针刺镇痛中内源性促肾上腺皮质激素释放激素的作用及机制探讨
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摘要
目的:观察电针(EA)夹脊穴对弗氏完全佐剂(CFA)诱发的炎症痛模型(AA)大鼠的镇痛治疗作用和神经内分泌免疫调节作用,探讨EA镇痛抗炎的作用机制;观察侧脑室注射抗5-羟色胺(5-HT)或抗β-内啡肽(β-EP)血清对内源性CRH在痛觉调制及EA镇痛作用的影响,以探讨内源性神经肽促肾上腺皮质激素释放激素(CRH)在针刺镇痛及免疫调节作用中与内阿片肽、经典神经递质之间的作用关系;观察电针及侧脑室注射抗CRH血清对AA大鼠下丘脑抑制性G蛋白(inhibitory GTP-binding protein,Gi)表达的变化及EA镇痛的影响,以初步探讨针刺镇痛中内源性CRH的细胞信号转导机制。
方法:以AA大鼠为研究对象,应用行为学痛阈观察、足跖容积测定、高效液相电化学法、放射免疫法等方法,观察EA夹脊穴和侧脑室注射抗CRH血清对AA大鼠痛阈、足跖容积、下丘脑和脊髓5-羟色胺(5-HT)、β-内啡肽(β-EP)含量、血清IL-1β和TNF-α水平的影响;应用免疫组化技术观察电针对AA大鼠下丘脑核转录因子NF-κB表达的影响及电针及侧脑室注射抗CRH血清对AA大鼠下丘脑室旁核CRH免疫阳性神经元表达的影响;观察侧脑室注射抗5-HT血清或抗β-EP血清对内源性CRH在AA大鼠及电针镇痛中的影响;用免疫印迹法观察电针夹脊穴及侧脑室注射抗CRH血清对AA大鼠下丘脑Gi蛋白表达的影响。
结果:EA夹脊穴可提高AA大鼠痛阈,降低足跖容积,降低AA大鼠下丘脑核转录因子NF-κB的表达;侧脑室注射抗CRH血清可降低AA大鼠痛阈及EA镇痛效果,降低下丘脑5-HT和β-EP的含量,降低血清TNF-α水平,使AA大鼠及电针大鼠下丘脑CRH的表达减少,电针则增强下丘脑CRH的表达。侧脑室注射抗5-HT血清可增强抗CRH对痛觉调制及电针镇痛的影响,侧脑室注射抗β-EP血清可使下丘脑CRH表达增多,对抗CRH在电针镇痛中影响不大;EA可降低AA大鼠下丘脑Gi蛋白表达,侧脑室注射抗CRH血清则导致其含量升高。
结论:
电针镇痛及抗炎治疗作用可能与调节下丘脑活化的NF-κB的表达有关。
电针镇痛及神经内分泌-免疫调节机制与G蛋白信号转导机制有关。
内源性CRH在痛觉调制及针刺镇痛中的作用与5-HT及β-EP相关。
内源性CRH在针刺镇痛及神经内分泌免疫调节中的作用与G蛋白信号转导机制有关。
Objective: to explore the mechanism of electro-acupuncture analgesia on CFA-induced arthritis inflammatory pain model (AA rats);To explore the the relation of CRH with 5-HT orβ-EP on the effect of acupuncture analgesia through the way of injecting anti-corticotropin-releasing-hormone serum (anti-serum of CRH) and anti-5-HT serum or antiβ-EP serum from the lateral ventricle of cerebrum on AA rats; to explore the mechanism of G-protein mediated signal transduction of CRH in pain and acupuncture analgesia by studying the changes of the expression of inhibitory GTP-binding protein (Gi) in hypothalamus after acupuncture or injecting anti-serum of CRH from the lateral ventricle of cerebrum on AA rats.
Methods: AA rats were used as the subjects of the study. By using methods of behavioral threshold of pain test, measurement of foot cubage,high performance liquid chromatographic(HPLC) and radioimmunoassay et al, we observed the influence of EA at EX-B2 on AA rats and injection of anti serum of CRH from the threshold of pain, foot cubage, the level of IL-1? and TNF-αin blood serum, the content of 5-HT andβ-endorphin in hypothalamus or spinal cord. Observe the changes of pain threshold after inject anti-serum of 5-HT orβ-endorphin on anti-serum of CRH or electro-acupuncture.The expression of NF-κB or CRH and Gi in hypothalamus was also observed by using methods of immunohistochemistry and western blot.
Results: The pain threshold of AA rats were improved and the foot cubage foot swelling rate were reduced by using electro-acupuncture at EX-B2. The amounts of 5-HT andβ-EP in hypothalamus and spinal cord increased after electro-acupuncture while the IL-1? and TNF-αdecreased in blood serum. We also observed that the expression of NF- κB in hypothalamus decreased remarkably after electro-acupuncture. And the expression of CRH increased .However,after injection of anti-serum of CRH weakened the pain threshold of AA rats and the effect of acupuncture analgesia. the amounts of 5-HT andβ-EP and the expression of CRH in hypothalamus decreased. If add injection the anti-serum of 5-HT ,the pain threshold mostly decreased than before.but when add injection the anti-serum ofβ-EP ,no difference on the pain threshold happened,and the expression of CRH increased than before.from the results of western -blot we can see that the inhibitory G-protein in hypothalamus decreased after electro-acupuncture while increased after inject the anti-serum of CRH .
