Variations of Canine Sperm Subpopulations Related with the Age in Fresh Ejaculates

Introduction - In the last years many studies have been made to obtain more information about the fertility capacity of the sperm (8). Strong evidences suggest that many different sperm populations coexist within the mammalian ejaculate and some authors such as Abaigaret al. (1,2) and Harrison (3) have suggested that capacitation and fertility capacity would change depending on the subpopulations. These subpopulations have been identified in some species, including the canine specie, but do not exist a lot of researches of the sperm subpopulation in dogs (5) and the number of subpopulations in dogs do not match with the number of subpopulations observed in other mammalians in some studies (4,6,7). Therefore, it is very important to determinate these sperm kinematic parameters in canines and, in our knowledge, no data exist regarding the existence of the relationship of these subpopulations with the age in canines, being these the aims of this study.

Materials and Methods - Twenty eight privately owned dogs of different breeds with ages ranging from 1 to 7 years old (y.o) were used and they were grouped in three different periods of age: 1 to ≤ 2 y.o; >2 to < 5 y.o; 5-7 y.o . The motility analysis was performed using a computed-assisted semen motility analysis (CASA, commercial system Sperm Class Analyzer; Integrated Sperm Analysis System V1.0, Proiser®, Valencia, Spain). Images were taken from 5 μl aliquots placed on slides covered with 20 × 20 mm coverslides at a magnification of 200× in a negative phase-contrast field and each image had moreover 50 spermatozoa. All procedure was performed at 37oC. A FASTCLUS procedure was applied to determinate the kinematic subpopulation structure of canine ejaculate. The same FASTCLUS was applied to determinate the changes in sperm subpopulations regarding to the age, but adding these parameters to the equation. After this, PROC GLM and LSMEANS procedures were applied to determinate if there were significant differences among the age periods and where these differences were, respectively.

Results - There were four statistically different (P < 0.05) sperm subpopulations in the general canine ejaculates. The spermatozoa of the subpopulation 1 showed the lowest curvilinear velocity (VCL: 47.4±0.8μm/s) and linearity (LIN: 43.1±0.5%). The spermatozoa of the subpopulation 2 showed a VCL of 131±0.8μm/s and a LIN of 47.2±0.5%. Subpopulation 3 showed a VCL of 176.6±1.0μm/s and a LIN of 66.6±0.7%. The subpopulation 4 showed the highest curvilinear velocity and linearity (VCL: 253.2±2.2μm/s; LIN: 77±1.5%). Regarding to the age, all four subpopulations showed a significant (P < 0.05) increase in VCL values and a significant (P < 0.05) decrease of the LIN values as the dogs became older.