电磁辐射对神经突触可塑性影响的分子机制研究
发布时间：2018-08-18 08:16
【摘要】：影响电磁辐射健康效应的主要参数包括频率和强度。不同频率的电磁波来源、用途不同,其健康效应也不尽相同。微波(300MHz-300GHz)电磁辐射属于非电离辐射,900MHz是国际通用的民用无线通讯频段。高强度微波电磁辐射(EMR)通过“致热效应”引起机体的健康危害已经得到有关研究的证实,而低强度微波辐射,尤其是900MHz微波电磁辐射,长期暴露的健康效应及其机理目前尚有争议。国际上关于微波电磁辐射的研究焦点主要是对中枢神经系统(CNS)肿瘤和行为功能影响的研究,流行病学调查表明,电磁辐射的过量接触可导致中枢神经系统和植物神经功能紊乱,且动物实验表明,电磁辐射可以引起学习记忆及认知能力障碍。 为了进一步探讨900MHz微波电磁辐射对中枢神经系统功能影响及其作用机理,本实验选用2000μW/cm2的辐射强度,采用体内动物实验及体外原代神经元细胞培养的方法,从突触结构可塑性(脑区突触数目和结构的改变等)和功能可塑性(神经元和突触部位的某些受体的变化)两方面入手,探讨电磁辐射对学习记忆影响的机制。 第一部分EMR后对大鼠学习记忆功能的影响及组织病理学观察 目的： 探讨2000μW/cm2电磁辐射对大鼠学习记忆功能的影响。 方法： 实验分为空白对照组,假辐射组,1h/d、2h/d、3h/d辐射组。将辐射组大鼠固定体位,头部接受功率密度为2000μW/cm2的近场辐射,连续辐射30d。通过Morris水迷宫检测大鼠的学习记忆能力,采用硫堇染色法观察大鼠海马组织结构的变化。 结果： 1行为学检测结果表明,假辐射组大鼠各项指标与空白对照组相比均无明显变化(P0.05)；而各辐射组大鼠的逃避潜伏期较空白对照组明显延长(P0.05),在空间探索实验中各辐射组大鼠的空间探索次数明显减少(P0.05)。 2形态学检测结果表明,假辐射组海马神经元形态与数目与空白对照组相比均无明显差异；而辐射组大鼠海马CA1区神经细胞数量显著减少(P0.05),且细胞排列紊乱,胞浆尼氏体明显减少,细胞核固缩。 结论： 2000μW/cm2电磁辐射可导致大鼠学习记忆功能下降,其机制可能与损伤大鼠海马神经元有关。 第二部分EMR对大鼠海马超微结构及凋亡相关因子表达的影响 目的： 探讨2000μW/cm2电磁辐射对大鼠海马神经元超微结构及凋亡相关因子表达的影响。 方法： 实验分为空白对照组,假辐射组,1h/d、2h/d、3h/d辐射组。将辐射组大鼠固定体位,头部接受功率密度为2000μW/cm2的近场辐射,连续辐射30d。通过透射电镜观察海马超微结构改变,采用免疫组织化学法检测大鼠海马组织Bcl-2、Bax的表达变化,Western blotting法检测海马组织Caspase-3蛋白的表达变化。 结果： 辐射组海马细胞核仁消失,染色质呈颗粒、块状凝集在核膜下,电子密度高,核内出现透明区等早期凋亡征象,突触界面结构出现明显病理性改变；与空白对照组比较,辐射组大鼠海马组织Bax、Caspase-3蛋白表达升高(P0.05),Bcl-2蛋白表达降低,Bax/Bcl-2比值显著升高(P0.05)。 结论： 电磁辐射可引起海马神经元细胞损伤和凋亡,而机制可能与凋亡相关因子Bcl-2、Bax、Caspase-3表达变化相关。 第三部分EMR对大鼠海马脑区NMDA受体蛋白及其mRNA表达的影响 目的： 探讨2000μW/cm2电磁辐射对大鼠海马(?)NMDA(NR1、NR2A和NR2B)受体蛋白及其mRNA水平表达的影响,揭示电磁辐射对大鼠学习记忆功能的损伤机制。 方法： 实验分为空白对照组,假辐射组,1h/d、2h/d、3h/d辐射组。将辐射组大鼠固定体位,头部接受功率密度为2000μW/cm2的近场辐射,连续辐射30d。采用免疫组化法和Western blotting法检测大鼠海马组织NMDA (NR1、NR2A和NR2B)受体蛋白表达的变化,RT-PCR法检测大鼠海马组织NMDA (NR1、NR2A和NR2B)mRNA表达的变化。 结果： 各辐射组大鼠海马神经元排列紊乱,NMDA (NR1、NR2A和NR2B)阳性细胞比率显著下降,海马组织NMDA (NR1、NR2A和NR2B)蛋白及其mRNA表达水平明显降低(P0.05)。 结论： 2000μW/cm2电磁辐射可导致大鼠学习记忆功能下降,其机制可能与大鼠海马组织NMDA (NR1、NR2A和NR2B)蛋白及其mRNA的表达降低有关。 第四部分EMR致海马神经细胞CREB和CaMKⅡ表达的影响 目的： 探讨2000μW/cm2电磁辐射对原代培养海马神经元细胞CREB、 CaMKⅡ表达的影响,揭示电磁辐射对大鼠海马神经细胞的损伤机制。 方法： 取原代培养7d的大鼠海马神经元,随机分为5组：分为空白对照组,假辐射组,1h/d、2h/d、3h/d辐射组。将辐射组海马神经细胞接受功率密度为2000μW/cm2的近场辐射,连续辐射5d。