【作者】 杨姝；

【导师】 唐勇；

【作者基本信息】 重庆医科大学 ， 神经病学， 2010， 博士

【摘要】 第一部分大鼠大脑白质及白质内有髓神经纤维老年改变的体视学研究目的:运用新的体视学方法对28月龄(老年)雌性大鼠进行体视学定量研究,以期进一步认识雌性大鼠大脑白质及白质内有髓神经纤维的老年改变规律。方法:雌性Long-Evans大鼠,青年组(6月龄)5只,中老年组(18月龄)5只,老年组(28月龄)6只。随机抽取4块大脑白质组织块制作成电镜标本,放大6000倍拍照。运用体视学方法计算大脑白质总体积、白质内有髓神经纤维总长度、白质内有髓神经纤维总体积、髓鞘总体积、有髓神经纤维平均直径。结果:老年组雌性大鼠大脑白质总体积分别比青年组和中老年组雌性大鼠显著性降低了42.7% (p<0.00001)和37.7% (p<0.0001);老年组雌性大鼠大脑白质内有髓神经纤维总长度分别比青年组和中老年组雌性大鼠显著性降低了52.5% (p<0.01)和48.6% (p<0.05);老年组雌性大鼠大脑白质内有髓神经纤维总体积分别比青年组和中老年组雌性大鼠显著性降低了26.7% (p<0.05)和43.0% (p<0.05);老年组雌性大鼠大脑白质内有髓神经纤维髓鞘总体积分别比青年组和中老年组雌性大鼠显著性降低了30.7% (p<0.05)和46.7% (p<0.05)。老年组雌性大鼠大脑白质内有髓神经纤维平均直径比青年组雌性大鼠显著性增加了37.5% (p<0.05)。结论:与青年组和中老年组雌性大鼠相比,老年雌性大鼠大脑白质及白质内有髓神经纤维发生了明显减少,本实验研究结果与我们以前的研究发现一起表明,雌性大鼠大脑白质老年性改变发生的时间晚于雄性大鼠。第二部分丰富生存环境对大鼠空间记忆能力及大脑白质内有髓神经纤维的作用目的:研究丰富生存环境对老年大鼠空间记忆能力的作用,并首次运用无偏体视学方法研究丰富生存环境能否延缓大脑白质及白质内有髓神经纤维的老年改变进程。方法:中老年(14月龄)SD大鼠,雌雄各24只,随机分为丰富生存环境干预组(雌雄各12只)和标准环境组(雌雄各12只);老年(24月龄)SD大鼠,雌雄各24只,随机分为丰富生存环境干预组(雌雄各12只)和标准环境组(雌雄各12只)。饲养时间4个月。干预结束后,用Morris水迷宫进行连续5天的空间记忆能力的测试,记录大鼠寻台时间。行为学测试后,从每组大鼠中随机抽取5只大鼠,随机抽取4块白质组织块制作成电镜标本,放大6000倍拍照,运用体视学方法计算出大鼠大脑白质总体积、白质内有髓神经纤维总长度、总体积和有髓神经纤维平均直径。结果:中老年雌性丰富生存环境组大鼠的寻台时间明显短于标准环境组大鼠(p<0.05)。与标准环境组大鼠相比,中老年雌性丰富生存环境组大鼠大脑白质总体积显著性增加了46.9% (p<0.001),老年雄性丰富生存环境组大鼠大脑白质总体积显著性增加了68.1% (p<0.05),老年雌性丰富生存环境组大鼠大脑白质总体积显著性增加了18.4% (p<0.001)。与标准环境组大鼠相比,中老年雄性丰富生存环境组大鼠大脑白质内有髓神经纤维的总长度显著性增加了34.1% (p<0.01),中老年雌性丰富生存环境组大鼠大脑白质内有髓神经纤维的总长度显著性增加了43.2% (p<0.05),老年雄性丰富生存环境组大鼠大脑白质内有髓神经纤维的总长度显著性增加了68.8% (p<0.01),老年雌性丰富生存环境组大鼠大脑白质内有髓神经纤维的总长度显著性增加了62.9% (p<0.001)。与标准环境组大鼠相比,丰富生存环境显著性地增加了中老年雌性大鼠大脑白质内有髓神经纤维总体积(54.0%, p<0.05)和老年雄性大鼠大脑白质内有髓神经纤维总体积(64.3%, p<0.001)。丰富生存环境对中老年及老年雌、雄性大鼠大脑白质内有髓神经纤维的平均直径没有明显作用。结论:短期丰富生存环境干预可以明显改善中老年雌性大鼠空间记忆能力,这一研究结果为将来寻找延缓老年大脑功能下降进程的手段提供了重要的科学依据。短期丰富生存环境干预可以不同程度增加中老年及老年大鼠大脑白质体积、白质内有髓神经纤维总长度和总体积,丰富生存环境干预所致的白质有髓神经纤维改变可能是丰富生存环境干预改善了老年大鼠空间记忆能力的重要结构基础之一。第三部分丰富生存环境对大鼠大脑白质有髓神经纤维髓鞘的作用目的:进一步认识短期丰富生存环境促进中老年及老年大鼠大脑白质总体积、白质内有髓神经纤维总长度和总体积增加的结构基础。方法:实验材料、丰富生存环境干预与标本取材同第二部分。将电镜标本放大20000倍拍照。运用体视学方法计算出大鼠大脑白质总体积、有髓神经纤维总长度、有髓神经纤维髓鞘总体积、有髓神经纤维轴突总体积、有髓神经纤维平均内径和平均外径、有髓神经纤维髓鞘平均内周长和平均外周长以及有髓神经纤维髓鞘平均厚度。结果:与标准环境组大鼠相比,中老年雄性丰富生存环境组大鼠大脑白质内有髓神经纤维髓鞘总体积显著性增加了27.1% (p<0.05),中老年雌性丰富生存环境大鼠有髓神经纤维髓鞘总体积显著性增加了39.7% (p<0.05),老年雌性丰富生存环境大鼠有髓神经纤维髓鞘总体积显著性增加了29.7% (p<0.05)。与标准环境组大鼠相比,中老年雌性丰富生存环境组大鼠大脑白质内有髓神经纤维轴突总体积显著性增加了73.2% (p<0.05),老年雄性丰富生存环境大鼠有髓神经纤维轴突总体积显著性增加了93.2% (p<0.05)。与标准环境组大鼠相比,各丰富生存环境组大鼠大脑白质内有髓神经纤维平均内径和平均外径、有髓神经纤维髓鞘平均内周长和平均外周长以及髓神经纤维髓鞘平均厚度没有显著性改变。丰富生存环境导致的大脑白质内有髓神经纤维总长度的增加主要是纤维直径小于0.6μm、髓鞘厚度小于0.16μm的有髓神经纤维的增多造成的。比较发生明显总长度改变的各直径段有髓神经纤维的内径-外径比(轴突直径-纤维直径比),丰富生存环境组平均为0.60,对照组平均为0.62。髓鞘厚度与轴突直径呈线性改变,在相同内径的情况下,丰富生存环境大鼠有髓神经纤维的髓鞘厚度在一定程度上大于对照组。结论:短期丰富生存环境导致了有髓神经纤维髓鞘的大量再生,其结果是细小直径、薄髓鞘的有髓神经纤维大量增多,此类型的有髓神经纤维集中分布在额叶白质,与大脑认知功能密切相关。短期丰富生存环境引起的再生的有髓神经纤维,其髓鞘厚度与轴突直径呈正相关关系,且内径-外径比为0.60左右。这种最佳的内径-外径比保证了神经冲动的最快传导,这为丰富生存环境干预改善老年大脑功能提供了形态学基础。大脑白质内有髓神经纤维的髓鞘为将来寻找延缓大脑衰老进程的手段提供了一个重要的“靶结构”。更多还原

