Sperm subpopulational dinamycs during the cryopreservation procedure in caprine (Capra aegagrus hircus) ejaculates

Abstract

The objective of the present research was to determine specific sperm subpopulational dynamics in different processing steps during cryopreservation process by using objective functional sperm kinematic descriptors in goat ejaculates. Fresh ejaculates (n=40) collected from eight bucks were analised for volume, concentration, sperm viability, acrosome integrity, and sperm motility using computer-assisted sperm analysis (CASA) system. Eight sperm kinematic descriptors (VCL, VSL, VAP, LIN, STR, BCF, ALH, and WOB) were assessed using CASA system after five different handling step (1st: fresh semen collection (F); 2nd: 1st washing/centrifugation step (1WC); 3rd: 2nd washing / centrifugation step (2WC); 4th: cooling step at 4ºC (CL); and 5th: post-thawing step at 37ºC (PT)) during a standard cryopreservation protocol for goat semen. The results obtained from the kinematic parameters were analysed by using Principal Component Analysis (PCA) and multivariate clustering procedures to identify specific kinematic subpopulations and establish the relationship between the distribution of the subpopulations found and the functional sperm motility in each step. Except for the 1st (SbpF1-SbpF3) and 4th (SbpCL1-SbpCL3) intervals, four sperm kinematic subpopulations (Sbp1LC1-Sbp1LC4, Sbp2LC1-Sbp2LC4 and SbpPD1-SbpPD4) were observed. Based on kinematic velocity parameters and the subpopulation disclosed, rapid, slow, vigorous, passive, non-progressive and progressive sperm were discerned. Moreover, based on kinematic linearity parameters and depending on the subpopulation uncovered, curvilinear, regular-linear, parabolic and erratic-non-linear trajectories were detected. Subpopulations remained varible throughout handling steps and multiple significant differences among the sperm kinematic parameters were observed (p<0.001) as well as in the frequency of distribution of kinematic subpopulations among steps (p<0.05). In conclusion, this study confirms the variability and heterogeneity of goat sperm kinematic patterns throughout the cryopreservation process. The relationships among sperm subpopulation structure and the different handling steps were particularlly relevant, specially after the cooling and the post-thawing steps, when effects derived from these critical handling steps were evident and altered drastically the sperm motion patterns

Keywords

• Sperm subpopulations
• cryopreservation
• semen
• caprine