Spermatogenesis in horses is the process by which spermatozoa are produced in the testes. This complex sequence of events involves the proliferation and differentiation of germ cells, supported by Sertoli cells and influenced by hormonal regulation. Key stages include spermatogonial mitosis, meiosis, and spermiogenesis, resulting in the formation of mature sperm cells. Hormones such as testosterone and follicle-stimulating hormone (FSH) are integral to regulating this process. This topic encompasses research studies and scholarly articles that explore the mechanisms, hormonal influences, and factors affecting spermatogenesis in horses, as well as its implications for equine reproduction and breeding management.
Salamone DF, Canel NG, Rodríguez MB.Intracytoplasmic sperm injection (ICSI) has become a useful technique for clinical applications in the horse-breeding industry. However, both ICSI blastocyst and offspring production continues to be limited for most farm and wild species. This article reviews technical differences of ICSI performance among species, possible biological and methodological reasons for the variable efficiency and potential strategies to improve the outcomes. One of the major applications of ICSI in animal production is the reproduction of high-value specimens. Unfortunately, some domestic species like the bovine s...
Ketchum CC, Larsen CD, McNeil A, Meyer-Ficca ML, Meyer RG.Chromatin remodeling during spermatogenesis culminates in the exchange of nucleosomes for transition proteins and protamines as an important part of spermatid development to give rise to healthy sperm. Comparative immunofluorescence analyses of equine and murine testis histological sections were used to characterize nucleoprotein exchange in the stallion. Histone H4 hyperacetylation is considered a key event of histone removal during the nucleoprotein transition to a protamine-based sperm chromatin structure. In the stallion, but not the mouse, H4 was already highly acetylated in lysine residu...
Aurich C, Ortega Ferrusola C, Peña Vega FJ, Schrammel N, Morcuende D, Aurich J.Spermatozoa contain polyunsaturated fatty acids (PUFA). Cryopreservation damages sperm membranes and they become less functional after thawing. We analysed the lipid composition of spermatozoa from Shetland stallions (n = 15) collected monthly from January to June and hypothesized that sperm lipid patterns change with season. In addition, one ejaculate per month was submitted to cryopreservation. Content of saturated palmytic and stearic acid decreased from January to March (p < 0.001) while content of the PUFA docosapentaenoic (p < 0.001) and arachidonic acid (p < 0.05) and total PUFA...
Ghallab AM, Shahat AM, Fadl AM, Ayoub MM, Moawad AR.The aim of the present study was to evaluate the effects of supplementation of semen extender with various non-enzymatic antioxidants on the quality of cooled or cryopreserved Arabian stallion spermatozoa. Semen collected from four pure Arabian stallions was centrifuged at 600g for 15 min. Spermatozoa were then diluted in INRA-82 extender supplemented with bovine serum albumin (BSA; 0, 10, 15 and 20 mg/mL) or trehalose (0, 75, 100 and 150 mM) or zinc sulphate (0, 100, 150 and 200 μM). The diluted semen was then either cooled at 5 °C or cryopreserved in 0.5-ml plastic straws. After cooli...
Costa GMJ, Avelar GF, Lacerda SMSN, Figueiredo AFA, Tavares AO, Rezende-Neto JV, Martins FGP, França LR.The establishment of proper conditions for spermatogonial stem cells (SSCs) cryopreservation and storage represents an important biotechnological approach for the preservation of the genetic stock of valuable animals. This study demonstrates the effects of different cryopreservation protocols on the survival rates and phenotypic expression of SSCs in horses. The cells were enzymatically isolated from testes of eight adult horses. After enrichment and characterization of germ cells in the suspension, the feasibility of several cryopreservation protocols were evaluated. Three different cryomedia...
Pricking S, Bollwein H, Spilker K, Martinsson G, Schweizer A, Thomas S, Oldenhof H, Sieme H.Accurate determination of the testes volume and prediction of the daily sperm output (DSO) is valuable information for reproductive management of a stallion. The aim of this study was to compare different methods for measuring the testes volume, including caliper, 2D and 3D ultrasound. Special emphasis was on feasibility of 3D volume analysis. First, 22 castrated testes were measured and derived volumes were compared with volumes determined via volume displacement in a graded cylinder with saline solution. Then, during the breeding season, testes sizes of 52 stallions were measured in vivo an...
