Diet-Induced Plasticity of Proopiomelanocortin Neurons

中央propropiomelanocortin （POMC）神经元是一种促进厌食行为和负能量平衡的回路，是最明确的调节代谢功能的神经网络之一。然而，POMC神经元在整个中枢神经系统中整合这些代谢信号的程度仍不清楚。POMC神经元位于下丘脑中基底，但由于缺乏明显的细胞学组织，使得它们相对难以进行解剖分析。同样，生物摄食状态的改变可以改变下丘脑回路，但饮食对POMC神经可塑性的影响尚未得到直接研究。为了完成中央POMC整合的解剖图谱，并检查饮食诱导的下丘脑回路重布线的影响，我们将利用转基因动物模型、跨突触病毒追踪和单细胞电生理学技术。在第一组实验中，将使用表达复制缺陷cr的犬腺病毒（CAV-Cre）载体，在Pomc的神经增强子模块中使用loxp -侧翼的neo卡带，功能性地重新激活神经特异性Pomc缺陷小鼠。由于CAV-Cre特异性地感染突触末端的受体，因此重新激活的POMC神经元的数量将取决于远端靶部位POMC神经支配的丰度。为了在细胞水平上直接检查POMC的形态和生理，我们将使用单细胞膜片钳技术测量基础神经活动，同时用神经生物素标记细胞。这些实验将采用不同的饮食模式，这将使我们能够检查急性和慢性摄食变化对POMC神经元形态和突触活动的影响。最后，我们将综合饲料对POMC网络的影响，将我们的饲养模式应用于表达Cre依赖的荧光树突和轴突标记的转基因POMC -Cre小鼠。这些小鼠将使我们能够研究POMC神经分布的回路水平变化，并使我们能够将中央POMC网络重建为完整的图谱。总之，破译中央POMC神经元的结构和功能将有助于深入了解能量稳态的神经控制，这对对抗全球肥胖流行病至关重要。

DESCRIPTION (provided by applicant): Central proopiomelanocortin (POMC) neuron, a circuit that promotes anorectic feeding behavior and negative energy balance, is one of the best-defined neural networks regulating metabolic function. Nevertheless, the extent to which POMC neurons integrate these metabolic signals throughout the CNS remains unclear. POMC neurons reside in the mediobasal hypothalamus, but a distinct lack of cytological organization has made them relatively intractable to anatomical analysis. Likewise, it is well-established that changes in the feeding state of an organism can rewire hypothalamic circuits, but the effects of diet on POMC neural plasticity has not been directly examined. To complete the anatomical map of central POMC integration and examine the effects of diet-induced rewiring of hypothalamic circuits, we will utilize transgenic animal models, trans-synaptic viral tracing, and single cell electrophysiology techniques. In the first set of experiments, replication-deficient Cr-expressing canine adenovirus (CAV-Cre) vectors will be used to functionally re- activate neural-specific Pomc deficient mice with a LoxP-flanked neo cassette in the neural enhancer module of Pomc. Because CAV-Cre specifically infects receptors at the synaptic terminal, the number of re-activated POMC neurons will be dependent on the abundance of POMC innervations at a distal target site. To directly examine POMC morphology and physiology at the cellular level, we will use single cell patch clamp techniques to measure basal neural activity and simultaneously label cells with neurobiotin. These experiments will be followed by different diet paradigms, which will allow us to examine the effects of acute versus chronic changes in feeding on the morphology and synaptic activity of POMC neurons. Finally, we will synthesize the effects of diet on the POMC network by applying our feeding paradigms to transgenic Pomc-Cre mice expressing Cre- dependent fluorescent dendritic and axonal markers. These mice will allow us to study circuit-level changes in POMC innervations, and will allow us to reconstruct the central POMC network into a complete atlas. Taken together, deciphering the structure and function of central POMC neurons will lend insight into neural control of energy homeostasis, critical for combating the worldwide obesity epidemic. PUBLIC HEALTH RELEVANCE: Obesity and related metabolic complications constitute over $150 billion in healthcare spending annually; therefore it is imperative for both the health and economy of the United States to develop therapeutics to combat this epidemic. Central proopiomelanocortin neurons are well known for their role in anorectic feeding behavior and energy expenditure, but the anatomy and plasticity of this circuit remains unclear. To facilitate an understanding of how the nervous system regulates metabolism and expedite the development of obesity-related therapeutics, the objective of this proposal is to elucidate the ful anatomical map of the proopiomelanocortin network and to determine how changes in diet can affect proopiomelanocortin physiology.