Evidence against a functional ATP-dependent calcium extrusion mechanism in bovine epididymal sperm

Bovine epididymal sperm resuspended in ionic buffers take up relatively large amounts of calcium. This uptake, which is almost entirely mitochondrial, apparently bypasses the sperm cytosol. The direct mitochondrial loading is an unusual aspect of sperm calcium uptake, which suggests that the plasma membrane region surrounding the mitochondria should be highly permeable to calcium, whereas the membrane domains surrounding the head and tail regions of sperm should be impermeable. This study was undertaken to determine the role of a plasma membrane calcium ATPase in sperm calcium homeostasis. Kinetics of calcium (45Ca2+) uptake into intact and permeabilized caudal epididymal sperm confirmed that mitochondrial calcium uptake occurs with virtually no resistance from the surrounding plasma membrane. Cytoplasmic calcium accumulation by sperm depleted of intracellular ATP, measured in the presence of mitochondrial calcium uptake inhibitors, showed no increase upon energy depletion as would be expected if an ATP-dependent calcium extrusion mechanism were present. Furthermore, lowering the incubation temperature to further reduce the activity of the calcium ATPase in these energy-depleted sperm was also without effect on calcium accumulation. The calcium ATPase inhibitor vanadate, even at high concentrations, failed to increase intracellular 45Ca2+ accumulation. However, vanadate was effective in inhibiting motility showing that the compound was accumulated into sperm to inhibit flagellar dyenin ATPase. Therefore, the lack of effect of vanadate on 45Ca2+ accumulation was not due to its inability to enter sperm. Other calcium ATPase inhibitors such as quercetin, thapsigargin, and cyclopiazonic acid, which readily demonstrate ATP-dependent calcium extrusion in other somatic cells, were also without effect on sperm calcium accumulation.(ABSTRACT TRUNCATED AT 250 WORDS)