Hidalgo M, Ortiz I, Dorado J, Morrell JM, Gosálvez J, Consuegra C, Diaz-Jimenez M, Pereira B, Crespo F.The aims of this study were to: 1) develop a new method for stallion sperm selection using a modified swim-up procedure through a colloid and 2) evaluate its impact in good quality ejaculates from bad freezers in comparison to methods involving centrifugation such as single layer centrifugation and sperm washing. Ejaculates were processed before freezing using three different procedures: sperm washing (SW), colloid single layer centrifugation (SLC) and a modified colloid swim-up (SU). After semen processing, sperm recovery rates were measured and sperm were frozen. Post-thaw sperm motility (as...
Ortiz-Rodriguez JM, Anel-Lopez L, Martín-Muñoz P, Álvarez M, Gaitskell-Phillips G, Anel L, Rodríguez-Medina P, Peña FJ, Ortega Ferrusola C.Testicular function is particularly susceptible to vascular insult, resulting in a negative impact on sperm production and quality of the ejaculate. A prompt diagnosis of testicular dysfunction enables implementation of appropriate treatment, hence improving fertility forecasts for stallions. The present research aims to: (1) assess if Doppler ultrasonography is a good tool to diagnose stallions with testicular dysfunction; (2) to study the relationship between Doppler parameters of the testicular artery and those of sperm quality assessed by flow cytometry and (3) to establish cut off values ...
Wach-Gygax L, Burger D, Malama E, Bollwein H, Fleisch A, Jeannerat E, Thomas S, Schuler G, Janett F.In this study annual fluctuations of DNA fragmentation and quality of cold-stored equine sperm were evaluated. Ejaculates were collected weekly during one year from 15 stallions. Ejaculate volume, sperm concentration and total sperm count were determined and semen was then extended and cold-stored for 48 h. Sperm motility was evaluated by CASA before and after 24 as well as 48 h of cold storage. In addition, the percentages of sperm with intact plasma membrane and acrosome (PMAI %) and with low intracellular Ca2+ level were determined in cold-stored semen (24 h, 48 h). SCSA™ was performe...
Schmidt K, Deichsel K, de Oliveira RA, Aurich J, Ille N, Aurich C.We hypothesized that housing of stallions in a thermoneutral temperature zone during autumn and winter does not only influence metabolism and hair shedding but also improves the characteristics of raw and processed semen. Fertile Shetland pony stallions were followed from October to June. This time coincided with the seasons autumn, winter and spring. Ponies were kept in outside paddocks (group CON, n = 8) or in indoor stables (group ST, n = 8) from October to March when ST stallions returned to outdoor paddocks, but ponies remained in the same groups. The rectal temperature was measured o...
Hall SE, Aitken RJ, Nixon B, Smith ND, Gibb Z.Oxidative stress is a major determinant of mammalian sperm function stimulating lipid peroxidation cascades that culminate in the generation of potentially cytotoxic aldehydes. The aim of this study was to assess the impact of such aldehydes on the functionality of stallion spermatozoa. The impact of exposure to exogenous acrolein (ACR) and 4-hydroxynonenal (4HNE) was manifested in a highly significant dose- and time-dependent increase in mitochondrial reactive oxygen species (ROS), total cellular ROS, a decrease in sperm motility, and a time-dependent increase in lipid peroxidation. Notably, ...
Jeannerat E, Janett F, Sieme H, Wedekind C, Burger D.The theory of ejaculate economics was mainly built around different sperm competition scenarios but also predicts that investments into ejaculates depend on female fecundity. Previous tests of this prediction focused on invertebrates and lower vertebrate, and on species with high female reproductive potential. It remains unclear whether the prediction also holds for polygynous mammals with low female reproductive potential (due to low litter size and long inter-birth intervals). We used horses (Equus caballus) to experimentally test whether semen characteristics are adjusted to the oestrous cy...
Pessoa GA, Martini AP, Trentin JM, Minela T, Fiorenza MF, Rubin MIB.The aim of this study was to compare the sperm separation technique using filtration through glass wool compared with just diluted cooled semen. Eighteen ejaculates were collected from 6 pony stallions of the Brazilian pony breed. Evaluations were done on pH, osmolarity, total motility, membrane functionality (HOST), membrane integrity (CFDA/PI), morphology and mitochondrial viability (MTT) in fresh, 24 and 48 h of cooled semen at 5°C. After dilution, the half of the extended semen was cooled (control group). The other half was cooled after filtration trough glass wool (filtered group). Reta...
