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Studies of the development and differentiation of mammalian embryos

Waugh, Esau Emanuel (1989) Studies of the development and differentiation of mammalian embryos. PhD thesis, Murdoch University.

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Abstract

Within this study programme, the metabolic capability of the preimplantation sheep conceptus as it develops and differentiates from the zygote to the 19th day of pregnancy was investigated. For comparative purposes, studies were also undertaken with the preimplantation mouse embryo and the morula/early-blastocyst stage of the cattle embryo. Short term incubation of freshly collected embryonic material in medium supplemented with radiolabelled glucose, lactate, acetate or amino acids was employed in the study. For experiments involving the 13- to 19- day sheep conceptus, the embryonic discs (day 13 and 15) or differentiating embryos (day 17 and 19) were separated from extraembryonic membranes and the membranes then separated by careful dissection. The metabolism of these embryos and extraembryonic membranes was assessed separately.

Initial experiments demonstrated the ability of cleavage stage embryos to utilize glucose, lactate and acetate. The catabolic utilization of acetate in the three species studied was very low compared to the utilization of glucose and lactate. The anabolic utilization of this substrate was also low and the results suggest that there is little acetate utilization either directly or indirectly by these embryos. Studies with the cleavage stage mouse embryo showed that lactate and glucose had no effect on the metabolism of acetate and that acetate had no effect on the metabolism of these other substrates. Furthermore, studies with the extraembryonic tissue of the late sheep conceptus failed to demonstrate a major effect of the presence of glucose on acetate utilization. In these advanced preimplantation stages there was a divergence in the utilization pattern of acetate between the embryonic structures as development progressed. In the differentiating embryo the rate of utilization of this substrate decreased with time, whilst in the emerging extraembryonic membranes there was a contrasting increase in the rate of utilization.

The studies of the catabolic utilization of glucose and lactate by tissues of the 13- to 19- day sheep conceptus showed that the rate of catabolism of these substrates decreased with development in all tissues. Both TCA cycle and glycolysis were active in these tissues with glycolysis becoming increasingly dominant with development.

The rate of catabolic utilization by trophoblastic tissue incubated in medium with lactate as sole energy source was lower than that when glucose was used as sole energy source. Further investigation through enzyme recycling techniques showed that lactate as sole substrate altered the redox state. Incubation in this substrate also lead to increased cell death.

Finally using incubation in labelled amino acids followed by polyacrylamide gel electrophoresis and fluorography, the synthesis and secretion of soluble proteins by the 13- to 19- day sheep trophoblast was investigated. The timing of the initiation and cessation of the synthesis of these proteins was examined as such proteins could function as embryonic messages in pregnancy recognition.

Item Type: Thesis (PhD)
Murdoch Affiliation: 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: repository@murdoch.edu.au. Thank you.
Supervisor(s): Wales, Ray
URI: http://researchrepository.murdoch.edu.au/id/eprint/53576
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