Regulation of Ca2+ influx in mouse oocytes and eggs during maturation and fertilization to improve assisted reproductive technologies and modulate fertility

调节小鼠卵母细胞和卵在成熟和受精过程中的 Ca2 流入，以改进辅助生殖技术并调节生育力

基本信息

批准号：
10401470
负责人：
Rafael Antonio Fissore
金额：
$ 25.33万
依托单位：
UNIVERSITY OF MASSACHUSETTS AMHERST
依托单位国家：
美国
项目类别：
财政年份：
2018
资助国家：
美国
起止时间：
2018-08-17 至 2024-05-31
项目状态：
已结题

项目摘要

There is a persistent gap in our knowledge of the molecules and mechanisms that mediate Ca2+ influx into oocytes and eggs. Ca2+ influx is required for egg activation and embryo development in all mammals, including human beings. This gap in our knowledge therefore represents a serious impediment, as until it is filled we cannot physiologically modulate Ca2+ influx or objectively diagnose and treat infertility associated with disturbances in Ca2+ homeostasis. The long-term goal is to understand how Ca2+ homeostasis is regulated in oocytes and eggs and identify its molecular effectors. The objective here is to identify the Ca2+ channel(s) that mediate Ca2+ influx during maturation and fertilization and characterize their regulatory mechanisms. Mouse oocytes/eggs are a great model because they display distinctive Ca2+ entry during maturation and fertilization, and Ca2+ release is required for egg activation. The central hypothesis is that expression and/or distinct regulation of underdetermined Ca2+ channels on the plasma membrane underlies Ca2+ influx in oocytes, its inactivation during maturation and its recovery after fertilization. This hypothesis was conceived based on extensive preliminary data. The rationale for this research is that once the channels are identified, a better understanding of the molecular determinants of oocyte maturation and fertilization will be gained. The findings here also have the potential to translate into therapeutic methods to assist infertile couples in this country. We plan to test our central hypothesis by pursuing the following specific aims: 1) Identify the Ca2+ channel(s) that mediate Ca2+ influx in oocytes prior to and during maturation; and 2) Identify the Ca2+ influx channel(s) that support oscillations after fertilization. Under Aim 1, Ca2+ imaging, pharmacology, conditional knockout mice, and electrophysiology will be used to identify the active channel(s) and to assess the impact of Ca2+ influx on maturation; the role of a TRPM7-like current recently discovered by the applicant and collaborators will be closely examined. Under Aim 2, the signaling mechanism(s) whereby fertilization stimulates Ca2+ influx and the contributing channel(s) will be determined. A novel approach that overcomes the inactivation of Ca2+ influx in eggs will facilitate these studies. Genetic models and electrophysiology will confirm the function of these channel(s) during fertilization. The research in this application is innovative because it combines electrophysiology, pharmacology and new KO lines, approach that has served to identify two new channels in oocytes, including TRPM7, whose global deletion is embryonic lethal at E7.5. The contribution of the proposed project is significant because it is expected to allow physiological modulation of Ca2+ entry in oocytes and eggs that will produce new conditions and activation protocols for use in the clinic. It will also expand our understanding of the impact of Ca2+ homeostasis on oocyte competence and infertility.

我们对介导Ca2+涌入的分子和机制的知识存在持续的差距进入卵母细胞和鸡蛋。在所有哺乳动物中，卵子激活和胚胎发育都需要Ca2+流入，包括人类。因此它填充了我们不能在生理上调节Ca2+涌入或客观地诊断和治疗不育症与CA2+稳态中的障碍有关。长期目标是了解CA2+如何稳态受到卵母细胞和卵的调节，并鉴定其分子效应子。这里的目的是确定在成熟和受精过程中介导Ca2+流入的Ca2+通道他们的监管机制。小鼠卵母细胞/鸡蛋是一个很好的模型，因为它们显示出独特的模型 Ca2+在成熟和受精过程中进入，鸡蛋激活需要Ca2+释放。中央假设是等离子体上不确定的Ca2+通道的表达和/或不同的调节膜是卵母细胞中Ca2+涌入的基础，其成熟过程中的失活以及其恢复受精。该假设是基于广泛的初步数据来构想的。理由研究是，一旦确定了通道，就可以更好地理解分子决定因素将获得卵母细胞成熟和受精的。这里的发现也有可能翻译进入治疗方法，以协助这个国家的不育夫妇。我们计划检验我们的中心假设通过追求以下特定目的：1）确定介导卵母细胞Ca2+涌入的Ca2+通道成熟之前和期间； 2）确定支持振荡之后振荡的CA2+涌入通道受精。在AIM 1，CA2+成像，药理学，有条件敲除小鼠和电生理学下将用于识别活性通道并评估Ca2+涌入对成熟的影响；这申请人和合作者最近发现的类似于TRPM7的电流的作用将紧密检查。在AIM 2下，施肥刺激Ca2+流入的信号传导机制将确定贡献渠道。一种克服CA2+失活的新方法卵中的涌入将有助于这些研究。遗传模型和电生理学将确认该功能在受精过程中这些通道。该应用程序中的研究具有创新性，因为它结合了电生理学，药理学和新的KO系，方法已经确定了两个新的卵母细胞中的通道，包括TRPM7，其全局缺失在E7.5处是胚胎致死的。这拟议项目的贡献很大，因为它有望允许生理调节在卵母细胞和卵中的Ca2+进入，这些卵母细胞和卵将产生新的条件和激活方案诊所。它还将扩展我们对CA2+稳态对卵母细胞能力的影响的理解和不育。

项目成果

期刊论文数量（11）

专著数量（0）

科研奖励数量（0）

会议论文数量（0）

专利数量（0）

SPERM FACTORS AND EGG ACTIVATION: ICSI and the discovery of the sperm factor and PLCZ1.

DOI：
10.1530/rep-21-0487
发表时间：
2022-05-23
期刊：
REPRODUCTION
影响因子：
3.8
作者：
Gupta, Neha;Akizawa, Hiroki;Lee, Hoi Chang;Fissore, Rafael A.
通讯作者：
Fissore, Rafael A.

Multidrug resistance transporter-1 dysfunction perturbs meiosis and Ca2+ homeostasis in oocytes.

DOI：
10.1530/rep-22-0192
发表时间：
2023-01-01
期刊：
REPRODUCTION
影响因子：
3.8
作者：
Nabi, Dalileh;Bosi, Davide;Gupta, Neha;Thaker, Nidhi;Fissore, Rafael;Brayboy, Lynae M.
通讯作者：
Brayboy, Lynae M.

Editorial: The Fertilization Success From the Oocyte's Perspective.