Darr CR, Moraes LE, Connon RE, Love CC, Teague S, Varner DD, Meyers SA.Mitochondrial DNA (mtDNA) copy number has been utilized as a measure of sperm quality in several species including mice, dogs, and humans, and has been suggested as a potential biomarker of fertility in stallion sperm. The results of the present study extend this recent discovery using sperm samples from American Quarter Horse stallions of varying age. By determining copy number of three mitochondrial genes, cytochrome b (CYTB), NADH dehydrogenase 1 (ND1) and NADH dehydrogenase 4 (ND4), instead of a single gene, we demonstrate an improved understanding of mtDNA fate in stallion sperm mitochond...
Talluri TR, Mal G, Ravi SK.To investigate various biochemical components of seminal plasma in Marwari stallions and Poitou Jacks and to find out their correlation with that of the seminal characteristics. Methods: In this study, semen was collected from six Marwari stallions and six Poitou jacks aged from 4 to 6 years and with known fertility status. The semen collection from the stallions were collected during the breeding season, i.e., between the months of April and June. From the collected semen ejaculates, we estimated the values of some biochemical components, viz., total protein content, total lipid content, and ...
Canesin HS, Brom-de-Luna JG, Choi YH, Ortiz I, Diaw M, Hinrichs K.We evaluated the meiotic and developmental competence of GV-stage equine oocytes vitrified under different conditions. In a preliminary study, using dimethyl sulfoxide (D), ethylene glycol (EG) and sucrose (S) as cryoprotectants, the maturation rate was higher for cumulus-oocyte complexes (COCs) held overnight before vitrification (37%) than for those vitrified immediately (14%; P < 0.05). Thereafter, all COCs were held overnight before vitrification. In Experiment 1 we compared 1 min (1m) and 4 min (4m) exposure to vitrification and warming solutions; oocytes that subsequently matured wer...
Berlinguer F, Pasciu V, Succu S, Cossu I, Caggiu S, Addis D, Castagna A, Fontani V, Rinaldi S, Passino ES.REAC technology (acronym for Radio Electric Asymmetric Conveyor) is a technology platform for neuro and bio modulation. It has already proven to optimize the ions fluxes at the molecular level and the molecular mechanisms driving cellular asymmetry and polarization. Methods: This study was designed to verify whether this technology could extend spermatozoa life-span during liquid storage, while preserving their functions, DNA integrity and oxidative status. At 0, 24, 48, and 72 h. of storage at 4 °C, a battery of analyses was performed to assess spermatozoa viability, motility parameters, a...
Zeng W, Alpaugh W, Stefanovski D, Schlingmann K, Dobrinski I, Turner RM.The study of spermatogenesis in the horse is challenging because of the absence of an in vitro system that is capable of reproducing efficient spermatogenesis and because of the difficulties and costs associated with performing well-controlled studies in vivo. In an attempt to develop novel methods for the study of equine spermatogenesis, we tested whether cells from enzymatically digested pre-pubertal equine testicular tissue were capable of de novo tissue formation and spermatogenesis following xenografting under the back skin of immunocompromised mice. Testes were obtained from normal pre...
Waddington B, Penitente-Filho JM, Neves J, Pinho RO, Chaya AY, Maitan PP, Silveira CO, Neves MG, Guimarães S, de Carvalho GR, Guimarães JD.This study was conducted to characterize the daily profile of testosterone secretion and its mean concentrations in the four seasons as well as to evaluate the semen characteristics and testicular biometry of Mangalarga Marchador stallions throughout the year in a tropical region. Three stallions were submitted to semen collections and evaluation of testicular biometry every 14 days along a year. Blood samples were collected once at the middle of each season, in a 20-min interval during 24 hr in order to evaluate the testosterone secretion profiles among seasons. Testosterone concentrations ...
Lee YS, Jung HJ, Yoon MJ.Putative markers for each specific germ cell stage can be a useful tool to study the fate and functions of these cells. Undifferentiated embryonic cell transcription factor 1 (UTF1) is a putative marker for undifferentiated spermatogonia in humans, rats and horses. The deleted in azoospermia-like (DAZL) protein is also expressed by differentiated spermatogonia and primary spermatocytes in several species. However, whether the expression patterns of these molecular markers are identical and applicable to donkeys remains to be elucidated. The objective of this study was to investigate the expres...
Bucci D, Spinaci M, Mislei B, Gadani B, Rizzato G, Love CC, Tamanini C, Galeati G, Mari G.Stallion semen storage for artificial insemination is mainly based on liquid cooled storage. In many stallions this technique maintains sperm quality for an extended period of time (24-72 hr) at 7°C. While this technique is commonly used in the horse industry, there can be a decline in fertility in some stallions, due to an inability of their sperm to tolerate the cool storage process. The aim of the present work was to evaluate the effect of two natural antioxidants (epigallocatechin-3-gallate (EGCG) at 20, 60 and 120 μm and green tea polyphenols, and p at .001, .01 and .1 mg/ml) on some...
