6-Hydroxydopamine (6-OHDA) is a neurotoxin used in the induction of experimental Parkinson's disease in both animals and PC12 cells, which are derived from rat pheochromocytoma tumors and have many properties similar to dopamine neurons. Biochemical and molecular approaches have shown that low doses of 6-OHDA induce apoptosis in PC12 cells and, in the processing of apoptosis, caspases are crucial mediators, and caspase inhibition is sufficient to rescue PC12 cells from apoptosis induced by 6-OHDA. However, because this caspase inhibition targets multiple caspases, it is not known whether a single caspase is primarily responsible for effecting cell death in this model. To assess the particular member (caspase-3) of the ced-3 family relevant to cell death and to position their activation within the apoptotic pathway, we constructed a hammerhead ribozyme directed against rat caspase-3, which could downregulate the expression of caspase-3 in vitro and in vivo, and transfer to PC12 cells. The results show that the ribozymes against caspase-3 could protect PC12 cells from apoptosis induced by low doses of 6-OHDA. The PC12 cell transfected with the ribozymes shows a significant decrease in caspase-3 activity compared with control cells at various time points. Parallel to the reduced caspase-3 protease activity, similar decreased levels of apoptotic cells and DNA fragmentation were also assessed by staining with Hoechst 33258 and ELISA, respectively. Overexpression of p35, a general caspase inhibitor, also protected PC12 cells from apoptosis. These results confirm that caspases play an important role in 6-OHDA-induced PC12 cell apoptosis and indicate that caspase-3 itself is one of the crucial mediators of neurotoxin-induced PC12 cell apoptosis.