DEVELOPMENT IN-VIVO OF RABBIT MORULAE AFTER FREEZING USING A 2-STEP COOLING METHOD

Development in vivo of rabbit compacted morulae after freezing using a two-step cooling method and then thawing was evaluated. Embryos were exposed during 2.5 min at room temperature to the freezing medium comp osed by dimethylsulphoxide (DMSO, 3.5 mol/l) and sucrose (0.25 mol/l). The embryos, placed into plastic straws, were maintained at -27 degre es C for 30-45 min and were then plunged into liquid nitrogen. After r apid thawing, frozen-thawed embryos were transferred into both oviduct s of recipient does (single transfer) or into one of the oviducts when fresh embryos (control) were transferred into the opposite one (combi ned transfer). Embryo viability was evaluated by the ability of the em bryo to develop to sites of implantation on day 17 of gestation, for a ll embryos, and to fullterm fetuses on day 28 of gestation or young ra bbits at birth, for combined or single transfer, respectively. A total of 98 (90.7%) frozen-thawed embryos were considered morphologically n ormal after applying the procedure of cryopreservation and 80 of these , together with 28 control embryos, were transferred to recipient does . The survival rate at day 17 of gestation for the frozen-thawed embry os was 29% (23/80), or 43% (23/54) for the embryos developed in the pr egnant does. The survival rate at term (full-term fetuses or young rab bits) for the frozen-thawed embryos was 25% (20/80), or 37% (20/54) fo r the embryos developed in the pregnant does. Overall, survival rates of the frozen-thawed embryos were significantly lower than those of th e control embryos. To summarize, rabbit embryos frozen using a two-ste p cooling method can continue their development in vivo, but they have about half the chance of survival compared to the fresh embryos.