三重脑震荡鼠多巴胺神经与胆碱能神经变化关系研究
发布时间：2018-08-18 12:13
【摘要】： 目的:建立三重脑震荡(multiple cerebral concussion,MCC)大鼠模型,观察MCC大鼠大脑多巴胺能神经元和胆碱能神经元、神经纤维变化的规律。探讨与MCC大鼠认知行为相关脑区多巴胺能神经元与胆碱能神经元之间的相互关系,以了解三重脑震荡鼠神经生物学基础。方法:应用金属单摆打击装置复制大鼠MCC模型36只,大鼠随机分为1天、2天、4天、8天、16天和24天组6个三重脑震荡组(n=6),另设一正常对照组(n=6)。用抗TH和抗CHAT抗体进行免疫组化双标S-P法检测大鼠中脑黑质致密区、前扣带皮质、尾壳核、伏隔核、前额叶皮质、基底前脑、脑干网状结构、Meynert基底神经核的多巴胺能神经和胆碱能神经表达变化及二者间的作用关系。用光镜对大鼠中脑黑质致密区、前扣带皮质、尾壳核、伏隔核、前额叶皮质、基底前脑、脑干网状结构、Meynert基底神经核的多巴胺能神经和胆碱能神经进行记数,并用图像分析系统分别对上述脑区的TH和CHAT阳性细胞、纤维进行光密度分析。结果:(1)MCC后脑黑质致密区TH阳性细胞计数分析结果:在黑质致密区TH阳性细胞数以伤后1天开始上升,8天达高峰,与对照组相比2天组、4天组、8天组、16天组有显著性差异(p＜0.05),24天组基本恢复。(2)MCC后脑黑质致密区TH阳性细胞图像分析结果:TH免疫反应阳性表达在黑质致密区以伤后1天开始上升,8天达高峰,与对照组相比8天有显著性差异(p＜0.05),24天组基本恢复。(3)大鼠脑前扣带皮质、尾壳核、伏隔核、前额叶皮质和内侧隔核、斜角带垂直支、水平支TH阳性纤维光密度分析结果:与对照组相比,各损伤组TH阳性纤维上升,8天组反应达高峰,24天组基本恢复正常。尾壳核8天组,内侧隔核8天组、16天组,斜角带垂直支8天组、16天组,斜角带水平支4天组、16天组与对照组相比有统计学差异(p＜0.05)。(4)MCC后不同脑区CHAT阳性细胞数:脑干网状结构、内侧隔核(MS)、斜角带垂直支(VDB)、左、右侧斜角带水平支(HDB)ChAT阳性细胞数在MCC后1～24d明显减少,然后逐渐增加至伤后24d组基本正常。与对照组比较,内侧隔核(MS)1～24d差异有明显统计学意义(P＜0.05)。左、右斜角带垂直支(VDB)在1d、2d、4d、8d有显著差异,左、右侧斜角带水平支(HDB)在1d、2d、4d、有显著差异(P＜0.05)。Meynert基底神经核ChAT阳性细胞数伤后1d、2d明显减少,然后逐渐增加至伤后24d组基本正常。与对照组比较Meynert基底神经核1d、2d差异有明显统计学意义(P＜0.05)。左、右侧相比差异无统计学意义(P＞0.05)。(5)MCC后在基底节前脑ChAT阳性细胞光密度值:ChAT阳性细胞光密度值在MCC后1d～24d均呈现下降趋势,至4天稍有升高后,8天下降至最低。与对照组相比,内侧隔核(MS)于MCC后16d组具有统计学意义(P＜0.05)。左、右侧斜角带垂直支(VDB)、内侧隔核和右侧斜角带水平支(HDB)于MCC后8d组、16天和24天组相比较差异具有统计学意义(P＜O.05)。内侧隔核、左、右侧斜角带垂直支(VDB)至伤后24d组仍未恢复正常。左侧斜角带水平支(HDB)8d、右侧斜角带水平支(HDB)24d和对照组比较有统计学意义(P＜0.05)。左、右侧Meynert基底神经核在MCC后8d时稍有上升后在16d组出现降低,至24天组有所恢复正常。Meynert基底神经核左右侧相比差异均无统计学意义(P＞0.05)。(6)MCC后脑干网状结构(BRF)ChAT阳性细胞和TH阳性纤维光密度值:脑干网状结构各组ChAT免疫反应阳性亦呈现下降趋势,至4天稍有上升后,8天下降至最低,至24d基本恢复正常。与对照组相比,在脑桥核平面1d到24d组均具有统计学意义(P＜0.05);而在斜方体核在伤后1d、2d、8d、16d组具有统计学意义(P＜0.05);橄榄核平面1d、2d、16d具有统计学意义(P＜0.05)。脑桥核、斜方体核和橄榄核平面在MCC后TH阳性纤维1d～24d均呈现上升趋势,8天组达高峰。与对照组相比,脑桥核8d、斜方体核16d和橄榄核平面8d有统计学意义(p＜0.05)。结论:1.采用金属单摆式机械打击装置能成功建立三重脑震荡大鼠模型:2.三重脑震荡后,大鼠大脑皮质、尾壳核、伏隔核、黑质致密区、内侧隔核、斜角带、脑干核平面TH阳性表达出现不同程度的上升,以伤后8天为重。3.MCC大鼠基底前脑、脑干网状结构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。4.三重脑震荡后,大鼠大脑皮质、基底前脑、脑干核平面多个与认知记忆相关的脑区出现TH阳性纤维表达增高,以及TH免疫反应阳性表达在黑质致密区伤后上升,而以之相关部位ChAT阳性表达下降,可能是由于脑损伤后多巴胺能神经兴奋,引起TH神经元合成及释放增加,中枢胆碱能神经和多巴胺能神经二者之间可能通过纤维与胞体或胞体与胞体进行突触联系,相互调节,并具有异同作用,通过多巴胺神经抑制乙酰胆碱能神经元释放乙酰胆碱。
[Abstract]:Objective: To establish a rat model of multiple cerebral concussion (MCC) and observe the changes of dopaminergic neurons, cholinergic neurons and nerve fibers in the brain of MCC rats. METHODS: Thirty-six MCC rats were randomly divided into 6 triple concussion groups (n=6) and a normal control group (n=6) on 1, 2, 4, 8, 16 and 24 days. The dense areas of substantia nigra and anterior cingulate were detected by double-labeled S-P immunohistochemical method with anti-TH and anti-CHAT antibodies. The expression of dopaminergic and cholinergic nerves in the cortex, caudate putamen nucleus, nucleus accumbens, prefrontal cortex, basal forebrain, brainstem reticular structure, Meynert basal nucleus and the relationship between them were studied. Dopaminergic and cholinergic nerves in the basal ganglia of Meynert were counted and the optical density of TH and CHAT positive cells and fibers in these areas were analyzed by image analysis system. Compared with the control group, there was a significant difference (p < 0.05) in 2 days group, 4 days group, 8 days group and 16 days group, and the positive expression of TH cells in substantia nigra compact area was basically restored in 24 days group. The optical density of TH-positive fibers in anterior cingulate cortex, caudate putamen nucleus, nucleus accumbens, prefrontal cortex and medial septal nucleus, perpendicular branch of oblique zone, and horizontal