Testicular sperm in mammals are morphologically differentiated but require a period of residence in the female tract to become fertilisation-competent, including the ability to undergo the acrosome reaction (a Ca²⁺-mediated secretory event that can be induced by progesterone from the cumulus). The physiological changes that occur to sperm during this period are collectively referred to as capacitation. This process can be induced in vitro in ‘capacitating’ media containing bicarbonate and BSA (cholesterol acceptor) and has been shown to include increased protein tyrosine phosphorylation. We investigated the effects of two in vitro media and capacitation duration on the progesterone-induced [Ca²⁺]_i increase in populations of human spermatozoa.

This work was carried out, according to local ethical guidelines, with the Birmingham Women’s Hospital (HFEA no. 0119). Donors gave informed consent. Spermatozoa were harvested by direct swim-up into capacitating medium [sEBSS containing CaCl₂ (1.8 mM), KCl (5.37 mM), NaCO₃ (26.20 mM), NaCl (116 mM), NaH₂PO₄ (1.02 mM), MgSO₄ (0.8 mM), sodium lactate (41.75 mM), sodium pyruvate (2.73 mM), glucose (5.55 mM) and 0.3 % BSA] or non-capacitating medium [CaCl₂ (2 mM), KCl (5 mM), NaCl (150 mM), MgCl₂ (1 mM), glucose (10 mM) and Hepes (10 mM)]. Cells were then labelled with Fura-2 and progesterone-induced [Ca²⁺]_i increases in the sperm were measured by fluorimetry.

After incubation in capacitating medium for 6 h (required for the cells to undergo progesterone-induced acrosome reaction), the peak amplitude of the P₄-induced [Ca²⁺]_i response (303.7 ± 45.2;n = 6) was significantly greater than that of cells incubated for an equivalent period in non-capacitating medium (89.3 ± 11.5; n = 10; P < 0.01; unpaired t). However, comparison of the [Ca²⁺]_i response to progesterone measured immediately after swim up into sEBSS with that obtained after 6 h incubation showed no significant difference (P > 0.05; unpaired t). When sperm incubated for 6 h in the non-capacitating medium were then re-suspended for 17 min in the capacitating medium, the peak amplitude (289.1 ± 25.2; n = 7) of the P₄-induced [Ca²⁺]_i increase was also significantly higher than that of cells in non-capacitating medium (P < 0.01; unpaired t). Conversely, if sperm incubated in the sEBSS for 6 h were transferred to non-capacitating medium for 17 min, the peak amplitude of the P₄-induced [Ca²⁺]_i increase (121.2 ± 13.1; n = 10) was significantly lower than that of the cells in sEBSS (P < 0.01; unpaired t).

We conclude that the cells can acquire or lose their competence to respond to progesterone with a [Ca²⁺]_i signal within 17 min of suspension in appropriate medium but that acquisition of competence to undergo AR requires other changes which require longer to develop.