Davila MP, Muñoz PM, Bolaños JM, Stout TA, Gadella BM, Tapia JA, da Silva CB, Ferrusola CO, Peña FJ.To investigate the hypothesis that oxidative phosphorylation is a major source of ATP to fuel stallion sperm motility, oxidative phosphorylation was suppressed using the mitochondrial uncouplers CCCP and 2,4,-dinitrophenol (DNP) and by inhibiting mitochondrial respiration at complex IV using sodium cyanide or at the level of ATP synthase using oligomycin-A. As mitochondrial dysfunction may also lead to oxidative stress, production of reactive oxygen species was monitored simultaneously. All inhibitors reduced ATP content, but oligomycin-A did so most profoundly. Oligomycin-A and CCCP also sign...
Lee G, Jung H, Yoon M.The molecular markers for specific germ cell stages can be utilized for identifying, monitoring, and separating a particular stage of germ cells. The RNA-binding protein Lin28 is expressed in gonocytes of human fetal testes. The Lin28 expression is restricted to a very small population of spermatogonial cells in human, mice, and monkey. The main objective of this study was to investigate the expression pattern of Lin28 in stallion testes at different reproductive stages. Based on the presence or absence of full spermatogenesis and lumina in seminiferous tubules, the testicular samples were cat...
Pinyopummin A, Mahasawangkul S, Kornkaewrat K, Rattanapirom S, Leartsang W, Kitkha S.The effects of seminal plasma (SP), derived from autologous, homologous and heterologous species (stallion, boar and dog) on chilled Asian elephant sperm quality, were determined. Semen was collected from eight males and samples with ≥30% motile spermatozoa were used in the study. Semen was diluted with Tris-glucose-egg yolk extender, supplemented with different SP types and preserved at 4°C for 48 hr. Experiment 1 (n = 31), showed that the presence of SP (autologous) helped to preserve sperm quality in terms of sperm motility and acrosome integrity (p < .05). Homologous SP did not ...
Gottschalk M, Sieme H, Martinsson G, Distl O.A high quality of stallion semen is of particular importance for maximum reproductive efficiency. In the present study, we estimated the relationships among estimated breeding values (EBVs) of semen traits and EBVs for the paternal component of the pregnancy rate per estrus cycle (EBV-PAT) for 100 German Warmblood stallions using correlation and general linear model analyses. The most highly correlated sperm quality trait was total number of progressively motile sperm (r = 0.36). EBV-PAT was considered in three classes with stallions 1 SD below (<80), around (80-120), and above (>120) the popu...
Ellinger I.Active placental transport of maternal serum calcium (Ca(2+)) to the offspring is pivotal for proper development of the fetal skeleton as well as various organ systems. Moreover, extracellular Ca(2+) levels impact on distinct processes in mammalian reproduction. The calcium-sensing receptor (CaSR) translates changes in extracellular Ca(2+)-concentrations into cellular reactions. This review summarizes current knowledge on the expression of CaSR and its putative functions in reproductive organs. CaSR was detected in placental cells mediating materno-fetal Ca(2+)-transport such as the murine int...
Bucci D, Giaretta E, Merlo B, Iacono E, Spinaci M, Gadani B, Mari G, Tamanini C, Galeati G.Alkaline phosphatase (AP) is present in equine seminal plasma and spermatozoa, but its functional role is not fully understood yet. Being that, sperm-oocyte interaction in equine species has been reported to be enhanced at a slightly basic pH, this work aimed at verifying whether exogenous alkaline phosphatase exerts any role on stallion spermatozoa and sperm-oocyte interaction at different pHs (7.4; 8.0; 9.0). Stallion spermatozoa were capacitated in Tyrode's medium at pH 7.4, 8.0, and 9.0 for 4 hours at 38 °C, 5% CO with 2.5-IU AP (AP group) or without AP (capacitated spermatozoa group); ...
Jung H, Kim N, Yoon M.The main objective of this study was to evaluate the efficacy of an additional cryoprotectant in 10% dimethyl sulfoxide (DMSO) on cryopreserving germ cells from stallions at different reproductive stages. Testicular samples were obtained from pre-pubertal (1-1.5 yr, n=6) and post-pubertal (3-7 yr, n=5) stallions. Germ cells were isolated using a two-enzyme digestion procedure and cryopreserved in minimal essential medium alpha containing 10% fetal bovine serum and 10% DMSO with or without addition of trehalose (50, 100, or 200mM) or polyethylene glycol (PEG, 2.5, 5, or 10%). Viability, cell po...
