VALDEBENITO ISLER, NEMESIO IVAN
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Effects of cryopreservation on mitochondrial function and sperm quality in fish
The development of cryopreservation techniques has led to important changes in animal reproductive biotechnology. However, these techniques are associated with cellular and molecular damage, affecting the mitochondrial function and quality of spermatozoa; moreover studies in fish are limited. In this work, the effects of cryopreservation on ultrastructure, mitochondrial function and antioxidant defences in Atlantic salmon (Salmo salar) spermatozoa were assessed, along with intracellular calcium (Ca2+)i, mitochondrial DNA sequence and sperm function (motility and fertilization rate). Significant ultrastructure alterations of the middle piece and mitochondria were observed in cryopreserved spermatozoa as compared to controls. Oxygen consumption and ATP were also significantly different in cryopreserved spermatozoa, and in spermatozoa incubated with electron transport chain (ETC) uncouplers and inhibitors. Mitochondrial membrane potential, motility, fertilization rate and Ca2+ i in cryopreserved spermatozoa displayed significant reductions compared to fresh spermatozoa. Mitochondrial potential correlated significantly with motility and fertilization rate. A redox imbalance was observed in frozen spermatozoa due to increased lipid peroxidation and superoxide anion production as compared to fresh spermatozoa. Likewise, increased activity of glutathione peroxidase and total glutathione (GSH/GSSG), as well as reduced catalase activity, were observed in comparison with fresh spermatozoa. Our results contribute to a better understanding of cryodamage to mitochondrial functions in fish spermatozoa, and enabled us to establish potential quality assessment indicators. The data suggest that cryopreservation induces a reduction in overall sperm quality and functionality through disruption of the mitochondrial ultrastructure and function, leading to energy depletion and increased oxidative stress. This knowledge may also lead to the identification of a potential biotechnological tool for improving reproductive efficiency in species of commercial and biological interest. © 2019 Elsevier B.V., All rights reserved.
The CatSper channel is present and plays a key role in sperm motility of the Atlantic salmon (Salmo salar)
Among all the Ca2+ channels, CatSper channels have been one of the most studied in sperm of different species due to their demonstrated role in the fertilization process. In fish sperm, the calcium channel plays a key role in sperm activation. However, the functionality of the CatSper channels has not been studied in any of the fish species. For the first time, we studied the relationship of the CatSper channel with sperm motility in a fish, using Atlantic salmon (Salmo salar) as the model. The results of our study showed that the CatSper channel in Salmo salar has chemical-physical characteristics similar to those reported for mammalian CatSper channels. In this work, it was shown that Salmo salar CatSper 3 protein has a molecular weight of approximately 55-kDa similar to Homo sapiens CatSper 3. In silico analyses suggest that this channel forms a heterotetramer sensitive to the specific inhibitor HC-056456, with a binding site in the center of the pore of the CatSper channel, hindering or preventing the influx of Ca (2+) ions. The in vitro assay of the sperm motility inhibition of Salmo salar with the inhibitor HC-056456 showed that sperm treated with this inhibitor significantly reduced the total and progressive motility (p < .0001), demonstrating the importance of this ionic channel for this cell. The complementation of the in silico and in vitro analyses of the present work demonstrates that the CatSper channel plays a key role in the regulation of sperm motility in Atlantic salmon.