition of GSK3 beta promotes PC12 cell survival after NGF-W. To determine whether these pathways interact from a signalling perspective, we compared the effects of BAF (a general caspase inhibitor), Li+ (a GSK3 beta inhibitor) and insulin on NGF-W induced PC12 cell death. Maximal increase in DNA fragmentation was observed 3 h after NGF-W and was inhibited by BAF (7.5 mu M), Li+ (IC50 = 2 mM) and insulin (IC50 = 100 nM). BAF inhibited caspase-3 activity and delayed cell death up to 6 h after NGF-W indicating that caspase inhibition is sufficient to prevent apoptosis. BAF had no major effect on GSK3 beta active site phosphorylation or activity suggesting the caspase pathway does not regulate GSK3 beta activity. Conversely, Li+ inhibited caspase activity by only 20% but promoted cell survival for 24 h after NGF-W. Overexpression of dominant negative mutants of GSK3 beta also inhibited apoptosis, but had only a minor effect on caspase activity after NGF-W. Taken together, these results suggest that GSK3 beta is upstream of caspase signalling, and exerts a small effect on the caspase pathway.