不同术式健侧颈7神经根移位对幼年大鼠全臂丛根性撕脱伤术后脑功能重塑影响的实验研究
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摘要
第一部分跨突触病毒示踪法研究大鼠颈7神经根中枢神经系统支配回路组成
目的研究猪假狂犬病病毒(PRV)减毒株颈7神经根注射后标记神经元在脑和脊髓的分布,为健侧颈7神经根移位术后的脑功能重塑研究奠定实验基础。
方法SD大鼠20只,根据脊髓和脑组织取材时间随机分为4组:6小时组、12小时组、24小时组、36小时组。在右侧颈7神经根注射猪假狂犬病病毒液2μl。动物存活6～36小时后,处死动物,使用抗病毒的多克隆抗体,通过免疫组织化学方法检测相应脊髓节段(C1～C7)和脑中感染神经元的分布。结果病毒注射后6～36小时,脊髓和脑中的一些结构及区域可见阳性标记细胞,主要分布在C1～C7节段脊髓灰质、外侧类巨细胞网状核、A5细胞、红核、第一二运动皮质、第一二感觉皮质等。随着动物存活时间的延长,脊髓和脑中被标记神经元的数目也明显增多。
结论本跨突触病毒示踪研究证实示踪剂标记的脊髓和脑组织结构与颈7神经根存在直接或(和)间接突触联系,可能直接或间接参与对颈7神经根的神经支配和调节。
第二部分跨突触病毒神经示踪法研究不同术式健侧颈7神经根移位术对幼年大鼠全臂丛根性撕脱伤术后脑功能重塑的影响
目的采用跨突触病毒神经示踪技术研究不同术式健侧颈7神经根移位对幼年大鼠术后脑功能重塑的影响。
方法建立三种不同术式健侧颈7神经根移位幼年大鼠动物模型,分别是:健侧颈7神经根移位至上干前股(A组);健侧颈7同时移位至肌皮和正中神经(B组);健侧颈7神经根移位至正中神经(C组)。分别于术后1.5月、3月、6月、9月和12月在受体神经注射猪假狂犬病病毒液2μl。动物存活30小时后,使用抗病毒的多克隆抗体,通过免疫组织化学方法检测运动皮层前肢支配区椎体细胞层阳性神经元分布,以各时间点左右侧运动皮层阳性神经元数目的比值为指标,分析各实验组间观测指标的差别。
结果随着术后时间的延长,各实验组大脑两侧运动皮层阳性神经元数量均逐渐增加。术后早期患肢同侧大脑运动皮层阳性神经元数量多于对侧,而后期则对侧多于同侧。各实验组术后各时间点左右两半球间阳性神经元的比值不同。健侧颈7神经根移位至上干前股组和同时移位至肌皮正中神经组术后脑功能发生跨半球重塑的速度和程度优于健侧颈7神经根移位至正中神经组(A和B之间t值为2.074,P＞0.05;A和C,B和C之间t值分别是6.494和4.420,P均＜0.01)。
结论1.幼年大鼠周围神经损伤后脑功能重塑过程中存在神经通路的改变,重塑可能与新的神经通路的形成有关。2.受体神经类型影响周围神经损伤后脑重塑,健侧颈7神经根移位至臂丛上干前股或者同时移位至肌皮和正中神经更有利于实现脑功能的跨半球重塑。
第三部分不同术式的健侧颈7神经根移位对幼年大鼠全臂丛根性撕脱伤术后脑功能重塑影响的电生理研究
目的皮层微电极刺激法研究健侧颈7神经根移位术后脑功能的重塑变化,比较不同术式健侧颈7神经根移位对幼年大鼠术后皮层重塑变化的影响。
方法建立三种不同术式健侧颈7神经根移位幼年大鼠动物模型,分别是:健侧颈7神经根移位至上干前股,健侧颈7神经根移位至肌皮和正中神经和健侧颈7神经根移位至正中神经。分别于术后1.5月、3月、6月、9月和12月,以微电极刺激技术检测患肢支配区在双侧大脑运动皮层的分布,比较各组术后脑功能重塑速度和程度的差异。
结果术后患肢支配区在运动皮层的分布是一个动态变化过程。无论是哪种健侧颈7神经根移位术式,最初患肢支配区位于同侧运动皮层;随后,患肢支配区位于双侧运动皮层;随着术后时间的进一步推移,最后患肢支配区仅位于对侧运动皮层,就术后跨半球重塑的速度和程度而言,健侧颈7神经根移位至上干前股或同时至肌皮正中神经优于健侧颈7至正中神经。
结论1.幼年大鼠健侧颈7神经根移位术后运动皮层患肢支配区可实现由同侧皮层到双侧最后到对侧皮层的跨大脑半球的功能重塑。2.健侧颈7神经根移位术受体神经的类型影响术后脑功能重塑,移位至臂丛上干前股或者同时移位至肌皮和正中神经更有利于实现脑功能的跨半球重塑。
PartⅠIdentification of CNS neural circuitry involved in the innervation of the C7 nerve root:a viral transsynaptic tracing study
Objective The distribution of labeled positive neurons in the brain and spinal cord was studied after injecting the attenuated strain of pseudorabies virus(PRV) into the C7 nerve root so as to provide a baseline for further studying of brain functional reorganization after contralateral C7 was transfered.
