Refrigerated storage of red deer epididymal spermatozoa in the epididymis, diluted and with vitamin C supplementation

We have approached the problem of refrigerated storage of epididymal sperm samples from red deer by comparing three options: storing the genital (testicles within the scrotum), diluting the semen in extender or diluting the semen in extender supplemented with an antioxidant. Twenty-nine pairs of testes, were collected. Spermatozoa from one of each of the pairs were immediately recovered, and diluted to 400106 sperm/mL in Tris-citrate-fructose with 20% egg yolk. Control group was stored as such, and Antioxidant group was supplemented with 0.8 mM vitamin C. The remaining epididymides and the diluted samples were stored at 5 ºC and spermatozoa were analyzed at 0, 24, 96 and 192 h for: motility (CASA), acrosomal integrity, sperm viability (eosine/nigrosine staining), normal tails and chromatin status (SCSA). In general, seminal quality decreased with storage time. Vitamin C better supported progressive motility at 24 h (median 42% vs. 23% Control and 15% epididymis), reduced the incidence of tail abnormalities and protected chromatin. Storing the semen in the epididymis slowed down motility loss, but slightly increased the occurrence of tail abnormalities and viability was lower at 192 h. However, regarding chromatin status, sperm stored in the epididymis was protected similarly to those diluted in the medium supplemented with vitamin C. Although the differences between the three groups were small, there were some advantages in supplementing the extender with vitamin C. Besides, refrigerating the epididymis may be a good option when immediate processing is not available