通过倒置显微镜观察海马神经元形态变化,MTT法检测神经元细胞活力,采用Western blotting法检测大鼠海马神经细胞CREB和CaMKII蛋白表达的变化,RT-PCR法检测大鼠海马神经细胞CREB和CaMKII mRNA表达的变化。 结果： 正常对照组大鼠海马神经元核规整,细胞突起明显；辐射组大鼠海马神经元核皱缩、突起回缩、变性。假辐射组大鼠各项指标与空白对照组相比均无显著差异(P0.05)；而各辐射组大鼠海马神经元细胞活力明显下降(P0.05)；CREB和CaMKⅡ蛋白及其mRNA表达水平明显降低(P0.05)。 结论： 2000μW/cm2电磁辐射致大鼠海马神经细胞损伤机制可能与CREB和CaMKⅡ蛋白及其mRNA的表达降低有关。
[Abstract]:The main parameters affecting the health effects of electromagnetic radiation include frequency and intensity. Different sources and uses of electromagnetic waves have different health effects. Microwave (300MHz-300GHz) electromagnetic radiation belongs to non-ionizing radiation, 900MHz is the international common civil wireless communication frequency band. High intensity microwave electromagnetic radiation (EMR) passes through "heating effect". "The health hazards caused by microwave radiation, especially 900MHz microwave electromagnetic radiation, have been confirmed by relevant studies. The long-term health effects and their mechanisms of exposure to microwave electromagnetic radiation are still controversial. Epidemiological studies have shown that excessive exposure to electromagnetic radiation can lead to functional disorders of the central nervous system and autonomic nervous system, and animal experiments have shown that electromagnetic radiation can cause learning and memory and cognitive impairment.
In order to further explore the effect of 900MHz microwave electromagnetic radiation on the function of the central nervous system and its mechanism, we used the radiation intensity of 2000 mu W/cm 2 to study the plasticity of synaptic structure (the number and structure of synapses in the brain area, etc.) and functional plasticity (neural plasticity) by in vivo animal experiment and in vitro primary culture of neurons. The mechanism of the effect of electromagnetic radiation on learning and memory was discussed.
Part one the effect of EMR on learning and memory in rats and its histopathological observation.
Objective:
Objective to investigate the effect of 2000 W/cm2 electromagnetic radiation on learning and memory function in rats.
