A decrease in phosphorylation of cAMP-response element-binding protein (CREBP) promotes retinal degeneration.

Excitotoxicity, induced either by N-Methyl-d-aspartate (NMDA) or kainic acid (KA), promotes irreversible loss of retinal ganglion cells (RGCs). Although the intracellular signaling mechanisms underlying excitotoxic cell death are still unclear, recent studies on the retina indicate that NMDA promotes RGC death by increasing phosphorylation of cyclic AMP (cAMP) response element (CRE)-binding protein (CREBP), while studies on the central nervous system indicate that KA promotes neuronal cell death by decreasing phosphorylation of CREBP, suggesting that CREBP can elicit dual responses depending on the excitotoxic-agent. Interestingly, the role of CREBP in KA-mediated death of RGCs has not been investigated. Therefore, by using an animal model of excitotoxicity, the aim of this study was to investigate whether excitotoxicity induces RGC death by decreasing Ser(133)-CREBP in the retina. Death of RGCs was induced in CD-1 mice by an intravitreal injection of 20 nmoles of kainic acid (KA). Decrease in CREBP levels was determined by immunohistochemistry, western blot analysis, and electrophoretic mobility gel shift assays (EMSAs). Immunohistochemical analysis indicated that CREBP was constitutively expressed in the nuclei of cells both in the ganglion cell layer (GCL) and in the inner nuclear layer (INL) of CD-1 mice. At 6 h after KA injection, nuclear localization of Ser(133)-CREBP was decreased in the GCL. At 24 h after KA injection, Ser(133)-CREBP was decreased further in GCL and the INL, and a decrease in Ser(133)-CREBP correlated with apoptotic death of RGCs and amacrine cells. Western blot analysis indicated that KA decreased Ser(133)-CREBP levels in retinal protein extracts. EMSA assays indicated that KA also reduced the binding of Ser(133)-CREBP to CRE consensus oligonucleotides. In contrast, intravitreal injection of CNQX, a non-NMDA glutamate receptor antagonist, restored the KA-induced decrease in Ser(133)-CREBP both in the GCL and INL, and inhibited loss of RGCs and amacrine cells. These results, for the first time, suggest that KA promotes retinal degeneration by reducing phosphorylation of Ser(133)-CREBP in the retina.