The effects of free radicals and hypotaurine on the development of bovine embryos produced by in vitro fertilization of in vitro-matured oocytes were examined. Embryos that developed to the 4- to 6-cell stage after in vitro fertilization were cultured without feeder cells in TCM199 medium supplemented with 1% calf serum (CS) under either 5% CO2 in air or 5% O2, 5% CO2 and 90% N2. The percentages of blastocysts (including early, expanding, and hatched stages) that developed under 5% O2 was higher (P < .01) than the percentage of those that developed under 20% O2. The respective percentages in 5% vs 20% O2 concentration were as follows: blastocysts (d 8), 49% vs 17%; expanded blastocysts (d 8), 19% vs 6%; hatched blastocysts (d 10), 16% vs 0%. The development of embryos to blastocysts was suppressed (P < .05) when oxygen radicals were generated in culture medium by 2,2'-azobis(2-amidinopropane) dihydrochloride (AAPH) under both 5% and 20% O2. The addition of hypotaurine permitted the development of embryos to blastocyst stage in medium with AAPH only under 5% O2 (P < .05), but superoxide dismutase (SOD) did not permit the development of embryos to blastocysts. Alternatively, when embryos were cultured in medium without AAPH, the rates of development into blastocysts under 20% O2 increased (P < .05) by the addition of hypotaurine and SOD. However, under 5% O2, the rates of blastocyst formation were not improved by addition of hypotaurine and SOD. Moreover, the cell numbers of blastocysts cultured in medium containing hypotaurine were greater (P < .01) than those of blastocysts cultured in medium without hypotaurine. It is concluded that hypotaurine may exert beneficial effects on in vitro development of bovine embryos under both 20% O2 and 5% O2 of gaseous conditions.

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lized in vitro by epididymal sperm of JCL:ICR strain were cultured for 72hr, transferred to the uteri of pseudopregnant recipients and the reproductive capacity of the young born from the recipients was examined by full-sib mating. Fertilization rate was 80.9%(418/517) as judged from the second polar body extrusion at 6hr after insemination. The developmental rate of these fertilized eggs to the morula and early blastocyst was 97% (393/405) after 72hr in culture under 5%CO2 in air. Transferred to the uteri of Day 3 or Day 4 pseudopregnant recipients were 245 of these embryos, resulting 29 male and 23 female normal newborn young. Of these young, 7 females were randomly selected and mated polygamously within the same litter at 2 to 3 months of age. All females became pregnant and delivered 37 males and 41 females. No gross anomalies were foundThe fertilization process has been accomplished in vitro in many species of mammals1). Criteria for normal fertilization, however, have not in most cases included the production of viable young following transfer of the embryos to suitable recipient females. We may conclude at this point that the development of eggs fertilized in vitro into viable young when embryos were transferred to pseudopregnant animals was comfirmed only in rabbits2-7), rats8), mice9-14) and most recently human15)None of the reports, however, have so far clarified the reproductive potentiality of the young derived from eggs fertilized in vitro.The purpose of this paper is to investigate the reproductive capacity of female and male mice developed from eggs fertilized in vitro. Materials and Methods 1)In vitro fertilization and culture of fertilized eggs ated by injection of 5 IU PMSG (Peamex, Sankyo Zoki), followed 48hr later by 5 IU hCG (Puberogen, Sankyo Zoki). Methods and medium for fertilization and subsequent culture in vitro used were as described by KASAI et al.16). The females were killed 15-16hr after the injection of hCG and their oviducts were removed and put into the Jap. J. Zootech. Sci., 50 (12): 885-890 885 1979

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Summary.The two-cell hamster embryos were transferred into the oviduct of pseudopregnant foster mothers through the infundibulum with a fine glass capillary pipette.Modified Krebs-Ringer's solution (280+Hepes) and tissue culture medium (Japan.