The physiology and developmental competence of cultured hamster embryos: Impact of the culture milieu

The efficiency of new reproductive technologies, such as genetic engineering and cloning, are dependent on the ability of cultured embryos to develop competently. Appropriate media formulation is critical for maintaining embryo viability in vitro. Previous work demonstrates that culture with glucose and phosphate (G/P_i) inhibits hamster embryo development in vitro, however the mechanism of action remains to be elucidated. The goal of this work therefore, was to examine potential mechanisms of the G/P_i block by investigating the effect of phosphate and glucose independently on the developmental and physiological competence of the preimplantation embryo.; The effect P_i on development, ionic homeostasis, metabolic profile and mitochondrial organization was examined. Treatment with ≥2.5 mM P_i reduced developmental competence. Arrested 2-cell embryos had elevated levels of both intracellular pH and intracellular free calcium. Although culture with P_i did not alter TCA cycle activity, it did increase glycolytic activity. Mitochondrial organization was disrupted by P_i in a dose-dependent manner. These data demonstrate that for hamster embryos alone alters embryo development. Additionally, inclusion of P_i in culture medium dramatically alters embryo physiology, including metabolic profile, ionic homeostasis and mitochondrial organization.; The effects of glucose, fructose and galactose on hamster embryo development in vitro were also examined. Following culture with increasing concentrations of hexoses embryo development, morula/blastocyst cell number and cell allocation, and fetal viability were assessed. Galactose significantly reduced embryo development. Although high concentrations of glucose inhibited development, culture with low concentrations of glucose dramatically increased fetal viability post-transfer. Fructose was not inhibitory at any concentration examined and resulting fetal viability was similar to that of low levels of glucose. These data demonstrate that morphology is a poor indicator of viability and that exposure of preimplantation embryos to glucose or fructose is important for embryo viability post-transfer.; Collectively, these studies give us further insight into how specific culture components impact embryo physiology. In addition to enhancing our knowledge of basic developmental processes, the studies presented here have resulted in the design of media that more effectively supports the development of embryos in vitro. The ethical implications of transferring this technology to human use are considered.