【作者】 赵圆宇；

【导师】 唐勇；

【作者基本信息】 重庆医科大学 ， 神经病学， 2011， 博士

【摘要】 目的:研究丰富生存环境对中老年雌性大鼠空间学习、记忆能力的作用,并运用无偏体视学方法研究短期丰富生存环境是否能延缓大脑胼胝体以及胼胝体内有髓神经纤维老年性改变的进程。方法:将中老年（14月龄）雌性SD大鼠24只,随机分为丰富生存环境干预组（12只）和标准环境组（12只）,饲养时间为4个月。干预结束后,用Morris水迷宫对两组大鼠空间学习、记忆能力进行连续5天的测试,分别记录两组大鼠的寻台时间。行为学测试后,分别从两组大鼠随机抽选5只大鼠,将其大脑制成1 mm厚的冠状连续脑片,用卡瓦列里原理计算每只大鼠胼胝体的体积。每只大鼠按等距随机的原则抽选4个脑片,从抽取的脑片中随机抽取4个胼胝体组织块。将抽选的组织块运用isector包埋法制作电镜切片,每张切片随机抽选4个视野,每个视野间无任何重叠,放大至6 000倍拍照,运用体视学方法测量并计算大鼠大脑胼胝体内有髓神经纤维和髓鞘的总体积、有髓神经纤维的总长度和平均直径。结果:丰富生存环境组大鼠隐藏平台实验的寻台时间显著短于标准环境组大鼠（p = 0.001）,显露平台实验的寻台时间无显著性差异（p = 0.069）。与标准环境组大鼠相比,丰富生存环境大鼠大脑胼胝体总体积显著性增加34.5%（p = 0.017）,胼胝体内有髓神经纤维和髓鞘的总体积分别显著性增加60.8%（p = 0.007）和57.7%（p = 0.008）,有髓神经纤维总长度显著性增加82.5%（p = 0.016）,而有髓神经纤维的平均直径显著性下降9.3%（p = 0.045）。丰富生存环境组大鼠大脑胼胝体内有髓神经纤维的增加以直径小于1μm,尤其是0.4 0.6μm的细小直径有髓神经纤维增加为主。结论:短期丰富生存环境干预能明显改善中老年雌性大鼠空间、学习记忆能力。本实验首次运用无偏体视学方法研究了短期丰富生存环境对中老年雌性大鼠胼胝体以及胼胝体内有髓神经纤维的作用。我们发现:短期丰富生存环境能显著增加大鼠大脑胼胝体总体积、胼胝体内有髓神经纤维和髓鞘的总体积、胼胝体内有髓神经纤维的总长度。短期丰富生存环境干预所致的胼胝体内有髓神经纤维的改变可能是短期丰富生存环境干预对中老年大鼠空间、学习记忆能力改善的重要结构基础之一。这些结果为进一步寻找延缓大脑衰老进程的手段提供了重要的科学依据。目的:研究丰富生存环境对中老年雌性大鼠空间学习、记忆能力的作用,并运用无偏体视学方法研究短期丰富生存环境对中老年大脑胼胝体内成熟型少突胶质细胞的作用。方法:实验动物分组与行为学测试同第一部分。行为学测试后,分别从两组大鼠随机抽选5只大鼠,将其大脑制成2 mm厚的冠状连续脑片,将所有脑片进行石蜡包埋并切片,切片厚度为14μm,每只动物等距随机抽选13 15张切片。采用CNPase标记成熟型少突胶质细胞,苏木精复染细胞核。利用体视学测试系统计数每张切片胼胝体内阳性细胞数量,利用光学分合法计算每只动物胼胝体内CNPase⁺细胞总数。结果:丰富生存环境组大鼠隐藏平台实验的寻台时间显著短于标准环境组大鼠（p = 0.002）,显露平台实验的寻台时间无显著性差异（p = 0.371）。与标准环境组大鼠相比,丰富生存环境大鼠大脑胼胝体内CNPase⁺细胞的总数显著性增加343.6%（p < 0.001）。结论:丰富生存环境干预能明显改善中老年雌性大鼠空间学习、记忆能力,能显著增加大鼠胼胝体内成熟型少突胶质细胞的总数。丰富生存环境干预所致的胼胝体内成熟型少突胶质细胞的改变可能是丰富生存环境干预改善中老年大鼠胼胝体内有髓神经纤维的原因之一。目的:研究丰富生存环境对中老年雌性大鼠空间学习、记忆能力的作用,并运用无偏体视学方法研究丰富生存环境对中老年大脑海马内成熟型少突胶质细胞的作用。方法:实验动物分组与行为学测试同第一部分。行为学测试后,分别从两组大鼠随机抽选5只大鼠,将其大脑制成2 mm厚的冠状连续脑片,将所有脑片进行石蜡包埋并切片,切片厚度为14μm,每只动物按等距随机原则抽选15 20张切片。采用免疫组织化学染色,采用CNPase标记成熟型少突胶质细胞,苏木精复染细胞核。利用体视学测试系统计数每张切片整个海马内、海马CA₁区、齿状回内阳性细胞的数量,利用光学分合法计算每只动物大脑海马以及海马各亚区内CNPase+细胞的总数。结果:行为学测试结果同第二部分。与标准环境组大鼠相比,丰富生存环境大鼠大脑海马内CNPase⁺细胞的总数显著性增加113%（p = 0.008）,海马CA₁区和齿状回内CNPase⁺细胞的总数显著性增加127%（p = 0.005）和127%（p = 0.006）。结论:短期丰富生存环境干预能显著增加大鼠大脑海马内以及海马CA₁区和齿状回内成熟型少突胶质细胞的总数。短期丰富生存环境干预所致的胼胝体内成熟型少突胶质细胞的改变可能是短期丰富生存环境干预改善中老年大鼠海马内有髓神经纤维的原因之一。更多还原

