Alenazi Fahaad S H, Ibrahim Baher A, Briski Karen P
Department of Basic Pharmaceutical Sciences, School of Pharmacy, The University of Louisiana, Monroe, LA 71201, USA.
Department of Basic Pharmaceutical Sciences, School of Pharmacy, The University of Louisiana, Monroe, LA 71201, USA.
Neuropeptides. 2015 Apr;50:29-33. doi: 10.1016/j.npep.2015.02.001. Epub 2015 Feb 24.
Fos immunocytochemistry is a valuable anatomical mapping tool for distinguishing cells within complex tissues that undergo genomic activation, but it is seldom paired with corroborative molecular analytical techniques. Due to preparatory requirements that include protein cross-linking for specimen sectioning, histological tissue sections are regarded as unsuitable for those methods. Our studies show that pharmacological activation of the hindbrain energy sensor AMPK by AICAR elicits estradiol (E)-dependent patterns of Fos immunolabeling of hypothalamic metabolic loci. Here, Western blotting was applied to hypothalamic tissue removed from histological sections of E- versus oil (O)-implanted ovariectomized (OVX) female rat brain to measure levels of metabolic transmitters associated with Fos-positive structures. In both E and O rats, AICAR treatment elicited alterations in pro-opiomelanocortin, neuropeptide Y, SF-1, and orexin-A neuropeptide expression that coincided with patterns of Fos labeling of structures containing neurons that synthesize these neurotransmitters, e.g. arcuate and ventromedial nuclei and lateral hypothalamic area. O, but not E animals also exhibited parallel augmentation of tissue corticotropin-releasing hormone neuropeptide levels and paraventricular nucleus Fos staining. Data demonstrate the utility of immunoblot analysis as a follow-through technique to capitalize on Fos mapping of transactivation sites in the brain. Findings that induction of Fos immunoreactivity coincides with adjustments in hypothalamic metabolic neuropeptide expression affirms that this functional indicator reflects changes in neurotransmission in pathways governing metabolic outflow.
Fos免疫细胞化学是一种用于区分复杂组织中经历基因组激活的细胞的有价值的解剖学绘图工具，但它很少与确证性分子分析技术相结合。由于包括用于标本切片的蛋白质交联在内的制备要求，组织学组织切片被认为不适用于那些方法。我们的研究表明，AICAR对后脑能量传感器AMPK的药理学激活引发了下丘脑代谢位点的Fos免疫标记的雌二醇（E）依赖性模式。在此，对从植入E或油（O）的去卵巢（OVX）雌性大鼠脑的组织学切片中取出的下丘脑组织进行蛋白质印迹分析，以测量与Fos阳性结构相关的代谢递质水平。在E组和O组大鼠中，AICAR处理均引起促阿片黑素皮质素原、神经肽Y、SF-1和食欲素-A神经肽表达的改变，这些改变与含有合成这些神经递质的神经元的结构的Fos标记模式一致，例如弓状核、腹内侧核和下丘脑外侧区。O组动物而非E组动物还表现出组织促肾上腺皮质激素释放激素神经肽水平和室旁核Fos染色的平行增加。数据证明了免疫印迹分析作为一种后续技术的实用性，可利用其对大脑中转录激活位点进行Fos绘图。Fos免疫反应性的诱导与下丘脑代谢神经肽表达的调整相一致的发现证实，这种功能指标反映了控制代谢流出的途径中神经传递的变化。
