Ovarian Hormone Regulation of LHRH Biosynthesis

描述(申请人提供)：这项研究的长期目标是确定卵巢类固醇如何触发黄体生成素释放激素(LHRH)和黄体生成素释放高峰，从而诱导排卵。在这些研究中，我们将研究光周期和雌激素信号相互作用来调节雌性前腹侧脑室周围核(AVPV)中新的双重表型GABA/谷氨酸神经元的机制。这个建议的具体目的是：1)为了验证GABA/谷氨酸神经元与LHR神经元的直接联系，我们将使用顺行和逆行追踪和免疫细胞化学方法；2)我们将使用通过GGP表达来识别LHRH神经元的电生理学研究来验证AVPV GABA/谷氨酸神经元在上午主要释放GABA，在下午主要释放谷氨酸；3)确定加压素(VP)和/或去甲肾上腺素(NA)是否触发与黄体生成素激增有关的中午GABA释放增加。我们将检测特定的VP和NA受体拮抗剂对GABA释放的标志物谷氨酸脱羧酶67(GAD67)mRNA表达的影响。我们还将确定当动物处于恒定光照下时，GAD67 mRNA水平的变化是否被阻断，如果这种变化，以及黄体生成素受体激动剂是否可以诱导这种变化；4)为了确定自动反馈是否在黄体生成素高峰之前抑制GABA合成同时刺激谷氨酸释放，我们将应用GABAB受体拮抗剂并检测AVPV细胞体、接触LHRH神经元的双表型终末的囊泡GABA转运体和囊泡谷氨酸转运体的GAD67 mRNA水平的变化；5)我们将确定VIP是否通过在黄体生成素峰的下午通过非依赖配体和依赖PR的PR激活来刺激cAMP依赖的NT基因的表达。这些研究的结果可能会为人类和非人类灵长类动物的研究提供新的靶点，并可能有助于更好地理解青春期、不孕症和下丘脑老化的神经方面。此外，确定GABA神经元能够从抑制性神经传递切换到兴奋性传递，将对理解GABA和谷氨酸在其他大脑区域的“双重信号”具有重要意义。最后，这些研究的结果可能为E2如何调节性别特有的神经发育、神经保护和神经退化等不同功能提供关键的见解。

DESCRIPTION (provided by applicant): The long-term objective of this research is to determine how ovarian steroids trigger luteinizing hormone-releasing hormone (LHRH) and LH surge release, thereby inducing ovulation. In these studies, we will investigate mechanisms through which photoperiod and estrogen signals interact to regulate novel dual-phenotype GABA/glutamate neurons in the anteroventral periventricular nucleus (AVPV) of females. The specific aims of this proposal are: 1) To verify that GABA/glutamate neurons communicate directly with LHR neurons we will use anterograde and retrograde tracing and immunocytochemistry in rats and mice; 2) We will use electrophysiological studies in which LHRH neurons are identified by GGP expression to verify that AVPV GABA/Glutamate neurons release predominantly GABA during the morning and predominantly glutamate in the afternoon; 3) To determine whether vasopressin (VP) and/or noradrenaline (NA) trigger the midday rise in GABA release linked to the LH surge. We will examine the effects of specific VP and NA receptor antagonists on expression of a marker of GABA release, glutamic acid decarboxylase 67 (GAD67) mRNA. We will also determine whether changes in GAD67 mRNA levels are blocked when animals are placed in constant light and if such changes, as well as LH surge release, can be induced with VP and/or NA receptor agonists; 4) To determine whether autofeedback inhibits GABA synthesis while stimulating glutamate release prior to the LH surge, we will administer GABAB receptor antagonists and measure changes in GAD67 mRNA levels in AVPV cell bodies, as well as vesicular GABA transporter and vesicular glutamate transporter in dual-phenotype terminals contacting LHRH neurons; 5) We will determine whether VIP stimulates cAMP-dependent NT gene expression through ligand-independent activation of PR on the afternoon of the LH surge. The results of these studies may provide new targets for examination in humans and nonhuman primates and may lead to a better understanding of the neural aspects of puberty, infertility and hypothalamic aging. In addition, establishing that GABA neurons are able to switch from inhibitory neurotransmission to excitatory transmission will have important implications for understanding GABA and glutamatergic "dual-signaling" in other brain regions. Finally, the results of these studies may provide key insights into how E2 regulates such diverse functions as sex-specific neurodevelopment, neuroprotection and neurodegeneration.