EGF-induced MAPK-mediated calpain2 activation triggers a neuroprotective priming mechanism against oxygen-glucose deprivation: implication of AMPAR downregulation and degradation of their scaffolding PDZ proteins

Abstract

Calpains are Ca++-dependent regulatory proteases that truncate various enzymes, cytoskeletal proteins, neurotransmitter receptors, and ion channels. However, under physiological conditions, Calpain2 (a.k.a. mCalpain) rapid activation depends on MAP Kinase and can be achieved in the presence of BAPTA-AM, an intracellular Ca++ chelator (Jourdi et al., 2010). Treatment of primary cortical neuronal cultures (PCNC) with EGF and EGF-like cytokines reduces AMPAR expression and levels of their scaffolding proteins GRIP1 and SAP97 (Yokomaku D et. al., 2005; Namba H et. Al., 2006). Here, we show that EGF treatment in primary neurons and cultured hippocampal slices causes the appearance of lower molecular weight bands representing calpain-mediated SAP97 and GRIP1 degradation products. The appearance of these degradation products is sensitive to calpain, MAPK and ErbB1 (a.k.a. EGF receptor) inhibition. EGF and related cytokines are neuroprotective against various neurotoxic conditions, including ischemia. Neuroprotection against ischemia can also be achieved by brief exposure of nervous tissues to ischemic conditions (ischemic preconditioning or priming). Thus, can EGF pretreatment exert neuroprotective priming-like effects against oxygen-glucose deprivation (OGD)? Indeed, EGF pretreatment followed by OGD does not cause additional accumulation of SBDP or SAP97 and GRIP1 degradation products, implying abrogated calpain activation during OGD. Equally important are the findings that calpain inhibition eliminates the neuroprotective effects of EGF pretreatment against OGD. The interaction of EGF pretreatment with calpain activation is also assessed in the context of a priming-like effect of EGF against OGD using acute hippocampal slices taken from mice lacking or over-expressing the endogenous calpain inhibitor, calpastatin. The data indicate that calpastatin expression levels in mutant mice are inversely correlated with calpain activation and SAP97 and GRIP1 degradation under OGD.