Biphasic activation of apoptosis signal-regulating kinase 1-stress-activated protein kinase 1-c-Jun N-terminal protein kinase pathway is selectively mediated by Ca²⁺-permeable alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionate receptors involving oxidative stress following brain ischemia in rat hippocampus

Stress-activated protein kinase/extracellular signal-regulated kinase-1 (SEK1/MKK4) was examined in a rat model of global brain ischemia. Western blot assay showed that SEK1 activation was biphasic in CA1 but not CA3/dentate gyrus. The second activation peak (3 days after ischemia) was prevented by pretreatment with l-naphthyl acetyl spermine (Naspm), a channel blocker of Ca²⁺-permeable alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionate (AMPA) receptors, or N-acetylcysteine (NAC), a free radical scavenger. Concomitantly, the late activation of apoptosis signal-regulating kinase 1 (ASK1) and c-Jun N-terminal protein kinase (JNK) was also prevented by Naspm or NAC. Moreover, phospho-SEK1 and phospho-JNK co-immunoprecipitated with ASK1 and the bindings peaked at 3 days of reperfusion. Together with previous results, these findings indicate that Ca²⁺-permeable AMPA receptors are important routes to mediate the late activation of ASK1-SEK1-JNK pathway involving oxidative stress in hippocampal CA1 region after ischemia.