CENTRAL/PERIPHERAL TARGETS FOR METABOLIC ACTIONS OF LEPTIN

该建议的总体目标是了解瘦素作用的分子机制，以快速增强胰岛素敏感性，而不依赖于其对食物摄入和体重的影响。我们将研究CNS介导的和直接作用在外周组织水平的贡献。在目标1中，我们将确定胰岛素和瘦素是否共享共同的细胞内信号转导途径。通过在大鼠中静脉内施用瘦素，我们将检验瘦素的胰岛素增敏作用涉及瘦素活化的和胰岛素活化的信号转导途径之间的协同作用的会聚的假设。我们将确定瘦素激活胰岛素受体和IRSs的磷酸化以及体内胰岛素靶组织（骨骼肌、BAT、WAT肝脏）中PI 3激酶、MAPK激酶和Stat 1和Stat 2的激活的时间过程。我们将研究瘦素和胰岛素的潜在叠加效应。在目标2中，我们将检验这样的假设，即瘦素对信号通路的早期激活是由于对靶组织水平的直接影响，但后来的影响也涉及CNS介导。我们将在一段时间内测量静脉注射或ICV瘦素后下丘脑中信号通路的激活。我们将确定骨骼肌或BT交感神经切除术后存在哪些影响。我们将确定哪些信号通路在体外脂肪细胞和肌肉中被激活。在目标3中，我们将检验瘦素的某些生物学作用直接在脂肪细胞水平发挥的假设。我们将使用aP 2启动子/增强子产生在脂肪中选择性表达瘦素受体的转基因db/db小鼠。我们将确定是否恢复长型瘦素受体专门在脂肪细胞改变脂肪细胞的生理，葡萄糖稳态，或胰岛素的作用。在im 4中，我们将检验特定的下丘脑核团参与瘦素的胰岛素增敏作用的假设。我们将在DMH、VMH和PVH中微量注射瘦素，并通过正葡萄糖钳夹和葡萄糖摄取来测量体内特定肌肉和脂肪库的葡萄糖处置。因此，我们将绘制胰岛素敏感性区域。这些研究将产生有关瘦素增强胰岛素敏感性的直接和CNS介导的机制的重要信息，并可能导致治疗肥胖和糖尿病的新途径。

The overall goal of this proposal is to understand the molecular mechanisms underlying the effect of leptin to rapidly enhance insulin sensitivity independent of its effects on food intake and body weight. We will investigate the contribution of CNS-mediated and direct effects at the level of peripheral tissues. In Aim 1 we will determine whether insulin and leptin share common intracellular signal transduction pathways. With i.v. administration of leptin in rats, we will test the hypothesis that the insulin- sensitizing effects of leptin involve convergence of synergism between leptin-activated and insulin-activated signal transduction pathways. We will determine the time course for leptin activation of phosphorylation of the insulin receptor and IRSs and activation of PI3 kinase, MAPK kinase and Stat1 and Stat in insulin target tissues (skeletal muscle, BAT, WAT liver) in vivo. We will investigate potential additive effects of leptin and insulin. In Aim 2 we will test the hypothesis that early activation of signaling pathways by leptin results from direct effects the level of the target tissues but later effects involve CNS mediation as well. We will measure activation of signaling pathways in the hypothalamus after iv or ICV leptin over a time course. We will determine which effects are present after sympathectomy of skeletal muscle or BT. We will determine which signaling pathways are activated in vitro in adipocytes and muscle. In Aim 3 we will test the hypothesis that some of the biological actions of leptin are exerted directly at the level of the adipocyte. We will generate transgenic db/db mice expressing the leptin receptor selectively in fat using the aP2 promoter/enhancer. We will determine whether restoration of long form leptin receptors exclusively in adipocytes alters adipocyte physiology, glucose homeostasis, or insulin action. In im 4 we will test the hypothesis that specific hypothalamic nuclei are involved in the insulin-sensitizing effects of leptin. We will microinject leptin into the DMH, VMH and PVH and measure glucose disposal by euglycemic clamp and glucose uptake into specific muscles and adipose depots in vivo. Thus, we will map the regions of insulin sensitivity. These studies will yield important information about the direct and CNS-mediated mechanisms by which leptin enhances insulin sensitively and may lead to new avenues for treatment of obesity and diabetes.