摘要
目的:通过观察下丘脑穹窿周区orexin-A神经元对不同刺激方式的反应特性探索能够激活该系统的高效而适宜的方法。方法:采用禁食、腹腔注射胰岛素和2-DG三种饥饿刺激方式,利用免疫组织化学染色及ELISA检测相结合的方法对大鼠穹隆周区orexin-A神经元的Fos的表达,脑脊液中orexin-A的浓度进行了对比分析。27只SD大鼠随机分为六组,分别进行禁食2 d,腹腔注射胰岛素或生理盐水存活5 h,腹腔注射2-脱氧-D-葡萄糖(2-DG)或生理盐水存活2 h和正常对照处理,动物的饮水量保持正常。结果:三种饥饿刺激引发的Fos表达比较类似,主要集中于下丘脑背内侧核,下丘脑外侧区和下丘脑后区,2-DG组的Fos表达最为浓密。三种刺激对orexin-A神经元的数量无明显影响,但orexin-A/Fos双标细胞数占所有orexin-A阳性细胞数的比例以2-DG组最高,为26%;禁食2 d组次之为21%;胰岛素组最低为14%。禁食组和2-DG组的双标细胞率与胰岛素组相比差异具有统计学意义(P<0.05)。ELISA检测结果显示禁食组脑脊液中orexin-A的含量显著高于对照组23%,而其它两组与对照组相比没有差别。结论:本研究提示orexin-A的功能状态与刺激方式密切相关:急性刺激如2-DG注射适于研究神经元的激活状态,而慢性刺激如禁食适于研究激活后导致的orexin-A表达变化。
Objective: To explore an efficient and adequate method to activate orexin-A system in hypothalamus,we observed the responsiveness of orexin-A containing neurons in perifornical area to differential stimuli.Methods:We adopted three differential stimuli and observed the resultant Fos expression and alterations of orexin-A level in cerebrospinal fluid(CSF) by immunohistochemical staining and ELISA assay,respectively.Twenty-seven Sprague Dawley rats were randomly divided into six groups and were applied fasting for 2 d,i.p.injection of insulin or saline and survived for 5 h,i.p.injection of 2-deoxy-D-glucose(2-DG) and survived for 2 h and normal control treatment.All animals were free to water supply ad libitum.Results: Three hunger stimuli induced similar Fos expression pattern in doromedial nucleus,lateral and posterior areas of hypothalamus.Fos expression in 2-DG group was most abundant compared with the other two experimental groups.The number of orexin-A containing neurons was comparable among all the groups.The rate of orexin-A/Fos double labeled neurons to total orexin-A containing neurons was 26% in 2-DG groups,21% in fasting group and 14% in insulin group.Fasting and 2-DG groups showed significant higher double labeling rate compared to insulin group(P〈0.05).On the other hand,ELISA assay revealed that orexin-A level in CSF of fasting group increased by 23% compared to normal control group,while that of insulin and 2-DG group showed no difference with saline injection group.Conclusion: These results suggest that the functional status of orexin-A containing neurons is closely related to the stimulating pattern;acute stimuli such as injection of 2-DG is suitable for observation of the activation of these neurons while chronic stimuli as fasting is more useful to check the expression and release of orexin-A.
出处
《神经解剖学杂志》
CAS
CSCD
北大核心
2011年第4期371-376,共6页
Chinese Journal of Neuroanatomy
基金
国家自然科学基金(30871004
31071012)