Equine blastocyst production under different incubation temperatures and different CO2 concentrations during early cleavage

Some basic parameters for equine in vitro embryo production have not yet been established, including the optimum temperature for maturation and embryo culture, and the optimum CO2 concentration and pH during early embryo development. To explore this, we first performed cultures in incubators set at 37.2 degrees C, 37.7 degrees C or 38.2 degrees C. At these temperatures, the corresponding maturation rates were 33%, 38% and 42%; cleavage rates were 84%, 86% and 88%; and blastocyst rates were 35%, 44% and 44% per injected oocyte. These rates did not differ significantly (P > 0.2). We then evaluated three different CO2 concentrations (6%, 6.5% or 7% CO2) in 5% O-2 for culture over Days 0-5 after intracytoplasmic sperm injection, using a commercial human embryo medium with added serum, at 38.2 degrees C. The pH values of these media were 7.36, 7.33 and 7.29 respectively. In the presence of 6%, 6.5% or 7% CO2, cleavage rates were 68%, 80% and 70% respectively, and blastocyst rates per injected oocyte were 42%, 54% and 27% respectively. The blastocyst rate for the 7% CO2 treatment was significantly lower than that for the 6.5% CO2 treatment (P < 0.05). We conclude that equine in vitro embryo production is equally effective within the range of 37.2-38.2 degrees C, but that equine early cleavage stage development is sensitive to small changes in CO2 atmosphere and/or medium pH.