Patricio Morales1 and Miguel Llanos2

1 P. Catholic University of Chile, Faculty of Biological Sciences. P. O. Box 114-D. Santiago, Chile.

2INTA, University of Chile, P. O. Box 138-11. Santiago, Chile.


In response to an oocyte-derived signal or signals, at the time of fertilization of the oocyte, the spermatozoon undergoes acrosomal exocytosis (37). The generally accepted view is that the fertilizing spermatozoon traverses the cumulus oophorus using the hyaluronidase activity present in the protein PH20, located on the plasma membrane of the posterior head (107). Then, the fertilizing spermatozoon, with its acrosome-intact, binds to the zona pellucida glycoprotein, ZP3. Following stimulation by ZP3, the spermatozoon undergoes the AR. Exposure to ZP3 leads to tyrosine phosphorylation of a 95 kDa protein (15, 28, 29) and activation of a guanosine triphosphate (GTP)-binding protein (68). Progesterone (P) trapped in or produced by the cumulus oophorus can also initiate the AR (46). P action is specific (54, 58), is mediated by a surface receptor (108), and leads to phosphorylation of a 95 kDa protein (60). G protein activation, however, does not take place (109).

9.1. Modulators of spermatozoa-zona pellucida binding

9.1.1. Progesterone

Interaction between P and the ZP has been reported in the mouse (110). During sperm activation, both agonists sequentially interact. As compared with simultaneous presence of both P and ZP or presence of agonists in the reverse order, spermatozoa treated sequentially with half maximal concentrations of P and ZP exhibited enhanced exocytosis (110). Based on these findings, it was concluded that P exerts a priming effect in the initiation of the AR. Whether a similar interaction between P and ZP takes place during human fertilization is not known. Regarding spermatozoa-ZP binding, P may (111) or may not (112) increase the total number of spermatozoa able to bind to the ZP. Recent experiments conducted in our laboratory have shown that P definitively increases the number of zona bound spermatozoa. Treatment of the spermatozoa with P (1 µg/ml) caused a significant increased (292±25%) in their capacity to bind to the ZP. This effect required presence of calcium in the culture medium (Morales and Llanos, unpublished results).

9.1.2. Fucoidin

Fucoidin is a heteropolisaccharide consisting primarily of alpha(1'2)- and ß(1'3)-linked L-fucose sulfate subunits. Treatment of the spermatozoa with fucoidin inhibits spermatozoa-ZP binding in humans (113). The specificity of the fucoidin, however, is ambiguous since incubation of ZP with fucoidin also caused inhibition of spermatozoa-zona binding (113).

9.1.3. Sialic acid

Removal of sialic acid may also play an important role in spermatozoa-zona binding since treatment of the spermatozoa with neuraminidase, an enzyme that cleaves alpha(2-->3)- or ß(2-->8)-linked sialic acid residues, enhanced attachment of spermatozoa to the human ZP (114). Removal of sialic acid from the sperm surface is associated with loss of net negative charges (115) and sperm capacitation (116).

9.1.4. D-Mannose

Previous studies have attributed a role to the sperm mannose-binding sites on the spermatozoa-ZP binding process (24-26, 117). However, Mori and colleagues observed only a 50% decrease in the number of spermatozoa bound to the ZP even at the highest mannose concentration tested (50 mmol/l) (25, 26). Zona penetration was completely inhibited (table I). A 30% inhibition of spermatozoa-ZP binding by mannose was observed by Oehninger et al. (118). Chen et al. found that treatment of the spermatozoa with mannose caused a 70% inhibition of spermatozoa-ZP binding (117). Inhibition of binding, however, was not observed during the initial phase of the spermatozoa-ZP interaction. Acrosin is one of the candidates to support binding of acrosome-reacted spermatozoa to ZP (119, 120). The action of human acrosin is inhibited by a variety of monosaccharides, especially D-fructose and D-mannose (121). Based on these observations, the data presented above can be reinterpret as follows. Mannose-treated spermatozoa can initiate binding to the human ZP followed by development of AR in the spermatozoa bound to the zona surface. Since acrosin activity is inhibited after AR, spermatozoa cannot perforate a path through the zona and such spermatozoa eventually detach from ZP.

9.1.5. Egg yolk and Milk

Egg yolk and milk were recently suggested to increase the number of spermatozoa that bind to the human ZP (122, 123). The mechanism of action and the biological significance of these observations are not yet clear.

9.1.6. Placental protein 14

Recently, it was reported that human placental protein 14, a glycoprotein synthesized and secreted by the endometrial cells, significantly reduced spermatozoa-zona binding (124). This effect was specific for this protein and not for other endometrial stromal products. Since placental protein 14 is also expressed in ectopic endometrium (125), the authors suggested that this may provide a mechanism for endometriosis-related infertility (124).

9.1.7. Gonadotropin hormone-releasing hormone (GnRH)

Some recent reports have indicated that GnRH and analogs (agonists and antagonists) may modulate spermatozoa-ZP binding in humans (126, 127). GnRH is a decapeptide of hypothalamic origin which causes the release of LH and FSH from the pituitary. GnRH or GnRH-like molecules have been detected in hFF (128) and human seminal plasma (129, 130). These molecules are probably synthesized in the gonads (131-134) and in the prostate (135).

Treatment of the spermatozoa with 20 nM GnRH for 5 min increased, by 357±14%, the number of zona bound spermatozoa in comparison to the control, saline treated spermatozoa (126). Busereline, a GnRH agonist, increased the binding by 326±32%. Treatment of the spermatozoa with the GnRH antagonist 4pF (20 nM) did not cause any significant change in the number of zona-bound spermatozoa. However, 4pF administered 5 min before the addition of GnRH completely blocked the stimulatory effect of GnRH. Treatment of the spermatozoa with GnRH does not modify the percentage of acrosome reactions or the pattern of sperm movement (126).

The above findings suggest that the spermatozoa may interact with GnRH, or GnRH-like molecules at different steps during the reproductive process: 1) during spermatogenesis by local, intratesticular production (136-139); 2) during sperm maturation in the epididymis; 3) during ejaculation, upon mixing with seminal plasma (129, 130, 135); and 4) during the ascent to the site of fertilization in the ampulla of the oviduct. In the oviduct, the spermatozoa may interact with GnRH secreted locally or transported by the products of ovulation (follicular fluid, granulose cells) from the ovary (128, 131, 133).

[Table of Contents ] [Next Section] [Previous Section]