Frequency of Semen Collection Affects Ram Sperm Cryoresistance

Palacin-Martinez, Cristina; Álvarez, M.; Montes-Garrido, Rafael; Neila-Montero, Marta; Anel-López, Luis; Paz, P. de; Anel, L.; Riesco, Marta F.

doi:10.3390/ani12121492

Cited by 8 publications

(4 citation statements)

References 64 publications

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“…CRDR has been considered an ROS marker in ram ( 35 , 50 ), bull ( 38 , 51 ) and stallion sperm ( 44 , 52 – 54 ). However, contradictory results have been published concerning their correlation with sperm quality in rams ( 42 , 55 , 56 ) and stallions ( 57 , 58 ). These results could suggest that there may be a species-specific effect, which we have decided to study in depth.…”

Section: Discussionmentioning

confidence: 99%

The characterization of CellROX™ probes could be a crucial factor in ram sperm quality assessment

Palacin-Martinez,

Anel-Lopez,

Alvarez

et al. 2024

Front. Vet. Sci.

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Several authors have demonstrated that low levels of reactive oxygen species (ROS) are necessary for the physiological functions of sperm, such as capacitation, hyperactivation, acrosomal reaction and fertilization. However, high levels of ROS are associated with oxidative stress and detrimental effects on fertility. Consequently, deep characterization of ROS presence using different fluorescent probes could be crucial. In this sense, the study of intracellular ROS localization and the relationships between ROS and other conventional parameters could improve the characterization of sperm quality for semen preservation protocols in rams. In this work, a multiparametric study was carried out by analyzing four experimental groups of ram sperm with different initial qualities: fresh semen (from both breeding and nonbreeding seasons), frozen-thawed semen and, a positive control group treated with hydrogen peroxide (300 μM) as a marker of extreme damage. Sperm analyses, including viability, apoptosis, lipid peroxidation, motility and kinetic parameters, were applied to compare several experimental groups with different sperm qualities. After that, the signals from two different ROS probes: CellROX™ Deep Red (CRDR) and Green (CRG), were examined by flow cytometry (percentage of cells that express ROS) and fluorescence microscopy (intracellular ROS location). Comparing conventional parameters, fresh samples from the breeding season showed the highest sperm quality, while the positive control samples showed the worst sperm quality. Concerning the ROS probes, the CRDR levels were higher in fresh samples from the breeding season than in the positive control and cryopreserved samples. Surprisingly, CRG presented its highest level (P < 0.05) in the positive control group treated with peroxide by flow cytometry. CRDR and CRG presented opposite labeling patterns that were corroborated by fluorescence microscopy, which determined that the probes localized in different parts of sperm. CRDR was found in the sperm mitochondrial region, while CRG was observed in the cell nucleus, suggesting that ROS localization is an important factor. Finally, our study indicates that CRDR is correlated with proper viability and sperm motility, and could be associated with high mitochondrial activity, while CRG is associated with sperm damage.

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Section: Discussionmentioning

confidence: 99%

The characterization of CellROX™ probes could be a crucial factor in ram sperm quality assessment

Palacin-Martinez,

Anel-Lopez,

Alvarez

et al. 2024

Front. Vet. Sci.

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“…The results of our morphometric assessments showed a wide variability prior to, during, and after the mating period. Effects on spermatogenesis can give rise to variations in the size of the spermatozoa and could explain the results observed for sperm head and midpiece size and shape variables that were low after mating; it could be specially related with frequency of collection/ejaculation [ 65 ] and a competition effect between males [ 66 , 67 ]. Moreover, Kondracki et al [ 68 ] reported in bulls that sperm head size increases as the sperm concentration of the ejaculate decreases.…”

Section: Discussionmentioning

confidence: 99%

Kinematic and Morphometric Assessment of Fresh Semen, before, during and after Mating Period in Brahman Bulls

