Exogenous Albumin Is Crucial for Pig Sperm to Elicit In Vitro Capacitation Whereas Bicarbonate Only Modulates Its Efficiency.

Abstract

Simple SummaryIn this work, we addressed if the presence of exogenous bicarbonate required for pig sperm capacitation, which is a necessary step to acquire fertilizing ability. While sperm incubated in media without BSA or BSA/bicarbonate did not achieve in vitro capacitation, those incubated with BSA reached that status under any bicarbonate concentration, even when bicarbonate was absent. Interestingly, there were differences related to the concentration of bicarbonate, since sperm incubated in media with BSA and with no bicarbonate or 5 mM bicarbonate showed lower overall efficiency in achieving in vitro capacitation than those incubated in the presence of BSA and higher concentration of bicarbonate. Additionally, at the end of the experiment, sperm incubated in the presence of BSA and 38 mM bicarbonate showed lower motility and plasma membrane integrity than those incubated in media with BSA and lower concentrations of bicarbonate. In conclusion, BSA is crucial in for pig sperm to elicit in vitro capacitation and trigger the subsequent progesterone-induced acrosome exocytosis. In contrast, although exogenous bicarbonate does not appear to be indispensable, it shortens the time needed to reach that capacitated status.This work sought to address whether the presence of exogenous bicarbonate is required for pig sperm to elicit in vitro capacitation and further progesterone-induced acrosome exocytosis. For this purpose, sperm were either incubated in a standard in vitro capacitation medium or a similar medium with different concentrations of bicarbonate (either 0 mM, 5 mM, 15 mM or 38 mM) and BSA (either 0 mg/mL or 5 mg/mL). The achievement of in vitro capacitation and progesterone-induced acrosomal exocytosis was tested through the analysis of sperm motility, plasma membrane integrity and lipid disorder, acrosome exocytosis, intracellular calcium levels, mitochondria membrane potential, O2 consumption rate and the activities of both glycogen synthase kinase 3 alpha (GSK3α) and protein kinase A (PKA). While sperm incubated in media without BSA or BSA/bicarbonate, they did not achieve in vitro capacitation; those incubated in media with BSA achieved the capacitated status under any bicarbonate concentration, even when bicarbonate was absent. Moreover, there were differences related to the concentration of bicarbonate, since sperm incubated in media with BSA and with no bicarbonate or 5 mM bicarbonate showed lower overall efficiency in achieving in vitro capacitation than those incubated in the presence of BSA and 15 mM or 38 mM bicarbonate. Additionally, at the end of the experiment, sperm incubated in the presence of BSA and 38 mM bicarbonate showed significantly (p < 0.05) lower values of motility and plasma membrane integrity than those incubated in media with BSA and lower concentrations of bicarbonate. In conclusion, BSA is instrumental for pig sperm to elicit in vitro capacitation and trigger the subsequent progesterone-induced acrosome exocytosis. Furthermore, while exogenous bicarbonate does not seem to be essential to launch sperm capacitation, it does modulate its efficiency.