The population of snow leopards (Uncia uncia) maintained in U.S. zoos is no longer sustainable due to poor reproductive success. Our objective was to assess reproductive traits in male snow leopards and identify factors (markers of oxidative stress in seminal fluid, surveys of husbandry practices, gonadal and adrenocortical activity, dietary intake of various nutrients, and genetics) that may affect ejaculate traits and subsequent fertility. Ejaculates (2.9 ± 0.2 ml) from 32 male snow leopards (9.8 ± 0.7 y; 38.6 ± 0.8 kg) housed at 27 institutions contained 119.2 + 26.0 x 106 spermatozoa, of which 75.1 ± 2.3% were motile and 28.6 ± 2.6% exhibited normal morphology. Overall, 34% of males produced <5 million spermatozoa and 27% of males produced spermatozoa with <20% normal morphology. Activity of superoxide dismutase (SOD) in the seminal fluid was negatively correlated (P<0.05, r2=0.90) with normal sperm morphology. Husbandry practices, mean concentrations of fecal androgen metabolites (fAM), and baseline concentrations of fecal glucocorticoid metabolites (fGM), inbreeding coefficients, and generations each male was removed from the founders in their lineages were not correlated (P>0.05) with the total number of spermatozoa or the proportion of spermatozoa with normal morphology. Total sperm count was positively correlated (P<0.05, R2=0.86) with weekly intake of polyunsaturated fatty acids (PUFA) and the proportion of spermatozoa with normal morphology tended (P<0.10, R2=0.31) to be positively correlated with copper intake. Altering the nutrient composition of snow leopard diets could provide managers with a possible method of improving reproductive traits in this endangered species.

According to the World Health Organization guidelines, ejaculatory abstinence (EA) of 2–7 days is recommended for semen analysis. This study aimed to determine how seminal quality may be affected by two EA periods from the same man. Seminal samples from 65 men were evaluated by conventional semen analysis and qualitative characteristics after 1 and 4 days of EA (two samples/man). The semen was qualitatively analyzed by examining oxidative activity (intracellular and seminal plasma), sperm function (acrosome integrity, mitochondrial activity, and nuclear DNA integrity), and epididymal function. As expected, samples collected after 1 day of EA showed a decrease in volume and sperm total number compared to samples collected after 4 days of EA. The sperm motility of the samples collected after 1 day of EA was better compared to samples collected after 4 days of EA. Oxidative activity measured was lower after 1 day of EA compared with those measured after 4 days of EA. With regards to sperm function, samples collected after 1 day of EA showed an increase in acrosome integrity, mitochondrial activity, and nuclear DNA integrity compared with samples collected after 4 days of EA. Epididymal function showed no difference between the two-time points. Although samples collected after 4 days of EA showed better results for sperm quantity, samples collected after 1 day of EA showed better qualitative results, including motility, oxidative activity, and sperm function. Thus, it can be concluded that sperm storage at the epididymal tail may make spermatozoa more susceptible to oxidative damage.

Undernutrition impairs reproductive success through suppression of gonadotropin-releasing hormone (GnRH), and subsequently luteinizing hormone (LH), secretion.  Given that kisspeptin and neurokinin B (NKB) neurons in the arcuate nucleus (ARC) of the hypothalamus are thought to play key stimulatory roles in the generation of GnRH/LH pulses, we hypothesized that feed restriction would reduce the ARC mRNA abundance and protein expression of kisspeptin and NKB in young, male sheep.  Fourteen wethers (castrated male sheep five months of age) were either fed to maintain (FM; n = 6) pre-study body weight or feed-restricted (FR; n = 8) to lose 20% of pre-study body weight over 13 weeks.  Throughout the study, weekly blood samples were collected and assessed for LH concentration using radioimmunoassay.  At Week 13 of the experiment, animals were euthanized, heads were perfused with 4% paraformaldehyde, and brain tissue containing the hypothalamus was collected, sectioned, and processed for detection of mRNA (RNAscope) and protein (immunohistochemistry) for kisspeptin and NKB.  Mean LH was significantly lower and LH inter-pulse interval was significantly higher in FR wethers compared to FM wethers at the end of the experiment (Week 13).  RNAscope analysis revealed significantly fewer cells expressing mRNA for kisspeptin and NKB in FR wethers compared to FM controls, and immunohistochemical analysis revealed significantly fewer immuno-positive kisspeptin and NKB cells in FR wethers compared to FM wethers.  Taken together, this data supports the idea that long-term feed restriction regulates GnRH/LH secretion through central suppression of kisspeptin and NKB in male sheep. 

Cryopreservation compromises sperm fertilising capacity due to a series of alterations in sperm structure and physiology; use of antioxidants such as melatonin, added to freezing media, may help to reduce sperm cryoinjury. To test the effect of melatonin on Bulldog [Canis lupus familiaris] sperm cryosurvival, spermatozoa were diluted in a standard freezing medium, cooled to 5°C, and added more freezing medium to obtain 200 x 106 cells/mL, and 5% glycerol. Diluted spermatozoa were added melatonin (0.0, 0.0005, 0.002, and 0.0035 mol/L), and packaged in 0.25 mL straws which were further cooled to -5°C before freezing in liquid nitrogen. Thawing was carried out at 70°C for 5 sec, and then spermatozoa (at 37°C) were assessed for progressive motility, viability, plasma membrane integrity, acrosome integrity, capacitation status, and plasma membrane fluidity; data was analysed by ANOVA to detect differences between melatonin doses. There were differences (P<0.05) in the percentage of sperm having hyper-fluid membranes, intact acrosome, capacitated acrosome-intact, and acrosome reacted, being the values for high (0.002 and 0.0035 mol/L) better than for low (0.0 and 0.0005 mol/L) melatonin doses. In conclusion, 0.002 and 0.0035 mol/L melatonin improved cryosurvival of sperm from Bulldog males.