The role of exogenous energy substrates in blastocoele fluid accumulation in the rat

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Abstract
Preimplantation mammalian development culminates in the formation of a fluid-filled cavity, the blastocoele, which is a prerequisite for successful implantation and further development. The blastocoele is enclosed by a single layer of polarised cells, the trophectoderm, which is the first epithelium formed in development. In embryos of the mouse and the rabbit, a basolaterally located Na+/K+-ATPase hydrolyses ATP to drive the vectorial transport of ions, which is responsible for the accumulation of blastocoele fluid. Using non-invasive assays of energy substrate consumption and blastocoele fluid accumulation, experiments were carried out on single preimplantation rat embryos, to establish: (1) the roles of the Na+/K+-ATPase and exogenous energy substrates, and (2) the relationship between the consumption and metabolism of energy substrates and fluid accumulation, during blastocoele cavity formation in this species. Ouabain 0.5 mM and energy-substrate-free medium both caused an inhibition in the number of embryos forming a blastocoele in culture, and also reduced the rate of fluid accumulation by day 5 blastocysts collapsed in cytochalasin-D and allowed to re-expand. Ouabain also reduced the consumption of glucose (but not pyruvate) and the production of lactate by re-expanding blastocysts. In the absence of the inhibitor, a direct relationship was seen between fluid accumulation and both glucose (but not pyruvate) consumption and lactate production. However, ouabain had no effect on intact, expanded blastocysts. These results suggest that (1) a basolaterally located, ouabain-inhibitable Na+/K+-ATPase is involved in rat blastocoele formation, (2) this process is dependent on exogenous energy substrates, and (3) there may be a direct relationship between the metabolism of glucose via glycolysis, and blastocoele fluid accumulation.