FIGURE 4.
Epac signaling mobilizes calcium in a PLC-dependent manner. Human sperm (nonpermeabilized) were loaded with 2 μm Fluo-3-AM and 0.02% pluronic acid and incubated for 30 min at 37 °C. At the indicated times (arrows) 50 μm 8-pCPT-2′-O-Me-cAMP (8pCPT) or 4 μm progesterone (Pg) were added with (B) or without (A) previous incubation with 1 μm U73122 or U73343. Maximal [Ca2+]i response was calibrated with 0.1% Triton X-100 (TX-100) at the end of the incubation period. Shown are traces representative of five experiments. The increase in fluorescence is expressed as (F/F0) − 1 ((maximum fluorescence intensity/initial fluorescence) − 1) versus time in seconds. Bars represent mean ± S.E. of five experiments; data were normalized against the calcium response to 4 μm progesterone (A) or 50 μm 8 pCPT-2′-O-Me-cAMP (B). ***, p < 0.001; *, p < 0.05 compared with 8 pCPT-2′-O-Me-cAMP.