Submitted to: The International Embryo Technology Society
Publication Type: Abstract Only
Publication Acceptance Date: 10/10/2000
Publication Date: N/A
Technical Abstract: Sperm sexing by flow cytometry requires expensive equipment, and utilization of sexed sperm for conventional AI is limited by the number of sperm that can be sorted, 5-15 million per h for 90% purity. In vitro fertilization (IVF) promises the most efficient means to use sexed sperm for in vitro embryo production. However, sexed sperm has yielded lower cleavage and blastocyst formation rates compared to unsorted frozen-thawed sperm. In order to improve the rate of embryo development sorted sperm (X and Y) was assigned to 3 treatments: 1) control (no treatment); 2) heparin (10 mg/ml); and 3) dbcAMP (1mM) during IVF. Cumulus-oocyte complexes (COC) were matured in TCM 199 supplemented with pyruvate (50 mg/mL), cysteamine (0.1 mM), cystine (0.1 mM), glutamine (0.25 mM), HEPES (10 mM), PVA (0.1%), NaHCO3 (26 mM), recombinant hFSH (0.1 IU/mL), and recombinant hIGF-I (5 ng/mL). Sperm and COC were co-incubated in mDM for 18 h. Then presumptive zygotes were cultured 54 h in g-SOF+NEA prepared without glucose, with threonine (0.4 mM) and reduced PVA (3 mg/mL). From 72 to 144 h post-insemination (pi) embryos were cultured in c-SOF+NEA. From 144 to 192 h pi cultures were in the maturation medium without hormones. Data were analyzed by Chi-square. No significant effects were found among treatments for Y-sperm when compared to controls. A significant (P< 0.001) improvement in the cleavage rate was found for X-sperm treated with cAMP (51/72; 73%) compared to untreated control (32/58; 55%). After culture, blastocysts were obtained. Treatment with dbcAMP enabled a better blastocyst formation rate (8/26; 31%) compared to control (3/31; 10%) and heparin (2/34; 6%). These data suggest dbcAMP to be an effective treatment of sorted sperm for production of female bovine embryos in vitro.