Ginkgolic acids induce neuronal death and activate protein phosphatase type-2C

The standardized extract from Ginkgo biloba (EGb 761) is used for the treat ment of dementia. Because of allergenic and a effects, ginkgolic acids are restricted in EGb 761 to 5 ppm. The question arises whether ginkgolic acids also have neurotoxic effects. In the present study, ginkgolic acids caused death of cultured chick embryonic neurons in a concentration-dependent man ner, in the presence and in the absence of serum. Ginkgolic acids-induced d eath showed features of apoptosis as we observed chromatin condensation, sh rinkage of the nucleus and reduction of the damage by the protein synthesis inhibitor cycloheximide, demonstrating an active type of cell death. Howev er, DNA fragmentation detected by the terminal-transferase-mediated ddUTP-d igoxigenin nick-end labeling (TUNEL) assay and caspase-3 activation, which are also considered as hallmarks of apoptosis, were not seen after treatmen t with 150 muM ginkgolic acids in serum-free medium, a dose which increased the percentage of neurons with chromatin condensation and shrunken nuclei to 88% compared with 25% in serum-deprived, vehicle-treated controls. This suggests that ginkgolic acid-induced death showed signs of apoptosis as wel l as of necrosis. Ginkgolic acids specifically increased the activity of pr otein phosphatase type-2C, whereas other protein phosphatases such as prote in phosphatases 1A, 2A and 2B, tyrosine phosphatase, and unspecific acid- a nd alkaline phosphatases were inhibited or remained unchanged, suggesting p rotein phosphatase 2C to play a role in the neurotoxic effect mediated by g inkgolic acids. (C) 2001 Elsevier Science B.V. All rights reserved.