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Topic:Sperm
Equine sperm refers to the male reproductive cells produced by stallions, essential for the process of fertilization and successful breeding in horses. The study of equine sperm encompasses various aspects, including morphology, motility, viability, and genetic integrity. These parameters are critical for assessing stallion fertility and improving breeding outcomes. Research in this field often focuses on understanding the factors that influence sperm quality, such as age, nutrition, and environmental conditions. Additionally, advancements in assisted reproductive technologies, such as artificial insemination and cryopreservation, rely heavily on the detailed study of sperm characteristics. This page compiles peer-reviewed research studies and scholarly articles that explore the biology, evaluation, and technological applications related to equine sperm.
Capacitation, acrosome function and chromatin structure in stallion sperm. In general, fertility in breeding stallions is lower and more variable than in the other farm animal species, primarily because selection is based on pedigree, looks and/or athletic performance, with little consideration of fertility or fertility potential. Moreover, because the average stallion breeds only a limited number of mares per year and in-field fertility is influenced significantly by non-stallion factors such as management and mare fertility, meaningful fertility data are hard to come-by. Unfortunately, generating usable figures would involve impractically high costs, time and numbe...
Characterization of an A-kinase anchor protein in equine spermatozoa and examination of the effect of semen cooling and cryopreservation on the binding of that protein to the regulatory subunit of protein kinase-A. To determine whether a homologue of A-kinase anchor protein 4 (AKAP4) is present and functional as an AKAP in equine spermatozoa and examine the effect of semen cooling and cryopreservation on binding of equine AKAP4 to the regulatory (RII) subunit of protein kinase-A (PK-A). Methods: Ejaculated semen collected from 2 fertile stallions, 3 bulls, and 3 humans. Methods: Identification of an equine homologue of AKAP4 was investigated via DNA sequencing. Protein was extracted from the spermatozoa of each species for immunoblot analysis to identify AKAP4 and its precursor protein, pro-AKAP4; immuno...
Low-dose insemination–why, when and how. The typical dose for insemination into the uterine body of the mare is > 300 x 10(6) progressively motile spermatozoa (PMS) and an insemination dose of > 200 x 10(6) PMS is recommended for frozen-thawed semen. Low-dose insemination techniques allow for a drastic reduction in the numbers of spermatozoa required to achieve pregnancy. Acceptable pregnancy rates can be achieved with doses ranging from 1 to 25 x 10(6) PMS in volumes ranging from 20 to 1000 microL. Two techniques have been described: hysteroscopic insemination and transrectally guided deep horn insemination using a pipette. Si...
Effect of caspase inhibitors on the post-thaw motility, and integrity of acrosome and plasma membrane of cryopreserved equine spermatozoa. The present study was designed to test the hypothesis that addition of anticaspase cocktails (inhibiting caspases and thus blocking apoptosis) to the extenders increases the post-thaw viability of equine spermatozoa. The addition of caspase inhibitors failed to improve the acrosome and plasma membrane integrity of spermatozoa, suggesting that in equine sperm cryopreservation protocols, the addition of these caspase inhibitors to cryopreservation medium may not be beneficial in protecting the sperm from the stress of cryopreservation.
Update on equine ICSI and cloning. Intracytoplasmic sperm injection (ICSI) has recently become efficient enough to be considered for clinical use. With ICSI, one spermatozoa is injected into a mature oocyte. Harvesting of an oocyte ex vivo, followed by ICSI and transfer of the fertilized oocyte to the oviduct, may be applicable when semen quality is insufficient for standard insemination. Sperm injection, followed by in vitro embryo culture to the blastocyst stage, may be used in cases where multiple oocytes are to be fertilized (e.g. when oocytes are collected post-mortem). Nuclear transfer (cloning) of horses is possible but ...
Influence of vedaprofen (Quadrisol) on quality and freezability of stallion semen. The objective of this study was to evaluate the effect of the non-steroidal anti-inflammatory drug (NSAID) vedaprofen (Quadrisol) on quality and freezability of stallion semen. Experiments were performed using 22 Franches Montagnes stallions from the National Stud in Avenches (Switzerland) randomly divided into a control and test group. Vedaprofen was given orally to all stallions of the test group at the recommended therapeutic dose (initial dose of 2mg/kg followed by 1mg/kg body weight every 12h) for 14 days. Control animals received the same amount of carrier substance. During treatment, bl...
