银杏叶提取物对发育期戊四氮点燃癫癎大鼠学习记忆与海马神经干细胞增殖、分化的影响

目的 探讨银杏叶提取物( EGb)对癫癎点燃模型幼鼠神经发生的影响及与学习记忆的关系.方法 选择21日龄SD大鼠,随机分为9g·L-1盐水对照组(NS组)、戊四氮(PTZ)点燃对照组(PTZ7d组、PTZ 14 d组)以及EGb治疗组(EGb7 d组、EGb 14d组).应用PTZ制作慢性点燃癫癎模型；使用Y-型电迷宫测试模型建立前后及EGb治疗前后大鼠的行为学变化；通过免疫组织化学法观察大鼠点燃成功后EGb 7 d或EGb 14 d治疗对海马区细胞增殖情况的影响；同时通过标记5-溴脱氧尿核苷(BrdU)、神经元核蛋白( NeuN)、胶质纤维酸性蛋白(GFAP)观察神经干细胞(NSCs)分化情况.结果 1.Y-型电迷宫测试:EGb治疗前,PTZ各点燃大鼠与NS组相比,达到学会标准所需的电击次数明显增多(P＜0.01);EGb(300mg·kg-1·d-1)治疗7d和14 d大鼠达到学会标准的次数均明显少于PIZ点燃对照组(Pa＜0.01).2.NSCs增殖情况:选择Nestin阳性细胞表达作为判断NSCs增殖的指标,PTZ点燃对照组Nestin阳性细胞表达量较NS组明显增多(P＜0.01)；EGb 7 d组和EGb 14 d组Nestin阳性细胞表达量均较相对应PTZ点燃对照组表达进一步增多(Pa＜0.05),随着疗程增加,EGb 14 d组Nestin阳性细胞数量要低于EGb7 d组(P＜0.05),增殖的Nestin阳性细胞均主要分布于海马CA1 、CA3区颗粒细胞层和颗粒细胞下层.3.EGb治疗后BrdU/NeuN阳性细胞数明显多于EGb治疗前,且EGb 14 d组BrdU/NeuN阳性细胞表达百分比又高于EGb 7 d组,差异均有统计学意义(Pa＜0.05),EGb治疗组有4％～5％共表达GFAP.结论 EGb可诱导NSCs增殖、分化,且NSCs的这种改变与改善大鼠学习记忆有关.

Effect of Ginkgo Biloba Extract on Learning and Memory of Developing Rats with Kindled Seizure by Pentetrazole and Proliferation and Differentiation of Hippocampal Neural Stem Cells

Objective To study the effect of Ginkgo biloba extract(EGb) on kindling rats at developmental phase and its relationship with learning and memory. Methods Postnatal 21 days(P21) Sprague-Dawley(SD) rats were grouped randomly into 5 groups including normal sodium controls(NS group),pentylenetetrazol(PTZ)kindling models(P7 group and P14 group,PTZ 35 mg·kg-1·d-1,ip) and EGb groups(EGb 7 d group and EGb 14 d group,EGb 300 mg·kg-1·d-1,po),which all rats was preliminary screened by Y-maze test.Then hippocampal neural stem cells(NSCs) proliferation of CA1 and CA3 granular cell layer was studied by immunohistochemical method for all experimental rats,which Nestin positive cells were regarded as marker of NSCs.Meanwhile,5-bromo-deoxy-uridine(BrdU) served as the marker of cell proliferation,NeuN which was a marker of neuron,GFAP which was a marker of astrocytes were observed for differentiation of proliferative NSCs. Results 1.Compared with the NS group,the times of every kindled group rats reaching academic standard were increased significantly by Y-maze test(P<0.01).The number of reaching the learning standards of rats after treatment of EGb for 7 days and 14 days were significantly less than that in PTZ kindling group(Pa<0.01).2.For NSCs proliferation,Nestin positive cells under or within granular cell layer of CA1 or CA3 in hippocampus of PTZ kindled rats increased significantly compared with NS group(P<0.01),meanwhile,that in EGb 7 d and EGb 14 d groups were more than that of corresponding PTZ kindled rats(P<0.05).The Nestin positive cell of CA1 and CA3 region of rats with EGb 7 d and EGb 14 d were compared and found that Nestin positive cells in EGb 14 d rats were less those that in EGb 7 d rats for NSCs differentiation(P<0.05).3.The number of BrdU/NeuN cells in PTZ kindled group was less than that in EGb treatment groups.The percentage of BrdU/NeuN positive cells in hippocampus in EGb 14 d group was higher than that in EGb 7 d group(Pa<0.05).Meanwhile,4%-5% co-expression of GFAP with BrdU was found after EGb treatment. Conclusions EGb can significantly improve the learning and memory function of kindled rats.The possible mechanism of EGb treatment for improving learning-memory dysfunction may be related to influence the NSCs proliferation and differentiation of hippocampus in kindled rats.