Estrogen Modulation of Bursting Activity in GnRH neurons

雌激素对 GnRH 神经元爆发活性的调节

基本信息

批准号：
7805411
负责人：
Oline Karin Rønnekleiv
金额：
$ 33.35万
依托单位：
OREGON HEALTH & SCIENCE UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2004
资助国家：
美国
起止时间：
2004-01-01 至 2014-03-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/7805411
关键词：
Agonist Animal Model Applications Grants Back Cell model Cell physiology Cells Complex Contraceptive methods Coupled Enkephalin, Ala(2)-MePhe(4)-Gly(5)-Estradiol Estrogen Receptors Estrogens Exhibits Feedback Feeds Female Fertility G Protein-Coupled Receptor 54 GTP-Binding Proteins Gene Expression Glutamates Hypogonadism Hypothalamic structure Infertility Ion Channel KISS1R gene Knowledge Lead Ligands Luteinizing Hormone Measurement Mediating Membrane Metabolic Molds Molecular Mutation Neurons Neuropeptides Neurosecretion Neurotransmitters Opioid Opioid Receptor Ovarian Ovulation Patients Phase Phosphorylation Pituitary Gland Play Regulation Research Reverse Transcriptase Polymerase Chain Reaction Reverse Transcription Role Sexual Development Signal Pathway Signal Transduction Signaling Molecule Steroids Synapses T-Type Calcium Channels Techniques Time Whole-Cell Recordings Work attenuation computerized data processing density experience feeding gamma-Aminobutyric Acid kisspeptin mRNA Expression mouse model neuronal excitability novel patch clamp postsynaptic presynaptic public health relevance receptor receptor coupling reproductive research study response

项目摘要

DESCRIPTION (provided by applicant): The overall objectives of this project are to ascertain the mechanisms by which the ovarian steroid 17 - estradiol (E2) regulates GnRH neuronal excitability, which controls GnRH neurosecretion and fertility. These neurons are located in the hypothalamus and constitute the final step in the regulation of pituitary luteinizing hormone (LH) secretion and ovulation in females. Importantly, E2 feeds back to alternately inhibit and stimulate GnRH neurosecretion, via negative and positive feedback, respectively. Extensive studies have demonstrated that these E2 actions are complex and involve multiple neurotransmitters and metabolic factors. However, we have limited knowledge about the cellular and molecular mechanisms by which GnRH neurons are regulated, and therefore, incomplete understanding of the control of fertility. Recent evidence suggests that E2 acts directly on GnRH neurons through estrogen receptor (ER) or a novel membrane ER (mER), as well as presynaptically through ER and mER. We have identified that inwardly rectifying K+ (Kir) conductances that are regulated by E2 play a major role in mediating GnRH cellular excitability and may be involved in negative feedback on GnRH secretion. In addition, we have discovered that the reproductively essential neuropeptide kisspeptin depolarizes GnRH neurons through inhibition of Kir channels and activation of nonselective cationic (TRPC-like) channels. The inhibition of Kir and activation of TRPC channels are important mechanisms by which kisspeptin abrogates inhibitory drive and depolarizes GnRH neurons at the time of the GnRH surge. In this proposal, we seek to further explore the mechanisms by which E2 governs GnRH neuronal excitability using whole-cell patch clamp and single cell reverse transcription PCR approaches, techniques with which we have extensive experience. We will focus on elucidating the mechanisms by which E2 modulates critical excitatory input (e.g. kisspeptin, glutamate) and inhibitory input (e.g. opioids, GABA). Our Specific Aims will examine important factors key to the understanding of GnRH excitability: (1) elucidate the signaling cascade by which E2 increases Kir channel activity in GnRH neurons; (2) elucidate the pre- and postsynaptic effects of - opioid receptor agonists on GnRH neurons; (3) elucidate E2 modulation of kisspeptin-GPR54 actions and the cellular signaling cascades activated by kisspeptin that increase TRPC channel activity in GnRH neurons; and (4) elucidate the E2 regulation of T-type calcium channels in GnRH neurons using the mER ligand STX. It is envisioned that the results from these studies will help in understanding the cellular actions of E2 that govern GnRH neuronal excitability, which is critical for pulsatile neurosecretion and ultimately ovulation in the female. PUBLIC HEALTH RELEVANCE: The focus of our research is to understand the cellular mechanisms by which GnRH neurons are regulated by estrogen during the negative and positive feedback phases of the female reproductive cycle. Functioning GnRH neurons are essential for survival of the species, and very little is known about the cellular and molecular mechanism by which these vital neurons are regulated. We have crafted a careful set of experiments to elucidate the signaling pathways and channels that are involved in estrogen mediated inhibition (negative feedback) and excitation (positive feedback) of GnRH neurons. It is envisioned that the results from the proposed experiments will help mold a cellular model of burst firing of GnRH neurons, pulsatile neurosecretion, and ultimately ovulation in the female.