Lipid peroxide formation in relation to membrane stability of fresh and frozen thawed stallion spermatozoa. In this study we used a new method to detect reactive oxygen species (ROS) induced damage at the level of the sperm plasma membrane in fresh and frozen-thawed stallion sperm. Lipid peroxidation (LPO) in sperm cells was assessed by a fluorescent assay involving the labeling of stallion sperm with the LPO reporter probe C11-BODIPY(581/591). The peroxidation dependent spectral emission shift of this membrane probe could be localized using inverted spectral confocal microscopy and quantified on living and deteriorated sperm cells using flow cytometry. Mass spectrometric analysis of the main endoge...
Assessment of the cryopreservation of equine spermatozoa in the presence of enzyme scavengers and antioxidants. To evaluate the effect of the addition of enzyme scavengers and antioxidants to the cryopreservation extender on characteristics of equine spermatozoa after freezing and thawing. Methods: 2 ejaculates collected from each of 5 stallions. Methods: Equine spermatozoa were cryopreserved in freezing extender alone (control samples) or with the addition of catalase (200 U/mL), superoxide dismutase (200 U/mL), reduced glutathione (10 mM), ascorbic acid (10 mM), alpha-tocopherol (25, 50, 100, or 500 microM or 1 mM), or the vehicle for alpha-tocopherol (0.5% ethanol). After thawing, spermatozoal motili...
Effect of alpha-tocopherol and tocopherol succinate on lipid peroxidation in equine spermatozoa. The objective of this study was to compare the effect of alpha-tocopherol and its ester, alpha tocopherol succinate, on lipid peroxidation and motility of equine spermatozoa. In experiment one, spermatozoa were incubated with dl-alpha-tocopherol (5, 25, 100 or 500 microM), DL-alpha tocopherol succinate (5, 25, 100 or 500 microM) or vehicle (0.5% ethanol) at 38 degrees C, and sperm motility was determined at 30, 60 and 120 min. In experiment two, spermatozoa loaded with the lipophilic probe, C11BODIPY(581/591), were incubated with dl-alpha-tocopherol (50 and 100 microM), DL-alpha-tocopherol suc...
Effect of seminal plasma on the cryopreservation of equine spermatozoa. Seminal plasma is generally removed from equine spermatozoa prior to cryopreservation. Two experiments were designed to determine if adding seminal plasma back to spermatozoa, prior to cryopreservation, would benefit the spermatozoa. Experiment 1 determined if different concentrations of seminal plasma affected post-thaw sperm motility, viability and acrosomal integrity of frozen/thawed stallion spermatozoa. Semen was washed through 15% Percoll to remove seminal plasma and spermatozoa resuspended to 350 x 10(6)sperm/mL in a clear Hepes buffered diluent containing either 0, 5, 10, 20, 40 or 80%...
Comparison of in vitro laboratory analyses with the fertility of cryopreserved stallion spermatozoa. Assessing the fertilizing potential of a semen sample is important for effective stallion management and for rapid progress in evaluating new cryopreservation technologies. Unfortunately, sperm motility does not estimate fertility well. These experiments established assays to measure cell viability, acrosomal integrity and mitochondrial function for cryopreserved stallion spermatozoa, using flow cytometry, and determined the variability associated with these assays. Correlations between results for these laboratory assays and stallion fertility were also determined. The inter-assay variability...
Application of MTT reduction assay to evaluate equine sperm viability. The assay of MTT reduction depends on the ability of metabolically active cells to reduce the tetrazolium salt (3[4,5-dimethylthiazol-2-y1]-2,5-diphenyltetrazolium bromide) to formazan. This study was conducted to examine and validate of a simple and less costly MTT test in determining equine sperm viability and compare the efficiency of this test with a flow cytometer. Fresh ejaculates from 11 stallions (warm blood) were included in this study. Semen was diluted to 100 million cells/ml in a Hepes 0.1% BSA. The rates of MTT reduction were measured in microtiter plates after incubation for 1 an...
Sperm motility patterns and metabolism in Catalonian donkey semen. The Sperm-Class Analyzer detected four subpopulations of spermatozoa with different motility characteristics in the ejaculate of the Catalonian donkey. Significant differences (P < 0.001) in the distribution of these subpopulations, as well as in total sperm number and percentage total motility, were seen in the diluted semen of four sampled donkeys. All the ejaculates evaluated showed excellent semen quality characteristics; the sperm they contained was more rapid than horse sperm. Principal components analysis showed sperm l-lactate production to be a good predictor of semen condition. Th...
