外周第Ⅲ组代谢型谷氨酸受体在CFA引起的脊髓星形胶质细胞活化和长时程痛敏中的作用
发布时间：2018-11-22 07:19
【摘要】： 大鼠脚掌注射完全弗氏佐剂(CFA)是一个被广泛应用的慢性炎性痛模型。在注射CFA后,动物会迅速出现一个对热刺激或机械刺激敏感的状态,表现为伤害性热刺激引起的缩足反射潜伏期明显缩短。由CFA导致的热痛敏的高峰出现在注射后2-6小时,并持续约2周左右。近年来的研究表明,脊髓胶质细胞的激活在慢性痛的发生发展中发挥重要作用。在此过程,脊髓小胶质细胞与星形胶质细胞均有激活,并且小胶质细胞在慢性痛敏的产生和诱导过程发挥重要作用,而星形胶质细胞对于痛敏状态的持续维持发挥作用。本研究室之前的观察已证实,外周第III组代谢型谷氨酸受体(mGluRs)在Formalin引起的急性炎性痛中发挥重要作用。由于另外的研究表明,脊髓胶质细胞上同样表达第III组mGluRs。本实验在大鼠脚掌注射CFA的基础上,通过痛行为观察(热辐射刺激缩足反射潜伏期,PWL)、对脊髓星形胶质细胞活化标志物(GFAP)的免疫组化检测,结合外周注射第III组mGluRs激动剂L-SOP,观察外周第III组mGluRs在CFA所致热痛敏及脊髓胶质细胞活化中的作用。 实验在Sprague-Dawley雄性大鼠(90~100g)进行,分为免疫组化实验和行为学观察两部分。60只大鼠分为两组,第一组(予先给药组)在CFA注射前15分钟预先脚掌皮下给予第III组mGluRs的激动剂L-SOP (250nmol),然后在CFA注射后的第1,2,3,4天分别重复给予同样剂量的L-SOP。第二组(晚期给药组)在给予CFA后的第4,5,6,7,8天分别重复给予相同剂量的L-SOP。两组均在CFA注射后8小时,1天、4天,8天,14天随机选择六只大鼠,进行痛行为学观察及脊髓星形胶质细胞激活标志物GFAP的免疫组化检测,分别对比背角两侧星形胶质细胞的免疫组化反应,进行图像分析及统计学处理。 行为学观察结果显示,大鼠足掌给予CFA(1:1,50μl)后,动物即刻出现痛行为反应,表现为PWL的缩短,4-8小时出现了热痛敏反应的高峰,观察至第14天仍未完全恢复正常。在早期给药组外周应用L-SOP(250nmol)后,PWL明显延长,与对照侧相比,8h、1天、4天具有显著性差异(P0.05),在晚期给药组,PWL的延长在4天、8天与对照侧相比有统计学差异(P0.05)。 免疫组化结果显示,大鼠足掌给予CFA后,脊髓背角星形胶质细胞在4天后表达增加,并持续到14天以后。在外周早期给予L-SOP后,星形胶质细胞GFAP阳性表达明显降低,在4、8、14天GFAP阳性表达的抑制率分别为18.61%、24.45%、20.54%,与对照侧相比均有显著性差异(P0.05)。在晚期给药组,GFAP阳性表达第8天时在用药侧有
[Abstract]:Rat foot injection of complete Freund's adjuvant (CFA) is a widely used chronic inflammatory pain model. After the injection of CFA, the animal will quickly appear to be sensitive to thermal or mechanical stimulation, showing that the latent period of foot contraction reflex induced by nociceptive heat stimulation is obviously shortened. The peak of heat sensitivity caused by CFA appears 2-6 hours after injection and lasts about 2 weeks. Recent studies have shown that the activation of glial cells plays an important role in the development of chronic pain. In this process, both microglia and astrocytes are activated, and microglia play an important role in the generation and induction of chronic hyperalgesia, while astrocytes play an important role in the sustained maintenance of hyperalgesia. Previous observations in this laboratory have confirmed that the metabolic glutamate receptor (mGluRs) in peripheral III group plays an important role in acute inflammatory pain induced by Formalin. As a result of other studies, the same expression of mGluRs. in group III was found in glial cells of the spinal cord. On the basis of the injection of CFA to the foot of rats, the pain behavior was observed by observing the immunocytochemistry of (GFAP), the activation marker of astrocytes of spinal cord, by, PWL), in latent period of reflex induced by thermal radiation. Combined with peripheral injection of L-SOP, mGluRs agonist L-SOP in III group, the role of mGluRs in CFA induced hyperthermia and activation of spinal glial cells in peripheral III group was observed. The experiment was carried out in male Sprague-Dawley rats (90g / 100g) and was divided into two parts: immunohistochemical experiment and behavioral observation. 60 rats were divided into two groups. The first group (pre-administration group) was subcutaneously given L-SOP (250nmol), the agonist of mGluRs in the III group, 15 minutes before the injection of CFA, and then repeated the same dose of L-SOP at 34th day after CFA injection. The second group (late administration group) was treated with the same dose of L-SOP on the 4th day, 5th day and 7th day after CFA. Six rats were randomly selected at 8 hours, 1 day, 4 days, 8 days and 14 days after CFA injection to observe the pain behavior and detect the GFAP of astrocyte activation marker in spinal cord. The immunohistochemical reaction of astrocytes on both sides of the dorsal horn was compared, and the image analysis and statistical analysis were carried out. The results of behavioral observation showed that after CFA (1: 1: 50 渭 l) was given to the palmar of the foot, the pain behavior of the rats appeared immediately, which was manifested by the shortening of the PWL, the peak of the thermal pain sensitivity at 4-8 hours, and the observation on the 14th day did not return to normal completely. In the early administration group, PWL was significantly prolonged after peripheral administration of L-SOP (250nmol). Compared with the control group, there was a significant difference at 8 h and 4 days (P0.05). In the late administration group, the PWL was prolonged at 4 days. There was statistical difference between 8 days and control side (P0.05). Immunohistochemical results showed that the expression of astrocytes in spinal dorsal horn increased 4 days after CFA and lasted for 14 days. After early L-SOP administration, the positive expression of GFAP in astrocytes decreased significantly, and the inhibitory rates of GFAP positive expression were 18.61 ~ 24.45 and 20.54 on day 14, respectively. Compared with the control side, there were significant differences (P0.05). In the advanced administration group, the positive expression of GFAP was found on the side of the drug on the 8th day.
