Effects of egg yolk-citrate and milk extenders on chromatin structure and viability of cryopreserved bull sperm.

Semen from four Holstein bulls was evaluated to compare effects of four extender treatments on postthaw semen quality. Extender fractions A and B, either heated whole milk or 20% egg yolk-citrate, were combined to yield the extender treatments 1) milk and milk, 2) milk and egg yolk-citrate, 3) egg yolk-citrate and milk, and 4) egg yolk-citrate and egg yolk-citrate. Semen was evaluated at thawing and after 30, 60, 120, and 180 min of incubation at 38.5 degrees C. Flow cytometry showed that acridine orange-stained sperm were most susceptible to in situ DNA denaturation when fraction A was milk. For sperm stained with rhodamine 123, flow cytometry showed that the proportion with intact mitochondrial membrane potential was lowest of all treatments at thawing but greatest at 180-min incubation with milk and milk extender. Flow cytometry of propidium iodine-stained sperm showed greatest proportion of cell membrane intact sperm when fraction A was egg yolk-citrate. Light microscopy showed the lowest proportion of cell membrane intact sperm with milk and milk extender after eosin-aniline blue vital staining. Postthaw motility scores tended to be reduced when both extender fractions were egg yolk-citrate. Results demonstrate differential extender effects on postthaw semen quality and indicate that altering extender composition or sequence of adding extender components may improve postthaw quality of cryopreserved sperm.