Drug therapy designed in the laboratory to protect ischemic brain has not translated to humans. For this reason attention has been focused on molecular mechanisms by which the brain can induce endogenous neuroprotective strategies. Such neuroprotection occurs when the brain is """"""""preconditioned"""""""" by exposure to brief ischemic stress resulting in """"""""tolerance"""""""" to subsequent severe ischemia. Such ischemic tolerance produces robust neuroprotection through protein-synthesis dependent mechanisms, which require 24 hours to evolve. These gene-based mechanisms of tolerance were the focus during our previous grant period. We now propose to investigate the cellular mechanisms involved in rapid ischemic tolerance, a protective mechanism, well know in the heart, which is protein synthesis independent, and inducible with in an hour, a time frame likely to have substantial clinical potential. We offer novel preliminary data describing the cell biology producing rapid ischemic tolerance in brain, resulting from the action of the constitutive anti apoptotic protein Bcl-w. We demonstrate its neuroprotective effects, the mechanism by which its function is rapidly potentiated in ischemia, offer experiments showing additional mechanisms to further regulate Bcl-w function and show a hitherto unknown function of an anti-apoptotic bcl-2 family member protein, that of modulating GABA channel currents which are additionally neuroprotective. Thus, we offer a new description of an endogenous neuroprotective cell biology strategy for ischemic brain which is rapidly effective. To further define this biology we offer the following aims:
Aim 1 :Investigate the role of Bcl-w in rapid ischemic tolerance Aim 2: Investigate the effect of over expression of Bcl-w on ischemia-induced cell death Aim 3: Investigate the role of 14-3-3 and CKs (casein kinases )in regulating Bcl-w function Aim 4: Investigate the role of Bcl-w-induced enhancement of GABA mediated currents in rapid ischemic preconditioning In order to investigate these aims we will use a variety of approaches, including immunoprecipitation, knockout mice deficient in the Bcl-w gene, HIV TAT-based protein transduction vectors for delivery of Bcl-w to neurons, electrophysiological recordings from neurons and site directed mutagenesis studies. Experiments will be predominantly performed using in vitro ischemia models using cortical neuronal cultures, and key observations replicated using in vivo focal ischemia models. As such, understanding the endogenous mechanisms that regulate rapid ischemic tolerance may lead to the identification of novel therapeutic targets for the treatment of stroke.

National Institute of Health (NIH)
National Institute of Neurological Disorders and Stroke (NINDS)
Research Project (R01)
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Special Emphasis Panel (ZRG1-BDCN-L (90))
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Silberberg, Shai D
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Emanuel Hospital and Health Center
United States
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Zhou, An; Simon, Roger P; David, Larry (2011) Nascent proteomes of ischemic-injured and ischemic-tolerant neuronal cells. Int J Comput Biol Drug Des 4:40-55
Thompson, Simon; Pearson, Andrea N; Ashley, Michelle D et al. (2011) Identification of a novel Bcl-2-interacting mediator of cell death (Bim) E3 ligase, tripartite motif-containing protein 2 (TRIM2), and its role in rapid ischemic tolerance-induced neuroprotection. J Biol Chem 286:19331-9
Ordonez, Andrea Nicole; Jessick, Veronica Joy; Clayton, Corrin Erin et al. (2010) Rapid ischemic tolerance induced by adenosine preconditioning results in Bcl-2 interacting mediator of cell death (Bim) degradation by the proteasome. Int J Physiol Pathophysiol Pharmacol 2:36-44
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Meller, Robert; Thompson, Simon John; Lusardi, Theresa Ann et al. (2008) Ubiquitin proteasome-mediated synaptic reorganization: a novel mechanism underlying rapid ischemic tolerance. J Neurosci 28:50-9
Doyle, Kristian P; Yang, Tao; Lessov, Nikola S et al. (2008) Nasal administration of osteopontin peptide mimetics confers neuroprotection in stroke. J Cereb Blood Flow Metab 28:1235-48
Maysami, Samaneh; Lan, Jin Quan; Minami, Manabu et al. (2008) Proliferating progenitor cells: a required cellular element for induction of ischemic tolerance in the brain. J Cereb Blood Flow Metab 28:1104-13
Sato, Yu; Meller, Robert; Yang, Tao et al. (2008) Stereo-selective neuroprotection against stroke with vitamin A derivatives. Brain Res 1241:188-92
Thompson, Simon J; Loftus, Liam T; Ashley, Michelle D et al. (2008) Ubiquitin-proteasome system as a modulator of cell fate. Curr Opin Pharmacol 8:90-5

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