Cross-talk between Leptin and Estrogen Signaling in Hypothalamic Arcuate Neurons

描述（由申请人提供）：本项目的主要目标是阐明kisspeptin神经元中瘦素和17-雌二醇信号之间的相互作用。先天性瘦素缺乏和/或瘦素突变导致的瘦素功能丧失可导致肥胖和低促性腺激素性性腺功能减退症。下丘脑性腺功能减退症及其相关紊乱可通过给予瘦素逆转。瘦素通过其同源受体瘦素受体（LRs）发出信号。长同种型（LRb）是受体的主要信号形式，并在下丘脑神经元中大量表达，包括弓形阿黑皮素原（POMC）和kisspeptin神经元，但不在促性腺激素释放激素（GnRH）神经元中。因此，瘦素对GnRH神经元的作用被认为是通过与GnRH神经元的突触间接介导的。下丘脑kisspeptin神经元在调节GnRH释放和生殖控制中起着关键作用。此外，肥胖和不育ob/ob小鼠中KiSS 1 mRNA减少，瘦素给药后Kiss 1 mRNA水平增加。此外，KiSS 1 mRNA和kisspeptin蛋白高度调节17-雌二醇（E2），最近，我们发现，E2差异调节弓状kisspeptin神经元在雌性豚鼠，抑制表达在负反馈和增强表达在正反馈。此外，我们已经发现，瘦素通过一种新的信号通路，耦合到典型的瞬时受体电位（TRPC）通道的激活POMC神经元去极化，和kisspeptin神经元可能受到类似的调节。因此，我们目前的工作集中在下丘脑弓状kisspeptin神经元和E2和瘦素之间的相互作用，通过多个信号级联影响kisspeptin神经元的兴奋性，并最终生殖周期。我们的多学科方法结合了一系列独特的细胞和分子工具以及我们的综合专业知识（电生理学，分子生物学，组织化学和整体动物生理学）。我们的工作假设是，kisspeptin神经元是女性GnRH神经元兴奋性驱动的“看门人”，并且这些神经元中E2和瘦素的复杂相互作用控制了进食和禁食状态下的排卵周期。因此，我们的具体目标如下：（1）研究去卵巢雌性豚鼠弓状核kisspeptin神经元的瘦素信号通路。(2)研究正反馈和负反馈时E2对弓状kisspeptin神经元的影响。(3)阐明E2对禁食与进食豚鼠弓状吻肽神经元正反馈的作用。(4)探讨E2和禁食对Kisspeptin神经元K-ATP通道表达的影响。了解弓状kisspeptin神经元中瘦素和E2信号的会聚将提供对这些激素在将代谢线索传递到生殖轴中的基本作用的深入了解。 公共卫生相关性：妇女过度运动、饮食失调或体重减轻可引起闭经，其特征是血清17-雌二醇水平低和无排卵。众所周知，在负能量平衡期间，从脂肪储存中释放的激素瘦素急剧减少，并可能导致下丘脑性闭经。然而，瘦素受体不表达在下丘脑促性腺激素释放激素（GnRH）神经元，这是直接负责控制排卵周期，但在kisspeptin神经元是必不可少的兴奋性输入到GnRH神经元。因此，目前的研究将描述在进食和禁食状态下kisspeptin神经元中瘦素和17-雌二醇信号之间的串扰。了解弓状kisspeptin神经元中瘦素和雌激素信号的会聚将提供对这些激素在将代谢线索传递到生殖轴中的基本作用的深入了解。

DESCRIPTION (provided by applicant): The primary goal of this project is to elucidate the cross-talk between leptin and 17-estradiol signaling in kisspeptin neurons. Congenital leptin deficiency and/or loss of leptin function due to mutations in leptin can cause obesity and hypogonadotropic hypogonadism. Hypothalamic hypogonadism and its associated disturbances can be reversed by administration of leptin. Leptin signals via its cognate receptors, leptin receptors (LRs). The long isoform (LRb) is the predominant signaling form of the receptor and is abundantly expressed in hypothalamic neurons, including arcuate proopiomelanocortin (POMC) and kisspeptin neurons, but not in gonadotropin releasing-hormone (GnRH) neurons. Therefore, the effects of leptin on GnRH neurons are thought to be mediated indirectly via neurons synapsing on GnRH neurons. Hypothalamic kisspeptin neurons play a critical role in modulating GnRH release and hence the control of reproduction. Moreover, KiSS1 mRNA is reduced in obese and infertile ob/ob mice, and the levels of Kiss1 mRNA increase after administration of leptin. Furthermore, KiSS1 mRNA and kisspeptin protein are highly regulated by 17- estradiol (E2), and recently, we have found that E2 differentially regulates arcuate kisspeptin neurons in the female guinea pig, inhibiting expression during negative feedback and augmenting expression during positive feedback. In addition, we have discovered that leptin depolarizes POMC neurons via a novel signaling pathway that is coupled to activation of canonical transient receptor potential (TRPC) channels, and kisspeptin neurons may be similarly regulated. Therefore, our current work focuses on hypothalamic arcuate kisspeptin neurons and the interaction between E2 and leptin acting through multiple signaling cascades to affect kisspeptin neuronal excitability and ultimately the reproductive cycle. Our multidisciplinary approach incorporates a unique array of cellular and molecular tools and our combined expertise (electrophysiology, molecular biology, histochemistry and whole animal physiology). Our working hypothesis is that the kisspeptin neurons are the "gate-keeper" of excitatory drive to GnRH neurons in the female, and it is the complex interaction of E2 and leptin in these neurons that control the ovulatory cycle in fed and fasted states. Therefore, our specific aims are the following: (1) To characterize the leptin signaling pathway in arcuate kisspeptin neurons in ovariectomized female guinea pigs. (2) To characterize the effects of E2 on arcuate kisspeptin neurons during positive feedback versus negative feedback. (3) To elucidate the actions of E2 on arcuate kisspeptin neurons during positive feedback in fasted versus fed guinea pigs. (4) To elucidate the effects of E2 and fasting on the expression of K-ATP channels in kisspeptin neurons. Understanding the convergence of leptin and E2 signaling in arcuate kisspeptin neurons will provide insight into the fundamental role of these hormones in conveying metabolic cues to the reproductive axis. PUBLIC HEALTH RELEVANCE: Excessive exercise, eating disorders or weight loss in women can cause amenorrhea, which is characterized by low levels of serum 17-estradiol and anovulation. It is known that during negative energy balance the hormone leptin that is released from fat stores is drastically reduced and may contribute to hypothalamic amenorrhea conditions. However, leptin receptors are not expressed in hypothalamic gonadotropin releasing hormone (GnRH) neurons, which are directly responsible for the control of the ovulatory cycle, but in kisspeptin neurons that are essential for the excitatory input to GnRH neurons. Therefore, the current studies will characterize the cross-talk between leptin and 17-estradiol signaling in kisspeptin neurons in fed and fasted states. Understanding the convergence of leptin and estrogen signaling in arcuate kisspeptin neurons will provide insight into the fundamental role of these hormones in conveying metabolic cues to the reproductive axis.