Effects of Some Respiratory and Glycolytic Inhibitors on MitochondrialFunctionality in Bovine Semen

摘要

Fertility potential of spermatozoa depends on maintenance of the mitochondrial membrane potential (ΔΨm)that provides energy for sperm hyperactivation immediately prior to successful fertilization. Mitochondrial structure andintegrity are associated with sperm motility and reduced fertility, and measurement of ΔΨm may be suitable fordetermining bull semen quality. Mitochondrial membrane potential in four commercial AI bulls was assessed using JC-1and propidium iodide in the presence of the glycolytic inhibitors 2-deoxy-D-glucose (DOG) and iodoacetamide (IAM)and the respiratory inhibitor valinomycin (VAL) to determine the maximum ΔΨm at minimum incubation. Flowcytometry recorded ΔΨm within a unified population for all treatments that represented sperm with low and high ΔΨmrespectively. Maximum ΔΨm was seen at 40 min incubation. Mean high fluorescence intensity (MFI) (orange) wassignificantly greater for untreated compared to the treated sperm, at 40 and 80 min incubation, in both fresh and frozenthawedsemen. In sperm treated with VAL and IAM, ΔΨm was lowered significantly, and the proportion of sperm withhigh: low ΔΨm ratio was higher in control and DOG- treated samples representing more active mitochondria. In samplestreated with VAL and IAM, the ratio was reversed, representing loss in activity. Cryopreservation increased the high:lowΔΨm ratio only in bull 1 by 30% and lowered it in bulls 2, 3 and 4 by 20% - 70% compared to fresh semen. The rise inbull 1 may relate to be the product of sperm demonstrating a capacitation- like effect of the freeze-thaw process whichstimulated sperm hyperactivity.We conclude that mitochondrial function was affected adversely on freeze-thaw process. The 40 min incubation issatisfactory for future studies. Furthermore, IAM, a known glycolytic inhibitor, has a similar effect to VAL, a recognizedrespiratory inhibitor, which should provide a basis for further research.