电针刺激对成年大鼠视神经切断后视网膜神经节细胞存活的保护及机制探讨
发布时间:2018-02-28 13:44
本文关键词: 视神经切断 电针 视网膜神经节细胞 神经元存活 谷氨酸 谷氨酸转运体-1 出处:《第四军医大学》2012年硕士论文 论文类型:学位论文
【摘要】:目的:研究不同穴位和频率的电针刺激对成年大鼠视神经切断后视网膜神经节细胞(节细胞)存活的作用,并探讨电针刺激对受损节细胞的保护机制。 实验一:随机选取54只SD大鼠,切断右侧视神经并用荧光金(FluoroGold, FG)逆行性标记节细胞后,随机分为9组:对照组(含2、7和14d对照组)及不同穴位、频率和伤后存活时间组合的电针组(含假穴4/20Hz电针7d组、百会穴4/20Hz电针7d组、睛明穴2Hz电针7d组、睛明穴4/20Hz电针2、7、14d组),每组6只动物。对应时间点处死动物取右眼视网膜后计数被荧光金标记的节细胞并计算相应的密度(细胞/mm2),数值用Mean±SEM表示。结果:(1)睛明穴4/20Hz电针7d组存活节细胞平均密度(1600±72)显著高于对照(1104±41)、假穴(1222±68)和百会穴(1168±56)组(P0.01),而假穴和百会穴组与对照组间无显著性差异;(2)睛明穴2Hz与睛明穴4/20Hz电针7d组节细胞密度虽无显著性差异,但均显著高于对照7d组(P0.01);(3)分别以4/20Hz电针刺激睛明穴2、7、14d,与对照组相同时间点相比,仅在伤后7d出现节细胞密度的显著增高(P0.01)。 实验二:随机选取60只大鼠并随机分为10组,即正常组、假手术2、7、14d组(手术暴露右侧视神经但不切断),,另外36只大鼠切断右侧视神经后再随机分为2、7、14d对照组和2、7、14d电针组(即2、7、14d睛明穴4/20Hz电针组),每组6只动物。于各组相应时间点麻醉动物后取动物右眼收集房水,并利用高效液相色谱检测其谷氨酸浓度(μmol/L),数值用Mean±SEM表示。结果:对照2、7、14d组(浓度分别为59.74±4.62、88.17±1.94、56.56±1.15)及电针2、7、14d组(浓度分别为47.62±4.29、54.72±1.98、52.27±1.16)房水谷氨酸浓度均显著高于正常组及2、7、14d假手术组(浓度分别为43.54±0.53、43.71±0.66、43.21±0.94、43.04±0.44)(P0.01);电针2、7d组房水谷氨酸浓度分别显著低于对照组2、7d组(P0.01);电针14d组房水谷氨酸浓度与对照14d组无差异。 实验三:随机选取54只大鼠切断右侧视神经后,随机分为6组:2、7、14d对照组和2、7、14d电针组(即2、7、14d睛明穴4/20Hz电针组),每组9只动物。分别利用免疫组织化学染色、Western-blot方法检测大鼠视网膜中分别参与谷氨酸重吸收与代谢的谷氨酸转运体-1和谷氨酰胺合成酶的表达。结果:电针7d组谷氨酸转运体-1表达显著高于对照7d组(P0.05);对照2、14d组分别与电针2、14d组的谷氨酸转运体-1表达无显著差异。视神经切断后电针组和对照组谷氨酰胺合成酶表达没有发生变化。 结论:切断成年大鼠视神经后每日以4/20Hz频率的电针刺激睛明穴1次,可在伤后7d促进视网膜谷氨酸转运体-1对伤侧眼房水内谷氨酸的重吸收,从而降低房水内谷氨酸的含量,减轻谷氨酸兴奋性毒性,延缓受损节细胞的死亡。
[Abstract]:Aim: to study the effects of electroacupuncture stimulation at different points and frequencies on the survival of retinal ganglion cells (ganglion cells) after optic nerve transection in adult rats and to explore the protective mechanism of electroacupuncture stimulation on injured ganglion cells. Experiment 1: Fifty-four Sprague-Dawley rats were randomly divided into 9 groups: control group (control group) and different acupoints after transection of the right optic nerve and fluorescent Fluoro Gold (FG) retrograde labeled ganglion cells. The frequency and survival time after injury were combined with electroacupuncture group (including 4 / 20 Hz electroacupuncture group for 7 days, Baihui point 4 / 20 Hz electroacupuncture group for 7 days, Jinming point 2 Hz electroacupuncture group for 7 days). There were 6 animals in each group after 4 / 20 Hz electroacupuncture for 14 days. The right eye retina was killed at the corresponding time point to count the fluorescent gold labeled ganglion cells and calculate the corresponding density (cell per mm2, expressed in Mean 卤SEM. The result was 1: 1). The mean cell density of surviving ganglion cells in the electroacupuncture group was significantly higher than that in the control group (1104 卤41g), the control group (1222 卤68) and the Baihui acupoint group (1168 卤56), but there was no significant difference between the sham acupoint group and Baihui acupoint group and the control group (P 0.01), although there was no significant difference in cell density between the Jingming acupoint (2Hz) and Jinming acupoint (4 / 20Hz) after 7 d electroacupuncture, although there was no significant difference in cell density between the sham acupoint and Baihui acupoint group and the control group. But it was significantly higher than that of the control group on the 7th day (P 0.01). Compared with the