【Abstract】 PART ONE STEREOLOGICAL INVESTIGATIONS OF THE AGE-RELATED CHANGES IN THE WHITE MATTER AND THE MYELINATED FIBERS IN THE WHITE MATTER OF RAT BRAINObjective: To investigate the changes of the white matter and the myelinated fibers in the white matter of 28-month (old-aged) female rats using new stereological methods in order to further understand in order to further understand how the white matter and the myelinated fibers in the white matter of old-aged female rats changed during aging process.Methods: Five young (6-month), five middle-aged (18-month) and six old-aged (28-month) female Long-Evans rats were used. Four white matter blocks were sampled in a systematic random fashion for electronic microscope use. Fields of view were randomly photographed with the magnification of 6000. The white matter volume, the total length of the myelinated fibers in the white matter, the total volume of the myelinated fibers in the white matter, the total volume of the myelin sheaths in the white matter and the mean diameter of the myelinated fibers in the white matter were investigated with the new stereological methods.Results: The white matter volume of old-aged female rats was significantly reduced by 42.7% (p<0.00001) when compared to young female rats and was significantly reduced by 37.7% (p<0.0001) when compared to middle-aged female rats, respectively. The total length of the myelinated fibers in the white matter of old-aged female rats was significantly decreased by 52.5% (p<0.01) when compared to young female rats and was significantly decreased by 48.6% (p<0.05) when compared to middle-aged female rats, respectively. The total volume of the myelinated fibers in the white matter of old-aged female rats was significantly decreased by 26.7% (p<0.05) when compared to young female rats and was significantly decreased by 43.0% (p<0.05) when compared to middle-aged female rats, respectively. The total volume of the myelin sheaths in the white matter of old-aged female rats was significantly decreased by 30.7% (p<0.05) when compared to middle-aged female rats and was significantly decreased by 46.7% (p<0.05) when compared to middle-aged female rats, respectively. The mean diameter of the myelinated fibers in the white matter of old-aged female rats was significantly increased 37.5% (p<0.05) when compared to young female rats.Conclusiton: When compared to young and middle-aged female rats, the white matter volume, the myelinated fiber length, the myelinated fiber volume and the myelin sheath volume of old-aged female rats were all significantly decreased. The present results together with our previous findings indicated that the age-related white matter changes in female rats happened much later than in male rats. PART TWO EFFECTS OF THE ENRICHED ENVIRONMENT ON THE SPATIAL MEMORY AND THE MYELINATED FIBERS IN THE WHITE MATTER OF RATObjective: To investigate the effects of enriched environment on the spatial learning ability of middle-aged rats and old-aged rats, and then for the first time use the new stereological methods to investigate if the enriched environment can delay the age-related changes of the white matter and the age-related changes of the myelinated fibers in the white