Method 20 adult Sprague Dawley rats were randomly divided into 4 groups according to the time of being sacrificed:6 hours,12 hours,24 hours and 36 hours groups.2ul of the virus was slowly injected into the right C7 nerve root. After survival times of 6-36 hours,the spinal cord(C1-C7) and brain were processed immunocytochemically using a polyclonal antibody against the PRV.
Result PRV labeled positive neurons were found in some spinal cord and brain regions from 6 hours to 36 hours post injection,such as the gray matter in the C1-C7 spinal cord,the lateral paragigantocellular nuclei,and the A5 cells,red nucleus,primary and secondary motor cortex,primary and secondary somatosensory cortex et al.Increasing the number in viral labeling from the spinal cord to brain regions became apparent with increasing survival time.
Conclusion The spinal cord and brain structures labelled from the C7 nerve root by the transsynaptic tracing technique are synaptically connected with C7 and presumably are involved in the neural regulation of C7.
PartⅡA viral transsynaptic tracing study on brain functional reorganization of different contralateral C7 oprative modes treat total brachial plexus avulsion in young rats
Objective To study the brain functional reorganization and compare the influence on functional reorganization by different operative modes on young rat models after contralateral C7 transfer opration by viral transsynaptic tracing technique.
Method The contralateral C7 was transferred to musculocutaneous nerve,both the musculocutaneous and median nerves and median nerve respectively in three young rat models groups 1.5,3,6,9 and 12 months later respectively,2ul PRV was injected to the recipient nerves.After 30-hours survival times,the forelimb representation of motor cortex was processed immunocytochemically,using a polyclonal antibody against the PRV to detect the positive neurons and study the difference of the amount of them among differert operative modes.
Result The amount of positive neurons increased in both motor cortex after the contralateral C7 transfer operation in all experimental groups.During earlier period,there were more positive neurons in the left motor cortex than that in the right one.Otherwise in the advanced stage the results were opposite.The funcional reorganization pace were faster in contralateral C7-to -musculocutaneous nerve group and contralateral C7-to-both the musculocutaneous and median nerves group than contralateral C7-to-median nerve group(P＜0.01).
Conclusion 1.There were changes of neural pathway in the brain functional reorganization after peripheral nerve injuries on young rats and the reorganization might be relevant with the formation of new neural pathway.2.the type of recipient nerve affected the brain functional reorganization after peripheral nerve injuries.To transfer contralateral C7 to musculocutaneous nerve or to both the musculocutaneous and median nerves were more beneficial to complete the functional reorganization across hemiphere.
PartⅢAn electrophysiologicai study on brain functional reorganization of different contralateral C7 oprative modes treat total brachial plexus avulsion in young rats
Objective To study the brain functional reorganization and compare the influence on functional reorganization by different operative modes in young rats after contralateral C7 transfer opration by microelectrode electrical stimulation.
Method The contralateral C7 was transferred to musculocutaneous nerve,both the musculocutaneous and median nerves and median nerve respectively in three young rat models groups.1.5,3,6,9 and 12months later respectively,cortical representation of repaired limb at both hemisphere motor cortex will be determined to study the difference in dynamic process of functional reorganization across hemisphere among differert operative modes.
Result A dynamic across hemispherical functional reorganization corresponding to the repaired limb was observed after operation.Cortical representation of repaired limb was firstly located at the ipsilateral cortex,and then at both sides. As the time paces,it was finally observed in the contralateral motor cortex.The funcional reorganization results were better in contralateral C7-to -musculocutaneous nerve group and contralateral C7-to-both the musculocutaneous and median nerves group.
Conclusion 1.Cortical representation of repaired limb can complete the across hemispherical functional reorganization after the contralateral C7 transfer operation in young rats.2.The pace and extent of the functional reorganization would be influnenced by the operative modes of the contralateral C7 transfer.To transfer contralateral C7 to musculocutaneous nerve or both the musculocutaneous and median nerves could lead to better functional reorganization results.
