329 COMPARING CHANGES IN MOTION PARAMETERS IN EPIDIDYMAL AND EJACULATED BOAR SPERMATOZOA UNDER THREE DIFFERENT TREATMENTS

The motion ability of mammalian spermatozoa is acquired during their epididymal transit but observed only upon dilution with seminal plasma (SP) at the time of ejaculation (Yanahimachi 1994 in The Physiology of Reproduction, New York: Raven Press). The bicarbonate present in seminal plasma activates multiple sperm functions, some of which are essential for the initiation of motility. Sperm hyperactivity has been observed in vitro in various mammalian species, especially if capacitation of spermatozoa was induced with Ca2+ and bicarbonate media, such as TALP (Harrison et al. 1996 Mol. Reprod. Dev. 45, 378–391). Computer-assisted sperm analysis (CASA) is a tool for the objective assessment of sperm motility. The aim of this study was to determine if there are differences in motility parameters of ejaculated (EJ) and epididymal (EP) boar spermatozoa under different treatments. Ejaculated and epididymal sperm cells from 10 different boars in each group were used. The sperm treatments were: washed in Dulbecco's PBS supplemented with 0.1p BSA (PBS-BSA), washed on a Percoll gradient (PG), and unwashed (UW: Control); the sperm samples were incubated in TALP medium at 38.5°C and 5p CO2 during the analysis. Motion parameters were determined using a computer-assisted sperm analysis (CASA) system. A 7-µL drop of the sample was placed on a warmed (37°C) slide. At least 4 fields per sample were evaluated, with a minimum of 100 spermatozoa counted per sub-sample. The CASA-derived motility characteristics studied were motility (MOT, p), progressive motility (PM, p), curvilinear velocity (VCL, µm s−1), straight-line velocity (VSL, µm s−1), average path velocity (VAP, µm s−1), linearity of the curvilinear trajectory (LIN, ratio of VSL/VCL, p), straightness (STR, ratio of VSL/VAP, p), amplitude of lateral head displacement (ALH, µm), wobble of the curvilinear trajectory (WOB, ratio of VAP/VCL, p), and beat cross-frequency (BCF, Hz). Data were analyzed by ANOVA. If we evaluated all of the data together (EJ vs. EP), EP sperm after treatment showed a higher motility (PM: 38.20p; MOT: 74.23p) than EJ sperm (PM: 29.27p; MOT: 63.24p), and all of the motion parameters related to velocities and ALH were higher in EP (VCL: 86.02; VSL: 41; VAP: 57.94; ALH: 3.21) than in EJ (VCL: 69.70; VSL: 34.67; VAP: 48.16; ALH: 2.54). No differences were found for LIN, STR, WOB, and BCF. The treatments significantly affected the VCL and ALH, with lower values for the PG treatment. When VCL was lower and the VSL and VAP were similar, consequently the LIN and WOB were significantly higher for the PG group. STR also was higher for the PG group. In conclusion, when both groups of sperm were incubated in TALP medium, the EJ sperm showed a decrease in the majority of motion parameters when compared with EP sperm. This work was supported by MEC (AGL2006-03495/GAN) and Fundacion Seneca (03018/PI/05).