Cryopreserved semen is widely used in assisted reproductive techniques. Post-thawing spermatozoa endure oxidative stress due to the high levels of reactive oxygen and nitrogen species, which are produced during the freezing/thawing process, and the depletion of antioxidants. To counteract this depletion, supplementation of sperm preparation medium with antioxidants has been widely applied. Melatonin is a hormone with diverse biological roles and a potent antioxidant, with an ameliorative effect on spermatozoa. In the present study, we assessed the effect of melatonin on thawed bovine spermatozoa during their handling. Cryopreserved bovine spermatozoa were thawed and incubated for 60 min in the presence or absence of 100 μΜ melatonin. Also, the effect of melatonin was assessed on spermatozoa further challenged by the addition of 100 μΜ hydrogen peroxide. Spermatozoa were evaluated in terms of kinematic parameters (CASA), viability (trypan blue staining) and antioxidant capacity (glutathione and NBT assay, determination of iNOS levels by Western blot analysis). In the presence of melatonin, spermatozoa presented better kinematic parameters, as the percentage of motile and rapid spermatozoa was higher in the melatonin group. They also presented higher viability and antioxidant status, as determined by the increased cellular glutathione levels and the decreased iNOS protein levels.

The polytene chromosomes of Drosophila triauraria and D. quadraria, two of the sibling species of D. auraria, were examined. The polytene chromosomes of all three species exhibit very clear homology. Unlike the stock of D. auraria that we studied, D. triauraria and D. quadraria carry heterozygous paracentric inversions. In both species, 2R and 3R are the arms where these inversions are concentrated. In addition, in D. quadraria, the 3L chromosome arm is very complicated because of heterozygous inversions. The mode of inheritance of these rearrangements was studied. A homozygous strain for all chromosome arms of D. triauraria was isolated, while a homozygous strain was obtained only for the arms X, 2L, 3L, and 4 of D. quadraria. Like D. auraria, both species show a large number of inverted tandem duplications in the paired condition, even in the chromosomes of their hybrids. Small deletions were also detected, one of which, in D. triauraria, is homozygous terminal. Hypotheses are discussed concerning the relationships of the species and the existence of inverted tandem duplications.Key words: Drosophila, duplications, inversions, deletions, evolution.

Reactive Oxygen Species (ROS) play an important role in sperm physiology. They are required in processes such as capacitation and fertilization. However, the exposure of spermatozoa to ROS generated from internal or external sources may create a potentially detrimental redox imbalance. Antioxidant supplementation in semen is now a rather common approach to protect spermatozoa from oxidative stress (OS) during their handling and/or cryopreservation. Supplementation with pterostilbene, a potent antioxidant, protects spermatozoa from OS and ameliorates their post-thawing characteristics and viability. In the present study, we used freezing/thawing as a model of natural ROS overproduction and investigated the molecular mechanisms modulated by pterostilbene. Specifically, bovine frozen/thawed spermatozoa were incubated with 10 or 25 μM pterostilbene for 60 min. Results have shown that in a dose-independent manner, pterostilbene decreased lipid peroxidation and increased intracellular GSH levels. Moreover, pterostilbene ameliorated energy production, as ATP and AMP/ATP levels were restored, and increased autophagy levels through AMP-activated protein kinase (AMPK) activation, which finally resulted in the inhibition of apoptotic cell death in bovine spermatozoa when exposed to OS. This study sheds light on spermatozoa redox state, the crosstalk between apoptotic and autophagic pathways, and its role in determining the beneficial or detrimental effect of ROS in spermatozoa.