三重脑震荡鼠认知行为障碍及中枢胆碱能神经元变化研究
发布时间：2019-06-22 15:43
【摘要】： 目的：建立三重脑震荡(multiple cerebral concussion，MCC)大鼠模型，观察MCC大鼠的认知功能以及基底前脑、脑干网状结构和其它有关核团(包括被盖背外侧核、迷走神经背核和脊髓前角运动神经元)胆碱能神经元的变化，探讨三重脑震荡大鼠认知功能障碍与相关脑区胆碱能神经元变化的关系，为运动性脑震荡(sports-related cerebralconcussion，SRCC)的防治提供理论依据。方法：采用金属单摆打击装置复制大鼠MCC模型48只，随机分为1天、2天、4天、8天、16天和24天组共6个三重脑震荡组(n=8)。另设一组正常对照组(n=8)。应用平衡木实验(beam balance test，BBT)及噪音横杆跑动实验(beam walking test，BWT)评价MCC大鼠平衡运动与认知功能。运用免疫组织化学的方法检测大鼠基底前脑、脑干网状结构、被盖背外侧核、迷走神经背核和脊髓前角运动神经元中乙酰胆碱合成过程中的催化酶——胆碱乙酰转移酶(ChAT)的表达，并在光镜下对其计数。结果：(1)BBT及BWT实验结果：BBT结果显示，与对照组相比，MCC组大鼠在伤后第1天平衡时间明显缩短，差异具有统计学意义(P＜0.05)，第2天开始恢复，至第4天恢复正常。BWT结果显示，伤后第1天步入潜伏期明显延长，第2天开始恢复，至第10天尚未完全恢复正常。除第9天外，其它各时间点与对照组相比，差异均具有统计学意义(P＜0.05)。BWT评分结果显示，伤后第1天评分较低，第2天开始逐渐升高，至第4天恢复正常，差异具有统计学意义(P＜0.05)。(2)胆碱乙酰转移酶(ChAT)免疫组织化学检测结果：与对照组相比，大鼠MCC后1～24d上述脑区ChAT阳性细胞数和ChAT免疫反应阳性均呈现下降趋势。①ChAT阳性细胞计数结果：内侧隔核(MS)和斜角带垂直支背侧(VDBD)ChAT阳性细胞数在伤后1～24d明显减少，，与对照组相比，差异具有统计学意义(P＜0.05)。左、右侧斜角带垂直支腹侧(VDBV)ChAT阳性细胞数在伤后1d组即减少，分别在伤后4d组和8d组减至最低，然后逐渐增加至伤后24d组基本恢复正常(左侧VDBV伤后4d组与对照组比较，P＜0.05；右侧VDBV伤后4d组和8d组与对照组比较，P＜0.05)。斜角带水平支(HDB)和Meynert基底神经核ChAT阳性细胞数伤后亦减少，但与对照组比较，差异无统计学意义(P＞0.05)。被盖背外侧核ChAT阳性细胞数伤后1d组出现一过性减少，然后逐渐增加，至伤后16d组又出现明显减少，24d组减至最低(伤后16d组和24d组与对照组相比，P＜0.05)。MCC后各时间点ChAT阳性细胞数在BRF三个平面、迷走神经背核和脊髓前角运动神经元与对照组相比，差异无统计学意义(P＞0.05)。②ChAT免疫反应阳性细胞灰度值半定量结果：内侧隔核(MS)和左、右侧斜角带垂直支腹侧(VDBV)ChAT免疫反应阳性从MCC后8d组开始，与对照组相比差异具有统计学意义(P≤0.05)，内侧隔核和右侧斜角带垂直支腹侧至伤后24d组基本恢复正常(P＞0.05)，而左侧斜角带垂直支腹侧至伤后24d组仍未恢复正常(P＜0.05)。斜角带垂直支背侧(VDBD)ChAT免疫反应阳性在伤后4d组即有显著性降低，随后略升高，至伤后24d组又出现显著性降低(P＜0.05)。左右侧Meynert基底神经核ChAT免疫反应阳性仅在MCC后16d组出现显著性降低(P＜0.05)。脑干网状结构(BRF)ChAT免疫反应阳性在脑桥核平面，与对照组相比差异在伤后8d组和16d组具有统计学意义(P＜0.05)；而在斜方体核和橄榄核平面差异仅在伤后8d组具有统计学意义(P＜0.05)。ChAT免疫反应阳性在被盖背外侧核、迷走神经背核、脊髓前角运动神经元和斜角带水平支(HDB)与对照组相比，差异无统计学意义(P＞0.05)。不同部位左右侧相比差异均无统计学意义(P＞0.05)。结论：1、单摆式闭合性机械打击装置成功建立MCC大鼠模型；2、MCC大鼠存在不同程度的平衡运动与认知功能障碍，平衡功能恢复快，协调运动与认知功能恢复较慢；3、MCC大鼠基底前脑、脑干网状结构、被盖背外侧核、迷走神经背核和脊髓前角运动神经元等部位胆碱能神经元数目和活性均呈现下降趋势，以伤后8天组和16天组为重；4、中枢胆碱能神经元数目减少和表达下降可能是导致MCC大鼠认知功能障碍的重要因素之一。
[Abstract]:Objective: To establish a model of multiple cerebral concussion (MCC) rats, to observe the cognitive function of MCC rats and the changes of the cholinergic neurons in the basal forebrain, the brain stem and other related nuclei (including the lateral nucleus of the covered back, the back of the vagus nerve and the motor neuron of the anterior horn of the spinal cord). To study the relationship between the cognitive impairment and the changes of cholinergic neurons in the relevant brain regions of the triple-concussion rats, and provide the theoretical basis for the prevention and treatment of the sports-related cerebral concussion (SRCC). Methods:48 rats were randomly divided into 1 day,2 days,4 days,8 days,16 days and 24 days, and 6 triple-concussion groups (n = 8) were randomly divided into 1 day,2 days,4 days,8 days,16 days and 24 days. A group of normal controls (n = 8) was set. The balanced exercise and cognitive function of MCC rats were evaluated by beam balance test (BBT) and beam walking test (BWT). Immunohistochemical method was used to detect the expression of the catalytic enzyme _ Choline-B-transferase (ChAT) in the process of the synthesis of B-choline in the nucleus of the basal forebrain and the brain stem of the rat, and to count it under the light microscope. Results: (1) The results of BBT and BWT: The results of BBT and BWT showed that the balance time of MCC group was significantly shortened at the first day after injury (P <0.05), and the difference was statistically significant (P <0.05). The results of BWT showed that the latency of the first day of injury was significantly prolonged, and the recovery from day 2 to the 10th day had not completely returned to normal. The difference between the other time points and the control group was statistically significant (P <0.05), except for the 9th