三重腦震蕩鼠認知行為障礙及中樞膽堿能神經(jīng)元變化研究
發(fā)布時間：2019-06-22 15:43
【摘要】： 目的：建立三重腦震蕩(multiple cerebral concussion，MCC)大鼠模型，觀察MCC大鼠的認知功能以及基底前腦、腦干網(wǎng)狀結構和其它有關核團(包括被蓋背外側核、迷走神經(jīng)背核和脊髓前角運動神經(jīng)元)膽堿能神經(jīng)元的變化，探討三重腦震蕩大鼠認知功能障礙與相關腦區(qū)膽堿能神經(jīng)元變化的關系，為運動性腦震蕩(sports-related cerebralconcussion，SRCC)的防治提供理論依據(jù)。方法：采用金屬單擺打擊裝置復制大鼠MCC模型48只，隨機分為1天、2天、4天、8天、16天和24天組共6個三重腦震蕩組(n=8)。另設一組正常對照組(n=8)。應用平衡木實驗(beam balance test，BBT)及噪音橫桿跑動實驗(beam walking test，BWT)評價MCC大鼠平衡運動與認知功能。運用免疫組織化學的方法檢測大鼠基底前腦、腦干網(wǎng)狀結構、被蓋背外側核、迷走神經(jīng)背核和脊髓前角運動神經(jīng)元中乙酰膽堿合成過程中的催化酶——膽堿乙酰轉移酶(ChAT)的表達，并在光鏡下對其計數(shù)。結果：(1)BBT及BWT實驗結果：BBT結果顯示，與對照組相比，MCC組大鼠在傷后第1天平衡時間明顯縮短，差異具有統(tǒng)計學意義(P＜0.05)，第2天開始恢復，至第4天恢復正常。BWT結果顯示，傷后第1天步入潛伏期明顯延長，第2天開始恢復，至第10天尚未完全恢復正常。除第9天外，其它各時間點與對照組相比，差異均具有統(tǒng)計學意義(P＜0.05)。BWT評分結果顯示，傷后第1天評分較低，第2天開始逐漸升高，至第4天恢復正常，差異具有統(tǒng)計學意義(P＜0.05)。(2)膽堿乙酰轉移酶(ChAT)免疫組織化學檢測結果：與對照組相比，大鼠MCC后1～24d上述腦區(qū)ChAT陽性細胞數(shù)和ChAT免疫反應陽性均呈現(xiàn)下降趨勢。①ChAT陽性細胞計數(shù)結果：內側隔核(MS)和斜角帶垂直支背側(VDBD)ChAT陽性細胞數(shù)在傷后1～24d明顯減少，，與對照組相比，差異具有統(tǒng)計學意義(P＜0.05)。左、右側斜角帶垂直支腹側(VDBV)ChAT陽性細胞數(shù)在傷后1d組即減少，分別在傷后4d組和8d組減至最低，然后逐漸增加至傷后24d組基本恢復正常(左側VDBV傷后4d組與對照組比較，P＜0.05；右側VDBV傷后4d組和8d組與對照組比較，P＜0.05)。斜角帶水平支(HDB)和Meynert基底神經(jīng)核ChAT陽性細胞數(shù)傷后亦減少，但與對照組比較，差異無統(tǒng)計學意義(P＞0.05)。被蓋背外側核ChAT陽性細胞數(shù)傷后1d組出現(xiàn)一過性減少，然后逐漸增加，至傷后16d組又出現(xiàn)明顯減少，24d組減至最低(傷后16d組和24d組與對照組相比，P＜0.05)。MCC后各時間點ChAT陽性細胞數(shù)在BRF三個平面、迷走神經(jīng)背核和脊髓前角運動神經(jīng)元與對照組相比，差異無統(tǒng)計學意義(P＞0.05)。②ChAT免疫反應陽性細胞灰度值半定量結果：內側隔核(MS)和左、右側斜角帶垂直支腹側(VDBV)ChAT免疫反應陽性從MCC后8d組開始，與對照組相比差異具有統(tǒng)計學意義(P≤0.05)，內側隔核和右側斜角帶垂直支腹側至傷后24d組基本恢復正常(P＞0.05)，而左側斜角帶垂直支腹側至傷后24d組仍未恢復正常(P＜0.05)。斜角帶垂直支背側(VDBD)ChAT免疫反應陽性在傷后4d組即有顯著性降低，隨后略升高，至傷后24d組又出現(xiàn)顯著性降低(P＜0.05)。左右側Meynert基底神經(jīng)核ChAT免疫反應陽性僅在MCC后16d組出現(xiàn)顯著性降低(P＜0.05)。腦干網(wǎng)狀結構(BRF)ChAT免疫反應陽性在腦橋核平面，與對照組相比差異在傷后8d組和16d組具有統(tǒng)計學意義(P＜0.05)；而在斜方體核和橄欖核平面差異僅在傷后8d組具有統(tǒng)計學意義(P＜0.05)。ChAT免疫反應陽性在被蓋背外側核、迷走神經(jīng)背核、脊髓前角運動神經(jīng)元和斜角帶水平支(HDB)與對照組相比，差異無統(tǒng)計學意義(P＞0.05)。不同部位左右側相比差異均無統(tǒng)計學意義(P＞0.05)。結論：1、單擺式閉合性機械打擊裝置成功建立MCC大鼠模型；2、MCC大鼠存在不同程度的平衡運動與認知功能障礙，平衡功能恢復快，協(xié)調運動與認知功能恢復較慢；3、MCC大鼠基底前腦、腦干網(wǎng)狀結構、被蓋背外側核、迷走神經(jīng)背核和脊髓前角運動神經(jīng)元等部位膽堿能神經(jīng)元數(shù)目和活性均呈現(xiàn)下降趨勢，以傷后8天組和16天組為重；4、中樞膽堿能神經(jīng)元數(shù)目減少和表達下降可能是導致MCC大鼠認知功能障礙的重要因素之一。
[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.
【學位授予單位】：昆明醫(yī)學院
【學位級別】：碩士
【學位授予年份】：2007
【分類號】：D919
本文編號：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.
【學位授予單位】：昆明醫(yī)學院
【學位級別】：碩士
【學位授予年份】：2007
【分類號】：D919
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