Method:
The rats in the radiation group were placed in a fixed position and received near-field radiation with a power density of 2000 mu W/cm2 for 30 days. The learning and memory abilities of the rats were measured by Morris water maze, and the hippocampal tissues were observed by thionine staining.
Result:
The results of behavioral tests showed that there were no significant changes in the indexes of rats in the sham radiation group compared with the blank control group (P 0.05), but the escape latency of rats in each radiation group was significantly longer than that in the blank control group (P 0.05), and the number of space explorations in each radiation group was significantly reduced (P 0.05).
The results of morphological examination showed that there was no significant difference in the morphology and number of hippocampal neurons between the sham radiation group and the blank control group, but the number of neurons in the CA1 area of the hippocampus in the radiation group was significantly reduced (P 0.05), and the cell arrangement was disordered, the cytoplasmic Nissl body was significantly reduced, and the nucleus was condensed.
Conclusion:
2 000 mu W/cm 2 electromagnetic radiation can lead to the decline of learning and memory function in rats, which may be related to the damage of hippocampal neurons.
The second part is the effect of EMR on the ultrastructure of hippocampus and the expression of apoptosis related factors in rats.
Objective:
Objective To investigate the effects of 2 000 mu W/cm 2 electromagnetic radiation on ultrastructure and expression of apoptosis-related factors in hippocampal neurons of rats.
Method:
The experiment was divided into three groups: control group, sham radiation group, 1h/d, 2h/d, 3h/d radiation group. The rats in radiation group were fixed in position and received near-field radiation with a power density of 2000 mu W/cm 2 for 30 days. Blotting method was used to detect the expression of Caspase-3 protein in hippocampus.
Result:
In radiation group, nucleolus disappeared, chromatin was granular, mass agglutinated in the subnuclear membrane, high electron density, transparent area appeared in the nucleus and other early signs of apoptosis, synaptic interface structure showed obvious pathological changes; compared with the blank control group, the expression of Bax, Caspase-3 protein in the hippocampus of radiation group increased (P 0.05), and the expression of Bcl-2 protein decreased (P 0.05). The Bax/Bcl-2 ratio increased significantly (P0.05).
Conclusion:
Electromagnetic radiation can induce injury and apoptosis of hippocampal neurons, and the mechanism may be related to the expression of apoptosis-related factors Bcl-2, Bax and Caspase-3.
The third part is the effect of EMR on the expression of NMDA receptor protein and mRNA in rat hippocampus.
Objective:
Objective To investigate the effects of 2 000 mu W/cm 2 electromagnetic radiation on the expression of (?) NMDA (NR1, NR2A and NR2B) receptor protein and its mRNA in rat hippocampus, and to reveal the mechanism of learning and memory impairment induced by electromagnetic radiation.
Method:
The experiment was divided into three groups: control group, sham radiation group, 1h/d, 2h/d, 3h/d radiation group. The rats in radiation group were fixed in position and received near-field radiation with a power density of 2000 mu W/cm2 for 30 days. The expression of NMDA (NR1, NR2A and NR2B) receptors in hippocampus was detected by immunohistochemistry and Western blotting, and RT-PCR. Changes of NMDA (NR1, NR2A and NR2B) mRNA expression in rat hippocampus.
Result:
The distribution of hippocampal neurons was disordered, the ratio of NMDA (NR1, NR2A and NR2B) positive cells decreased significantly, and the expression levels of NMDA (NR1, NR2A and NR2B) protein and its mRNA in hippocampus decreased significantly (P 0.05).
Conclusion:
2000 mu W/cm2 electromagnetic radiation can induce the decline of learning and memory function in rats. The mechanism may be related to the decrease of NMDA (NR1, NR2A and NR2B) protein and its mRNA expression in rat hippocampus.
The fourth part is the effect of EMR on the expression of CREB and CaMK II in hippocampal neurons.
Objective:
Objective To investigate the effect of 2 000 mu W/cm 2 electromagnetic radiation on the expression of CREB and CaMK II in primary cultured hippocampal neurons and to reveal the mechanism of injury of electromagnetic radiation on hippocampal neurons in rats.