branch in rats were analyzed. Compared with the control group, the TH-positive fibers in each injury group increased, the reaction in 8 days group reached the peak, and the reaction in 24 days group basically returned to normal. There were significant differences in the number of CHAT positive cells in different brain regions after MCC: brainstem reticular structure, medial septal nucleus (MS), diagonal vertical branch (VDB), left and right diagonal horizontal branch (HDB) ChAT positive cells. Compared with the control group, the medial septal nucleus (MS) was significantly different (P The number of ChAT positive cells in the basal ganglia decreased significantly one day and two days after injury, and then gradually increased to normal 24 days after injury. Compared with the control group, the medial septal nucleus (MS) was significantly higher at 16 days after MCC (P < 0.05). The vertical branch of left and right diagonal band (VDB), medial septal nucleus and horizontal branch of right diagonal band (HDB) were significantly lower at 8 days, 16 days and 24 days after MCC. The medial septal nucleus, the vertical branch of left and right diagonal band (VDB) did not return to normal until 24 days after injury. The horizontal branch of left diagonal band (HDB) 8 days, the horizontal branch of right diagonal band (HDB) 24 days compared with the control group were statistically significant (P < 0.05). There was no significant difference between the left and right sides of the basal nucleus of Meynert (P > 0.05). (6) Optical density values of ChAT positive cells and TH positive fibers in brainstem reticular structure (BRF) after MCC: ChAT immunoreactivity in each group of brainstem reticular structure also showed a downward trend, and increased slightly after 4 days, 8 days. Compared with the control group, there were significant differences in pontine nucleus level from 1 day to 24 days (P < 0.05), in trapezoidal nucleus at 1 day, 2 days, 8 days and 16 days after injury (P < 0.05), and in olivary nucleus level at 1 day, 2 days and 16 days (P < 0.05). TH-positive fibers showed an upward trend from 1D to 24D after MCC, reaching a peak at 8D. Compared with the control group, the pontine nucleus at 8d, the trapezoidal nucleus at 16d and the olivary nucleus at 8D had statistical significance (p < 0.05). CONCLUSION: 1. Triple concussion rat model was successfully established by using metal pendulum mechanical percussion device: 2. TH positive expression in putamen nucleus, nucleus accumbens, substantia nigra dense area, medial septal nucleus, diagonal zone and brain stem nucleus increased in varying degrees. The number and activity of cholinergic neurons in basal forebrain and brainstem reticular structure of MCC rats decreased at 8 days and 16 days after injury. 4. After triple concussion, the number and activity of cholinergic neurons in the brain of MCC rats decreased. The increased expression of TH-positive fibers in the cortex, basal forebrain and brainstem nucleus, and the increased expression of TH-immunoreactive fibers in the substantia nigra dense area were observed, whereas the decrease of ChAT-positive expression in the related areas might be due to the increased synthesis and release of TH neurons after brain injury. Additionally, the central cholinergic nerve and the dopaminergic nerve may have synaptic connections with the cell body or the cell body through the fibers, and they may regulate each other, and have similar and different effects. The dopaminergic nerve inhibits the release of acetylcholine from the acetylcholinergic neurons through the dopaminergic nerve.
