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目的研究兴奋性氨基酸转运体2(EAAT2)过表达对癫痫发作及SE诱导的海马神经元死亡的作用。方法实验采用野生型或者EAAT2转基因FVB/N小鼠,腹腔注射匹鲁卡品诱导癫痫持续状态(SE)。SE后3d,取脑、固定、切片,进行EAAT2、生长抑素的免疫组化染色以及甲酚紫染色,分别对海马CA1和齿状回门区阳性神经元进行计数。结果与野生型动物相比,EAAT2在转基因小鼠海马中表达显著增加。野生型小鼠达到SE或者死亡所需的总匹鲁卡品剂量为344±40.3mg/kg,,而EAAT2转基因小鼠达到同等效应所需剂量为657±119.9mg/kg,显著高于野生型所用剂量(P<0.05)。SE后3d,野生型小鼠海马CA1区锥体细胞层神经元相对数量为0.56,而转基因动物中为0.9,显著高于野生型动物(P<0.05)。同时,野生型和转基因小鼠癫痫后齿状回门区中间神经元相对数量分别为0.11和0.67,转基因组数量显著高于野生型组(P<0.05)。野生型小鼠癫痫后齿状回门区生长抑素阳性神经元数量为0,但是,在EAAT2转基因小鼠,数量为0.4,显著高于野生型(P<0.05)。结论EAAT2过表达对SE产生及其诱导的神经元死亡具有显著保护作用。过表达的EAAT2可能通过加强细胞外谷氨酸转运而调控其兴奋毒性。
Objective To investigate the effect of excitatory amino acid transporter 2 (EAAT2) overexpression on the seizure and SE-induced neuronal death in hippocampus. Methods Wild-type or EAAT2 transgenic FVB / N mice were injected intraperitoneally with pilocarpine to induce status epilepticus (SE). Three days after SE, brain, fixed and sectioned were taken for immunohistochemical staining of EAAT2 and somatostatin, and cresyl violet staining. Positive neurons of hippocampal CA1 and dentate gyrus were counted. Results EAAT2 was significantly increased in the hippocampus of transgenic mice compared to wild-type animals. The dose of total pilocarpine required to reach SE or death in wild-type mice was 344 ± 40.3 mg / kg, whereas the dose required for EAAT2 transgenic mice to achieve the same effect was 657 ± 119.9 mg / kg, significantly higher than wild-type The dose used (P <0.05). Three days after SE, the relative number of neurons in hippocampal CA1 pyramidal cell layer of wild-type mice was 0.56, while that of transgenic mice was 0.9, which was significantly higher than that of wild-type animals (P <0.05). At the same time, the relative number of interneurons in the dentate gyrus was 0.11 and 0.67 respectively in wild type and transgenic mice, and the number of transgenic groups was significantly higher than that in wild type (P <0.05). In wild-type mice, the number of somatostatin positive neurons in the dentate gyrus was 0 in epileptic rats, however, the number was 0.4 in EAAT2 transgenic mice, which was significantly higher than that in wild type (P <0.05). Conclusion EAAT2 overexpression has a significant protective effect on SE production and its induction of neuronal death. Overexpression of EAAT2 may regulate its excitotoxicity by enhancing extracellular glutamate transport.