Effect of cryopreservation protocol on postthaw characteristics of stallion sperm

Three ejaculates from each of eight stallions were subjected to cryopreservation in a milk/egg yolk-based freezing extender or an egg yolk-based freezing extender. Semen was exposed to a fast prefreeze cooling rate (FAST; semen immediately subjected to cryopreservation) or a slow prefreeze cooling rate (SLOW; semen pre-cooled at a controlled rate for 80 min prior to cryopreservation). Postthaw semen was diluted in initial freezing medium (FM) or INRA 96 (IMV Technologies, L'Aigle, France) prior to analysis of 10 experimental end points: total motility (MOT; %), progressive motility (PMOT; %), curvilinear velocity (VCL; μm/s), linearity (LIN; %), intact acrosomal and plasma membranes (AIMI; %), intact acrosomal membranes (AI; %), intact plasma membranes (MI; %), and DNA quality. Eight of 10 experimental endpoints (MOT, PMOT, average-path velocity [VAP], mean straight-line velocity [VSL], LIN AIMI, AI, and MI) were affected by extender type, with egg yolk-based extender yielding higher values than milk/egg yolk-based extender (P < 0.05). Exposure of extended semen to a slow prefreeze cooling period resulted in increased values for six of eight endpoints (MOT, PMOT, VCL, AIMI, AI, and MI), as compared with a fast prefreeze cooling period (P < 0.05). As a postthaw diluent, INRA 96 yielded higher mean values than FM for MOT, PMOT, VCL, average-path velocity, and mean straight-line velocity (P < 0.05). Treatment group FM yielded slightly higher values than INRA 96 for LIN and MI (P < 0.05). In conclusion, a slow prefreeze cooling rate was superior to a fast prefreeze cooling rate, regardless of freezing extender used, and INRA 96 served as a satisfactory postthaw diluent prior to semen analysis.