Stage-dependent development of rat 1-cell embryos in a chemically defined medium after fertilization in vivo and in vitro

The present study was conducted to examine the development of rat 1-cell embryos cultured in a chemically defined medium (mR1ECM) soon after penetration of eggs in vivo or in vitro. When eggs were recovered from naturally mated females at 0600-0900 h on the following day, no penetrated eggs with male pronuclei were observed. However, the percentage of pronuclear eggs had increased significantly at 1:000-1100 h (17%) and 1200-1300 h (65%). When penetrated eggs recovered at 0600-0900 h were cultured in mR1ECM, blastocyst formation (22-46%) was significantly less frequent than in those (79-93%) recovered at 1000-1300 h. After collection at 0600-0800 h, preculture of penetrated eggs in modified Krebs-Ringer bicarbonate solution (mKRB) for up to 1200-1300 h improved their development to the blastocyst stage from 42% to 78%. Pronuclei were formed in almost all (98-100%) of the penetrated eggs examined after 6-20 h of insemination in mKRB. When penetrated eggs were transferred from mKRB into mR1ECM after 4-30 h of insemination, the percentages (47-64%) of blastocyst formation were significantly higher than those (1-20%) for eggs transferred after 1-3 h or after 40 h of insemination. When a total of 70 morulae or early blastocysts that had been produced by in vitro fertilization and developed in vitro were transferred to 7 pseudopregnant rats, 5 recipients into which 48 embryos had been transferred maintained their pregnancies and 25% of the embryos developed into late fetuses or pups. The results of the present study indicate that rat 1-cell embryos recovered from oviducts before pronuclear formation, or produced by in vitro fertilization, can develop to the blastocyst stage in vitro and that one or more factors in mKRB are necessary to maintain their development in mR1ECM.