Less is more: Reduced nutrient concentration during in vitro culture improves embryo production rates and morphophysiology of bovine embryos

Abstract

Reproducing the environment to which the embryo is naturally exposed may be an alternative to improve viability of embryos produced in vitro. In the first part of this work, we describe a novel culture media, namely Embryonic Culture Supplementation (ECS100). The composition of this media was based on the contents of carbohydrates and amino acids found in oviductal and uterine fluids. Because it was a new formulation, we investigated the performance of ECS100 in comparison with conventionally used SOFaa, and possible benefits to embryo development. Embryo production rates (cleavage, morula and blastocyst conversion, blastocyst and hatching rates) and morphophysiological parameters (total cell number, cell allocation, Mitochondrial membrane potential (MMP), Reactive Oxygen Species (ROS), NADH, FAD+ and ATP content) were similar between ECS100 and SOFaa. Next, we tested if a reduction of ECS100 concentration could positively contribute to embryo viability by resembling the more dynamic availability of nutrients that reach the embryos in vivo. Therefore, embryos were cultured in ECS100 or in its serial dilution (ECS75, 50 and 25). Despite the fact that the lowest concentration (ECS25) still supported blastocyst formation, halving the concentration of metabolites (ECS50) actually improved embryo production rates. Thus, embryos produced in ECS100 or ECS50 were submitted to further analyses on Days 4 and 7. Embryos cultured in ECS50 presented better developmental rates and morphophysiological profile than embryos cultured in ECS100. Additionally, physiological traits (MMP, ROS and NADH levels) of embryos cultured in ECS50 presented the expected pattern for embryos produced in vivo. In conclusion, we presented a novel, more personalized and effective culture media for bovine IVP embryos. And although the ECS media formulation was based on the contents of female reproductive fluids, it is worth mentioning that adaptations must be specifically directed for in vitro conditions rather than reproduced exactly from in vivo state.