【Abstract】 Objective: To investigate the effects of enriched environment on the spatial learning ability of middle-aged female rats and then investigate the effects of short-term enriched environment on the age-related changes of the corpus callosum （CC） and the age-related changes of the myelinated fibers in the corpus callosum by means of unbiased stereologlcal methods.Methods:24 female SD rats （14 months old） were divided randomly into two groups: 12 rats lived in an enriched environment （EE） and 12 control rats were reared in standard environment （SE）. The rats were housed for 4 months under either EE or SE. The spatial memory of aged rats was examined with Morris water maze for five consecutive days. The time latency was record. After the behavioral tests, five rats of each group were selected randomly for stereological study. The brains were cut into 1mm-thick serial sections, and the volume of corpus callosum was calculated according to Cavalieri’s principle. Four brain slices were sampled according to systematic random sampling fashions. Four tissue blocks were sampled randomly from the sampled brain slices and embedded for electronic microscopy following isector process. From each section, four visional fields were photographed randomly at a magnification of 6 000×. The total volume of myelinated nerve fibers and the total volume of myelin sheath, the total myelinated fiber length and mean diameter of the myelinated fibers in the corpus callosum were quantitatively investigated using stereological methods.Results: Compared with SE group, the time latency of hidden-platform test was shortened significantly in EE group （p = 0.001）. There was no significant difference of mean myelinated fiber diameter between EE group and SE group （p = 0.069）. When compared to SE rats, the total volume of corpus callosum was significantly increased by 34.5% （p = 0.017）. Compared to SE rats, the total volumes of the myelinated nerve fibers and the total volume of the myelin sheaths in the corpus callosum were significantly increased by 60.8% （p = 0.007） and 57.7% （p = 0.008）, respectively. Compared to SE rats, the total myelinated fiber length in CC of EE rats was significantly increased by 82.5% （p = 0.016）. In contrast, the mean diameter of the myelinated fibers in the CC of EE rats was significantly decreased by 9.3% （p = 0.045） when compared to that of SE rats. The the enrichment-induced changes of the myelinated nerve fibers in the CC of EE rats were mainly due to the marked increases of the myelinated nerve fibers with diameter less than 1μm, especially the fibers with diameter of 0.4 0.6μmConclusion: The increase of the myelinated fibers in enriched rat corpus callosum might provide one of the structural bases for the enrichment-induced improvement of the spatial memory. The present study provided, to the best of our knowledge, the first evidence of environmental enrichment-induced increases of the corpus callosum volume and the myelinated fibers in the corpus callosum of aged rats. Objective: To investigate the effects of enriched environment on the spatial learning ability of middle-aged female rats and then investigate the effects of short-term enriched environment on the age-related changes of mature oligodendrocytes in the corpus callosum by means of unbiased stereological methods.Methods: The enriched environment protocol and behavioral tests were the same as in part one. After the behavioral tests, five rats of each group were selected randomly for stereological study. The brains were cut into 2 mm-thick serial slabs. The slabs of the randomly selected hemisphere were embedded in paraffin. Serial coronal sections with thickness of 14μm were cut with a sliding microtome. One section was systematically sampled every 30th section. On average, 13 15 sections were sampling per rat. CNPase was used to mark the mature oligodendrocytes. The nuclei were satined by haematoxylin. The optical disector was used to count the number of positive cells in the corpus callosum of each section. Then, the total number of CNPase positive cells in each rat corpus callosum was calculated. Results: For hidden-platform test, the mean latency of EE group from day 1 to 4 was significantly shortened when compared to SE group （p = 0.002）, and there was no significant change for visible-platform test between them （ p = 0.371）. When compared to SE group, the total number of CNPase⁺ cells was significantly increased by 343.6% （p < 0.001）.Conclusion: The increase of the CNPase⁺ cells in enriched rat corpus callosum might provide one of the structural bases for the enrichment-related improvement of the myelinated nerve fibers. Objective: To investigate the effects of enriched environment on the spatial learning ability of middle-aged female rats and then investigate the effects of short-term enriched environment on the age-related changes of mature oligodendrocytes in the hippocampus by means of unbiased stereological methods.Methods: The enriched environment protocol, material and behavioral tests were the same as in part one. One section was systematically sampled every 10th section. On average, 15 20 sections were sampling per rat. The number of positive cells in the whole hippocampus, hippocampal CA₁ and DG of each section was counted. Then, the total number of CNPase positive cells in each rat hippocampus was calculated.Results: The results of Morris water maze tests were the same as in part two. When compared to SE group, the total number of CNPase⁺ cells was significantly increased by 113% （p = 0.008）, 127% （p = 0.005） and 127% （p = 0.006） in the whole hippocampus, CA₁ and DG, respectively.Conclusion: The increase of the CNPase⁺ cells in enriched rat whole hippocampus and subdivisions in hippocampus might provide one of the structural bases for the enrichment-induced improvement of the myelinated nerve fibers in hippocampus.更多还原