Oxidative stress in cryopreserved semen of sex-reversed female and normal male rainbow trout

Abstract

The aim of the current study was to determine the sperm quality of sex-reversed female (SRF) and normal male rainbow trout measured as ROS+ (percentage of spermatozoa exhibiting reactive oxygen species) production and TAC (total antioxidant capacity) in semen cryopreserved at different glucose concentration [optimal (0.15 M), low (0.11 M) and high (0.19 and 0.21 M for SRF and normal male rainbow trout, respectively)] in 7.5% methanol (glucose-methanol extender). Our hypothesis assumes that at non-optimal glucose concentrations, production of ROS+ is induced, resulting in damage to spermatozoa which should be characteristic for normal males and SRF rainbow trout. Such approach would allow to describe the characteristics of the response of spermatozoa of normal males and SRF to oxidative stress which is prerequisite for better understanding the mechanism of oxidative stress of normal males and SRF. This knowledge can be important for improvement of protocols of short-term storage and cryopreservation of SRF and normal males semen. We established baseline values for ROS+ and TAC for fresh semen from SRF and normal male rainbow trout and found a negative regression between those two characteristics in SRF. We determined the effect of glucose concentration on ROS+ production, TAC and sperm quality parameters of cryopreserved semen. TAC activity was monitored at the same time, and its relationship to oxidative stress was assessed. The glucose concentration significantly affected sperm motility, curvilinear velocity and viability after thawing. Cryopreservation resulted in increased ROS+ values in both tested groups. On the other hand, TAC values were significantly lower in cryopreserved samples than in equilibrated samples of SRF rainbow trout semen at all tested glucose concentrations. Our results strongly suggest that increasing the osmolality of the extender induces oxidative stress in spermatozoa of SRF and normal male rainbow trout. Most changes are related to freezing and thawing; however, some changes may occur during equilibration. Specific changes in oxidative stress characteristics occur in fresh and cryopreserved semen of SRF and normal males. An inverse relationship between ROS+ and TAC is likely caused by consumption of antioxidants by ROS+. Our results can be useful for optimization of cryopreservation protocols for salmonid fish. Further studies are necessary to elucidate the mechanisms that lead to differences in oxidative stress levels in semen from SRF and normal male rainbow trout.