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Studies of embryo metabolism during preimplantation development

Du, Zheng Feng (1992) Studies of embryo metabolism during preimplantation development. PhD thesis, Murdoch University.

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The experiments reported in this thesis were undertaken to study two aspects of the biology of mammalian preimplantation embryos. The first of these dealt with the utilization of the energy substrate, glucose, by the sheep embryo up to and including day 19 of pregnancy. The second dealt with the utilization of glutamine by mammalian embryos and the reciprocal effects of glutamine and glucose on each others metabolism. This latter study was prompted by the recent reports of the beneficial effects of medium containing glutamine, but excluding glucose, on the ability of mouse zygotes to undergo full preimplantation development in vivo . Prior to undertaking the metabolic studies with glutamine a series of experiments were undertaken to study the capacity of the mouse zygotes of different genetic background to develop in vitro and the constituents of the medium that may influence this development.

Initial experiments were carried out to examine the effects of reducing oxygen concentration upon the catabolic and anabolic utilization of glucose by the preimplantation sheep conceptus as it develops and differentiates from the cleaving stage to the 19th day of pregnancy. The results of this study demonstrated that lowering O2 to 5% substantially altered both the catabolic and anabolic metabolism of glucose but the pattern of response depended on the stage at which the conceptus was explanted. Next the activity of the pentose phosphate pathway was estimated in both cleaving embryos and the advanced conceptus. There was a substantial utilization of glucose through this pathway and the activity of the pathway changed with development and differentiation. As it has been proposed that trophoblastic vesicles formed by in vitro culture of fresh trophoblast may be used as a model system for studying the biochemistry and physiology of the preimplantation embryos, a study was undertaken to examine glucose metabolism by sheep trophoblastic vesicles formed from day 13 trophoblast and cultured for 6 days. By assessing catabolic and anabolic metabolism of glucose, it was shown that these vesicles were active in the utilization of glucose but the pattern of metabolism did not mimic that seen with fresh tissue samples.

The second major aspect dealt with the role of glutamine in the biology and metabolism of the embryo during preimplantation development. The first experiment investigated the development of mouse zygotes of different genotype through the "2-cell block" and the effect of medium components including glutamine, glucose and EDTA upon this development. The results showed that the glutamine containing medium, (CZB medium), significantly improved the development of embryos with more than 50% of zygotes from blocking strains (randombred) reaching the blastocyst stage. By using reciprocal crosses between blocking and non-blocking strains it was shown that the maternal genome is mainly responsible for the "2-cell block". As regards media constituents it was found that EDTA was essential for the 1-cell embryo to pass the 2-cell block. Glutamine, in general, did not exert a major effect upon development of mouse zygotes but appeared to benefit development at early stages. Glucose was necessary for successful development of embryos at least from the post-block stages and had no detrimental effects when added from the commencement of culture. Subsequent studies were undertaken to investigate the metabolism of glutamine and glucose by in vivo and in vitro derived mouse embryos. The data showed that glutamine can be utilized catabolically as an energy source by oxidation to CO2. Embryos from the blocking strain metabolised significantly less glutamine than did those from the non-blocking strain but glucose metabolism was similar between the embryos from the two strains, suggesting that strain differences in embryonic development in vitro may be related to the metabolism of glutamine rather than glucose.

Finally, a study of glutamine utilization and its reciprocal relationship with glucose by the sheep conceptus was undertaken. These experiments showed that, as found in the mouse, glutamine can be substantially utilized by the conceptus via oxidation to CO2 through the TCA cycle. However, there was increasing preferential utilization of glucose for energy metabolism with development from the cleaving embryo through to the day 19 conceptus.

Item Type: Thesis (PhD)
Murdoch Affiliation(s): School of Veterinary Studies
Notes: Note to the author: If you would like to make your thesis openly available on Murdoch University Library's Research Repository, please contact: Thank you.
Supervisor(s): Wales, Ray
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