Conclusion: the mechanism of electro-acupuncture analgesia and the function of immune regulation may be related with the activity of NF-κB.
the mechanism of acupuncture analgesia and regulation of NEI may be related with the G-protein mediated signal transduction pathway.
The endogenous of CRH may be related with the 5-HT orβ-EP during the process of electro-acupuncture analgesia.
The endogenous of CRH may be related with the G-protein mediated signal transduction pathway during the process of electro-acupuncture analgesia.
方法:以AA大鼠为研究对象,应用行为学痛阈观察、足跖容积测定、高效液相电化学法、放射免疫法等方法,观察EA夹脊穴和侧脑室注射抗CRH血清对AA大鼠痛阈、足跖容积、下丘脑和脊髓5-羟色胺(5-HT)、β-内啡肽(β-EP)含量、血清IL-1β和TNF-α水平的影响;应用免疫组化技术观察电针对AA大鼠下丘脑核转录因子NF-κB表达的影响及电针及侧脑室注射抗CRH血清对AA大鼠下丘脑室旁核CRH免疫阳性神经元表达的影响;观察侧脑室注射抗5-HT血清或抗β-EP血清对内源性CRH在AA大鼠及电针镇痛中的影响;用免疫印迹法观察电针夹脊穴及侧脑室注射抗CRH血清对AA大鼠下丘脑Gi蛋白表达的影响。
结果:EA夹脊穴可提高AA大鼠痛阈,降低足跖容积,降低AA大鼠下丘脑核转录因子NF-κB的表达;侧脑室注射抗CRH血清可降低AA大鼠痛阈及EA镇痛效果,降低下丘脑5-HT和β-EP的含量,降低血清TNF-α水平,使AA大鼠及电针大鼠下丘脑CRH的表达减少,电针则增强下丘脑CRH的表达。侧脑室注射抗5-HT血清可增强抗CRH对痛觉调制及电针镇痛的影响,侧脑室注射抗β-EP血清可使下丘脑CRH表达增多,对抗CRH在电针镇痛中影响不大;EA可降低AA大鼠下丘脑Gi蛋白表达,侧脑室注射抗CRH血清则导致其含量升高。
结论:
电针镇痛及抗炎治疗作用可能与调节下丘脑活化的NF-κB的表达有关。
电针镇痛及神经内分泌-免疫调节机制与G蛋白信号转导机制有关。
内源性CRH在痛觉调制及针刺镇痛中的作用与5-HT及β-EP相关。
内源性CRH在针刺镇痛及神经内分泌免疫调节中的作用与G蛋白信号转导机制有关。
Objective: to explore the mechanism of electro-acupuncture analgesia on CFA-induced arthritis inflammatory pain model (AA rats);To explore the the relation of CRH with 5-HT orβ-EP on the effect of acupuncture analgesia through the way of injecting anti-corticotropin-releasing-hormone serum (anti-serum of CRH) and anti-5-HT serum or antiβ-EP serum from the lateral ventricle of cerebrum on AA rats; to explore the mechanism of G-protein mediated signal transduction of CRH in pain and acupuncture analgesia by studying the changes of the expression of inhibitory GTP-binding protein (Gi) in hypothalamus after acupuncture or injecting anti-serum of CRH from the lateral ventricle of cerebrum on AA rats.
Methods: AA rats were used as the subjects of the study. By using methods of behavioral threshold of pain test, measurement of foot cubage,high performance liquid chromatographic(HPLC) and radioimmunoassay et al, we observed the influence of EA at EX-B2 on AA rats and injection of anti serum of CRH from the threshold of pain, foot cubage, the level of IL-1? and TNF-αin blood serum, the content of 5-HT andβ-endorphin in hypothalamus or spinal cord. Observe the changes of pain threshold after inject anti-serum of 5-HT orβ-endorphin on anti-serum of CRH or electro-acupuncture.The expression of NF-κB or CRH and Gi in hypothalamus was also observed by using methods of immunohistochemistry and western blot.
Results: The pain threshold of AA rats were improved and the foot cubage foot swelling rate were reduced by using electro-acupuncture at EX-B2. The amounts of 5-HT andβ-EP in hypothalamus and spinal cord increased after electro-acupuncture while the IL-1? and TNF-αdecreased in blood serum. We also observed that the expression of NF- κB in hypothalamus decreased remarkably after electro-acupuncture. And the expression of CRH increased .However,after injection of anti-serum of CRH weakened the pain threshold of AA rats and the effect of acupuncture analgesia. the amounts of 5-HT andβ-EP and the expression of CRH in hypothalamus decreased. If add injection the anti-serum of 5-HT ,the pain threshold mostly decreased than before.but when add injection the anti-serum ofβ-EP ,no difference on the pain threshold happened,and the expression of CRH increased than before.from the results of western -blot we can see that the inhibitory G-protein in hypothalamus decreased after electro-acupuncture while increased after inject the anti-serum of CRH .
Conclusion: the mechanism of electro-acupuncture analgesia and the function of immune regulation may be related with the activity of NF-κB.
the mechanism of acupuncture analgesia and regulation of NEI may be related with the G-protein mediated signal transduction pathway.
The endogenous of CRH may be related with the 5-HT orβ-EP during the process of electro-acupuncture analgesia.
The endogenous of CRH may be related with the G-protein mediated signal transduction pathway during the process of electro-acupuncture analgesia.
引文
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