CNS Fatty Acid Metabolism in Regulation of Food Intake

描述(由申请人提供)： 该提案的目的是确定抑制脂肪酸合成酶(FasI)和通过刺激肉碱棕榈酰转移酶-1(CPT-1s)增强β-氧化改变下丘脑能量状态和调节食物摄入量的行为和细胞机制。应聘者将(1)在静态培养中获得原代下丘脑神经元培养和急性下丘脑外植体的经验，(2)学习一系列生化分析，(3)在高效液相色谱、免疫组织化学和信使核糖核酸检测方面的经验。培训将促进候选人向独立研究的过渡。实验将在约翰霍普金斯大学医学院进行。主要的导师是加布里埃勒·罗内特博士，他是一位使用神经元培养的神经学家和生化学家。共同导师是蒂莫西·莫兰博士，他是喂养神经生理学方面的专家。候选人将与一个跨学科团队互动，该团队调查脂肪酸代谢在包括肥胖在内的一系列人类健康问题中的作用。C75是一种Fasi/CPT-1s，当给啮齿动物中枢给药时，它可以减少食物摄入量，并改变下丘脑摄食相关神经肽的基因表达。我们发现，icv c75通过减少进食频率而不是大小来减少大鼠的进食，这表明Fasi或CPT-1，或者两者都减少了开始进食的动力，而不是增加饱腹感。C75影响原代培养的下丘脑神经元的能量状态，改变细胞内信号以调节神经肽的产生。针对Fas和CPT-1的新化合物可用于研究Fas和CPT-1在摄食调节中的单独作用。总体假设是，下丘脑神经元中脂肪酸代谢的改变改变了它们的能量状态，并导致食物摄入量的动态平衡改变。实验将针对两个特定的目标：(1)确定大脑中的Fasi或CPT-1如何改变食物摄入量和基因表达：在小鼠的实验中，将测量ICV C75、Fasi或CPT-1之后的食物摄入量、进食模式、大脑c-Fos的表达，以及与摄食相关的神经肽的mRNA表达。第二个目标是(2)确定Fasi和CPT-1s如何改变下丘脑神经元的能量状态和摄食调节神经肽的产生：这些化合物将应用于(A)原代下丘脑神经元培养和(B)急性下丘脑外植体。我们将测量细胞内AMP/ATP比率和AMP激活的蛋白激酶活性，摄食相关神经肽的mRNAs水平，以及神经肽的产生和释放。

DESCRIPTION (provided by applicant): The objective of the proposal is to identify behavioral and cellular mechanisms by which inhibition of fatty acid synthase (FASi) and enhanced beta-oxidation by stimulation of carnitine-palmitoyltransferase-1 (CPT-1s) alter hypothalamic energy status and adjust food intake. The candidate will (1) gain experience with primary hypothalamic neuronal cultures and acute hypothalamic explants in static incubation, (2) learn a range of biochemical assays, and (3) build upon experience with HPLC, immunohistochemistry, and mRNA detection. The training will facilitate the candidate's transition to independent research. Experiments will be conducted at the Johns Hopkins University School of Medicine. The primary mentor is Dr. Gabriele Ronnett, a neuroscientist and biochemist who uses neuronal cultures. The co-mentor is Dr. Timothy Moran, an expert in neurophysiology of feeding. The candidate will interact with an interdisciplinary team that investigates roles of fatty acid metabolism in a range of human health problems, including obesity. C75 is a FASi/CPT-1s that reduces food intake and alters gene expression for hypothalamic feeding-related neuropeptides when given centrally to rodents. We find that icv C75 decreases feeding in rats by reducing meal frequency, not size, suggesting that FASi or CPT-1s, or both, reduce drive to initiate feeding, rather than enhance satiety. C75 affects energy status in primary cultures of hypothalamic neurons, altering intracellular signaling to modulate neuropeptide production. Novel compounds selective for potent FASi or CPT-1s are available to us to investigate individual roles of FAS and CPT-1 in feeding regulation. The overall hypothesis is that altered fatty acid metabolism in hypothalamic neurons changes their energy state and leads to homeostatic alterations in food intake. Experiments will address two Specific Aims: (1) Identify how FASi or CPT-1s in the brain alter food intake and gene expression: Experiments in mice will measure food intake, meal patterns, brain c-Fos expression, and expression of mRNA for feeding-related neuropeptides after icv C75, FASi, or CPT-1s. The second aim is to (2) Identify how FASi and CPT-1s change hypothalamic neuronal energy status and production of feeding-regulating neuropeptides: The compounds will be applied to (a) primary hypothalamic neuronal cultures and (b) acute hypothalamic explants. We will measure changes in intracellular AMP/ATP ratio and activity of AMP-activated protein kinase, levels of mRNAs for feeding-related neuropeptides, and neuropeptide production and release.