Seminal plasma addition attenuates the dilution effect in bovine sperm

Dilution of semen to low cell numbers/dose can result in a bull-dependent r eduction in the post-thaw viability of cryopreserved bovine spermatozoa. It is possible that essential seminal plasma components are lacking at the gr eater dilution rates, thereby contributing to the deleterious effects of se men dilution. Ejaculates of 6 Holstein bulls were diluted to 120 x 10(6) sp erm/mL in an egg yolk citrate extender (EYC). Split samples were further di luted to 80, 40, 20 and 4 x 10(6) sperm/mL in EYC extender with (+SP) and w ithout (-SP) the addition of frozen/thawed seminal plasma previously obtain ed from a vasectomized bull. Serial dilutions for the +SP treatments were c alculated and performed such that each dilution contained a volume of semin al plasma equal to the original 120 x 10(6) sperm/mL dilution. Samples were then loaded into 0.5-mL French straws yielding final sperm concentrations of 30, 20, 10, 5 and 1 x 10(6)/dose. Straws from each dilution were analyze d using 2 stain combinations: the sperm viability stain, SYBR-14 and propid ium iodide (PI); or the mitochondrial-specific, membrane potential-dependen t stain JC-1 along with PI. Split-plot analysis of variance indicated that within bulls, there were greater proportions of viable spermatozoa,oa in al iquots containing added seminal plasma than in aliquots without added semin al plasma (P < 0.05). Contrast analyses showed that sperm viability signifi cantly decreased as sperm concentration decreased in the -SP samples. Altho ugh the dilution effect was also observed in the +SP samples, the magnitude of the Effect was less than in the -SP samples. At most sperm concentratio ns, the proportions of spermatozoa that stained with JC-1 were correlated ( r > 0.84; P < 0.05) with the percentages of SYBR-14 stained spermatozoa. Fu rthermore, the proportions of JC-l-stained spermatozoa were greater in the +SP aliquots than in the -SP samples at a concentration of 10 x 10(6) sperm /0.5 mt. These results suggest that the addition of seminal plasma can be b eneficial to sperm viability when semen is diluted to low cell numbers/dose . (C) 2001 by Elsevier Science Inc.