Method:
The primary cultured hippocampal neurons were randomly divided into five groups: blank control group, sham radiation group, 1h/d, 2h/d, 3h/d radiation group. Western blotting was used to detect the expression of CREB and CaMKII protein in rat hippocampal neurons. RT-PCR was used to detect the expression of CREB and CaMKII mRNA in rat hippocampal neurons.
Result:
In the normal control group, the hippocampal neurons had regular nuclei and distinct cellular processes; in the irradiated group, the hippocampal neurons had shrinkage, retraction and degeneration; there was no significant difference in all the indexes between the sham-irradiated group and the blank control group (P 0.05); however, the activity of the hippocampal neurons in the irradiated groups decreased significantly (P 0.05); the CREB and CaMK II proteins in the irradiated group were And the expression level of mRNA decreased significantly (P0.05).
Conclusion:
The injury mechanism of hippocampal neurons induced by 2 000 mu W/cm 2 electromagnetic radiation may be related to the decrease of CRB and CaMK II protein and its mRNA expression.
【学位授予单位】：承德医学院
【学位级别】：硕士
【学位授予年份】：2011
【分类号】：R363
本文编号：2188899
[Abstract]:The main parameters affecting the health effects of electromagnetic radiation include frequency and intensity. Different sources and uses of electromagnetic waves have different health effects. Microwave (300MHz-300GHz) electromagnetic radiation belongs to non-ionizing radiation, 900MHz is the international common civil wireless communication frequency band. High intensity microwave electromagnetic radiation (EMR) passes through "heating effect". "The health hazards caused by microwave radiation, especially 900MHz microwave electromagnetic radiation, have been confirmed by relevant studies. The long-term health effects and their mechanisms of exposure to microwave electromagnetic radiation are still controversial. Epidemiological studies have shown that excessive exposure to electromagnetic radiation can lead to functional disorders of the central nervous system and autonomic nervous system, and animal experiments have shown that electromagnetic radiation can cause learning and memory and cognitive impairment.
In order to further explore the effect of 900MHz microwave electromagnetic radiation on the function of the central nervous system and its mechanism, we used the radiation intensity of 2000 mu W/cm 2 to study the plasticity of synaptic structure (the number and structure of synapses in the brain area, etc.) and functional plasticity (neural plasticity) by in vivo animal experiment and in vitro primary culture of neurons. The mechanism of the effect of electromagnetic radiation on learning and memory was discussed.
Part one the effect of EMR on learning and memory in rats and its histopathological observation.
Objective:
Objective to investigate the effect of 2000 W/cm2 electromagnetic radiation on learning and memory function in rats.
Method:
The rats in the radiation group were placed in a fixed position and received near-field radiation with a power density of 2000 mu W/cm2 for 30 days. The learning and memory abilities of the rats were measured by Morris water maze, and the hippocampal tissues were observed by thionine staining.
Result:
The results of behavioral tests showed that there were no significant changes in the indexes of rats in the sham radiation group compared with the blank control group (P 0.05), but the escape latency of rats in each radiation group was significantly longer than that in the blank control group (P 0.05), and the number of space explorations in each radiation group was significantly reduced (P 0.05).
The results of morphological examination showed that there was no significant difference in the morphology and number of hippocampal neurons between the sham radiation group and the blank control group, but the number of neurons in the CA1 area of the hippocampus in the radiation group was significantly reduced (P 0.05), and the cell arrangement was disordered, the cytoplasmic Nissl body was significantly reduced, and the nucleus was condensed.
Conclusion:
2 000 mu W/cm 2 electromagnetic radiation can lead to the decline of learning and memory function in rats, which may be related to the damage of hippocampal neurons.
The second part is the effect of EMR on the ultrastructure of hippocampus and the expression of apoptosis related factors in rats.
Objective:
Objective To investigate the effects of 2 000 mu W/cm 2 electromagnetic radiation on ultrastructure and expression of apoptosis-related factors in hippocampal neurons of rats.
