Metabolic regulation of in-vitro-produced bovine embryos. I. Effects of metabolic regulators at different glucose concentrations with embryos produced by semen from different bulls
Jose Fernando De La Torre-Sanchez A C , Kimberly Preis A B and George E. SeidelA Animal Reproduction and Biotechnology Laboratory, Colorado State University, Fort Collins, CO 80523-1683, USA.
B Colorado Center for Reproductive Medicine, 799 E. Hampden Av., Englewood, CO 80113, USA.
C Present address: Centro de Investigación Regional ‘Pacifico Centro’, INIFAP, Interior Parque Los Colomos S/N Col. Providencia, GuadalaJara, Jalisco, Mexico, CP 44660.
D Corresponding author. Email: gseidel@colostate.edu
Reproduction, Fertility and Development 18(5) 585-596 https://doi.org/10.1071/RD05063
Submitted: 8 June 2005 Accepted: 7 March 2006 Published: 24 May 2006
Abstract
The toxic and/or beneficial effects of four metabolic regulators on embryo development were evaluated. In-vitro-produced compact morulae were cultured for 3 days in a chemically defined medium + bovine serum albumin (BSA; CDM-2) plus regulators (4991 total embryos). Phenazine ethosulfate (PES), phloretin (PL), pyrroline-5-carboxylate (P5C), and sodium azide (NaN3) were evaluated at four doses each in factorial combinations with four concentrations of glucose: 0, 0.5, 2, and 8 mm. Phenazine ethosulfate at 0.9 μm resulted in poorer development than lower or no PES. Phloretin was, in general, detrimental for embryo development, but most markedly at the highest dose (270 µm). Pyrroline-5-carboxylate had little effect on post-compaction embryos at the doses studied, 9 to 81 μm. Sodium azide at the concentrations used (3, 9, and 27 μm) had little effect on embryo development compared with controls. Concentrations of glucose had little effect on development of embryos. A fifth metabolic regulator, 2,4-dinitrophenol (DNP), was studied at various doses at pre-morula or morula-blastocyst stages cultured in 2 mm glucose. Embryos (2189 total) cultured in 90 µm DNP developed more slowly and were darker than embryos cultured at lower doses. Embryos cultured in 30 µm DNP had a higher blastocyst rate (48.3%) than controls (34.9%). In the last experiment using G1.2/G2.2 media, DNP (30 μm) resulted in a marked decrease in embryonic development when embryos were exposed at the zygote to 8- to 16-cell stages but had little effect when morulae were exposed for 2 days. The dose–response information for these metabolic regulators is crucial for designing future experiments.
Acknowledgments
Numerous students and personnel in the Embryo Transfer Laboratory at Colorado State University assisted with this research, which was funded in part by the Colorado State University Experiment Station via the United States Department of Agriculture Regional Project W-1171, as well as NRI grant no. 2003-35203-13705 of the United States Department of Agriculture Cooperative State Research, Education and Extension Service. G1.2 and G2.2 media were provided by the Colorado Center for Reproductive Medicine. J. F. De La Torre-Sanchez was supported by a scholarship from CONACyT, and the Instituto Nacional de Investigaciones Forestales, Agricolas y Pecuarias, Mexico.
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