Relationship of seminal plasma level and extender type to sperm motility and DNA integrity. The relationship between seminal plasma level (0, 10, or 20%) and extender type [Kenney type (EZ-Mixin-CST) or Kenney-modified Tyrodes-KMT] to the susceptibility of sperm DNA to denaturation and sperm motility measures were investigated in cooled (5 degrees C) stallion sperm. Three ejaculates from each of three fertile stallions were collected in an artificial vagina and processed as follows: diluted one part uncentrifuged semen with four parts of extender to a final concentration of 20% seminal plasma in either CST or KMT (20% CST; 20% KMT); diluted to a final concentration of 25 million sper...
High pressure flow cytometric sorting damages sperm. Sexing sperm by high-speed flow cytometry subjects them to high pressure. The routine operating pressure of the MoFlo SX flow cytometer for sperm sorting for commercial production has been 50 pounds/square inch (psi), with a standard 70 microm standard nozzle tip. It was hypothesized that lowering the sorting pressure could reduce sperm damage. Therefore, a series of experiments using semen from six bulls, sorted with three MoFlo SX sorters, was conducted to determine optimal pressure. An additional experiment was done with stallion spermatozoa. In Experiment 1, sorting at 30 psi compared to 5...
Activity of glutathione peroxidase, superoxide dismutase and catalase and lipid peroxidation intensity in stallion semen during storage at 5 degrees C. Sperm cell membranes are susceptible to peroxidative damage by an excess of reactive oxygen species (ROS). Antioxidative defence systems consisting of glutathione peroxidase (GSH-Px), superoxide dismutase (SOD) and catalase (CAT) physiologically control the balance between ROS production and neutralization. In the present study the hypothesis was tested that lipid peroxidation occurs during storage of semen at 5 degrees C and that semen extender has positive effects on the antioxidative potential of equine semen. The aim of the study was to determine the activity of GSH-Px, SOD and CAT and the...
Freezability of equine semen after glass beads column separation. The success rate of artificial insemination following the freezing of stallion semen is limited; therefore, improving the stallion semen quality after the freezing and thawing process is a necessary objective. Objective: To investigate the influence of glass bead column separation on the freezability of stallion semen. Objective: Glass beads in a column separator remove damaged and dead spermatozoa in the ejaculate during centrifugation. Methods: In total, 50 ejaculates from 6 Lipizzaner stallions were studied. Each ejaculate was divided into 2 parts, one half processed following standard proc...
SNARE proteins and caveolin-1 in stallion spermatozoa: possible implications for fertility. Proteins implicated in the "SNARE hypothesis" for membrane fusion have been characterized in the acrosome of several mammalian species, and a functional role for these proteins during the acrosome reaction has been proposed. We have investigated the presence of SNAREs in equine sperm, using semen samples obtained from stallions with varying fertility. Immunocytochemical analysis revealed that members of different SNARE families can be detected on the acrosome of equine sperm, notably in the acrosomal cap and equatorial segment. These proteins include the t-SNARE syntaxin, the v-SNARE synaptobr...
Expression and subcellular localization of the mu-opioid receptor in equine spermatozoa: evidence for its functional role. The development of fertilizing ability in sperm cells is associated with changes in the plasma membrane. However, to date the exact nature of sequentially activated primary receptors and channels and the signal transduction pathways derived from these remains elusive. We analyzed the expression and localization of the mu-opioid receptor in equine spermatozoa. A transcript corresponding to the third extracellular loop that selectively binds mu agonists was amplified, sequenced and compared with the known sequences in humans, rats and cattle. The amplification product showed a high degree of nuc...
Effects of glutamine on post-thaw motility of stallion spermatozoa: an approach of the mechanism of action at spermatozoa level. The cryoprotective effect of l-glutamine and an approach of its mechanism of action, in preserving motility of stallion spermatozoa during the freezing-thawing process, were studied. In Experiment 1, thirty-six ejaculates were collected from six stallions (two good, two middle, and two of poor sperm freezability) and were diluted with 10 different freezing media derived from INRA 82 medium supplemented with 20 mM HEPES and 2% (v/v) centrifuged egg yolk (BM). After thawing, sperm motility was evaluated by a computer-assisted semen motility analyser. The effects of glutamine and glycerol at diff...