目的研究猪假狂犬病病毒(PRV)减毒株颈7神经根注射后标记神经元在脑和脊髓的分布,为健侧颈7神经根移位术后的脑功能重塑研究奠定实验基础。
方法SD大鼠20只,根据脊髓和脑组织取材时间随机分为4组:6小时组、12小时组、24小时组、36小时组。在右侧颈7神经根注射猪假狂犬病病毒液2μl。动物存活6～36小时后,处死动物,使用抗病毒的多克隆抗体,通过免疫组织化学方法检测相应脊髓节段(C1～C7)和脑中感染神经元的分布。结果病毒注射后6～36小时,脊髓和脑中的一些结构及区域可见阳性标记细胞,主要分布在C1～C7节段脊髓灰质、外侧类巨细胞网状核、A5细胞、红核、第一二运动皮质、第一二感觉皮质等。随着动物存活时间的延长,脊髓和脑中被标记神经元的数目也明显增多。
结论本跨突触病毒示踪研究证实示踪剂标记的脊髓和脑组织结构与颈7神经根存在直接或(和)间接突触联系,可能直接或间接参与对颈7神经根的神经支配和调节。
第二部分跨突触病毒神经示踪法研究不同术式健侧颈7神经根移位术对幼年大鼠全臂丛根性撕脱伤术后脑功能重塑的影响
目的采用跨突触病毒神经示踪技术研究不同术式健侧颈7神经根移位对幼年大鼠术后脑功能重塑的影响。
方法建立三种不同术式健侧颈7神经根移位幼年大鼠动物模型,分别是:健侧颈7神经根移位至上干前股(A组);健侧颈7同时移位至肌皮和正中神经(B组);健侧颈7神经根移位至正中神经(C组)。分别于术后1.5月、3月、6月、9月和12月在受体神经注射猪假狂犬病病毒液2μl。动物存活30小时后,使用抗病毒的多克隆抗体,通过免疫组织化学方法检测运动皮层前肢支配区椎体细胞层阳性神经元分布,以各时间点左右侧运动皮层阳性神经元数目的比值为指标,分析各实验组间观测指标的差别。
结果随着术后时间的延长,各实验组大脑两侧运动皮层阳性神经元数量均逐渐增加。术后早期患肢同侧大脑运动皮层阳性神经元数量多于对侧,而后期则对侧多于同侧。各实验组术后各时间点左右两半球间阳性神经元的比值不同。健侧颈7神经根移位至上干前股组和同时移位至肌皮正中神经组术后脑功能发生跨半球重塑的速度和程度优于健侧颈7神经根移位至正中神经组(A和B之间t值为2.074,P＞0.05;A和C,B和C之间t值分别是6.494和4.420,P均＜0.01)。
结论1.幼年大鼠周围神经损伤后脑功能重塑过程中存在神经通路的改变,重塑可能与新的神经通路的形成有关。2.受体神经类型影响周围神经损伤后脑重塑,健侧颈7神经根移位至臂丛上干前股或者同时移位至肌皮和正中神经更有利于实现脑功能的跨半球重塑。
第三部分不同术式的健侧颈7神经根移位对幼年大鼠全臂丛根性撕脱伤术后脑功能重塑影响的电生理研究
目的皮层微电极刺激法研究健侧颈7神经根移位术后脑功能的重塑变化,比较不同术式健侧颈7神经根移位对幼年大鼠术后皮层重塑变化的影响。
方法建立三种不同术式健侧颈7神经根移位幼年大鼠动物模型,分别是:健侧颈7神经根移位至上干前股,健侧颈7神经根移位至肌皮和正中神经和健侧颈7神经根移位至正中神经。分别于术后1.5月、3月、6月、9月和12月,以微电极刺激技术检测患肢支配区在双侧大脑运动皮层的分布,比较各组术后脑功能重塑速度和程度的差异。
结果术后患肢支配区在运动皮层的分布是一个动态变化过程。无论是哪种健侧颈7神经根移位术式,最初患肢支配区位于同侧运动皮层;随后,患肢支配区位于双侧运动皮层;随着术后时间的进一步推移,最后患肢支配区仅位于对侧运动皮层,就术后跨半球重塑的速度和程度而言,健侧颈7神经根移位至上干前股或同时至肌皮正中神经优于健侧颈7至正中神经。
结论1.幼年大鼠健侧颈7神经根移位术后运动皮层患肢支配区可实现由同侧皮层到双侧最后到对侧皮层的跨大脑半球的功能重塑。2.健侧颈7神经根移位术受体神经的类型影响术后脑功能重塑,移位至臂丛上干前股或者同时移位至肌皮和正中神经更有利于实现脑功能的跨半球重塑。
PartⅠIdentification of CNS neural circuitry involved in the innervation of the C7 nerve root:a viral transsynaptic tracing study
Objective The distribution of labeled positive neurons in the brain and spinal cord was studied after injecting the attenuated strain of pseudorabies virus(PRV) into the C7 nerve root so as to provide a baseline for further studying of brain functional reorganization after contralateral C7 was transfered.