matter.Methods: Forty-eight (24 females and 24 males) middle-aged (14 month) SD rats and forty-eight (24 females and 24 males) old-aged (24 month) SD rats were randomly divided into enriched environment (EE) group and standard environment (SE) group. Rats were reared under either EE or SE for following 4 months. Spatial memory was examined with Morris water maze tasks for five consecutive days. Time latency was recorded. After the behavioral tests, five rats of each group were randomly sampled for stereological study. Four white matter blocks were sampled in a systematic random fashion for electronic microscope use. Fields of view were randomly photographed with the magnification of 6000. The white matter volume, the total length of the myelinated fibers, the total volume of the myelinated fibers and the mean diameter of the myelinated fibers in the white matter were investigated with the new stereological methods.Results: The time latency in Morris water maze tasks of middle-aged female rats in EE group was significantly shorter than that of controls in SE group (p<0.05). When compared to SE rats, the white matter volume was significantly increased by 46.9% (p<0.001) in middle-aged female EE rats, 68.1% (p<0.05) in old-aged male EE rats and 18.4% (p<0.001) in old-aged female EE rats, respectively. When compared to SE rats, the myelinated fiber length in the white matter was significantly increased by 34.1% (p<0.01) in middle-aged EE males, 43.2% (p<0.05) in middle-aged EE females, 68.8% (p<0.01) in old-aged EE males and 62.9% (p<0.001) in old-aged EE females, respectively. When compared to SE rats, the myelinated fiber volume in the white matter was significantly increased by 54.0% (p<0.05) in middle-aged female EE rats and 64.3% (p<0.001) in old-aged male EE rats, respectively. There were no significant effects of enriched environment on the mean diameters of the myelinated fibers in the white matter between SE rats and EE rats.Conclusion: The spatial memory of middle-aged female rats was significantly improved by the exposure to short-term enriched environment, which provided an important scientific basis for the future study to search strategy to delay the progress of brain function decline with aging. The short-term enriched environment induced the increases in the white matter volume, the total volume and total length of the myelinated fibers in the white matter middle-aged rats and old-aged rats. The changes of the myelinated fibers in the white matter induced by enriched environment might provide one of the important structural bases for the enriched environment-induced improvement of the spatial learning ability of aged rats. PART THREE THE EFFECTS OF THE ENRICHED ENVIRONMENT ON THE MYELIN SHEATHS OF THE MYELINATED FIBERS IN THE WHITE MATTER OF RAT BRAINObjective: To further investigate the structural bases of the short-term enriched environment induced significant increases of the white matter volume, myelinated fiber length and myelinated fiber volume in the white matter