Heparin had a more ubiquitous effect and severely decreased binding of all biotinylated carbohydrates tested (P < 0.001, n = 3; see Table 2 and Fig. 8C). Higher levels of glucose in the T+BIC+Hep buffer did not significantly reduce binding of biotinylated carbohydrates (Table 2; P > 0.05). However, the oligosaccharide binding of mannan-BSA-biotin remained unaltered after T+BIC treatment (P > 0.05, n = 3) and only partly decreased after T+BIC+Hep and T+BIC+Hep+Glu capacitation incubations (P < 0.005, n = 3) when compared with T—BIC treatment (Table 2).
Competition of SPM-Carbohydrate Binding by Follicular Fluid and Corresponding Isolated Follicular Glycosaminoglycans
The presence and localization of sulfated GAG in the oviduct was detected with toluidine staining, which, in the presence of GAG, changes its color from blue to pink. GAGs were highly abundant in both the isthmus (Fig. 9, A and B) and the ampulla (Fig. 9, C and E), and localized on the luminal surface of the oviductal mucosa. GAGs were equally distributed over the apical plasma membranes of the secretory and ciliated oviductal epithelial cells, in the form of a thin layer.
FIG. 9. Light microscopic micrograph of toluidine blue staining of cow oviducts. Pink-stained glycosaminoglycans (GAG) can be found on the apical surface of the epithelium equally distributed over the whole luminal side (arrows) of the isthmus (A, B, C) and the ampulla (D, E). Within the ampulla (E) GAG were detected within the apical protrusion of the secretory cells (circle) and so indicating their origin.
TABLE 2. Effects of different capacitation media on glycoconjugate binding to the bovine sperm after 4 h of incubation.*
* The effect of bicarbonate, heparin, or glucose addition to the Tyrode medium is given as mean of percentage fluorescence detected compared with the control medium ± SD (n = 3).
Significant decreases in binding when compared with T-BIC are indicated with ** (P < 0.05), *** (P < 0.01), and **** (P < 0.001).