The Inositol Phosphate Cascade as a Regulator of FSH and LH actions in the Ovary

描述（由申请人提供）：促性腺激素诱导的颗粒细胞中芳香化酶的表达依赖于激活的信号通路，这是由每种促性腺激素受体的密度决定的。我们假设低促性腺激素受体密度支持芳香化酶的表达，因为只有cAMP和Akt通路被激活，而高促性腺激素受体密度不支持芳香化酶的表达，因为它们也刺激肌醇磷酸盐和二酰基甘油的积累，导致表调节蛋白的释放和EGFR和ERK1/2的磷酸化。据报道，LHR参与刺激肌醇磷酸级联作为芳香酶表达的抑制剂，仅使用在颗粒细胞原代培养中表达的促性腺激素受体突变体。因此，我们建议使用转基因小鼠模型来确定LHR激活卵巢肌醇磷酸级联对于排卵后芳香化酶表达的下调是否必要。Aim 1将生成一个颗粒细胞特异性缺失Gq/11敏感肌醇磷酸级联的小鼠模型，并表征这些小鼠的Gaq和Ga11的表达。Aim 2将决定由Aim 1产生的小鼠的生殖表型。我们将对未成熟雌性小鼠注射PMSG后再注射hCG的卵巢芳香化酶表达进行量化，以证实我们的假设。我们还将测量未成熟和3至6月龄KO和wt小鼠的血清LH， FSH， E2和P4。分析单独注射PMSG或PMSG与hCG的未成熟雌性卵巢组织学和E2、P4血清浓度。如果我们发现KOmice中hCG后芳香化酶的表达仍然很高（正如预测的那样），我们将使用这些小鼠的颗粒细胞来测量芳香化酶反应中所提出的信号通路的不同组成部分。无论所提出的实验结果如何，具有颗粒细胞特异性肌醇磷酸级联缺失的小鼠将使我们能够测试肌醇磷酸级联在卵巢生理学上的其他潜在作用。

DESCRIPTION (provided by applicant): The gonadotropin-induced expression of aromatase in granulosa cells is dependent on the signaling pathways that are activated, which is dictated by the density of each gonadotropin receptor. We hypothesize that low gonadotropin receptor densities support aromatase expression because only cAMP and Akt pathways are activated, while higher gonadotropin receptor densities do not support aromatase expression because they also stimulate the accumulation of inositol phosphates and diacylglycerol leading to the release of epiregulin and phosphorylation of EGFR and ERK1/2. Involvement of LHR to stimulate the inositol phosphate cascade as an inhibitor of aromatase expression has been reported only using gonadotropin receptor mutants expressed in primary cultures of granulosa cells. Therefore, we propose to use genetically modified mouse models to determine if activation of the ovarian inositol phosphate cascade by LHR is necessary for down regulation of aromatase expression that occurs after ovulation. Aim 1 will generate a mouse model with a granulosa cell- specific deletion of the Gq/11-sensitive inositol phosphate cascade and characterize the expression of Gaq and Ga11 of these mice. Aim 2 will determine the reproductive phenotype of mice produced from Aim 1. Ovarian aromatase expression will be quantified in immature female mice injected with PMSG followed by hCG to confirm our hypothesis. We will also measure serum LH, FSH, E2 and P4 of immature and 3 to 6 month old KO and wt mice. Ovarian histology and E2 and P4 serum concentrations of immature females injected with PMSG alone or PMSG and hCG will also be analyzed. If we find that aromatase expression remains high after hCG (as predicted) in the KOmice, we will then use granulosa cells from these mice to measure the different components of the proposed signaling pathway involved in the aromatase response. Regardless of the results of the proposed experiments having mice with a granulosa cell-specific deletion of the inositol phosphate cascade will allow us to test for other potential roles of the inositol phosphate cascade on ovarian physiology. PUBLIC HEALTH RELEVANCE: Understanding common and divergent effects of the FSH/FSHR and LH/LHR pairs on ovarian physiology will aid in the use of FSH and LH/hCG as infertility drugs and how they regulate the major hormones (estradiol and progesterone) that control reproduction.