Estrogen Modulation of Bursting Activity in GnRH Neurons

描述（由申请人提供）：本项目的总体目标是确定17 β-雌二醇（E2）调节GnRH神经元兴奋性的机制。GnRH神经元对物种的生存至关重要，E2和神经递质对这些神经元的神经分泌特性至关重要。然而，只有有限的知识是可用的GnRH神经元调节的细胞机制。最近开发的转基因小鼠与绿色荧光蛋白（EGFP）标记的GnRH神经元，大大促进了这些重要的神经元的研究。最近的证据表明，E2通过ER β直接作用于GnRH神经元，以及通过ER α间接作用于GABA神经元，从而影响GnRH的产生和/或释放。此外，E2改变神经递质的效力，可能直接或间接影响GnRH神经元。尽管越来越多的人认识到，E2对GnRH神经元的调节还不完全清楚。为了进一步探索GnRH神经元的不同电特性，管理GnRH兴奋性和E2如何调节这些特性，我们将专注于选择性离子通道和受体的基础上的模型，在GnRH神经元的突发放电。我们的工作假设是，E2调节突触输入和离子通道功能急性通过膜信号事件，并在一个较长的时间内改变特定的受体和离子通道在GnRH神经元的表达。这些变化将增强或抑制GnRH神经元活性和GnRH释放，这取决于女性的类固醇环境，最终调节生育能力。 本研究的主要目的是：（1）研究GnRH神经元在LH分泌正反馈过程中ATP敏感性钾通道亚基mRNA的表达，并探讨E2是否改变了这种表达。(2)测定E2对K-ATP通道功能的急性作用，并探讨其细胞机制。(3)确定E2是否增加α 1-肾上腺素能受体mRNA表达和蛋白。(4)测定E2对小电导钙激活K+（SK）电流的α 1-肾上腺素能抑制的影响，该电流是中等后超极化的基础。(5)确定E2是否增加钙T通道亚单位的mRNA表达，并增加T通道活性，从而增强爆发性放电。这些研究将提供新的和重要的信息，雌激素如何改变下丘脑GnRH神经元的内在电导，以及雌激素一般如何修改突触输入，从而促进不同的放电模式的GnRH神经元，这是生殖能力的关键。

DESCRIPTION (provided by applicant): The overall objectives of this project are to ascertain the mechanisms by which 17 beta-estradiol (E2) regulates GnRH neuronal excitability. GnRH neurons are crucial for the survival of the species, and both E2 and neurotransmitters are critical for the neurosecretory properties of these neurons. However, only limited knowledge is available on the cellular mechanisms by which GnRH neurons are regulated. The recent development of transgenic mice with green fluorescent protein (EGFP)-tagged GnRH neurons has greatly facilitated studies of these important neurons. Recent evidence suggests that E2 through ERbeta acts directly on GnRH neurons, as well as indirectly through ERalpha on GABA neurons to affect GnRH production and/or release. In addition, E2 alters the potency of neurotransmitters that may directly or indirectly affect GnRH neurons. Despite this growing appreciation, E2 regulation of GnRH neurons is incompletely understood. To further explore the distinct electrical properties of GnRH neurons that govern GnRH excitability and how E2 modulates these properties, we will focus on selective ion channels and receptors based on a model for burst firing in GnRH neurons. Our working hypothesis is that E2 modulates synaptic input and ion channel function acutely through membrane signaling events and over a longer time period alters the expression of specific receptors and ion channels in GnRH neurons. These changes will enhance or inhibit GnRH neuronal activity and GnRH release depending on the steroid mileu of the female, which ultimately regulates fertility. Our Specific Aims focus on key issues regulating GnRH neurons during positive feedback of LH (GnRH) secretion: (1) To measure the mRNA expression of ATP-sensitive potassium channel subunits, and determine whether E2 alters this expression. (2) To measure the acute effects of E2 on K-ATP channel function, and determine the cellular mechanism(s). (3) To ascertain whether E2 increases alpha1-adrenergic receptor mRNA expression and protein. (4) To measure the effects of E2 on the alpha1-adrenergic inhibition of a small conductance calcium-activated K+ (SK) current that underlies the medium afterhyperpolarization. (5) To ascertain whether E2 increases the mRNA expression of calcium T-channel subunits, and increases T-channel activity leading to enhanced burst firing. These studies will provide new and important information on how estrogen alters intrinsic conductances of hypothalamic GnRH neurons and how estrogen in general modifies synaptic input and thereby facilitates distinct firing patterns in GnRH neurons, which is critical for reproductive competence.