day. The results of BWT showed that the score of the first day after injury was lower, the second day began to increase gradually, and returned to normal in the fourth day, and the difference was of statistical significance (P <0.05). (2) Immunohistochemistry of ChAT (ChAT): Compared with the control group, the number of ChAT-positive cells and ChAT-positive reaction in the above-mentioned brain regions of the rat's MCC decreased in 1 to 24 days after MCC. The results showed that the number of ChAT positive cells in the medial septal nucleus (MS) and the oblique side (VDBD) of the oblique belt was significantly reduced from 1 to 24 days after the injury, and the difference was statistically significant (P <0.05) compared with the control group. The number of ChAT positive cells (VBV) in the left and right oblique sides was reduced at the first day after injury, and then decreased to the lowest in the 4th day and the 8th day after injury, and then gradually increased to 24 days after the injury (the left VDBV and the control group were compared with the control group, P <0.05). The VBV in the right side was compared with the control group at the 4th day and the 8th day after the injury (P <0.05). The number of ChAT positive cells in both the horizontal branch (HDB) and the Meynert basal nerve nucleus decreased after the injury, but the difference was not statistically significant (P> 0.05) compared with the control group. There was a transient decrease in the 1d group after the number of ChAT-positive cells in the lateral nucleus of the back of the cap, and then gradually increased to the lowest in the 16-d group after the injury (P <0.05, compared with the control group at the 16-day and the 24-day post-injury group). The number of ChAT positive cells at all time points after MCC was not significant (P> 0.05) in the three plane of BRF, the back nucleus of the vagus nerve and the motor neuron of the anterior horn of the spinal cord in comparison with the control group. The results of the semi-quantitative analysis of the gray value of the positive-positive cells of ChAT-ChAT: the positive side of the medial septal nucleus (MS) and the left and right oblique-sided (VBV) ChAT immunoreaction positive from the 8th day after the MCC, and the difference was statistically significant compared with the control group (P <0.05). The right angle of the medial septal nucleus and the right oblique angle was normal (P> 0.05) from the ventral side to the 24-day post-injury group, while the left oblique angle band was still not normal (P <0.05) from the ventral side to the 24-day post-injury group. The expression of ChAT in the vertical branch of the oblique band (VBD) was significantly lower in the 4-day post-injury group, and then slightly higher, and there was a significant decrease in the 24-d group after the injury (P <0.05). At the left and right side, the positive reaction of ChAT was significantly lower in the 16-day group after MCC (P <0.05). The immune response of the brain stem reticular structure (BRF) ChAT was positive in the nuclear plane of the pontine, and the difference between the control group and the control group was statistically significant (P <0.05). The difference between the plane of the oblique nucleus and the nucleus of the olive was statistically significant (P <0.05). The positive of ChAT was not statistically significant (P> 0.05) than that of the control group (P> 0.05) in the lateral nucleus of the dorsal nucleus, the dorsal nucleus of the vagus nerve, the motor neuron of the anterior horn of the spinal cord and the horizontal branch of the oblique band (HDB). There was no significant difference between the left and right of different parts (P> 0.05). Conclusion:1. The single pendulum closed mechanical