In the ampulla, they were also found within apical protrusions of the secretory cells, indicating their secretory origin (Fig. 9). The secretory activity of the ampulla is maximal around ovulation and probably increases the amount of GAG in the isthmus, where it may facilitate sperm release. A concentration of 403.5 |xg/ml of sulfated GAG was found in the fluid of dominant follicles. Increasing concentrations of isolated GAG from follicular fluids as well as complete corresponding follicular fluid were highly potent in inhibiting fucose-PAA biotin binding to SPM (Fig. 10).
This inhibitory effect was most predominant with whole follicular fluid, where inhibition was already visible at concentrations lower than 0.01 |xg GAG/ ml and reached its maximum at about 0.3 |xg GAG/ml. The inhibitory effect of the isolated GAG fraction from native glycosylated components in follicular fluid was visible at concentrations from 1 GAG/ml with EC50 values of 47.2 ± 6.7 ^g/ml, which was about a 50 times higher level than the EC50 values for the native follicular fluid GAG (0.25 ± 0.04 ^g/ml; Fig. 10).
FIG. 10. Inhibition of fucose-PAA-biotin binding to the SPM from uncapacitated sperm cells by complete follicular fluid or by GAG isolated from follicular fluid. SPM (0.5 |xg/well) were incubated with 0.25 |xg/ml of fucose-PAA-biotin in the presence of increasing concentrations of either follicular fluid (FF) or GAG (GAG). The data indicated in the figure are corrected for nonspecific binding of biotinylated carbohydrate to noncoated wells. Results shown are representative of three independent experiments, each done in triplicate. Mean values ± SD (n = 3) are indicated.