大鼠外伤性视神经损伤的闪光电生理特性和Tau蛋白及其磷酸化研究

Research on flash visual evoked potential and dynamic changes of Tau and its phosphorylation in rat model of traumatic optic neuropathy

目的:运用闪光视觉诱发电位(FVEP)技术检测外伤性视神经损伤(TON)大鼠视觉通路障碍的变化情况,探究TON大鼠视网膜及视神经中Tau及其磷酸化蛋白的变化。方法:将30只SD大鼠进行FVEP造模,1wk后构建TON模型,左眼进行视神经夹持损伤,右眼进行视神经暴露假手术。TON模型鼠分别于视神经夹持损伤后1、3、7、14、28d进行FVEP检测。FVEP检测完成后处死大鼠,分离左眼视网膜及视神经,并采用Western blot检测其中Tau及其Ser 396/Ser 404位点磷酸化蛋白的表达水平。结果:TON模型大鼠假手术眼出现典型的FVEP波形;相较于假手术眼,手术眼各检测时间点FVEP的N2波发生显著延迟,N2-P2波振幅发生大幅缩减,但视神经夹持损伤后各时间点N2波潜伏期延迟时间和N2-P2振幅缩减差异不明显。TON大鼠手术眼视网膜总Tau含量在夹持损伤1d后急剧下降,7d时短暂恢复后直至28d维持于略低于正常水平;pTau-Ser396/404的变化与总Tau的变化一致,其中Ser 396位点为Tau蛋白主要磷酸化位点。TON大鼠手术眼视神经总Tau含量逐渐增多,夹持损伤14d时达到高峰并维持至28d,pTau-Ser396/404的变化与总Tau的变化类似,7d时出现增长并抵达高峰,其中Ser 404位点为主要磷酸化位点。结论:FVEP的N2及P2相关指标可用于检测TON大鼠视觉功能障碍水平,TON大鼠视网膜与视神经Tau蛋白水平的变化截然不同,不同部位pTau-Ser396/404的变化跟随总Tau含量的变化而变化,但不同部位Tau蛋白主要改变的磷酸化位点不同。

AIM:To detect the visual dysfunction,and investigate the changes of Tau and its phosphorylated Ser396/Ser404 forms in retinas and optic nerves in traumatic optic neuropathy(TON)model rats by using FVEP technique.METHODS:Totally 30 SD rats were conducted FVEP electrode implantation.One week later,all rates were implemented TON operation with the optic nerve of left eye crushed and the optic nerve of right eye exposed(sham-operated).FVEP detections were performed respectively in these TON model rats at 1,3,7,14,and 28d post crush,with 5 rats tested at each time point.After FVEP tests were taken,rats were sacrificed and then retinas and optic nerves of left eyes were separated for detecting the expression levels of Tau and pTau-Ser396/404 by Western Blot.RESULTS:Typical FVEP waves were observed in the sham-operated eyes.Compared to the sham group,the N2 waves were significantly delayed and the amplitude of N2-P2 were greatly reduced at each time point in the operation eyes.However,the differences of N2 wave and the amplitude reduction of N2-P2 were not significant at each time point after crush.The contents of total Tau protein in retinas of TON rats sharply decreased at 1d post crush,briefly recovered at 7d post crush,and remained a slightly lower level than normal condition till 28d.The changes of pTau-Ser396/404 were consistent with the changes of total Tau in retains and the Ser396 was the main phosphorylation site.However,the total Tau contents in optic nerves of TON rats increased gradually,and peaked at the 14d post crush and remained till 28d.The changes of pTau-Ser396/404 were similar to the changes of total Tau in optic nerves,which peaked at 7d post crush.However,Ser404 was the main phosphorylation site of Tau in optic nerves.CONCLUSION:The related indexes of N2 and P2 waves in FVEP can be used to detect the visual dysfunction in TON rats.After TON,the content changes of total Tau in retinas and optic nerves were much different while the changes of pTau-Ser396/404 followed the alterations of total Tau in the two locations.However,the main phosphorylation site of Tau was differnet according to the locations.