MODELING THE FERTILIZATION CALCIUM WAVE IN MOUSE EGGS

这项提议的总体目标是开发一个现实的小鼠卵母细胞和卵子的空间模型，这将使我能够检查成熟和受精的机制。然后我将用这个模型来回答一个非常简单的问题：在成熟过程中，未成熟的卵母细胞发生了什么，使其可以受精，一旦受精，这些成熟诱导的变化对随后的钙信号是如何重要的？已知在成熟过程中，卵母细胞释放钙的能力增强，并提出了许多可能的机制：L)内质网钙隔离能力的增加；2)内质网通过IP3受体数量和/或密度的增加而增加内质网释放钙的能力；3)由于内质网结构的重排，内质网在特定的空间区域差异地隔离和/或释放钙的能力；4)由于内流导致细胞内总钙的增加；以及5)IP3受体敏感性的改变。借助于该模型结合成熟和受精机制，我将检验这些变化对受精前和受精后卵子的影响，并将它们与实验进行比较，以检验这些假设。此外，我将使用该模型来研究受精波的启动和传播机制，类似于我之前对非洲爪哇所做的工作。最后，在合理的情况下，我将进行有限的实验室实验，收集数据用于模型校准和可能的模拟结果比较，而这些数据在文献中无法获得。建模和仿真将与莱斯·勒夫博士和CBIT的其他人合作进行，而与劳琳达·贾菲博士的实验室成员合作将极大地帮助实验工作。

The overall goal of this proposal is to develop a realistic spatial model of the mouse oocyte and egg, that will allow me to examine the mechanisms underlying maturation and fertilization. I will then use this model to answer a very simple question: what happens to the immature oocyte during maturation that makes it fertilizable, and once it is fertilized, how are these maturation-induced changes important for the ensuing calcium signal? It has been known for some time that during maturation, the oocyte increases its capability to release calcium, and a number of potential mechanisms have been proposed: l) an increase in the ER's calcium-sequestering capability; 2) an increase in the ER's ability to release calcium via an increase in the number and/or density of Ip3 receptors; 3) an increase in the ER's ability to sequester and/or release calcium differentially in specific spatial regions as a result of structural rearrangement of the ER; 4) an increase in total intracellular calcium due to influx; and 5) a change in the IP3 receptor sensitivity. By dint of the model's incorporation of both maturation and fertilization mechanisms, I will test these hypotheses by examining the effects of such changes on both the pre-fertilization and post-fertilization egg, and comparing them to experiment. Additionally, I will use the model to investigate the mechanisms underlying the initiation and propagation of the fertilization wave, in a manner similar to my previous work with Xenopus. Finally, I will perform limited laboratory experiments, when reasonable, to collect data for both model calibration and, possibly, comparison of simulation results, when such data are not available in the literature. The modeling and simulation will be performed in collaboration with Dr. Les Loew and others in CBIT, and the experimental work will be greatly aided by collaboration with members of Dr. Laurinda Jaffe's lab.