Cell bioenergetics and ATP production of boar spermatozoa.

Cellular metabolism is an important feature of spermatozoa that deserves more insights to be fully understood, in particular in porcine semen physiology. The present study aims to characterize the balance between glycolytic and oxidative metabolism in boar sperm cells. Agilent Seahorse technology was used to assess both oxygen consumption rate (OCR), as an oxidative metabolism index, and extracellular acidification rate (ECAR), as a glycolytic index. Different metabolic parameters were studied on freshly ejaculated sperm cells (identified as day zero sample, d0) and after one day of storage at 17 °C in Androhep extender (d1). Mitochondrial ATP production rate (MitoATP) was higher than the glycolytic ATP production rate (glycoATP) at both d0 and d1 while at d1 the amount of ATP production decreased, in particular, due to OXPHOS reduction. Conversely, glycoATP was not significantly different between d0 and d1. Interestingly, OCR profile showed no different bioenergetic parameters (i.e. ATP turnover, basal or maximal respiration, and spare respiration) between d0 and d1, thus indicating that sperm cell metabolism was reversibly decreased by preservation conditions. Other metabolic parameters showed the same trend, irrespective of the storage time: under stressed conditions (oligomycin plus FCCP), spermatozoa showed an increase in mitochondrial respiration while the metabolic potential of glycolysis did not undergo variations when compared to baseline metabolism. The rate of oxidation of fuel substrates - glucose, fatty acids, and glutamine - showed that sperm reliance on glucose oxidation to maintain baseline respiration was higher than fatty acids or glutamine. Interestingly spermatozoa demonstrated to have a low "capacity" parameter, which indicates that they cannot use only a single fuel substrate to produce energy. This feature of sperm metabolism to be unable to increase oxidation of a particular fuel to compensate for inhibition of alternative fuel pathway(s) was demonstrated by the negative value of "flexibility". Our results showed that ATP production in boar sperm cells relied on mitochondrial oxidative metabolism in freshly ejaculated cells, while, under liquid storage conditions, their oxidative metabolism decreased while the glycolysis remained constant. These results open new fields of research in the preservation techniques of boar sperm cells.