earticle

영어

Artificial insemination with frozen-thawed epdidymal sperm showed low motility and conception rate compared to those with ejaculated frozen-thawed sperm. Cauda epididymal sperm has not contained seminal plasma. We hypothesized that addition of semenial plasma will affects epididymal sperm motility. Therefore, we examined the effect of different seminal plasma concentrations on frozen-thawed epididymal sperm motility. Two testicles were recovered by castration from one Hanwoo bull (age=441 days, scrotal circumference (cm)=31, weight (kg)=420). Epididymis with vas deferens in each bull was isolated from testis and blood vessels on surface of cauda epididymis. Cauda epididymal sperm were recovered by flushing and floating methods with semen freezing medium (OptixCell, IMV, France). Sperm concentration adjusted to 40×106 cells/mL, semen dilution loaded to 0.5 mL straw and crypreserved in LN2 tank until use. Seminal plasma was collected from a Hanwoo bull. In brief, semen collected by artificial vagina and centrifuged at 5,000 ×g for 10 min. The supernatant recovered and centrifuged 5,000 ×g for 10 min for 30 min. The supernatant was cryopreserved at − 80°C until use. Six straws of frozen-semen were thawed at 37°C for 40 sec, mixed and divided into 5 groups (500 uL/group). Frozen-thawed seminal plasma added to prepared 5 frozen-thawed epididymal sperm groups with 0, 1, 2, 5 and 10% of seminal plasma. Sperm motility was evaluated at 0, 2, 4, 6 and 8 h of incubation periods using a computer assisted sperm analysis system (MicroOptic, Spain). Five replicates in each groups were conducted. Addition of 2, 5 and 10% of seminal plasma to frozen- thawed epididymal sperm improved VSL (μM/sec), VAP (μM/sec), LIN (%) and decreased BCF (Hz) compared to those of 0% of seminal plasma group at 4 to 8 h of incubation periods. In conclusion, addition of seminal plasma to frozen-thawed epididymal sperm will improve longevity of sperm in vitro. It suggested that artificial insemination with epididymal sperm and seminal plasma will improve conception rate in vivo.