Ischemic preconditioning via epsilon protein kinase C activation requires cyclooxygenase-2 activation in vitro

The signaling pathway of cyclooxygenase-2 (COX-2) induction following ischemic preconditioning (IPC) in brain remains undefined. To determine role of COX-2 in ischemic preconditioning, we used two in vitro models: mixed cortical neuron/astrocyte cell cultures and organotypic hippocampal slice cultures. We simulated IPC by exposing cell or slice cultures to 1 h or 15 min of oxygen/glucose deprivation (OGD), respectively, 48 h prior to ischemia. To mimic ischemia in vitro, we exposed cell or slice cultures to OGD of 4 h or 40 min, respectively. In cell cultures, these experiments revealed that COX-2 induction peaked at 24 In following IPC in cell culture. Inhibition of COX-2 activation with 50 mu M NS-398 (a COX-2 selective inhibitor) abolished IPC-mediated neuroprotection in both in vitro models. Next, we tested whether e protein kinase C (EPKC) and extracellular signal regulated kinase 1/2 (ERK1/2) activation was involved in IPC-mediated neuroprotection and COX-2 expression in cell culture. Cell cultures were treated with an epsilon PKCspecific activating peptide (psi epsilon RACK, 100 nM) for 1 h, and 48 h later were exposed to OGD. epsilon PKC activation increased ERK1/2 phosphorylation and COX-2 induction and conferred neuroprotection similar to IPC. Additionally, inhibition of either EPKC or ERK1/2 activation abolished COX-2 expression and neuroprotection due to ischemic preconditioning. These results demonstrate a crucial role for the epsilon PKC -> ERK1/2 -> COX-2 pathway in the induction of neuroprotection via ischemic preconditioning. (c) 2007 IBRO. Published by Elsevier Ltd. All rights reserved.