of middle-aged rats and old-aged rats.Methods: The materials, enriched environment protocol and sampling were the same as in part two. Fields of view were randomly photographed with the magnification of 20000. The white matter volume, the total length of the myelinated fibers, the total volume of the myelin sheaths, the total volume of the axons, as well as the mean inner diameter and mean outer diameter of the myelinated fibers, the mean inner perimeter and mean outer perimeter of the myelin sheaths, and the mean thickness of the myelin sheaths in the white matter were investigated with the new stereological methods.Results: When compared to SE rats, the total volume of the myelin sheaths in the white matter was significantly increased by 27.1% (p<0.05) in middle-aged male EE rats, 39.7% (p<0.05) in middle-aged female EE rats and 29.7% (p<0.05) in old-aged female EE rats, respectively. When compared to SE rats, the total volume of the axons in the white matter was significantly increased by 73.2% (p<0.05) in middle-aged female EE rats and 93.2% (p<0.05) in old-aged male EE rats. There were no significant differences in the mean inner diameter and mean outer diameter of the myelinated fibers, in the mean inner perimeter and mean outer perimeter of the myelin sheaths and in the mean thickness of the myelin sheaths in the white matter between SE rats and EE rats, respectively. The enrichment-induced significant increase of the myelinated fiber length in the white matter was mainly due to the marked increases of the myelinated fibers with the size of less than 0.6μm and with the myelin sheath thickness of less than 0.16μm. For those myelinated fibers that their lengths were significantly changed, the ratio between inner diameter and outer diameter (axonal diameter and myelinated fiber diameter) was computed. The average value was 0.62 in SE groups and 0.60 in EE groups. The myelin sheath thickness was linearly proportional to the axonal diameter. The myelin sheath thicknesses in the white matter of EE rats were more or less thicker than those of SE rats.Conclusion: Short-term enriched environment induced marked increases of the myelin sheaths, which resulted in the significant increases of the myelinated fibers with small size and with thin myelin sheaths. This type of myelinated fibers was mostly located in frontal white matter, which was closely related to the cognition. The myelin sheath thickness of the remyelinated fibers induced by short-term enriched environment was positively correlated to the axonal diameter of the remyelinated fibers. The ratio between axonal diameter and fiber diameter was around 0.60, which was considered to be optimal ratio to maximize the conduction velocity. The results might provide the structural basis for the enrichment-induced functional improvement of aging brain. The myelin sheaths of the myelinated fibers in the white matter were one of the important targets for the future study to search the new ways to delay the process of brain aging.更多还原