Hysteroscopic or rectally guided, deep-uterine insemination of mares with spermatozoa stored 18 h at either 5 degrees C or 15 degrees C prior to flow-cytometric sorting. Practical application of sex-selected spermatozoa in the horse industry would be greatly improved by the ability to develop simplified methods for shipping, storing, and inseminating sex-selected spermatozoa. Acceptable pregnancy rates have been achieved using fresh sex-sorted stallion sperm, however many stallion owners are reluctant to send their stallions to the sorter location for collection during the breeding season. Furthermore, the technology would be more applicable if the hysteroscopic insemination technique was not necessary for adequate pregnancy rates. Hysteroscopic insemination r...
Step-wise dilution for removal of glycerol from fresh and cryopreserved equine spermatozoa. Osmotic stress is an important component of the damage to spermatozoa during cryopreservation. Osmotic injury, due to hyperosmolar freezing extenders, changes in relative solute concentration in the extra cellular medium during freezing and differences in the relative permeabilities of penetrating cryoprotectants, such as glycerol, and water occur when cryopreserved spermatozoa are diluted into isosmotic media or when spermatozoa are placed in the female reproductive tract. The purpose of the study reported here was to evaluate the effect of step-wise dilution for the removal of the permeating...
Evaluation of alternative cryoprotectants for preserving stallion spermatozoa. Although use of cryopreserved stallion spermatozoa is currently accepted by many breed registries, utilization of this technique remains limited due to poor fertility for some stallions. One reason for these results is osmotic stress that spermatozoa experiences when the cryoprotectant (glycerol) is added to the cells prior to freezing and removal from the cells after thawing. In an effort to minimize osmotic damage, alternative cryoprotectants, having lower molecular weights and greater membrane permeability than glycerol, were evaluated to determine their effectiveness for cryopreserving sta...
Low dose insemination in the mare: an update. The generally recommended minimum number of spermatozoa required for conventional artificial insemination in the mare is in excess of 200 x 10(6) progressively motile spermatozoa. Recent developments in different insemination techniques such as deep uterine, hysteroscopic and oviductal insemination, which have been designed to use significantly fewer spermatozoa, are reviewed in this paper. A number of studies have demonstrated that ultrasound guided deep uterine insemination of 5 x 10(6) fresh spermatozoa can produce satisfactory pregnancy rates. The use of hysteroscopic insemination enables ...
Oocyte transfer and gamete intrafallopian transfer in the mare. Methods for the collection and transfer of equine oocytes have been developed, and uses of these techniques have resulted in new clinical and research possibilities. Because oocyte transfer avoids reproductive problems associated with the oviduct, uterus, and cervix, pregnancies can be produced from many mares that cannot carry a pregnancy or produce embryos. Oocytes for clinical transfers are usually collected from preovulatory follicles and cultured for a short interval or transferred directly into a recipient's oviduct. For oocyte transfer, the recipient is inseminated within the uterus. A ...
[The influence of centrifugation on quality and freezability of stallion semen]. The aim of the present study was to investigate the influence of various centrifugation methods on sperm loss and quality of frozen-thawed semen. From at a total of 8 Warmblood stallions of the National Stud Farm in Avenches, 3 ejaculates each were collected and seminal plasma was removed using 3 different centrifugation regimes. In method I (reference method) centrifugation occurred by a speed of 600 x g during 10 minutes. In method II 1000 x g was used during 2 minutes while in method III centrifugation was performed by 2000 x g during 2 minutes. After centrifugation 90%, of the supernatant ...
Effect of L-carnitine administration on the seminal characteristics of oligoasthenospermic stallions. The effect of orally administered l-carnitine on the quality of semen obtained from stallions with different semen qualities was investigated. Four stallions with proven fertility (high motility group, HM) and with normal seminal characteristics (>50% progressive motility and > 80 x 10(6) spermatozoa/ml), and four questionable breeders (low motility group, LM) with <50% of sperm progressive motility and < 80 x 10(6) spermatozoa/ml, received p.o. 20 g of l-carnitine for 60 days. Blood and semen samples were collected before treatment (T0) and after 30 (T1) and 60 days (T2). Semen ev...
