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Viability of Stored Equine Embryos¹

Colorado State University² Ft. Collins 80523

Abstract

Equine embryos were recovered nonsurgically 6.5 d after ovulation (Exp. 1) and those >200 µm were stored in one of three media: 1) Ham's F10 + 10% fetal calf serum (FCS) under 5% C0₂, 5% 0₂ and 90% N₂ at 24 C (Ham's F10); 2) Minimal Essential Medium with Hank's balanced salts + 10% FCS in air (MEM) at 24 C or 3) MEM at 5 C n=10/treatment). Embryos 200 µm (n=10) were bisected microsurgically; one-half of each embryo was stored in Ham's F10 and the other half in either Dulbecco's phosphate-buffered saline + 10% FCS in air at 24 C (DPBS), or MEM in air at 24 C. At 0, 12 and 24 h, embryos were: 1) measured; 2) assigned a developmental score of 1 to 4 (1=tight morula, 4=expanding blastocyst) and 3) assigned a quality score of 1 to 5 (1=excellent, 5=degenerate). Whole embryos stored in MEM at 5 C or 24 C did not (P>.05) advance in development by 24 h, whereas those stored in Ham's F10 at 24 C were more (P<.05) advanced (i.e., higher developmental score) by 24 h. From 0 to 24 h, 1 of 10, 6 of 10 and 7 of 10 whole embryos developed when stored in MEM 5 C, MEM 24 C and Ham's F10 24 C, respectively. Embryo quality was better at 24 h (P<.05) for embryos stored in Ham's F10 at 24 C compared with MEM at 5 C. Quality of bisected embryos was better (P<.05) for those stored in Ham's F10 compared with MEM or DPBS. Experiment 2 evaluated the viability of embryos stored in Ham's F10 at 5, 24 or 37 C for 12 h and 37 C for an additional 12 h (n=10/treatment). Embryos cultured at 5 or 24 C did not (P<.05) develop or grow during the initial 12 h, but upon culture at 37 C they advanced (P<.05) in development. Experiment 3 was designed to determine pregnancy rates for embryos transferred nonsurgically after storage in Ham's F10 at 24 C for 1 h or 12 h. Treatments were: 1) embryos transferred 1 h into recipients that ovulated 1 d prior (+1) to 3 d after (–3) the donor mare; 2) transferred 1 h into progestin-treated recipients; 3) transferred after 12 h into naturally synchronized recipients (+1 to –3 d in relation to donor) and 4) transferred after 12 h into progestin-treated recipients. Progestin-treated recipients were administered .044 mg of altrenogest/kg body weight daily beginning the day after the donor ovulated. Thirteen of 32 synchronized recipients became pregnant (d 30) compared with 0 of 17 for progestin-treated recipients (P<.05). Pregnancy rates for embryos stored for 12 h were nearly identical (P>.05) to those transferred in 1 h. Therefore, embryo viability was maintained for 12 h by storage in Ham's F10 at 24 C.

¹ Supported in part by Van Camp Foundation and Colorado State Univ. Exp. Sta. Altrenogest was donated by Hoechst-Roussel-Agri Vet. Somerville, NJ. We thank Dr. Takeda for bisecting the embryos and assistance with culturing embryos.

² Anim. Reprod. Lab.