199 EFFECT OF HEPARIN CONCENTRATION ON BOVINE PREIMPLANTATIONAL DEVELOPMENT IN VITRO USING SEX-SORTED SPERM

Abstract

The objective of this study was to examine the effect of heparin on bovine IVF and to improve the efficiency of IVF production by using sex-sorted sperm. The fertility performance of sex-sorted and unsorted semen from 4 bulls was compared to determine the optimal heparin concentration during preimplantational embryo development. A total of 7615 matured bovine oocytes were randomly allocated among different heparin concentrations (0, 2.5, 5, 10, 20, 40, 60, 80, and 100 μg mL–1) in Brackett-Oliphant medium and coincubated with either sex-sorted or unsorted sperm for 6 h. Presumptive zygotes were cultured in CR1aa+ 6 mg mL–1 of BSA in 5% O2 , 5% CO2 and 90% N2 at 39°C until Day 8 (Day 0, culture post-IVF). Cleavage rates at Day 2 and embryo development to blastocyst (BL) at Day 8 were recorded. Data (4 replicates) were analyzed by a general linear model (SPSS 11.0, SPSS Inc., Chicago, IL). The optimal heparin concentration for each treatment was determined as the lowest value from those groups that resulted in the highest BL rates. The results (Table 1) demonstrated that a differential requirement of heparin concentration was important for the highest preimplantational BL development between sexed sperm and unsorted control within each bull. By optimizing heparin concentration, in 3 out of 4 (75%) bulls, the in vitro BL development with sex-sorted sperm could be increased to a level that was comparable to the highest BL rate from unsorted sperm (bulls A, B, and C, P > 0.05). A higher heparin concentration was required for optimal BL development in bulls A and C; however, a lower concentration was desirable for bulls B and D, indicating that a partial capacitation to the sperm may have taken place in bulls B and D during the sorting process, as reported by Lu and Seidel (2004 Theriogenology 62, 819–830). The fertility of sorted sperm from bull D (1 out of 4, 25%) was adversely affected, even after heparin optimization for BL development (P < 0.05). This result suggests that sperm sorting could affect the IVF fertility of sorted sperm in a bull-specific manner, but it was not significant for all bulls. Table 1. Blastocyst (BL) development in bovine IVF after heparin optimization using sorted and unsorted sperm This project was supported by the SBIR program under a USDA Cooperative State Research, Education, and Extension Service (CSREES) grant to F. Du (USDA #2006-03069).