control group at the same time point, there was a significant increase in the cell density of the ganglion on the 7th day after injury (P 0.01) after 4 / 20 Hz electroacupuncture stimulation on the Jingming acupoint for 2 ~ 7 ~ 14 days, respectively, and compared with the control group at the same time point, there was a significant increase in the cell density. Experiment 2: 60 rats were randomly selected and randomly divided into 10 groups, that is, the normal group. The rats in the sham operation group (the right optic nerve was exposed without transection of the right optic nerve, and the other 36 rats were randomly divided into two groups: the control group, the control group, and the electroacupuncture group, which were treated with electroacupuncture on the 2nd day, 7d, 14d, respectively), each group had 6 animals in each group, and the other 36 rats were divided into two groups: the control group, the control group, and the electroacupuncture group, which had 4 / 20 Hz electroacupuncture on the right optic nerve. After anaesthetized animals were anesthetized, aqueous humor was collected from the right eye of the animals. The concentration of glutamic acid (渭 mol / L) was measured by high performance liquid chromatography (渭 mol / L) (Mean 卤SEM). Results: the concentration of glutamate in the control group (59.74 卤4.62n 88.17 卤1.94 卤56.56 卤1.15) and electroacupuncture group (47.62 卤4.2954.72 卤1.98 卤52.27 卤1.16) were significantly higher than those in the control group and the control group for 714d. The concentration of glutamate in aqueous humor in the operation group (43.54 卤0.53U 43.71 卤0.66U 43.21 卤0.94U 43.04 卤0.44 P0.01A), in the electroacupuncture group of 2d 7d was significantly lower than that in the control group (P 0.01g), and there was no difference between the 14 d group and the control group (14d) in the concentration of glutamate in aqueous humor. Experiment 3: 54 rats were randomly selected after the right optic nerve was transected. The rats were randomly divided into 6 groups: control group (n = 6) and control group (n = 14) and electroacupuncture group (n = 9, n = 9 in each group). Immunohistochemical staining was used to detect glutamate reabsorption and metabolism in the retina of rats. Results: the expression of glutamate transporter-1 in electroacupuncture group was significantly higher than that in control group on 7 d group (P 0. 05), but there was no significant difference between control group and electroacupuncture group on day 214 and electroacupuncture group on day 214. the expression of glutamate transporter-1 in electroacupuncture group was significantly higher than that in control group on 7 d group (P < 0. 05). After optic nerve transection, the expression of glutamine synthase did not change in electroacupuncture group and control group. Conclusion: after transection of the optic nerve of adult rats, stimulation of Jinming point with 4 / 20Hz frequency electroacupuncture once a day can promote the reabsorption of glutamate transporter 1 in the injured eyes on the 7th day after injury, thereby reducing the content of glutamate in aqueous humor. Reduce glutamate excitotoxicity and delay the death of damaged ganglion cells.
【学位授予单位】:第四军医大学
【学位级别】:硕士
【学位授予年份】:2012
【分类号】:R245;R774.1
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