發(fā)布時間：2018-11-22 07:19

【摘要】： 大鼠腳掌注射完全弗氏佐劑(CFA)是一個被廣泛應用的慢性炎性痛模型。在注射CFA后,動物會迅速出現(xiàn)一個對熱刺激或機械刺激敏感的狀態(tài),表現(xiàn)為傷害性熱刺激引起的縮足反射潛伏期明顯縮短。由CFA導致的熱痛敏的高峰出現(xiàn)在注射后2-6小時,并持續(xù)約2周左右。近年來的研究表明,脊髓膠質細胞的激活在慢性痛的發(fā)生發(fā)展中發(fā)揮重要作用。在此過程,脊髓小膠質細胞與星形膠質細胞均有激活,并且小膠質細胞在慢性痛敏的產生和誘導過程發(fā)揮重要作用,而星形膠質細胞對于痛敏狀態(tài)的持續(xù)維持發(fā)揮作用。本研究室之前的觀察已證實,外周第III組代謝型谷氨酸受體(mGluRs)在Formalin引起的急性炎性痛中發(fā)揮重要作用。由于另外的研究表明,脊髓膠質細胞上同樣表達第III組mGluRs。本實驗在大鼠腳掌注射CFA的基礎上,通過痛行為觀察(熱輻射刺激縮足反射潛伏期,PWL)、對脊髓星形膠質細胞活化標志物(GFAP)的免疫組化檢測,結合外周注射第III組mGluRs激動劑L-SOP,觀察外周第III組mGluRs在CFA所致熱痛敏及脊髓膠質細胞活化中的作用。 實驗在Sprague-Dawley雄性大鼠(90~100g)進行,分為免疫組化實驗和行為學觀察兩部分。60只大鼠分為兩組,第一組(予先給藥組)在CFA注射前15分鐘預先腳掌皮下給予第III組mGluRs的激動劑L-SOP (250nmol),然后在CFA注射后的第1,2,3,4天分別重復給予同樣劑量的L-SOP。第二組(晚期給藥組)在給予CFA后的第4,5,6,7,8天分別重復給予相同劑量的L-SOP。兩組均在CFA注射后8小時,1天、4天,8天,14天隨機選擇六只大鼠,進行痛行為學觀察及脊髓星形膠質細胞激活標志物GFAP的免疫組化檢測,分別對比背角兩側星形膠質細胞的免疫組化反應,進行圖像分析及統(tǒng)計學處理。 行為學觀察結果顯示,大鼠足掌給予CFA(1:1,50μl)后,動物即刻出現(xiàn)痛行為反應,表現(xiàn)為PWL的縮短,4-8小時出現(xiàn)了熱痛敏反應的高峰,觀察至第14天仍未完全恢復正常。在早期給藥組外周應用L-SOP(250nmol)后,PWL明顯延長,與對照側相比,8h、1天、4天具有顯著性差異(P0.05),在晚期給藥組,PWL的延長在4天、8天與對照側相比有統(tǒng)計學差異(P0.05)。 免疫組化結果顯示,大鼠足掌給予CFA后,脊髓背角星形膠質細胞在4天后表達增加,并持續(xù)到14天以后。在外周早期給予L-SOP后,星形膠質細胞GFAP陽性表達明顯降低,在4、8、14天GFAP陽性表達的抑制率分別為18.61%、24.45%、20.54%,與對照側相比均有顯著性差異(P0.05)。在晚期給藥組,GFAP陽性表達第8天時在用藥側有
[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.
【學位授予單位】：山西醫(yī)科大學
【學位級別】：碩士
【學位授予年份】：2006
【分類號】：R33

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