Mitochondrial function and ram sperm fertility.

These experiments investigated the effect of freezing on mitochondrial function in ram sperm, the effectiveness of current freezing procedures in protecting mitochondria, and the role of mitochondrial respiration in cervical penetration and transit by ram sperm. Only sperm with functioning mitochondria (assessed by rhodamine 123 staining) after freezing and thawing were motile in a viscous medium (P < 0.05). A simplified rhodamine 123 uptake assay was developed to monitor sperm mitochondrial function. The results of this procedure were highly correlated (r2 = 0.98) with the proportion of damaged sperm in the semen sample. A semen freezing procedure commonly used by industry was compared with newer methods, and with freezing without cryoprotectants. None of the freezing protocols produced sperm with higher post-thaw levels of mitochondrial integrity than unprotected sperm. Merino ewes were inseminated with semen treated with metabolic inhibitors. Glycolytic inhibition did not affect fertility. Mitochondrial inhibition reduced fertility in cervically (P < 0.05), but not laparoscopically inseminated ewes. It is concluded that mitochondrial respiration plays an important part in penetration of the cervix by ram sperm. Mitochondrial injury during freezing is likely to be implicated in the poor fertility of frozen ram semen used for cervical insemination.