Interaction of Estrogen and AMPK in the Hypothalamus

描述(申请人提供)：本项目的目标是确定雌激素影响哺乳动物下丘脑摄食途径的机制。肥胖在西方社会越来越受到关注，主要是因为它的相关病理，包括糖尿病、动脉粥样硬化和心脏病。在女性中，雌二醇的减少发生在更年期，可能会导致吞噬过度和肥胖。雌二醇对啮齿动物和人类都有厌食作用。食物摄入量受作用于下丘脑的各种外源性神经肽(神经肽Y，NPY；刺鼠相关蛋白，AgRP)和厌食性神经肽(阿片黑素皮质素原，POMC)的调节。所有这些神经肽似乎都是受雌激素调节的，表达这些神经肽的神经元也表达雌激素受体。NPY、AgRP和POMC也受细胞能量感应酶AMP激活的蛋白激酶(AMPK)的调节，AMPK被肝激酶B1(LKB1)磷酸化(PAMPK)，当AMP：ATP比率降低时激活，表明内源能量较低。AMPK的激活导致食物摄入量的增加。去卵巢的雌性大鼠(缺乏雌二醇)变得肥胖，下丘脑pAMPK表达增加；然而，雌二醇影响下丘脑AMPK的机制尚不清楚。该建议的假设是，雌激素通过雌激素受体(ER)作用，减少LKB1的表达，从而减少AMPK的磷酸化和激活，从而减少食物的摄入量。为了验证这一假说，我们将利用细胞培养、转基因、Western blotting、定量RT-PCR、卵巢切除等方法，在体外和体内检测LKB1和AMPK基因和蛋白在雌二醇存在和不存在时的表达变化。 以及脑室内注射。初步数据显示，雌激素对N-38下丘脑细胞的治疗降低了AMPK活性和非活性的比例，并降低了NPY的相对丰度，从而有效地发出了高可用能量环境的信号。这些数据表明，雌二醇通过阻止AMPK的磷酸化和激活来影响能量平衡的潜在机制。因此，本项目的目的是验证雌激素通过对AMPK的作用而影响下丘脑摄食途径的假说，从而阐明肥胖的细胞途径，并为可能的药物干预与肥胖相关的病理提供信息。 公共卫生相关性：截至2010年，大约三分之一的美国人口被认为肥胖(BMIe30)，没有一个州的成年人口肥胖率低于20%(CDC BRFSS 2010)。肥胖个体患心脏病、糖尿病、某些癌症、中风和生殖问题的风险增加，与肥胖相关的健康问题每年花费约1470亿美元(Finkelstein等人)。2009年)。拟议的项目将调查蜂窝 食物摄入量和生殖激素之间相互作用的潜在机制，以及结果可能有助于减轻某些类型肥胖的病理影响。

DESCRIPTION (provided by applicant): The goal of this project is to determine the mechanism by which estradiol affects the food intake pathway in the mammalian hypothalamus. Obesity is of growing concern in Western society, mainly due to its associated pathologies, including diabetes, atherosclerosis, and heart disease. In women, decreases in estradiol occur during menopause and may lead to hyperphagia and obesity. Estradiol is anorexigenic in rodents and humans. Food intake is regulated by a variety of exigenic neuropeptides (neuropeptide Y, NPY; agouti-related protein, AgRP) and anorexigenic neuropeptides (pro- opiomelanocortin, POMC) that act in the hypothalamus. All of these neuropeptides appear to be estrogen-regulated and neurons expressing these peptides also express estrogen receptors. NPY, AgRP, and POMC are also regulated by the cellular energy sensing enzyme AMP-activated protein kinase (AMPK), which is phosphorylated (pAMPK) by the enzyme liver kinase B1 (LKB1) and activated when a decrease in the AMP:ATP ratio signals low endogenous energy. Activation of AMPK results in an increase in food intake. Ovariectomized female rats (lacking estradiol) become obese and have increased expression of hypothalamic pAMPK; however, the mechanism by which estradiol affects hypothalamic AMPK is unknown. The hypothesis of this proposal is that estradiol, acting through estrogen receptor ¿ (ER¿), decreases expression of LKB1, thereby decreasing phosphorylation and activation of AMPK and reducing food intake. To test this hypothesis, changes in gene and protein expression of LKB1 and AMPK will be examined both in vitro and in vivo in the presence and absence of estradiol, utilizing cell culture, transfection, Western blotting, quantitative RT-PCR, ovariectomy and intracerebroventricular injection. Preliminary data suggest that estradiol treatment in N-38 hypothalamic cells decreases the proportion of active to inactive AMPK, and decreases relative abundance of NPY, in effect signaling an environment of high available energy. These data point to a potential mechanism for estradiol to impact energy balance by preventing the phosphorylation and activation of AMPK. Therefore, the purpose of this project is to test the hypothesis that estradiol impacts the hypothalamic food intake pathway through its actions on AMPK, thereby shedding light on the cellular pathways underlying obesity and providing information for possible pharmacological interventions on the pathologies associated with the condition of obesity. PUBLIC HEALTH RELEVANCE: As of 2010, approximately one third of the U.S. population was considered obese (BMIe30), and no state had less than 20% obesity in its adult population (CDC BRFSS 2010). Obese individuals are at increased risk for heart disease, diabetes, certain cancers, stroke and reproductive problems, and health problems related to obesity cost about $147 billion per year (Finkelstein et al. 2009). The proposed project will investigate the cellular mechanisms underlying the interaction between food intake and reproductive hormones, and the results may be useful in mitigating the pathological effects of certain types of obesity.