Davolli GM, Ball BA, Esteller-Vico A, Claes AN, Canisso IF, Fedorka CE, Woodward EM, Troedsson MH, Squires EL.The GnRH antagonist, acyline, has not yet been investigated in the stallion. Our study aimed to: (1) evaluate the downregulation of the stallion hypothalamic-pituitary-gonadal axis by acyline through assessment of seminal parameters, testicular volume, and sexual behavior; (2) assess hormonal response of acyline-treated stallions to GnRH stimulation; and (3) verify reversibility after treatment. Stallions were assessed pretreatment and subsequently treated (every five days) for 50 days: acyline (n = 4; 330 μg/kg acyline) or control (n = 4, vehicle). The stallions were then monitored for 62...
Muñoz PM, Ferrusola CO, Lopez LA, Del Petre C, Garcia MA, de Paz Cabello P, Anel L, Peña FJ.Stallion-to-stallion variability in the quality of cryopreserved ejaculates postthaw affects the commercial acceptability of frozen semen and thus is a major constraint for the equine industry. In recent years, the molecular mechanisms associated with sperm damage during cryopreservation have become better understood. Identification of the freezability of the ejaculates before the freezing process is initiated will have a major impact on the equine industry. We studied three markers of oxidative stress in sperm, including 8-iso-PGF2alpha, 8-OH guanosine, and 4-hydroxynonenal (4-HNE); the prese...
Tanaka Y, Taniyama H, Tsunoda N, Shinbo H, Nagamine N, Nambo Y, Nagata S, Watanabe G, Herath CB, Groome NP, Taya K.Immunolocalization of the inhibin (a) and inhibin/activin (beta3A and betaB) subunit proteins in equine fetal testes was investigated to determine the ability of the fetal testis to produce inhibins at 120, 150, 200, and 250 days of gestation. In addition, concentrations of immunoreactive (ir)-inhibin, inhibin pro-aC, and inhibin A in both the maternal and fetal circulation were measured. It was found that plasma concentrations of ir-inhibin, inhibin pro-alphaC, and inhibin A were much higher (P < .05) in the fetal than in the maternal circulation at any stage of gestation examined. Similar...
Swegen A, Smith ND, Gibb Z, Curry BJ, Aitken RJ.Serine proteases are emerging as important players in the spermatozoon's acquisition of functional competence. This study aimed to characterize the serine protease testisin (PRSS21) in stallion spermatozoa, examining its surface expression, possible origins in the testis and epididymis, and changes in response to capacitation and acrosome reaction, as well as its capacity to form high molecular weight complexes and interact with other proteins. The role of serine proteases in spontaneous capacitation and acrosome reaction of stallion spermatozoa was established using the serine protease inhibi...
Rota A, Furzi C, Panzani D, Camillo F.This study on extended, cooled stallion spermatozoa aimed to compare the ability of three extenders to maintain sperm motility during 24 h of preservation, and to describe pregnancy and foaling rates after artificial insemination (AI) of stallion spermatozoa stored and transported in the extender chosen from the in vitro study. After 6 and 24 h of preservation, motility, both subjective and evaluated by the motility analyzer (total, progressive and rapid), was lower in non-fat, dried skim milk-glucose than in both other extenders: dried skim milk-glucose added to 2% centrifuged egg yolk, and u...
Lee G, Jung H, Yoon M.The molecular markers for specific germ cell stages can be utilized for identifying, monitoring, and separating a particular stage of germ cells. The RNA-binding protein Lin28 is expressed in gonocytes of human fetal testes. The Lin28 expression is restricted to a very small population of spermatogonial cells in human, mice, and monkey. The main objective of this study was to investigate the expression pattern of Lin28 in stallion testes at different reproductive stages. Based on the presence or absence of full spermatogenesis and lumina in seminiferous tubules, the testicular samples were cat...
Samper JC, Hellander JC, Crabo BG.Studies were designed to investigate whether sperm motility determined with a Hamilton-Thorn HTM-2000 motility analyzer (HTM), or the percentage of spermatozoa that passed through glass wool (GW), Sephadex (S), or glass wool/Sephadex (GWS) filters could be used to predict the fertilizing potential of fresh or frozen semen. In the fresh semen study, 10 randomly selected ejaculates from 4 stallions exclusively used for A.I. breeding were assayed during the season. The 521 mares used were inseminated with 500 x 10(6) motile spermatozoa after gynaecological examination every 2 days. In the frozen ...