【学位授予单位】：昆明医学院
【学位级别】：硕士
【学位授予年份】：2008
【分类号】：R651.15;D919
本文编号：2189436
[Abstract]:Objective: To establish a rat model of multiple cerebral concussion (MCC) and observe the changes of dopaminergic neurons, cholinergic neurons and nerve fibers in the brain of MCC rats. METHODS: Thirty-six MCC rats were randomly divided into 6 triple concussion groups (n=6) and a normal control group (n=6) on 1, 2, 4, 8, 16 and 24 days. The dense areas of substantia nigra and anterior cingulate were detected by double-labeled S-P immunohistochemical method with anti-TH and anti-CHAT antibodies. The expression of dopaminergic and cholinergic nerves in the cortex, caudate putamen nucleus, nucleus accumbens, prefrontal cortex, basal forebrain, brainstem reticular structure, Meynert basal nucleus and the relationship between them were studied. Dopaminergic and cholinergic nerves in the basal ganglia of Meynert were counted and the optical density of TH and CHAT positive cells and fibers in these areas were analyzed by image analysis system. Compared with the control group, there was a significant difference (p < 0.05) in 2 days group, 4 days group, 8 days group and 16 days group, and the positive expression of TH cells in substantia nigra compact area was basically restored in 24 days group. The optical density of TH-positive fibers in anterior cingulate cortex, caudate putamen nucleus, nucleus accumbens, prefrontal cortex and medial septal nucleus, perpendicular branch of oblique zone, and horizontal branch in rats were analyzed. Compared with the control group, the TH-positive fibers in each injury group increased, the reaction in 8 days group reached the peak, and the reaction in 24 days group basically returned to normal. There were significant differences in the number of CHAT positive cells in different brain regions after MCC: brainstem reticular structure, medial septal nucleus (MS), diagonal vertical branch (VDB), left and right diagonal horizontal branch (HDB) ChAT positive cells. Compared with the control group, the medial septal nucleus (MS) was significantly different (P The number of ChAT positive cells in the basal ganglia decreased significantly one day and two days after injury, and then gradually increased to normal 24 days after injury. Compared with the control group, the medial septal nucleus (MS) was significantly higher at 16 days after MCC (P < 0.05). The vertical branch of left and right diagonal band (VDB), medial septal nucleus and horizontal branch of right diagonal band (HDB) were significantly lower at 8 days, 16 days and 24 days after MCC. The medial septal nucleus, the vertical branch of left and right diagonal band (VDB) did not return to normal until 24 days after injury. The horizontal branch of left diagonal band (HDB) 8 days, the horizontal branch of right diagonal band (HDB) 24 days compared with the control group were statistically significant (P < 0.05). There was no significant difference between the left and right sides of the basal nucleus of Meynert (P > 0.05). (6) Optical density values of ChAT positive cells and TH positive fibers in brainstem reticular structure (BRF) after MCC: ChAT immunoreactivity in each group of brainstem reticular structure also showed a downward trend, and increased slightly after 4 days, 8 days. Compared with the control group, there were significant differences in pontine nucleus level from 1 day to 24 days (P < 0.05), in trapezoidal nucleus at 1 day, 2 days, 8 days and 16 days after injury (P < 0.05), and in olivary nucleus level at 1 day, 2 days and 16 days (P < 0.05). TH-positive fibers showed an upward trend from 1D to 24D after MCC, reaching a peak at 8D. Compared with the control group, the pontine nucleus at 8d, the trapezoidal nucleus at 16d and the olivary nucleus at 8D had statistical significance (p < 0.05). CONCLUSION: 1. Triple concussion rat model was successfully established by using metal pendulum mechanical percussion device: 2. TH positive expression in putamen nucleus, nucleus accumbens, substantia nigra dense area, medial septal nucleus, diagonal zone and brain stem nucleus increased in varying degrees. The number and activity of cholinergic neurons in basal forebrain and brainstem reticular structure of MCC rats decreased at 8 days and 16 days after injury. 4. After triple concussion, the number and activity of cholinergic neurons in the brain of MCC rats decreased. The increased expression of TH-positive fibers in the cortex, basal forebrain and brainstem nucleus, and the increased expression of TH-immunoreactive fibers in the substantia nigra dense area were observed, whereas the decrease of ChAT-positive expression in the related areas might be due to the increased synthesis and release of TH neurons after brain injury. Additionally, the central cholinergic nerve and the dopaminergic nerve may have synaptic connections with the cell body or the cell body through the fibers, and they may regulate each other, and have similar and different effects. The dopaminergic nerve inhibits the release of acetylcholine from the acetylcholinergic neurons through the dopaminergic nerve.
【学位授予单位】：昆明医学院
【学位级别】：硕士
【学位授予年份】：2008
【分类号】：R651.15;D919
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