Influences of retrieval stages and glutathione addition on post-thaw viability of quick frozen mouse morula during in vitro culture

Effects of the embryo retrieval stages and addition of glutathione (GSH) on post-thaw development of mouse morula were evaluated in 2 consecutive expe riments. In the first experiment, 1-, 2-, 3- to 4- and 5-to 8-cell stage em bryos were collected and cultured to the morula stage in Whitten's medium c ontaining 0.1 mM ethylenediaminetetraacetic acid (EDTA). The development ra te of 1-cell embryos to the morula stage was lower than that of the other s tages (P<0.01). The post-thaw development rate of the morulae obtained from in vitro culture of 1-, 2-, 3- to 4-, and 5- to 8-cell embryos and from in vivo embryos (control) to the blastocyst stage was 55.5, 84.9, 87.4, 90.1 and 90.8 %, respectively. The post-thaw development rate of morula obtained from in vitro produced 1-cell embryos was significantly lower than from th e other stages or from the in vivo counterparts (P<0.0001). In Experiment 2 , the impact of GSH supplementation of the culture medium in the presence o r absence of EDTA was evaluated for embryo development to the morula stage and post-thaw survival, using in the 2 x 2 factorial design. Although EDTA supplementation increased development rates to the morulae (P<0.01) stage, GSH did not have an influence on morula development. However, the presence of either GSH or EDTA in the culture medium supported development to the bl astocyst stage (P<0.01) of in vitro produced morulae. These data demonstrat e that 1-cell embryos from a blocking-strain mouse cultured in vitro to the morula stage have a lower development rate following freezing and thawing than embryos collected at the 2-cell or later stages. Addition of EDTA or G SH, individually or in combination, to the culture medium may improve the d evelopment rate of morula to blastocyst stage following cryopreservation. ( C) 1999 by Elsevier Science Inc.