Ionic control of sperm motility and trials for the improvement of pufferfish (Takifugu alboplumbeus) sperm extenders

Abstract

[EN] Seminal plasma characteristics, such as ionic composition and pH, are important for sperm maturation and further motility. In this work, a first batch of experiments evaluated the effect of the absence of several ions (Na+, K+, Mg2+) in the artificial seminal plasma, as well as the effect of artificial seminal plasma pH and seawater pH, on the sperm activation in the pufferfish (Takifugu alboplumbeus, named before Takifugu niphobles). In all the experiments, total motility, progressive motility, curvilinear velocity, straight line velocity, and average path velocity were measured. The absence of Na+, K+ or Mg2+ ions did not reduce the sperm motility in comparison with the control. In contrast, artificial seminal plasma pH had a strong effect on sperm motility. Samples diluted in artificial seminal plasma at pH 6.5 showed a high reduction of sperm motility in comparison with higher pHs (7.5, 8.5 and 9.5), and this inhibition was reversible. Where the effect of the pH of artificial seawater in sperm motility (used as activation medium) was concerned, there were no significant differences between the control (pH 8.2) and pH 6.5, indicating that pufferfish spermatozoa can swim in acidic environments. Finally, nigericin (that make equal the intracellular pH and extracellular pH) was added, and the highest sperm motility was found at pH 7.0, which suggested that the optimal intracellular pH of pufferfish for sperm motility is 7.0.

Our second batch of experiments was developed to optimize the pufferfish sperm extenders to achieve longer preservation under refrigeration, assessing the addition of several concentrations of magnesium and NaHCO3 (used in extenders for other fish species), as well as different pHs. From the extender experiments it was observed that bicarbonate reduced the sperm motility after 7 days of incubation, and it can be concluded that the use of magnesium 2 mM as part of the extender composition, in combination with the pH measured in the pufferfish sperm (7.5), seems to be the best option for the short-medium term storage of the sperm of this species.