Effects of calcium ions on the malate-aspartate shuttle in slow-cooled boar spermatozoa

The effects of intracellular free calcium (Ca2+) ions on the malate-aspartate shuttle were studied in slow-cooled boar spermatozoa. The capacity of the malate-aspartate shuttle was assessed by an indirect method on the basis of accumulation of lactate relative to pyruvate when ethanol is provided as substrate. The capacity of the malate-aspartate shuttle at 37 degrees C was dependent on the presence of Ca2+ ions and was stimulated by an influx of Ca2+ ions induced by the combination of the Ca2+ ionophore A23187 and 100 microM CaCl2. When washed spermatozoa were cooled slowly to 15 degrees C, the percentage of progressive motile spermatozoa was about half that at 37 degrees C in 100 microM Ca(2+)-containing medium, while the capacity of the malate-aspartate shuttle remained equal to that at 37 degrees C. The motility decreased further at higher concentrations of Ca2+ ions. Spermatozoa in EGTA-containing medium barely moved at 15 degrees C and the capacity of the malate-aspartate shuttle decreased. Even in Ca(2+)-containing medium, LaCl3 caused a decrease in the capacity of the malate-aspartate shuttle at 15 degrees C. These results suggest that an influx of a low concentration of Ca2+ ions activates the malate-aspartate shuttle at 15 degrees C, with a subsequent increase in the proportion of cells that maintain progressive motility.