Early cellular and molecular mechanisms underlying ischemia-induced neuronal death
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Global ischemia is a neuronal insult that induces delayed cell death. A hallmark event in the post-ischemic period is activation of signaling cascades which lead to enhanced permeability of mitochondrial membranes and release of cytochrome c. However, the precise mechanisms by which ischemia-induced signaling elicits channel activity and disrupts functional integrity of the mitochondrial membrane are unclear. Here I examine two aspects of the molecular and cellular mechanisms underlying global ischemia-induced neuronal death. The first of these projects focuses on the role of the PI3K/Akt signaling in ischemia induced neuronal death. Global ischemia in intact rats triggers expression and activation of the Akt inhibitor CTMP (Carboxyl-Terminal Modulator Protein) in hippocampal neurons that CTMP binds and extinguishes Akt activity and that CTMP is essential to ischemia-induced neuronal death. Whereas ischemia induces a dramatic phosphorylation and nuclear translocation of Akt, p-Akt in postischemic neurons is not active. RNA-interference-mediated depletion of CTMP in a clinically relevant model of stroke restores Akt activity and rescues hippocampal neurons.;In a second project, I examine a role for an N-terminally cleaved form of Bcl-xL in ischemia induced neuronal death. A critical step in mitochondrial demise in postischemic CA1 neurons is the formation of large channels in the outer mitochondrial membrane in the early postischemic period. Global ischemia promotes cleavage of Bcl-xL to generate DeltaN-Bcl-xL and elicits large channel activity in the outer membrane of mitochondria isolated from the hippocampal CA1. The Bcl-xL inhibitor, ABT-737 markedly inhibits channel activity and attenuates global ischemia-induced neuronal death. Introduction of recombinant DeltaN-Bcl-xL via the patch pipette to control mitochondria mimics the channel activity observed in ischemic mitochondria. Expression of DeltaN-Bcl-xL in primary neurons or in fibroblasts from bax-/- /bak-/- double-knockout mice induces apoptotic cell death that is blocked by ABT-737. Furthermore, we use knock-in mice expressing caspase cleavage resistant Bcl-xL. Ischemia induced death of CA1 neurons is attenuated in organotypically cultured hippocampal slices from cleavage-resistant Bcl-xL mice. These findings reveal a novel mechanism by which ischemic insults alter the integrity of the outer mitochondrial membrane and implicate Bcl-xL and/or its cleavage fragments, in the delayed cell death of hippocampal neurons.