Reproduction Abstracts

Ca²⁺ signalling is critical for regulation of sperm motility. [Ca²⁺]_i oscillations, which may underlie observed ‘switching’ of sperm behaviors, occur in human spermatozoa stimulated with progesterone. Our work aimed to investigate the potential contribution of changes in membrane potential, leading to cyclical activation of voltage dependent Ca²⁺-influx pathways, to [Ca²⁺]_i oscillations. Spermatozoa were incubated with fluo4-AM and clamping of membrane potential to E_K was performed using K+ ionophore valinomycin (VLN–1 μM) before or during progesterone (P4–3 μM) treatment. The cell population exhibiting [Ca²⁺]_i oscillations after P4 stimulation was 29.8 and 25.4% (4 and 7 h capacitation, respectively). Interestingly, two different oscillations patterns were observed: ‘rapid’ transients with amplitude similar to the initial progesterone response and slower transients with lower amplitude. Pre-treatment with VLN abolished ‘rapid’ [Ca²⁺]_i oscillations in 95% of cells, however, ‘slow’ transients were not sensitive to VLN. Moreover, VLN did not prevent nor alter the amplitude of the initial transient response induced by P4. Similar results were obtained when VLN was added after P4 stimulation. 98% of cell population had [Ca²⁺]_i oscillations inhibited in the presence of VLN but ‘slow’ transients were resistant to membrane potential hyperpolarization. Both Ca²⁺ oscillations patterns were completely recovered when VLN was removed. Our results showed that membrane potential contributes to [Ca²⁺]_i oscillations generation but only to high amplitude transients, suggesting that CatSper may be in involved in this oscillation profile, potentially triggering CICR. Low amplitude transients may be generated by a second Ca²⁺ signalling pathway.

Financial Support: CAPES.