striking device successfully established the MCC rat model;2. The MCC rats had different degrees of balance movement and cognitive dysfunction, the balance function was fast, the coordination movement and the cognitive function were slow to recover, and 3, the basal forebrain and the brain stem net structure of the MCC rat. The number and activity of cholinergic neurons in the lateral nucleus, the back nucleus of the vagus nerve and the motoneurons of the anterior horn of the spinal cord showed a downward trend, and the number and activity of the cholinergic neurons in the back of the dorsal nucleus of the vagus nerve and the anterior horn of the spinal cord showed a downward trend, The reduction of the number of central cholinergic neurons and the decrease in expression may be one of the most important factors leading to the cognitive dysfunction in the MCC rats.
【学位授予单位】：昆明医学院
【学位级别】：硕士
【学位授予年份】：2007
【分类号】：D919
本文编号：2504730
[Abstract]:Objective: To establish a model of multiple cerebral concussion (MCC) rats, to observe the cognitive function of MCC rats and the changes of the cholinergic neurons in the basal forebrain, the brain stem and other related nuclei (including the lateral nucleus of the covered back, the back of the vagus nerve and the motor neuron of the anterior horn of the spinal cord). To study the relationship between the cognitive impairment and the changes of cholinergic neurons in the relevant brain regions of the triple-concussion rats, and provide the theoretical basis for the prevention and treatment of the sports-related cerebral concussion (SRCC). Methods:48 rats were randomly divided into 1 day,2 days,4 days,8 days,16 days and 24 days, and 6 triple-concussion groups (n = 8) were randomly divided into 1 day,2 days,4 days,8 days,16 days and 24 days. A group of normal controls (n = 8) was set. The balanced exercise and cognitive function of MCC rats were evaluated by beam balance test (BBT) and beam walking test (BWT). Immunohistochemical method was used to detect the expression of the catalytic enzyme _ Choline-B-transferase (ChAT) in the process of the synthesis of B-choline in the nucleus of the basal forebrain and the brain stem of the rat, and to count it under the light microscope. Results: (1) The results of BBT and BWT: The results of BBT and BWT showed that the balance time of MCC group was significantly shortened at the first day after injury (P <0.05), and the difference was statistically significant (P <0.05). The results of BWT showed that the latency of the first day of injury was significantly prolonged, and the recovery from day 2 to the 10th day had not completely returned to normal. The difference between the other time points and the control group was statistically significant (P <0.05), except for the 9th day. The results of BWT showed that the score of the first day after injury was lower, the second day began to increase gradually, and returned to normal in the fourth day, and the difference was of statistical significance (P <0.05). (2) Immunohistochemistry of ChAT (ChAT): Compared with the control group, the number of ChAT-positive cells and ChAT-positive reaction in the above-mentioned brain regions of the rat's MCC decreased in 1 to 24 days after MCC. The results showed that the number of ChAT positive cells in the medial septal nucleus (MS) and the oblique side (VDBD) of the oblique belt was significantly reduced from 1 to 24 days after the injury, and the difference was statistically significant (P <0.05) compared with the control group. The number of ChAT positive cells (VBV) in the left and right oblique sides was reduced at the first day after injury, and then decreased to the lowest in the 4th day and the 8th day after injury, and then gradually increased to 