Developmental Potential of In Vitro Fertilization-Derived Mouse Zygotes Following Vitrification: Effects of Superovulation Method

Cryopreservation of pronuclear stage embryos from superovulated mice is beneficial to safeguard genetically modified (GM) mouse strains as well as the efficient production of novel GM mouse strains. C57BL/6J female mice were superovulated with either anti-inhibin serum (AIS) or equine chorionic gonadotropin (eCG), and the resulting oocytes were inseminated via in vitro fertilization (IVF) to produce pronuclear stage embryos. A subset of fresh embryos was cultured in vitro to assess their developmental potential to the blastocyst stage, and the remaining embryos derived from either AIS or eCG superovulation methods were cryopreserved via vitrification and subsequently assessed to determine both in vitro and in vivo developmental competence. The percentage of IVF-derived fresh embryos that developed to 2-cell (92.9 ± 4.1 compared with 92.4 ± 4.2) and the blastocyst stage (91.9 ± 3.84 compared with 91.8 ± 7.9) from eCG and AIS, respectively, was not different ( P = 0.89). The percentage of the vitrified pronuclear embryos that were intact after warming for eCG (93.08 ± 5.96) and AIS (88.0 ± 6.63) mice was different ( P = 0.039). However, the percentage of IVF-derived vitrified warmed zygotes that developed to 2-cell (82.23 ± 7.18 compared with 83.9 ± 7.22) and the blastocyst stage (71.35 ± 7.76 compared with 74.52 ± 5.57) from eCG and AIS, respectively, was not different. Vitrified pronuclear embryos produced after either AIS or eCG-administered mice were in vitro cultured to 2-cell and surgically transferred into CD-1 surrogate mothers to compare pregnancy rates and live birth rates. There were no differences in percent pregnancy rates between eCG (85.7) and AIS (85.7) superovulation methods. Similarly, there were no differences in the percentage of live offspring between eCG (27.6) and AIS (23.2) superovulation methods. This study suggests that eCG and AIS superovulation methods yield similar in vitro and in vivo embryonic development rates following vitrification, and thus, AIS may be preferred, especially for the strains with difficulty in obtaining large quantities of oocytes or embryos for the production of GM mice or genome banking.