Pregnancy Outcome Using Highly Concentrated, Cooled Stored Stallion Semen and Different Dilution Protocols With Autologous Seminal Plasma Before Insemination

Abstract

Reducing seminal plasma (SP) is essential in stallion semen preservation; however, SP has an important role in sperm fertility as well as sperm protection and transportation in the female genital tract. In the present study, semen storage at high concentrations and low volumes and the effect of SP added immediately before insemination, volume, and deposition (corpus or uterine horn) of the insemination dose were evaluated. Semen processing protocols were investigated in experiment 1: dilution to a final volume of 20 mL (control); or centrifugation and dilution of the sperm pellet to a volume of 1.5 mL containing 500 × 106 sperm. Immediately before insemination, centrifuged samples were diluted with INRA96 extender containing 0%, 5%, 20%, or 80% SP to a final volume of 20 mL. In experiment 2, concentrated semen was not further processed before insemination. Pregnancy rates were lowest when adding 20% SP to the semen extender (100%, 89%, 67%, 30%, and 67% for control; 0%, 5%, 20%, and 80% SP, respectively; P < .05). Embryonic growth between days 12 and 14 was highest using 0% SP (292%, 515%, 290%, 343%, and 404% for control; 0%, 5%, 20%, and 80% SP, respectively; P < .05). Uterine fluid accumulation did not differ among groups (P > .05). The number of progressively motile sperm per dose was similar in pregnant (median, range: 347, 217–452) and nonpregnant cycles (314, 195–369; P > .05). Deep intrauterine insemination did not improve pregnancy outcome when using 500 × 106 sperm (P > .05). Stallion semen can be stored at 5°C for 24 hours highly concentrated with low volumes of extender, yielding normal pregnancy rates. Adding SP to ejaculated sperm immediately before insemination is not necessary after 24 hours of cooled storage.