Method:
The experiment was divided into three groups: control group, sham radiation group, 1h/d, 2h/d, 3h/d radiation group. The rats in radiation group were fixed in position and received near-field radiation with a power density of 2000 mu W/cm 2 for 30 days. Blotting method was used to detect the expression of Caspase-3 protein in hippocampus.
Result:
In radiation group, nucleolus disappeared, chromatin was granular, mass agglutinated in the subnuclear membrane, high electron density, transparent area appeared in the nucleus and other early signs of apoptosis, synaptic interface structure showed obvious pathological changes; compared with the blank control group, the expression of Bax, Caspase-3 protein in the hippocampus of radiation group increased (P 0.05), and the expression of Bcl-2 protein decreased (P 0.05). The Bax/Bcl-2 ratio increased significantly (P0.05).
Conclusion:
Electromagnetic radiation can induce injury and apoptosis of hippocampal neurons, and the mechanism may be related to the expression of apoptosis-related factors Bcl-2, Bax and Caspase-3.
The third part is the effect of EMR on the expression of NMDA receptor protein and mRNA in rat hippocampus.
Objective:
Objective To investigate the effects of 2 000 mu W/cm 2 electromagnetic radiation on the expression of (?) NMDA (NR1, NR2A and NR2B) receptor protein and its mRNA in rat hippocampus, and to reveal the mechanism of learning and memory impairment induced by electromagnetic radiation.
Method:
The experiment was divided into three groups: control group, sham radiation group, 1h/d, 2h/d, 3h/d radiation group. The rats in radiation group were fixed in position and received near-field radiation with a power density of 2000 mu W/cm2 for 30 days. The expression of NMDA (NR1, NR2A and NR2B) receptors in hippocampus was detected by immunohistochemistry and Western blotting, and RT-PCR. Changes of NMDA (NR1, NR2A and NR2B) mRNA expression in rat hippocampus.
Result:
The distribution of hippocampal neurons was disordered, the ratio of NMDA (NR1, NR2A and NR2B) positive cells decreased significantly, and the expression levels of NMDA (NR1, NR2A and NR2B) protein and its mRNA in hippocampus decreased significantly (P 0.05).
Conclusion:
2000 mu W/cm2 electromagnetic radiation can induce the decline of learning and memory function in rats. The mechanism may be related to the decrease of NMDA (NR1, NR2A and NR2B) protein and its mRNA expression in rat hippocampus.
The fourth part is the effect of EMR on the expression of CREB and CaMK II in hippocampal neurons.
Objective:
Objective To investigate the effect of 2 000 mu W/cm 2 electromagnetic radiation on the expression of CREB and CaMK II in primary cultured hippocampal neurons and to reveal the mechanism of injury of electromagnetic radiation on hippocampal neurons in rats.
Method:
The primary cultured hippocampal neurons were randomly divided into five groups: blank control group, sham radiation group, 1h/d, 2h/d, 3h/d radiation group. Western blotting was used to detect the expression of CREB and CaMKII protein in rat hippocampal neurons. RT-PCR was used to detect the expression of CREB and CaMKII mRNA in rat hippocampal neurons.
Result:
In the normal control group, the hippocampal neurons had regular nuclei and distinct cellular processes; in the irradiated group, the hippocampal neurons had shrinkage, retraction and degeneration; there was no significant difference in all the indexes between the sham-irradiated group and the blank control group (P 0.05); however, the activity of the hippocampal neurons in the irradiated groups decreased significantly (P 0.05); the CREB and CaMK II proteins in the irradiated group were And the expression level of mRNA decreased significantly (P0.05).
Conclusion:
The injury mechanism of hippocampal neurons induced by 2 000 mu W/cm 2 electromagnetic radiation may be related to the decrease of CRB and CaMK II protein and its mRNA expression.
【学位授予单位】：承德医学院
【学位级别】：硕士
【学位授予年份】：2011
【分类号】：R363
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