Manipulation of bovine sperm metabolism and motility using anoxia and phosphodiesterase inhibitors

Bovine sperm that were subjected to extended anoxia (2.5 h) in the absence of glycolytic substrates then diluted into oxygenated medium were immotile but metabolically active, producing ATP from lactate via oxidative phosphorylation. In response to anoxia sperm ATP titers dropped from 15–20 μmoles/10⁸ cells to 1–2 μmoles/10⁸ cells in the first 5 min then remained extremely low until reoxygenation. Cyclic AMP titers declined slowly over the anoxic period, but did not show the same scale of depression as ATP. After dilution and re‐oxygenation ATP recovered to pre‐anoxia levels within 1 min, and cAMP rose to about the pre‐anoxia levels. However, motility, which varied quantitatively and qualitatively between ejaculates prior to anoxic treatment, was substantially depressed after extended anoxia in all cases; progressive motility was almost non‐existent in post‐anoxic sperm. Addition of isobutylmethylxanthine or Cibacron Blue F3GA, both putative phosphodiesterase inhibitors, stimulated a transient peak of cAMP, which was accompanied by motility stimulation. These techniques provide a protocol to manipulate and dissect the biochemical pathways of motility initiation in mammalian sperm.