Cross-talk between Estrogen and Metabolic Hormone Signaling in Kisspeptin Neurons

项目摘要 我们研究计划的长期目标是阐明代谢状态 和17β-雌二醇（E2）调节弓状核kisspeptin（Kiss 1ARH）神经元回路，这些神经元回路对于 协调女性的能量平衡和生殖。众所周知，E2是致癌的， Kiss 1神经元直接受E2调节，对青春期发育和成年生殖至关重要 成功然而，它们在控制其他稳态功能方面的作用才刚刚出现。早些时候，我们发现， 瘦素和胰岛素通过经典瞬时受体电位（TRPC）5通道兴奋Kiss 1ARH神经元 信号，并提出他们可能作为一个重要的枢纽，在控制能量稳态。最近， 我们发现，Kiss 1ARH神经元的高频光遗传学刺激释放谷氨酸， 促食欲的前阿黑皮素（POMC）神经元，但抑制食欲神经肽Y/刺鼠相关的 肽（AgRP）神经元在女性和男性。E2增加囊泡谷氨酸转运蛋白2（VEGF 2） 雌性Kiss 1ARH神经元的mRNA表达和谷氨酸释放增强POMC兴奋， AgRP抑制。相比之下，VEGF 2 mRNA表达和谷氨酸释放在去势大鼠中增加， 与完整的男性相比，说明了谷氨酸合成和释放的重要性别差异。关键 兴奋性阳离子通道被E2上调，导致兴奋性和突触能增加。 传输最近，我们发现选择性膜雌激素受体（GqmER）激动剂STX 增加Kiss 1ARH神经元的兴奋性而不下调肽的表达。它也降低 女性和男性的食物摄入量。因此，我们假设Kiss 1ARH神经元中的雌激素信号 对于增加Kiss 1ARH神经元兴奋性和维持对以下方面至关重要的稳态功能是重要的 繁殖成功我们的多学科方法结合了一套强大的细胞，分子，遗传 和光遗传学工具，以及我们在分子生物学，电生理学和整个动物 （1）测量雌激素介导的兴奋性增加， 用GCaMP 6和Voltron记录Kiss 1ARH神经元;（2）阐明雌激素对Kiss 1ARH神经元的调节作用， 使用光遗传学刺激从Kiss 1ARH到下丘脑室旁核神经元的突触输入， 其对E2（STX）处理的雌性和STX处理的雄性的食物摄入的影响;和（3）阐明雌激素 Kiss 1ARH神经元对下丘脑背内侧核神经元突触输入的调制及其作用 E2（STX）处理的雌性和STX处理的雄性的能量消耗。解释电路和信号 E2和STX作用的基础级联将提供一种神经生理学/神经药理学机制， 一个框架，更彻底地了解Kiss 1ARH神经元协调的细胞机制 自我平衡功能与生殖。

Project Summary The long range goals of our research program has been to elucidate the mechanism(s) by which metabolic states and 17β-estradiol (E2) regulate arcuate nucleus kisspeptin (Kiss1ARH) neuronal circuits that are critical for coordinating energy homeostasis and reproduction in females. It is well known that E2 is anorexigenic, and that Kiss1 neurons, which are directly regulated by E2, are essential for pubertal development and adult reproductive success. However, their role in the control of other homeostatic functions is just emerging. Earlier, we found that Kiss1ARH neurons are excited by leptin and insulin via canonical transient receptor potential (TRPC) 5 channel signaling and proposed that they may serve as an important hub in the control of energy homeostasis. Recently, we found that high frequency optogenetic stimulation of Kiss1ARH neurons releases glutamate to excite the anorexigenic proopiomelanocortin (POMC) neurons but inhibit the orexigenic neuropeptide Y/agouti-related peptide (AgRP) neurons in both females and males. E2 increases vesicular glutamate transporter 2 (Vglut2) mRNA expression and glutamate release from female Kiss1ARH neurons to augment the POMC excitation and AgRP inhibition. In contrast, Vglut2 mRNA expression and glutamate release are increased in castrates as compared to intact males, illustrating an important sex difference in the synthesis and release of glutamate. Key excitatory cationic channels are upregulated by E2 leading to increased excitability and glutamatergic synaptic transmission. Recently, we have found that the selective membrane estrogen receptor (GqmER) agonist STX increases the excitability of Kiss1ARH neurons without downregulating the peptide expression. It also decreases food-intake in both females and males. Therefore, we hypothesize that estrogenic signaling in Kiss1ARH neurons is important for increasing Kiss1ARH neuronal excitability and maintenance of homeostatic functions critical for reproductive success. Our multidisciplinary approach incorporates a powerful set of cellular, molecular, genetic and optogenetic tools, and our combined expertise in molecular biology, electrophysiology, and whole animal physiology to address the following aims: (1) to measure the estrogenic-mediated increase in excitability of Kiss1ARH neurons using GCaMP6 and Voltron recordings; (2) to elucidate the estrogenic modulation of the synaptic input from Kiss1ARH to hypothalamic paraventricular nucleus neurons using optogenetic stimulation and its effects on food intake in E2 (STX)-treated females and STX-treated males; and (3) to elucidate the estrogenic modulation of synaptic input from Kiss1ARH neurons to hypothalamic dorsomedial nucleus neurons and its effects on energy expenditure in E2 (STX)-treated females and STX-treated males. Elucidating the circuits and signaling cascades underlying the actions of E2 and STX will provide a neurophysiological/neuropharmacological framework for a more thorough understanding of the cellular mechanisms by which Kiss1ARH neurons coordinate homeostatic functions with reproduction.