描述（由申请人提供）：该项目的总体目标是确定卵巢类固醇 17 - 雌二醇 (E2) 调节 GnRH 神经元兴奋性的机制，从而控制 GnRH 神经分泌和生育能力。这些神经元位于下丘脑，构成调节女性垂体黄体生成素 (LH) 分泌和排卵的最后一步。重要的是，E2 分别通过负反馈和正反馈交替抑制和刺激 GnRH 神经分泌。大量研究表明，这些 E2 作用非常复杂，涉及多种神经递质和代谢因素。然而，我们对 GnRH 神经元调节的细胞和分子机制了解有限，因此对生育力的控制了解不完全。最近的证据表明，E2 通过雌激素受体 (ER) 或新型膜 ER (mER) 以及通过 ER 和 mER 在突触前直接作用于 GnRH 神经元。我们已经发现，受 E2 调节的内向整流 K+ (Kir) 电导在介导 GnRH 细胞兴奋性中发挥着重要作用，并且可能参与 GnRH 分泌的负反馈。此外，我们还发现生殖必需神经肽 Kisspeptin 通过抑制 Kir 通道和激活非选择性阳离子（TRPC 样）通道来使 GnRH 神经元去极化。 Kirspeptin 的抑制和 TRPC 通道的激活是重要机制，Kisspeptin 通过其在 GnRH 激增时消除抑制性驱动并使 GnRH 神经元去极化。在本提案中，我们寻求使用全细胞膜片钳和单细胞逆转录 PCR 方法，以及我们拥有丰富经验的技术，进一步探索 E2 控制 GnRH 神经元兴奋性的机制。我们将重点阐明 E2 调节关键兴奋性输入（例如 Kisspeptin、谷氨酸）和抑制性输入（例如阿片类药物、GABA）的机制。我们的具体目标将研究理解 GnRH 兴奋性的关键因素：（1）阐明 E2 增加 GnRH 神经元中 Kir 通道活性的信号级联； (2)阐明阿片受体激动剂对GnRH神经元的突触前和突触后作用； (3) 阐明 Kisspeptin-GPR54 作用的 E2 调节以及 Kisspeptin 激活的细胞信号级联，从而增加 GnRH 神经元中 TRPC 通道的活性； (4) 使用 mER 配体 STX 阐明 E2 对 GnRH 神经元 T 型钙通道的调节。预计这些研究的结果将有助于了解 E2 控制 GnRH 神经元兴奋性的细胞作用，这对于女性的脉动神经分泌和最终排卵至关重要。公共健康相关性：我们研究的重点是了解女性生殖周期的负反馈和正反馈阶段 GnRH 神经元受雌激素调节的细胞机制。发挥功能的 GnRH 神经元对于物种的生存至关重要，但人们对调节这些重要神经元的细胞和分子机制知之甚少。我们精心设计了一组实验来阐明参与雌激素介导的 GnRH 神经元抑制（负反馈）和兴奋（正反馈）的信号通路和通道。预计所提出的实验结果将有助于塑造 GnRH 神经元爆发放电、脉动神经分泌以及最终女性排卵的细胞模型。