Herrera-Luna CV, Scarlet D, Walter I, Aurich C.The aim of the present study was to characterise receptors for LH and FSH (LHR and FSHR, respectively) and aromatase in epididymal and testicular tissue from stallions of different ages (prepubertal, young, mature and old). Gene and protein expression were assessed by real-time quantitative polymerase chain reaction (real-time qPCR), immunohistochemistry and multiple immunofluorescence labelling. There were no differences in LHR mRNA expression in epididymal and testicular parenchyma in stallions of different age. In contrast, expression of FSHR and CYP19A1 in caput, corpus and cauda epididymi...
Campos JR, Breheny P, Araujo RR, Troedsson MH, Squires EL, Timoney PJ, Balasuriya UB.Equine arteritis virus (EAV) is the causal agent of equine viral arteritis (EVA), a respiratory and reproductive disease of equids. Some strains of EAV can cause fever, leukopenia, and dependent edema of the limbs, scrotum, and preputium in the acutely infected stallion. We hypothesized that fever and scrotal edema observed during the acute phase of the infection, but not the presence of EAV, have an adverse effect on semen quality. A group of seven stallions were intranasally inoculated with the Kentucky 84 (KY84) strain of EAV. Stallions were monitored for clinical signs of EVA until 42 days...
Alghamdi AS, Funnell BJ, Bird SL, Lamb GC, Rendahl AK, Taube PC, Foster DN.We performed a series of comparative studies of bull and stallion seminal plasma (SP) and its role on sperm-neutrophil binding as well as the interaction between semen extender and seminal DNase. Because of contrasting roles of SP on sperm-neutrophil binding between horses and cattle, it was suspected there were some species-specific differences on sperm interaction with SP proteins due to the variations in the natural location of semen deposition (uterus compared to vagina). Bull frozen-thawed sperm removed from egg yolk extender showed similar results to fresh sperm, but this also caused ext...
Ruiz AJ, Tibary A, Heaton RA, Hargreaves IP, Leadon DP, Bayly WM.Although coenzyme Q10 (CoQ10) serves as an antioxidant and energy source for spermatozoa when added to stallion semen before cooling or freezing, the effects of feeding CoQ10 on semen quality have not been studied. We assessed the effects of daily oral ingestion of CoQ10-ubiquinol by stallions on their plasma CoQ10 concentrations and semen quality. Seven mature Andalusian stallions ate 1g ubiquinol/day for 4 weeks followed by a 4-week washout period. Four horses initially completed an additional 4-week control period without ubiquinol. Blood was sampled weekly for determination of plasma CoQ1...
Schmidt K, Deichsel K, de Oliveira RA, Aurich J, Ille N, Aurich C.We hypothesized that housing of stallions in a thermoneutral temperature zone during autumn and winter does not only influence metabolism and hair shedding but also improves the characteristics of raw and processed semen. Fertile Shetland pony stallions were followed from October to June. This time coincided with the seasons autumn, winter and spring. Ponies were kept in outside paddocks (group CON, n = 8) or in indoor stables (group ST, n = 8) from October to March when ST stallions returned to outdoor paddocks, but ponies remained in the same groups. The rectal temperature was measured o...
Miller CD.This manuscript is a review of current protocols, advantages, and disadvantages of breeding mares with frozen-thawed equine semen. Issues affecting pregnancy rates are discussed, including proper mare selection, induction of ovulation, insemination dose, timing of insemination (single-dose versus multiple-dose insemination), methods of insemination (transrectal-guided deep-horn versus hysteroscopic insemination), and post-insemination mare management procedures. In a retrospective analysis of breeding records, a single-dose of frozen-thawed semen was inseminated within 6h post-ovulation; the p...
Suliman Y, Becker F, Tuchscherer A, Wimmers K.Horses are seasonal breeders with a natural breeding season beginning in spring and extending through midsummer. In this study, quantitative and qualitative parameters of chilled stallion semen were compared between fertile and subfertile stallions and between the breeding and the non-breeding season. Semen quality parameters compared included ejaculate volume, sperm concentration, total sperm number, sperm morphology, and computer-assisted semen analysis (CASA)-derived sperm movement characteristics obtained from two groups of warmblood stallions ( ; four fertile stallions and four subfert...
