Immunohistochemical Detection of Bax, Caspase-3 and Bcl-2 Proteins following Ischemia in a Gerbil Model of Ischemic Tolerance

It has been demonstrated that ischemic preconditioning prevents delayed neuronal death following transient ischemia. Although both apoptosis and necrosis have been shown to contribute to neuronal cell death, the ability of ischemic preconditioning to prevent apoptosis remains unknown. To test the hypothesis that ischemic preconditioning reduces irreversible cerebral ischemic injury in part by decreasing apoptosis, neuronal apoptotic events and changes in apoptosis-specific molecules Bax, Bcl-2 and caspase-3 were studied in gerbil model of transient global ischemia with and without ischemic preconditioning. The time-course expression of Bax, Bcl-2 and caspase-3 were examined immunohistochemically in the hippocampal CA1 region at 1, 3 and 7 days after ischemia. Following transient global ischemia, levels of Bax and caspase-3 proteins, identified as proapoptotic molecules, were increased in CA1 neurons. A decrease in immunoreactivity against Bax and caspase-3 in the ischemic preconditioning group was correlated with the CA1 neuronal protection. A time course study revealed that time dependent increase of these proapoptotic proteins, peaked at 3 days. The expression of antiapoptotic Bcl-2 oncoprotein was observed in CA1 neurons 3 days after ischemia in ischemic preconditioning group. Considering the expression of apoptosis related proteins and DNA strand breaks put together, CA1 neurons die through an apoptotic process after transient forebrain ischemia. This study demonstrated that the antiapoptotic Bcl-2 proteine plays a neuropro-tective role in neuronal cells which acquire ischemic tolerance. Although the detailed relationship between the function of bcl-2 protein and ischemic tolerance is still unclear, our results provide new evidence which indicates that ischemic preconditioning reduces neuronal death in part by decreasing apoptosis.