Abstract
Subsequent to binding to the zona pellucida, mammalian sperm undergo a regulated sequence of events that ultimately lead to acrosomal exocytosis. Like most regulated exocytotic processes, a rise in intracellular calcium is sufficient to trigger this event although the precise mechanism of how this is achieved is still unclear. Numerous studies on mouse sperm have indicated that a voltage‐operated Ca^2+^ channel plays some immediate role following sperm binding to the zona pellucida glycoprotein ZP3. However, there is also evidence that the mammalian sperm acrosome contains a high density of IP~3~ receptors, suggesting that the exocytotic event involves the release of Ca^2+^ from the acrosome. The release of Ca^2+^ from the acrosome may directly trigger exocytosis or may activate store‐operated Ca^2+^ channels in the plasma membrane. To test the hypothesis that the acrosome is an intracellular store we loaded mammalian sperm with the membrane permeant forms of three Ca^2+^‐sensitive fluorescent indicator dyes: fura‐2, indo‐1, and Calcium Green‐5N. Fluorescence microscopy revealed that the sperm were labeled in all intracellular compartments. When fura‐2 labeled sperm were treated with 150 μM MnCl~2~ to quench all fluorescence in the cytosol, or when the sperm were labeled with the low affinity dye Calcium Green‐5N, there was a large Ca^2+^ signal in the acrosome. Consistent with the acrosome serving as an intracellular Ca^2+^ reservoir, the addition of 20 μM thapsigargin, a potent inhibitor of the smooth endoplasmic reticular Ca^2+^‐ATPase (SERCA), to populations of capacitated sperm resulted in nearly 100% acrosomal exocytosis within 60 min (τ~1/2~ ∼ 10 min), in the absence of extracellular Ca^2+^. Additionally, treatment of sperm with 100 μM thimerosal, an IP~3~ receptor agonist, also resulted in acrosomal exocytosis. Taken together, these data suggest that the mouse sperm acrosome is a Ca^2+^ store that regulates its own exocytosis through an IP~3~ Ca^2+^ mobilization pathway. © 2004 Wiley‐Liss, Inc.