Melatonin affects red deer spermatozoa motility and physiology in capacitating and non-capacitating conditions

Melatonin affects sperm physiology, possibly through membrane receptors. Effects were tested at low concentrations (1 pM, 100 pM, 10 nM and 1 mu M) in red deer epididymal spermatozoa as a model for high-seasonality species. Samples were incubated with melatonin as uncapacitated or capacitating conditions (heparin) and evaluated for motility and physiology (flow cytometry). Most effects occurred at low concentrations (nM-pM), mainly protecting from apoptosis and maintaining acrosomal integrity, suggesting a role for membrane receptors rather than a direct antioxidant effect. Intracellular calcium was not affected, differing from other studies and perhaps because of the epididymal origin. This study supports the relevance of melatonin on sperm physiology and could contribute to the application of reproductive technologies in wild ruminants

Melatonin Non-Linearly Modulates Bull Spermatozoa Motility and Physiology in Capacitating and Non-Capacitating Conditions

[EN] Bull spermatozoa physiology may be modulated by melatonin. We washed ejaculated spermatozoa free of melatonin and incubated them (4 h, 38 °C) with 0-pM, 1-pM, 100-pM, 10-nM and 1-µM melatonin in TALP-HEPES (non-capacitating) and TALP-HEPES-heparin (capacitating). This range of concentrations encompassed the effects mediated by melatonin receptors (pM), intracellular targets (nM–µM) or antioxidant activity (µM). Treatment effects were assessed as motility changes by computer-assisted sperm analysis (CASA) of motility and physiological changes by flow cytometry. Melatonin effects were more evident in capacitating conditions, with 100 pM reducing motility and velocity (VCL) while increasing a “slow” subpopulation. All concentrations decreased apoptotic spermatozoa and stimulated mitochondrial activity in viable spermatozoa, with 100 pM–1 µM increasing acrosomal damage, 10 nM–1 µM increasing intracellular calcium and 1 pM reducing the response to a calcium-ionophore challenge. In non-capacitating media, 1 µM increased hyperactivation-related variables and decreased apoptotic spermatozoa; 100 pM–1 µM increased membrane disorders (related to capacitation); all concentrations decreased mitochondrial ROS production. Melatonin concentrations had a modal effect on bull spermatozoa, suggesting a capacitation-modulating role and protective effect at physiological concentrations (pM). Some effects may be of practical use, considering artificial reproductive techniquesSIThis research was funded by MINECO (Spain), Grant Number AGL2013-4332

Single layer centrifugation (SLC) for bacterial removal with Porcicoll positively modifies chromatin structure in boar spermatozoa

The storage of boar semen samples at 17 degrees C for artificial insemination (AI) doses enables the proliferation of the bacteria, making antibiotics necessary. This can contribute to the development of antimicrobial resistance (AMR). This study tested bacterial presence and sperm chromatin structure after using a low-density colloid (Porcicoll) as an antibiotic alternative to eliminate bacteria. Ejaculates (8 boars, 3 ejac-ulates each) were split as control and low-density colloid centrifugation (single layer centrifugation, SLC, 20%, and 30% Porcicoll) into 500 ml tubes. Analyses were carried out at days 0, 3, and 7 (17 degrees C) for microbial presence and sperm chromatin structure analysis: %DFI (DNA fragmentation) and %HDS (chromatin immaturity), monobromobimane (mBBr; free thiols and disulfide bridges), and chromomycin A3 (CMA3; chromatin compaction). Besides comparing bacterial presence (7 species identified) and chromatin variables between treatments, the associations between these sets of variables were described by canonical correlation analysis (CCA). Results showed a significant decrease of some bacteria or a complete removal after SLC (especially for P30). SLC also caused a decrease of %HDS and an increase of disulfide bridges and low and medium mBBr populations, suggesting the removal of immature sperm (poor chromatin compaction). CCA showed an association pattern compatible with the degradation of sperm chromatin parameters with bacterial contamination, especially Enterobacteria, P. aeuriginosa, and K. variicola. In conclusion, bacterial contamination affects sperm chromatin beyond DNA fragmentation; SLC with low-density colloid not only removes bacteria from boar semen, but also chromatin structure is enhanced after selection.(c) 2023 Published by Elsevier Inc