Neurochemistry/Physiology of Proopiomelanocortin Neurons

描述（由申请人提供）：肥胖及其相关的代谢性疾病是世界范围内的主要健康问题。虽然肥胖率迅速上升的原因是多方面的，但很明显，大脑中的神经回路在感知能量储存和调节能量平衡方面发挥着重要作用。这些神经通路是开发旨在逆转肥胖流行病的新药物疗法的候选靶点，因此我们必须详细了解它们的功能，包括与其他神经系统相互连接的完整地图以及它们对多种神经递质和细胞内信号通路的利用。这个项目的重点是大脑能量平衡回路的关键组成部分，位于下丘脑和脑干的前阿黑皮素（POMC）神经元。大脑POMC功能的遗传缺失导致严重肥胖和代谢综合征，其特征在于极度摄食过多和基础代谢率降低。然而，POMC神经元在许多方面都是异质的，越来越多的证据表明，不同的神经元亚群调节不同的神经过程，共同导致热量平衡的正常或病理控制。该项目的总体目标是确定这些神经元亚群的特定功能以及它们所利用的神经解剖学和分子通路。具体目标1包括一系列行为和药理学研究，以探索POMC缺陷小鼠模型中导致摄食过多的潜在组成过程和神经底物。这些实验利用一种新开发的膳食模式分析方法，并将测试黑皮质素信号协调调节刻板的运动，奖励和享乐方面的喂养行为的假设，这是特别相关的人类问题周围的食物选择和膳食规模的肥胖症的临床发病机制的过程。具体目标2将进一步定义连接弓状POMC神经元与边缘前脑核的神经回路。一种新的逆行追踪方法，涉及位点特异性显微注射的犬腺病毒载体表达Cre重组酶到突变小鼠与可逆沉默POMC基因等位基因将被用来映射轴突侧支的特定组合的目标网站。由POMC终末支配的边缘区中的二级神经元将通过用在POMC细胞体中表达并从POMC细胞体顺行转运的麦胚凝集素进行跨突触标记来鉴定。我们还将使用遗传技术和多标记免疫组织化学来研究POMC神经元的意外复杂的树突，从远端站点接收突触输入。最后，在具体目标3中，我们将使用互补遗传方法来研究空间上不同的POMC神经元亚群或发育改变的POMC基因表达在预防或减轻肥胖中的独特功能作用。所涉及的技术是聚集嵌合体的形成和Cre重组酶介导的POMC表达的神经元特异性和可逆沉默的POMC等位基因的再激活。公共卫生相关性： 目前对公共健康的最大威胁是超重、肥胖、糖尿病和代谢综合征的发病率不断增加。一组复杂的神经回路将热量需求和利用之间的平衡与进食相关的行为和心理过程相结合。该项目集中在大脑进食回路的关键组成部分，propiomelanocortin神经元，以解释这些神经元如何协调调节食欲，膳食开始和终止，以及代谢率，以正常维持体重在严格控制的范围内。

DESCRIPTION (provided by applicant): Obesity and its associated metabolic diseases are major health problems world-wide. Although the reasons for the rapid increase in rates of obesity are multifactorial, it is clear that neural circuits in the brain play a major role in sensing energy stores and regulating energy balance. These neural pathways are candidate targets for the development of new pharmacotherapies aimed at reversing the obesity epidemic and therefore it is essential that we understand their function in detail, including the complete map of interconnections with other neural systems and their utilization of multiple neurotransmitters and intracellular signaling pathways. This project focuses on a key component of the brain's energy balance circuitry, the proopiomelanocortin (POMC) neurons located in the hypothalamus and brainstem. Genetic deletion of POMC function from the brain results in profound obesity and a metabolic syndrome characterized by extreme hyperphagia and reduced basal metabolic rate. However, POMC neurons are heterogeneous in many aspects and accumulating evidence suggests that different subpopulations of the neurons regulate separate neurological processes that together result in normal or pathological control of caloric balance. The overall goals of this project are to identify specific functions of these neuronal subpopulations and the neuroanatomic and molecular pathways that they utilize. Specific aim 1 includes a series of behavioral and pharmacological studies to probe the underlying component processes and neural substrates contributing to hyperphagia in POMC-deficient mouse models. These experiments utilize a newly developed method for meal pattern analysis and will test the hypothesis that melanocortin signaling coordinately modulates stereotyped motor, reward, and hedonic aspects of feeding behavior; processes which are particularly relevant to human issues surrounding food choice and meal size in the clinical pathogenesis of obesity. Specific aim 2 will further define the neurocircuitry connecting arcuate POMC neurons to limbic forebrain nuclei. A novel retrograde tracing method involving site-specific microinjections of a canine adenoviral vector expressing Cre recombinase into mutant mice with a reversibly silenced POMC gene allele will be used to map axon collaterals to specific combinations of target sites. Second order neurons in limbic areas innervated by POMC terminals will be identified by trans-synaptic labeling with wheat germ agglutinin expressed in, and transported anterogradely from, POMC cell bodies. We will also use genetic techniques and multilabel immunohistochemistry to study the unexpectedly complex dendrites of POMC neurons that receive synaptic inputs from distal sites. Finally, in Specific aim 3 we will use complementary genetic approaches to study the unique functional role of spatially distinct POMC neuron subpopulations or developmentally altered POMC gene expression in the prevention or mitigation of obesity. The techniques involved are aggregation chimera formation and Cre recombinase-mediated reactivation of POMC expression from a neuron-specific and reversibly silenced POMC allele. PUBLIC HEALTH RELEVANCE: Among the greatest current threats to public health are the continually increasing rates of overweight, obesity, diabetes, and the metabolic syndrome. A complex set of neural circuits integrates the balance between caloric demand and utilization with the behavioral and psychological processes related to feeding. This project centers on a key component of the brain's feeding circuits, propiomelanocortin neurons, to explain how these neurons coordinately regulate appetite, meal initiation and termination, and metabolic rate to normally maintain body weight within tightly controlled limits.