Departamento de Bioquímica, Centro de Ciências Biológicas, Universidade Federal de Santa Catarina, Trindade, Florianópolis, SC, Brasil. molz.s @hotmail.com
J Neurosci Res. 2011 Sep;89(9):1400-8. doi: 10.1002/jnr.22681. Epub 2011 Jun 10.
Excitotoxicity and cell death induced by glutamate are involved in many neurodegenerative disorders. We have previously demonstrated that excitotoxicity induced by millimolar concentrations of glutamate in hippocampal slices involves apoptotic features and glutamate-induced glutamate release. Guanosine, an endogenous guanine nucleoside, prevents excitotoxicity by its ability to modulate glutamate transport. In this study, we have evaluated the neuroprotective effect of guanosine against glutamate-induced toxicity in hippocampal slices and the mechanism involved in such an effect. We have found that guanosine (100 μM) was neuroprotective against 1 mM glutamate-induced cell death through the inhibition of glutamate release induced by glutamate. Guanosine also induced the phosphorylation and, thus, activation of protein kinase B (PKB/Akt), a downstream target of phosphatidylinositol-3 kinase (PI3K), as well as phosphorylation of glycogen synthase kinase 3β, which has been reported to be inactivated by Akt after phosphorylation at Ser9. Glutamate treated hippocampal slices showed increased inducible nitric oxide synthase (iNOS) expression that was prevented by guanosine. Slices preincubated with SNAP (an NO donor), inhibited the protective effect of guanosine. LY294002 (30 μM), a PI3K inhibitor, attenuated guanosine-induced neuroprotection, guanosine prevention of glutamate release, and guanosine-induced GSK3β(Ser9) phosphorylation but not guanosine reduction of glutamate-induced iNOS expression. Taken together, the results of this study show that guanosine protects hippocampal slices by a mechanism that involves the PI3K/Akt/GSK3β(Ser9) pathway and prevention of glutamate-induced glutamate release. Furthermore, guanosine also reduces glutamate-induced iNOS by a PI3K/Akt-independent mechanism.
谷氨酸引起的兴奋毒性和细胞死亡与许多神经退行性疾病有关。我们之前已经证明，海马切片中毫摩尔浓度的谷氨酸引起的兴奋毒性涉及凋亡特征和谷氨酸诱导的谷氨酸释放。鸟苷，一种内源性鸟嘌呤核苷，通过调节谷氨酸转运来防止兴奋毒性。在这项研究中，我们评估了鸟苷对海马切片中谷氨酸诱导的毒性的神经保护作用及其涉及的机制。我们发现，鸟苷（100μM）通过抑制谷氨酸诱导的谷氨酸释放，对 1mM 谷氨酸诱导的细胞死亡具有神经保护作用。鸟苷还诱导蛋白激酶 B（PKB/Akt）的磷酸化，从而激活 PKB/Akt，Akt 的下游靶点是磷脂酰肌醇-3 激酶（PI3K），以及糖原合酶激酶 3β的磷酸化，后者据报道在 Ser9 磷酸化后被 Akt 失活。用谷氨酸处理的海马切片显示诱导型一氧化氮合酶（iNOS）表达增加，而鸟苷可防止这种增加。用 SNAP（一种 NO 供体）预处理的切片抑制了鸟苷的保护作用。LY294002（30μM），一种 PI3K 抑制剂，减弱了鸟苷诱导的神经保护作用、鸟苷对谷氨酸释放的预防作用以及鸟苷诱导的 GSK3β（Ser9）磷酸化，但不减弱鸟苷对谷氨酸诱导的 iNOS 表达的减少。总之，这项研究的结果表明，鸟苷通过涉及 PI3K/Akt/GSK3β（Ser9）途径和预防谷氨酸诱导的谷氨酸释放的机制来保护海马切片。此外，鸟苷还通过 PI3K/Akt 独立的机制减少谷氨酸诱导的 iNOS。
