【作者】 席艳；

【导师】 邓锦波；

【作者基本信息】 河南大学 ， 神经生物学， 2010， 硕士

【摘要】 孕期酒精暴露(prenatal alcohol exposure, PAE)会导致胎儿流产、死产、胎儿颜面部畸形、神经系统发育畸形以及出生后认知障碍等。本研究建立孕期酒精暴露(PAE)模型,研究孕期酒精暴露对小鼠视皮质突触数量的影响。利用免疫荧光染色技术标记对照组与PAE模型组(低剂量和高剂量)子鼠出生后0、7、14、30天视皮质突触前体的synaptophysin荧光颗粒,观察其数密度变化,并利用Western blotting方法对各实验组子代小鼠视皮质synaptophysin的表达量进行半定量分析。具体结果如下：synaptophysin荧光颗粒代表的突触数量随子鼠孕期酒精暴露剂量升高而下降,同时随子鼠年龄的增长而升高。横向比较(剂量依赖性比较),自对照组、低剂量组、高剂量组突触数密度呈下降趋势；纵向比较(长时程效应比较),随年龄增加突触数密度呈上升趋势。对照组内P0、P7、P14和P30间存在显著差异P<0.05,但是在同一水平的酒精处理组内P0、P7、P14和P30的个别组无显著性差异P>0.05。年龄与剂量两个因素之间存在交互作用,而且剂量(MS=9.75,F=140.44,P<0.05)的影响比年龄(MS=6.55,F=94.26,P<0.05)的影响作用稍大。Fishier LSD检验在有显著差异的两组之间进行均数差值比较,发现年龄增高使对照组与酒精处理组的差异增大,并且对照组与高剂量组的差异大于对照组与低剂量组的差异。利用Western blotting对PAE模型组和对照组子鼠P30、P14、P0进行synaptophysin半定量检测,以阳性条带与(3-actin条带光密度比值作为synaptophysin的相对表达值,结果显示：(1) synaptophysin在胶片上显示为分子量为38KD的活化片段；(2) synaptophysin蛋白表达量随子鼠年龄增长而增高,各年龄组组间差异显著(P<0.05)；(3)酒精处理组低于对照组(P<0.05),高剂量组低于低剂量组(P<0.05),即孕期酒精暴露使子鼠视皮质synaptophysin蛋白表达量下降且具有剂量依赖性。总之,孕期酒精暴露将造成子代突触数量的丢失,并且其对子代神经系统的影响具有剂量相关性及长时程放大效应。一方面我们应该充分认识到孕期饮酒对胎儿发育及家庭社会带来的不利影响,要求孕妇避免饮酒；另一方面我们也应该看到孕期酒精暴露对子代神经系统影响的机制、FAS的产前诊断、孕期酒精暴露出生后胎儿治疗有关的药物开发等诸多领域都大有可为。更多还原

【Abstract】 In order to understand the alcohol’s toxicity to the quantitative alternations of synapses in mouse visual cortex, the expression of synaptophysin after prenatal alcohol exposure was investigated. In present study, the experimental mice at PO, P7, P14 and P30 were grouped, according to different treatments, such as control,2g/kg alcohol treatment and 4g/kg alcohol treatment as well. The pre-synaptic elements which were used to represent synapses were marked with synaptophysin (a synaptic vesicle associated protein) immunocytochemistry. The synaptophysin positive boutons in layer VI of visual cortex were imaged under laser confocal microscope. With stereological methods, the number cal density of synapse in visual cortex was calculated in different groups at various ages. Moreover, Western-blotting was carried out to detect the expressions of synaptophysin in visual cortex as well. Our results showed that alcohol prenatal exposure could cause synaptic loss with dose dependency and long-term effect. For instance, there were significant difference among the different treatment groups with dose dependency at PO, P14 and P30 as well (P＜0.05). Western blotting supported the results of immunofluorescent labeling as well. In conclusion, prenatal alcohol exposure can induce the synaptic loss with dose dependency and long term effect. Our findings implicate that the synaptic loss with long term effect in CNS probably contributes to the lifelong mental retardation and memorial lowliness associated with childhood FAS.更多还原