The follicular fluid regulates sperm capacitation and the acrosome reaction in the donkey while alleviating oxidative stress

The follicular fluid (FF) is a complex and dynamic biological medium that surrounds the oocytes during their development and growth. This fluid provides the necessary nutrients to both the oocyte and the surrounding cells, enabling the proper maturation of the oocyte. Additionally, the FF is involved in key sperm function processes, such as capacitation, which occurs when sperm reach the oviduct and are exposed to this fluid. Nevertheless, how this happens in donkeys has not been investigated. For this reason, the present study aimed to evaluate the influence of the FF on sperm capacitation in the donkey. Preovulatory FF was obtained from five different Catalan jennies (two follicles per animal), and nine ejaculates from fertile Catalan donkeys were collected. Sperm were resuspended in four different treatments: non-capacitating medium (TBM), non-capacitating medium supplemented with 20 % FF (TBM+FF), capacitating medium (TCM), and capacitating medium supplemented with 20 % FF (TCM+FF). Samples were incubated at 38 °C for 120 min, and at 60 min, progesterone was added to induce the acrosome reaction. After 0, 60, 70, and 120 min of incubation, sperm motility, membrane lipid disorder, acrosome integrity, mitochondrial membrane potential (MMP), and intracellular concentrations of calcium, reactive oxygen species (ROS) and superoxides were evaluated. These results suggest that the preovulatory FF promotes sperm capacitation in donkeys, as it increases intracellular calcium, membrane lipid disorder, and acrosome exocytosis, while maintaining sperm motility and reducing ROS concentration. Although the results are not entirely conclusive and further tests are needed, this study supports that the FF modulates sperm capacitation and alleviates the oxidative stress associated with this process.