Propogating waves of Ca2+ have been observed by physiologists in a wide variety of cellular systems, ranging from secretory cells in the pancreas to neurons. These waves appear to function as part of the second messenger role of Ca2+ in transmitting signals from hormone receptors and other molecular detectors in the plasma membrane. In mature oocytes Ca2+ waves are initiated by the egg-sperm fusion event and have been observed in every species studied. In the mature Xenopus laevis oocyte, as in mammalian species, the inositol phospholipid cascade has been demonstrated to be the major signal transduction pathway involved in these waves. The proposed research investigates the mechanism of initiation, propogation, and termination of the fertilization wave in the mature Xenopus laevis oocyte by developing computer-based models of the primary Ca2+ handling mechanisms in this cell. Development and analysis of these kinetic mechanisms will be modular, i.e., individual steps in the overall mechanism will be fine-tuned to experimental measurements before being combined into a 3-dimensional model. The 3-dimensional model, necessitated by the large size of the oocyte (1.2mm), will include diffusion of Ca2+ in the presence of exogenous and endogenous buffers, diffusion of inositol 1,4,5-trisphosphate (IP3), and Ca2+ handling by the endoplasmic reticulum, mitochondria, and plasma membrane. To insure that model parameters are chosen correctly and to test predictions of the model, experiments that establish the rate of Ca2+ handling will be performed in the stratified mature Xenopus laevis oocyte, a novel preparation that permits separation of cellular organelles in vivo. Simulations will be carried out to help test hypotheses regarding possible mechanisms of initiation, propogation and termination of the fertilization wave. Additional calculations will be performed to help interpret on-going experiments with the unstratified egg. Because IP3-induced Ca2+ release from the endoplasmic reticulum is the major process involved in the fertilization wave in mammalian eggs, the African frog, Xenopus laevis, is a good biological model for fertilization in mammals. After dormancy for many years, fertilization stimulates events that activate the metabolism and development of an egg cell. This research will help clarify the role that the Ca2+ wave plays in these important physiological processes.
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