Morrell JM, Mari G, Kútvölgyi G, Meurling S, Mislei B, Iacono E, Rodriguez-Martinez H.Some stallions produce ejaculates of low quality and/or low fertility when used for artificial insemination (AI). The purpose of these five case studies was to use Single Layer Centrifugation (SLC) to select the best spermatozoa from 'problem' ejaculates for subsequent use in AI. Sperm quality, in terms of motility, morphology and chromatin integrity, was improved in the SLC-selected samples compared to the corresponding uncentrifuged samples, with the exception of one stallion thought to have ampullary stasis. In this stallion, neither the incidence of spermatozoa with detached heads nor the ...
Pricking S, Bollwein H, Spilker K, Martinsson G, Schweizer A, Thomas S, Oldenhof H, Sieme H.Accurate determination of the testes volume and prediction of the daily sperm output (DSO) is valuable information for reproductive management of a stallion. The aim of this study was to compare different methods for measuring the testes volume, including caliper, 2D and 3D ultrasound. Special emphasis was on feasibility of 3D volume analysis. First, 22 castrated testes were measured and derived volumes were compared with volumes determined via volume displacement in a graded cylinder with saline solution. Then, during the breeding season, testes sizes of 52 stallions were measured in vivo an...
Urra JA, Villaroel-Espíndola F, Covarrubias AA, Rodríguez-Gil JE, Ramírez-Reveco A, Concha II.Dopamine is a catecholamine with multiple physiological functions, playing a key role in nervous system; however its participation in reproductive processes and sperm physiology is controversial. High dopamine concentrations have been reported in different portions of the feminine and masculine reproductive tract, although the role fulfilled by this catecholamine in reproductive physiology is as yet unknown. We have previously shown that dopamine type 2 receptor is functional in boar sperm, suggesting that dopamine acts as a physiological modulator of sperm viability, capacitation and motility...
Chandley AC, Short RV, Allen WR.A detailed investigation of testicular meiosis in a mule, a hinny and a Przewalski horse/domestic horse hybrid were made. Abnormalities of pairing were observed in the mule and hinny in most germ cells at the pachytene stage of meiotic prophase, and spermatogenesis was alsmot totally arrested. A few mature spermatozoa were recovered from the ejaculate and epididymal flushings of the hinny. The Przewalski horse/domestic horse hybrid was fertile and showed normal spermatogenesis. Chromosome banding studies showed a close homology between the karyotypes of the Prezwalski horse (Equus przewalskii,...
Choi YH, Love CC, Chung YG, Varner DD, Westhusin ME, Burghardt RC, Hinrichs K.We investigated the use of direct nuclear injection using the Piezo drill and activation by injection of stallion sperm cytosolic extract for production of cloned equine embryos. When metaphase II horse oocytes were injected with either of two dosages of sperm extract and cultured 20 h, similar activation rates (88% vs. 90%) and cleavage rates (49% vs. 46%) were obtained. The successful reconstruction rate of horse oocytes with horse somatic cell donor nuclei after direct injection using the Piezo drill was 82%. Four dosages of sperm extract (containing 59, 176, 293, or 1375 microg/ml protein)...
Magdaleno L, Gasset M, Varea J, Schambony AM, Urbanke C, Raida M, Töpfer-Petersen E, Calvete JJ.HSP-3 is a member of the cysteine-rich secretory protein (CRISP) family from stallion seminal plasma. We report a large-scale purification protocol for native HSP-3. This protein is a non-glycosylated polypeptide chain with a pI of 8-9 and an isotope-averaged molecular mass of 24987 +/- 3 Da. The molecular mass of HSP-3, determined by equilibrium sedimentation, is 26 kDa, showing that the protein exists in solution as a monomer. The concentration of HSP-3 in the seminal plasma of different stallions ranged from 0.3 to 1.3 mg/ml. On average, 0.9-9 million HSP-3 molecules/cell coat the postacros...
Burger D, Dolivo G, Marti E, Sieme H, Wedekind C.Odours of vertebrates often contain information about the major histocompatibility complex (MHC), and are used in kin recognition, mate choice or female investment in pregnancy. It is, however, still unclear whether MHC-linked signals can also affect male reproductive strategies. We used horses (Equus caballus) to study this question under experimental conditions. Twelve stallions were individually exposed either to an unfamiliar MHC-similar mare and then to an unfamiliar MHC-dissimilar mare, or vice versa. Each exposure lasted over a period of four weeks. Peripheral blood testosterone levels ...
