Sperm melatonin receptors, seminal plasma melatonin and semen freezability in goats.

Goat bucks are seasonal breeders that show variation in sperm quality, endogenous melatonin (MLT), and presumably in the expression of MLT receptors on the sperm throughout the year, which may modify sperm freezability. The aim of this study was to determine whether sperm freezability is associated with (i) endogenous melatonin levels in seminal plasma and (ii) the expression of sperm plasma membrane melatonin receptors (MT1, MT2). To evaluate this, spermatozoa from seven Saanen goat bucks were cryopreserved throughout the year in Mexico using a standard freezing protocol. Seminal plasma MLT concentrations were determined by ELISA and the expression and localization of MT1 and MT2 were detected by immunocytochemistry and confirmed by western blotting. The recovery rate of progressive motility after thawing was higher in spring than autumn and winter; in contrast, the F pattern (CTC assay) was higher in winter than in the other seasons. A proportional increase in the AR pattern (CTC assay) was smaller in winter than in the other seasons and the proportion of sperm showing high plasma membrane fluidity was higher in spring than in summer and autumn. The seminal plasma MLT concentrations showed no significant interseasonal differences. The MT1 receptor was immunolocalised at the apical region of the sperm head, while MT2 was mainly localised in the neck. The relative expression of MLT receptors showed significant differences between summer and winter for all bands, except at 75 kDa of MT2. In conclusion, there was an association between the relative expression of MT1 and MT2 receptors throughout the year and sperm freezability in goat bucks in México. Post-thaw sperm quality is enhanced in semen samples collected during breeding season.

Implementation of a method for sperm cryopreservation in sceloporine lizards.

Actual loss of lizard biodiversity continues, even with the implementation of conventional conservation programs. An approach including assisted reproductive techniques such as sperm cryopreservation may contribute to the management of endangered species. We developed a method for sperm cryopreservation in sceloporine lizards and compared the response among the studied species. Prior to the mating season, we obtained semen from adult males of Sceloporus aeneus (n = 21), Sceloporus grammicus (n = 20) and Sceloporus torquatus (n = 21) via pressure of the genital papilla. Volume and sperm concentration were measured before semen dilution in a Tris-egg yolk (TEY) medium to evaluate progressive motility, sperm viability, morphology, plasma membrane and acrosome integrity. Then, we cooled the remaining volumes to 5°C at a rate of 0.1°C per minute to incorporate glycerol (8% v/v) in two fractions. Immediately afterwards, we placed 40 µl of the mix on solid CO2 to form pellets and immersed them in liquid nitrogen for storage. We thawed the pellets at 29°C for 3 minutes and diluted them 1:1 (v/v) in TEY medium to assess sperm quality. We found a positive relationship between body weight and seminal volume in S. grammicus and S. torquatus and a negative correlation with sperm concentration in S. grammicus (P < 0.05). Moreover, we observed that the freezing-thawing process decreased sperm quality in the three species, mostly affecting motility and viability. However, S. torquatus and S. aeneus showed a higher sperm tolerance than S. grammicus.