24 days after the injury (the left VDBV and the control group were compared with the control group, P <0.05). The VBV in the right side was compared with the control group at the 4th day and the 8th day after the injury (P <0.05). The number of ChAT positive cells in both the horizontal branch (HDB) and the Meynert basal nerve nucleus decreased after the injury, but the difference was not statistically significant (P> 0.05) compared with the control group. There was a transient decrease in the 1d group after the number of ChAT-positive cells in the lateral nucleus of the back of the cap, and then gradually increased to the lowest in the 16-d group after the injury (P <0.05, compared with the control group at the 16-day and the 24-day post-injury group). The number of ChAT positive cells at all time points after MCC was not significant (P> 0.05) in the three plane of BRF, the back nucleus of the vagus nerve and the motor neuron of the anterior horn of the spinal cord in comparison with the control group. The results of the semi-quantitative analysis of the gray value of the positive-positive cells of ChAT-ChAT: the positive side of the medial septal nucleus (MS) and the left and right oblique-sided (VBV) ChAT immunoreaction positive from the 8th day after the MCC, and the difference was statistically significant compared with the control group (P <0.05). The right angle of the medial septal nucleus and the right oblique angle was normal (P> 0.05) from the ventral side to the 24-day post-injury group, while the left oblique angle band was still not normal (P <0.05) from the ventral side to the 24-day post-injury group. The expression of ChAT in the vertical branch of the oblique band (VBD) was significantly lower in the 4-day post-injury group, and then slightly higher, and there was a significant decrease in the 24-d group after the injury (P <0.05). At the left and right side, the positive reaction of ChAT was significantly lower in the 16-day group after MCC (P <0.05). The immune response of the brain stem reticular structure (BRF) ChAT was positive in the nuclear plane of the pontine, and the difference between the control group and the control group was statistically significant (P <0.05). The difference between the plane of the oblique nucleus and the nucleus of the olive was statistically significant (P <0.05). The positive of ChAT was not statistically significant (P> 0.05) than that of the control group (P> 0.05) in the lateral nucleus of the dorsal nucleus, the dorsal nucleus of the vagus nerve, the motor neuron of the anterior horn of the spinal cord and the horizontal branch of the oblique band (HDB). There was no significant difference between the left and right of different parts (P> 0.05). Conclusion:1. The single pendulum closed mechanical striking device successfully established the MCC rat model;2. The MCC rats had different degrees of balance movement and cognitive dysfunction, the balance function was fast, the coordination movement and the cognitive function were slow to recover, and 3, the basal forebrain and the brain stem net structure of the MCC rat. The number and activity of cholinergic neurons in the lateral nucleus, the back nucleus of the vagus nerve and the motoneurons of the anterior horn of the spinal cord showed a downward trend, and the number and activity of the cholinergic neurons in the back of the dorsal nucleus of the vagus nerve and the anterior horn of the spinal cord showed a downward trend, The reduction of the number of central cholinergic neurons and the decrease in expression may be one of the most important factors leading to the cognitive dysfunction in the MCC rats.
【学位授予单位】：昆明医学院
【学位级别】：硕士
【学位授予年份】：2007
【分类号】：D919
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