Araya-Zúñiga,

Sevilla,

Molina-Montero

et al. 2023

Animals

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The objective of the present study was to determine the effects that the reproductive season has on the motility, kinematics, morphology, and sperm morphometry of Brahman bulls evaluated with a commercial CASA system. The experiment was carried out at the Costa Rica Institute of Technology from March to August 2021. A total of eight Brahman bulls were used. A total of 28 ejaculates were collected in the pre-mating period (PMP), during it (DMP), and after it (AMP) using an electroejaculator. The sperm concentration was measured with the Accuread photometer. The motility was measured using a Spermtrack® counting chamber. The analyses were performed with the CASA-Mot ISAS®v1 system. The morphology was analyzed using a microscope with a negative phase contrast objective. Morphometry was evaluated with the CASA-Morph. The sperm concentration did not present differences between the PMP and AMP; however, it was significantly higher than DMP (p > 0.05). Regarding the progressiveness variables, linearity on forward progression (LIN), straightness (STR), and wobble (WOB) were higher (p < 0.05) DMP. A kinematic principal component analysis grouped all the variables into three factors and an effect on the reproductive period was found (p < 0.05) in the parameters of the head and middle part of the sperm, such as width and perimeter, which were greater in the PMP. The length of the sperm head in the PMP and DMP did not show differences; however, both were larger (p < 0.05) than AMP. The insertion distance of the middle piece of the sperm was significantly greater than DMP. Finally, the PMP contained cells with a larger insertion angle (p < 0.05) than AMP. These findings are important to understand the implications of reproductive status on sperm quality and to consider them in andrological evaluations.

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“…According to Montes-Garrido et al [41], sperm motility and functionality were lower at 6 h post-collection (the time at which vaginal artificial insemination is performed) in the spermatozoa of rams that were overworked. Moreover, the commercial potential of overworked rams is also reduced for frozen-thawed doses, since previous authors have described the lower cryotolerance of spermatozoa produced in a high regime of collection due to a higher oxidative stress through the activity of glutathione peroxidase and superoxide dismutase [51]. Despite all this, it would be interesting to consider the economic impact of specific rams and the need to use them to accelerate genetic improvement.…”

Section: Discussionmentioning

confidence: 99%

Does Size Matter? Testicular Volume and Its Predictive Ability of Sperm Production in Rams

Montes-Garrido,

Anel-Lopez,

Riesco

et al. 2023

Animals

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Over the years, testicular volume has been used to evaluate the reproductive capacity of rams and the effects of different factors related to reproductive performance. The aim of this study was to determine the most suitable tool and formula to calculate testicular volume under field conditions to guarantee a more accurate determination of sperm production. First, testicles from 25 rams (n = 50) were measured in vivo and postmortem using calipers and ultrasonography during the breeding season (BS). The accurate testicular volume (ATV) was calculated through water displacement. In addition, the sexual status of donor rams was evaluated during a period of four years in a reproduction center, and the three most crucial groups in terms of genetic value and seminal collections were studied in the second part of this experiment: ER-NBS (Elite rams during the non-breeding season), ER-BS-S (Elite rams with a standard frequency of seminal collection), and ER-BS-O (Elite rams with a high frequency of seminal collection). The total testicular volume (TTV), testosterone (T), and total spermatozoa obtained from two consecutive ejaculates in the same day (SPERM) were measured, and the relationship between SPERM and TTV and T was analyzed to predict SPERM. Although all published formulas revealed statistically significant differences (p ≤ 0.05) from the ATV, our proposed formula (ItraULE) (Testicular volume = L × W × D × 0.61) did not show significant differences. In the second part of the study, in the ER as a model donor ram for its high genetic value and high demand from farmers, TTV and T showed strong positive correlations with SPERM (r = 0.587, p = 0.007 NBS; r = 0.684, p = 0.001 BS-S; r = 0.773, p < 0.0001 BS-O). Moreover, formulas were established to predict SPERM in these practical scenarios. In conclusion, the use of ultrasonography and a new formula adapted to rams could improve the prediction of SPERM considering crucial factors such as season and semen collection frequency.

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Frequency of Semen Collection Affects Ram Sperm Cryoresistance

Cited by 8 publications

References 64 publications

The characterization of CellROX™ probes could be a crucial factor in ram sperm quality assessment

The characterization of CellROX™ probes could be a crucial factor in ram sperm quality assessment

Kinematic and Morphometric Assessment of Fresh Semen, before, during and after Mating Period in Brahman Bulls

Does Size Matter? Testicular Volume and Its Predictive Ability of Sperm Production in Rams

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