Comparison of sperm velocity, motility and fertilizing ability between firstly and secondly activated spermatozoa of common carp (Cyprinus carpio)

The objective of the study was to compare carp sperm motility performances (sperm velocity and motility rates) from 10 males including fertilizing ability (hatching rates from 10 males and eight females) as a function of time elapsed after sperm exposure to activation medium in two situations: firstly activated sperm and sperm which had terminated swimming and was 're-activated' after incubation in a K+ rich (200 mm KCl) non-swimming solution. In case of both initial (first) and secondly activated spermatozoa, the motility was triggered in hatchery solution (HAS, 11.2 mOsmol) and in carp activation solution (CAS, 128.9 mOsmol) containing 45 mm NaCl, 5 mm KCl, 30 mm Tris-HCl while also adjusted to a pH of 8.0. First time activated sperm showed significantly higher relative motility, sperm velocity and fertilizing ability compared to re-activated sperm. The carp spermatozoa (in either first or second activation) rapidly lost their fertilizing ability as a function of exposure time of sperm to diluents prior to addition to eggs: this shows that spermatozoa must be in contact with eggs as soon as their motility is triggered. When sperm was firstly activated in CAS and also activated a second time in CAS (labeled CASCAS) the hatching rate was significantly higher at egg contact after 10, 20, 30, and 120 s of activation. Also at 20 s after the second activation of the sperm higher sperm motility was observed compared to the first activation. This study showed that incubation of spermatozoa in a K+-rich incubation medium can mitigate the affects of structural damages occurring in re-activated sperm, which may help spermatozoa to increase their motility and fertilization. To our knowledge, the results presented in this study document for the first time that fertilization can be achieved with sperm re-activated a second time while being exposed to a incubation medium that permits ATP reloading within the flagellum. Previous studies have show the potential for recovery of motility, however, the effect on possible fertilization is hitherto unknown. It critical outcome of the study clearly indicated the need for avoiding the use of different, subsequent activation media (e.g. first and second activation) but only on the same medium for both steps (see above CASCAS).