MODELING THE FERTILIZATION CALCIUM WAVE IN MOUSE EGGS

本提案的总体目标是建立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能够受精，一旦受精，这些成熟诱导的变化对随后的钙信号有何重要意义？一段时间以来，人们已经知道在成熟过程中，卵母细胞释放钙的能力增加，并提出了许多潜在的机制：1)内质网钙隔离能力的增加；2)内质网释放钙的能力通过Ip3受体数量和/或密度的增加而增强；3)内质网的结构重排导致内质网在特定空间区域内螯合和/或释放钙的能力差异增加；4)细胞内总钙因内流而增加；5) IP3受体敏感性的变化。由于该模型结合了成熟和受精机制，我将通过检查这些变化对受精前和受精后卵子的影响来检验这些假设，并将它们与实验进行比较。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Xenopus的研究。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，如果文献中没有这样的数据，可能还会比较模拟结果。建模和仿真将与Les Loew博士和CBIT的其他人合作进行，实验工作将得到Laurinda Jaffe博士实验室成员的大力协助。

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.