Phospholipase A2 (PLA2) activity has been shown to be involved in the sperm acrosome reaction (AR), but the molecular identity of PL A2 isoforms has remained elusive. Here, we have tested the role of two intracellular (iPL A2 β and cytosolic PL A2 α) and one secreted (group X) PLA2s in spontaneous and progesterone (P4)-induced AR by using a set of specific inhibitors and knock-out mice. iPL A2β is critical for spontaneous AR, whereas both iPL A2 β and groupXsecreted PL A2 are involved in P4-induced AR. Cytosolic PL A2 α is dispensable in both types of AR. P4-induced AR spreads over 30 min in the mouse, and kinetic analyses suggest the presence of different sperm subpopulations, using distinctPL A2 pathways to achieve AR. At low P4 concentration (2M), sperm undergoing early AR (0-5 min post-P4) rely on iPL A2 β , whereas sperm undergoing late AR (20-30 min post-P4) rely on group X secreted PL A2. Moreover, the role of PL A2 s in AR depends on P4 concentration, with the PLA2s being key actors at low physiological P4 concentrations (<2 μM) but not at higher P4 concentrations (10 μM).