【学位授予单位】：山西医科大学
【学位级别】：硕士
【学位授予年份】：2006
【分类号】：R33
本文编号：2348480
[Abstract]:Rat foot injection of complete Freund's adjuvant (CFA) is a widely used chronic inflammatory pain model. After the injection of CFA, the animal will quickly appear to be sensitive to thermal or mechanical stimulation, showing that the latent period of foot contraction reflex induced by nociceptive heat stimulation is obviously shortened. The peak of heat sensitivity caused by CFA appears 2-6 hours after injection and lasts about 2 weeks. Recent studies have shown that the activation of glial cells plays an important role in the development of chronic pain. In this process, both microglia and astrocytes are activated, and microglia play an important role in the generation and induction of chronic hyperalgesia, while astrocytes play an important role in the sustained maintenance of hyperalgesia. Previous observations in this laboratory have confirmed that the metabolic glutamate receptor (mGluRs) in peripheral III group plays an important role in acute inflammatory pain induced by Formalin. As a result of other studies, the same expression of mGluRs. in group III was found in glial cells of the spinal cord. On the basis of the injection of CFA to the foot of rats, the pain behavior was observed by observing the immunocytochemistry of (GFAP), the activation marker of astrocytes of spinal cord, by, PWL), in latent period of reflex induced by thermal radiation. Combined with peripheral injection of L-SOP, mGluRs agonist L-SOP in III group, the role of mGluRs in CFA induced hyperthermia and activation of spinal glial cells in peripheral III group was observed. The experiment was carried out in male Sprague-Dawley rats (90g / 100g) and was divided into two parts: immunohistochemical experiment and behavioral observation. 60 rats were divided into two groups. The first group (pre-administration group) was subcutaneously given L-SOP (250nmol), the agonist of mGluRs in the III group, 15 minutes before the injection of CFA, and then repeated the same dose of L-SOP at 34th day after CFA injection. The second group (late administration group) was treated with the same dose of L-SOP on the 4th day, 5th day and 7th day after CFA. Six rats were randomly selected at 8 hours, 1 day, 4 days, 8 days and 14 days after CFA injection to observe the pain behavior and detect the GFAP of astrocyte activation marker in spinal cord. The immunohistochemical reaction of astrocytes on both sides of the dorsal horn was compared, and the image analysis and statistical analysis were carried out. The results of behavioral observation showed that after CFA (1: 1: 50 渭 l) was given to the palmar of the foot, the pain behavior of the rats appeared immediately, which was manifested by the shortening of the PWL, the peak of the thermal pain sensitivity at 4-8 hours, and the observation on the 14th day did not return to normal completely. In the early administration group, PWL was significantly prolonged after peripheral administration of L-SOP (250nmol). Compared with the control group, there was a significant difference at 8 h and 4 days (P0.05). In the late administration group, the PWL was prolonged at 4 days. There was statistical difference between 8 days and control side (P0.05). Immunohistochemical results showed that the expression of astrocytes in spinal dorsal horn increased 4 days after CFA and lasted for 14 days. After early L-SOP administration, the positive expression of GFAP in astrocytes decreased significantly, and the inhibitory rates of GFAP positive expression were 18.61 ~ 24.45 and 20.54 on day 14, respectively. Compared with the control side, there were significant differences (P0.05). In the advanced administration group, the positive expression of GFAP was found on the side of the drug on the 8th day.
【学位授予单位】：山西医科大学
【学位级别】：硕士
【学位授予年份】：2006
【分类号】：R33
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