The effects of different temperatures and incubation times after thawing on progressive motility, kinematic parameters and mitochondrial membrane potential in frozen bull spermatozoa

The aim of the present study was to investigate the effect of different incubation times and thermal conditions on sperm high mitochondrial membrane potential, sperm motility and kinetic parameters after thawing in frozen bull semen. In the study, frozen semen samples, in straws of 0,25 ml, from four different bulls were used. Straws were thawed for 25 seconds in water baths at 38°C. After thawing, the sperm motility and kinetic parameters determined using a CASA system and the sperm mitochondrial membrane potential examined using JC-1 by flow cytometry. Total and progressive motility, sperm kinetic parameters and high mitochondrial membrane potential were investigated in different thermal conditions (35°C, 22°C and 5°C) and incubation times (immediately, 10 and 20 minutes) after thawing. Significant individual differences (p<0.05) were observed between bulls in terms of total and progressive motility after thawing. No significant difference (p˃0.05) was observed with respect to all criteria between 35°C and 22°C at different incubation times. On the other hand, the sperm total and progressive motility, the sperm kinematics as VCL, VAP and VSL significantly reduced (p<0.05) at 10 and 20 minutes after thawing in 5°C. Similarly, the proportion of spermatozoa with high mitochondrial membrane potential also decreased (p<0.05) significantly at 5°C during the same time periods compared to other temperatures. There was a high correlation among progressive motility, VCL, VAP, VSL and high mitochondrial membrane potential (p<0.01). In conclusion, sperm motility, sperm kinetic parameters and high mitochondrial membrane potential negatively affected during the incubation period of 20 minutes after thawing in a low-temperature environment such as 5°C.