基于多元统计分析的针刺效应对不同脑区CREB/p-CREB表达时空分布特征分析

Expression of CREB/p-CREB in different brain areas with manual acupuncture: A time-dependent manner based on multivariate statistical

目的:观察CREB及p-CREB在针刺穴位后的不同时间点及不同脑区的分布特点,以探讨针刺效应的中枢作用特点。方法:选用健康清洁级SD大鼠30只,分为5组(空白组、外关组及非穴组,其中外关组及非穴组按不同取材时间分为即刻组及20min组2个亚组),每组6只大鼠。空白组大鼠仅模拟抓取动作,外关组及非穴组分别针刺外关穴及旁开非穴20min,每5min捻针1次。处理组大鼠分别于针刺结束后即刻及20min后取相应皮质扣带回及小脑水平作冠状位切面,作免疫组化观察分析不同脑区的CREB及p-CREB的表达特点。结果:针刺后即刻及20min在小脑扁桃体区外关组及非穴组CREB表达强度增强,与空白组比较差异有统计学意义(P<0.05);而扣带回仅在起针后20min表达增强且与空白组比较存在显著性差异,且外关组与非穴组之间无显著差异。而p-CREB在针刺穴位后表现出了较强的特异性,不论在小脑抑或皮质扣带回,即刻针刺外关组的p-CREB的表达均明显增强,且针刺外关组较针刺非穴组有显著差异(P<0.05),同时20min后扣带回的p-CREB表达较针刺即刻明显减少(P<0.01)。而偏相关分析及轮廓分析的结果表明针刺后小脑及扣带回的不同脑区存在着随时间相关的分布特征(P<0.01)。结论:针刺外关穴使得对小脑及皮质扣带回区p-CREB特异性表达增强,且随着时间变化呈下降趋势,且在不同脑区之间存在较为一致的变化规律。

Objective： To observe the expression of CREB and p-CREB in different brain areas by acupuncture with different duration to investigate the characteristics of acupuncture effect in central nervous system. Methods： Thirty SD rats were divided 5 groups： control group, Waiguan group, Feixue group and according to the different time for sampling, Wauguan and Feixue group were both divided to Jike subgroup and 20rain subgroup. There were 6 rats in each group. Acupuncture was applied manual at Waiguan （SJ 5） of SD rats for 20 minutes with slowly rotating every 5 minute. After acupuncture instantly or 20 minutes later, the rats in Jike and 20rain groups were both sacrificed and immunohistochemistry was used to observe the expression of CREB and p-CREB in different encephalic region. Results： Application of acupuncture at the point significantly increased the expression of p-CREB in the tonsil of cerebellum compared with the control group （P〈0.05）. In addition, after 20 minutes later, the expression of p-CREB decreased significantly in the cingulate cortex and tonsil of cerebellum areas （P〈0.05）, and there was no significant difference of expression of CREB between acupuncture group and control group after acupuncture instantly or 20 minutes later. The expression of p-CREB in epencephalon and cingulate cortex were increased significantly of Waiguan group, and there were significant differences between Walguan group and Feixue group. But after 20 minutes later, the p-CREB expression in epencephalon and cingulate cortex was decreased significantly compared with acupuncture instantly （P〈0.01）. Furthermore, the expression of p-CREB showed a time-dependent manner after acupuncture with the contour analysis （P〈0.01）. Conclusion：Acupuncture on Waiguan point can make the specific expression of p-CREB in epencephalon and cingulate cortex increase, and on a declining curve with the change of time. The change rules in different encephalic region are more consistent.