Phosphorylant capacity study and lactate mitochondrial oxidation in frozen bovine sperm.

Frozen-stored bovine sperm-pellets of proven fertility were used, and the response to respiratory chain effectors was studied, thus demonstrating the energy conservation capacity. It was further observed that the assayed suspensions used lactate oxidatively, which proves the LDH-X mitochondrial activity (the presence of oxidative substrates is fundamental in capacitation and acrosome reaction processes). The suspensions were treated with 10mM phosphate buffer hypotonic medium to eliminate plasmalema and cytoplasmic content. Lactate respiration was sensitive to respiratory chain effectors, such as oligomycin and antimycin. To evaluate the LDH-X contribution to mitochondrial respiration, lipoate dehydrogenase was inhibited through 5-methoxyindole-2-carboxylic acid (MICA) in the presence of pyruvate-malate and citrate-malate, obtaining with the addition of lactate, oxygen uptakes of 18% and 51% with respect to respiration with the mentioned substrates. In the MICA dose-effect curve, a major sensitivity to inhibitor in active state mitochondrial respiration is obtained when pyruvate-malate is used. Lactate competence with pyruvate by mitochondrial LDH-X was observed. The results obtained would allow the thorough study of the necessity of oxidative energy in the capacitation and fertilization processes, and of the LDH-X role in frozen-stored bovine sperm.