Method 20 adult Sprague Dawley rats were randomly divided into 4 groups according to the time of being sacrificed:6 hours,12 hours,24 hours and 36 hours groups.2ul of the virus was slowly injected into the right C7 nerve root. After survival times of 6-36 hours,the spinal cord(C1-C7) and brain were processed immunocytochemically using a polyclonal antibody against the PRV.
Result PRV labeled positive neurons were found in some spinal cord and brain regions from 6 hours to 36 hours post injection,such as the gray matter in the C1-C7 spinal cord,the lateral paragigantocellular nuclei,and the A5 cells,red nucleus,primary and secondary motor cortex,primary and secondary somatosensory cortex et al.Increasing the number in viral labeling from the spinal cord to brain regions became apparent with increasing survival time.
Conclusion The spinal cord and brain structures labelled from the C7 nerve root by the transsynaptic tracing technique are synaptically connected with C7 and presumably are involved in the neural regulation of C7.
PartⅡA viral transsynaptic tracing study on brain functional reorganization of different contralateral C7 oprative modes treat total brachial plexus avulsion in young rats
Objective To study the brain functional reorganization and compare the influence on functional reorganization by different operative modes on young rat models after contralateral C7 transfer opration by viral transsynaptic tracing technique.
Method The contralateral C7 was transferred to musculocutaneous nerve,both the musculocutaneous and median nerves and median nerve respectively in three young rat models groups 1.5,3,6,9 and 12 months later respectively,2ul PRV was injected to the recipient nerves.After 30-hours survival times,the forelimb representation of motor cortex was processed immunocytochemically,using a polyclonal antibody against the PRV to detect the positive neurons and study the difference of the amount of them among differert operative modes.
Result The amount of positive neurons increased in both motor cortex after the contralateral C7 transfer operation in all experimental groups.During earlier period,there were more positive neurons in the left motor cortex than that in the right one.Otherwise in the advanced stage the results were opposite.The funcional reorganization pace were faster in contralateral C7-to -musculocutaneous nerve group and contralateral C7-to-both the musculocutaneous and median nerves group than contralateral C7-to-median nerve group(P＜0.01).
Conclusion 1.There were changes of neural pathway in the brain functional reorganization after peripheral nerve injuries on young rats and the reorganization might be relevant with the formation of new neural pathway.2.the type of recipient nerve affected the brain functional reorganization after peripheral nerve injuries.To transfer contralateral C7 to musculocutaneous nerve or to both the musculocutaneous and median nerves were more beneficial to complete the functional reorganization across hemiphere.
PartⅢAn electrophysiologicai study on brain functional reorganization of different contralateral C7 oprative modes treat total brachial plexus avulsion in young rats
Objective To study the brain functional reorganization and compare the influence on functional reorganization by different operative modes in young rats after contralateral C7 transfer opration by microelectrode electrical stimulation.
Method The contralateral C7 was transferred to musculocutaneous nerve,both the musculocutaneous and median nerves and median nerve respectively in three young rat models groups.1.5,3,6,9 and 12months later respectively,cortical representation of repaired limb at both hemisphere motor cortex will be determined to study the difference in dynamic process of functional reorganization across hemisphere among differert operative modes.
Result A dynamic across hemispherical functional reorganization corresponding to the repaired limb was observed after operation.Cortical representation of repaired limb was firstly located at the ipsilateral cortex,and then at both sides. As the time paces,it was finally observed in the contralateral motor cortex.The funcional reorganization results were better in contralateral C7-to -musculocutaneous nerve group and contralateral C7-to-both the musculocutaneous and median nerves group.
Conclusion 1.Cortical representation of repaired limb can complete the across hemispherical functional reorganization after the contralateral C7 transfer operation in young rats.2.The pace and extent of the functional reorganization would be influnenced by the operative modes of the contralateral C7 transfer.To transfer contralateral C7 to musculocutaneous nerve or both the musculocutaneous and median nerves could lead to better functional reorganization results.
引文
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