Sperm binding to the human zona pellucida and calcium influx in response to GnRH and progesterone

In this study the effect of the sequential exposure of spermatozoa to progesterone and gonadotrophin-releasing hormone (GnRH) upon zona binding and the intracellular free Ca²⁺ concentration was evaluated. Sperm aliquots were treated as follows: (a) 0.7 μmol l^-1 progesterone or 0.1% DMSO (progesterone solvent) followed by 50 nmol l^-1 of GnRH; (b) 50 nmol l^-1 of GnRH or distilled water (GnRH solvent) followed by 0.7 μmol l^-1 of progesterone. Additional aliquots were incubated with DMSO or distilled water (controls) and with 0.7 μmol l^-1 of progesterone or 50 nmol l^-1 of GnRH. All treatments were for 5 min. Motile spermatozoa were incubated in modified Tyrode's medium, at 37°C, 5% CO₂, 10 × 10⁶ spermatozoa ml^-1, for 4.5 h. Intracellular Ca²⁺ concentration and sperm-zona binding was evaluated using fura 2 and the hemizona assay, respectively. GnRH and progesterone increased sperm-zona binding and the Ca²⁺ concentration. Regarding zona binding, the effect of GnRH was significantly greater when the spermatozoa had been previously treated with progesterone (progesterone → GnRH = 185 ± 116 zona-bound spermatozoa versus DMSO → GnRH = 99 ± 15, P < 0.001). On the other hand, previous treatment with GnRH did not modify their subsequent response to progesterone (GnRH → progesterone = 114 ± 19 zona-bound spermatozoa versus distilled water → progesterone = 108 ± 22, NS). The results regarding intracellular Ca²⁺ showed a similar pattern. These findings suggest a priming effect of progesterone upon a GnRH-induced increase in sperm-zona binding and intracellular Ca²⁺.