Gamboa S, Rodrigues AS, Henriques L, Batista C, Ramalho-Santos J.A group of stallions with different reproductive indexes were used to study seasonal variations in sperm quality (Equus caballus). Semen samples were collected from late September to July and analyzed according to four seasonal periods: late September-December, January-March, late March-May, and June-July. Parameters monitored included sperm concentration, sperm motility, sperm morphology, sperm viability, acrosomal status, plasma membrane stability, and sperm mitochondrial membrane potential. Overall, seminal parameters monitored are affected mostly by time period, followed by animal and last...
Ebel F, Vallejos A, Gajardo G, Ulloa O, Clavel E, Rodríguez-Gil JE, Ramírez-Reveco A.The aim of this study was to evaluate seasonal changes in basic parameters of sperm quality and freezability behaviour of ejaculates from 10 fertile heavy draft stallions. A total of 140 ejaculates were collected, processed and evaluated during both the breeding (September-November) and non-breeding seasons (April-June). Fresh semen was evaluated for volume, concentration, total spermatozoa per ejaculate, plasma membrane integrity and total sperm motility. Cryopreserved samples were evaluated for plasma membrane integrity and sperm motility by the CASA system, and for the freezability index (F...
Han H, Chen Q, Gao Y, Li J, Li W, Dang R, Lei C.In the biological process of testicular spermatogenesis, the expression and interaction of many genes are regulated by microRNAs (miRNAs). However, comparisons of miRNA expression between descended testes (DTs) and undescended testes (UDTs) are rarely done in horses. In this study, we selected two UDTs (CKY2b and GU4b) from Chakouyi (CKY) and Guanzhong (GU) horses and eight DTs (GU1-3, CKY1, CKY3, CKY2a, GU4a, and GU5). Three groups were compared to evaluate expression patterns of testicular miRNA in stallion testes. Group 1 compared normal CKY horses and GU horses (CKY1 and CKY3 vs. GU1-3). G...
D'Anza E, Albarella S, Galdiero G, Tafuri S, Del Prete C, Cocchia N, Ciani F, Mastellone V, Pasolini MP, Carotenuto D, Selvaggi M, Ciotola F....The reproductive performances of livestock play an essential role in the economic management of the farm. The improvement of semen quantity and quality through the use of food supplements that lack substances which are forbidden in animal feeding, or that may have detrimental effects, is an important goal. Maca (Lepidium meyenii) is a plant that has been used for centuries in the Andes for nutrition and fertility enhancement in humans and animals. The aim of this study was to evaluate the effects of food supplementation of stallions with maca during the breeding season on spermatozoa parameter...
Leme DP, Papa FO.Fifteen stallions of different breeds, age 3-11 years, had their right testicles evaluated by fine needle aspiration cytology (FNAC). Cytological analysis showed the following spermatogenic cell types: spermatogonia (1.6% +/- 1.1); spermatocyte I (3.4% +/- 2.2); spermatocyte II (0.8% +/- 0.7); early spermatids (25.5% +/- 9.5); late spermatids (37.0% +/- 9.3). Spermatozoal numbers were expressed as the spermatic index (SI = 31.5% +/- 8.5) and Sertoli cells were expressed as the Sertoli cell index (SEI = 20.9% +/- 17.0) (means +/- s.d). Identification of cell types was relatively easy and no imm...
McDonnell SM.In summary, important events of ejaculation include emission of sperm and the accessory gland fluids into the urethra, simultaneous closure of the bladder neck, and forceful ejaculation of the combined semen through the urethra. Emission and bladder neck closure are primarily alpha-adrenergically mediated thoracolumbar sympathetic reflex events with supraspinal modulation. Ejaculation is a sacralspinal reflex mediated by the pudendal nerve. In stallions, the most common ejaculation disorders are emission and ejaculation failure, and urine contamination of semen. Rare disorders are azoospermia ...
Lemazurier E, Moslemi S, Sourdaine P, Desjardins I, Plainfosse B, Seralini GE.The steroid content of semen from a total of 11 mature fertile stallions was studied during two breeding seasons and one winter. The levels of free and conjugated substrates (testosterone and androstenedione), and products (estradiol and estrone), of aromatase were measured by radioimmunoassay with a validated method. The results were seasonally and monthly highly variable with characteristic peaks. The concentrations of free and conjugated estrogens were always higher in the gel-free ejaculate than in the gel except in one subfertile stallion used as comparison. Furthermore, the steroid produ...
Gottschalk M, Sieme H, Martinsson G, Distl O.In the present study, systematic effects on semen quality traits were investigated in 381 stallions representing 22 breeds. All stallions were used for AI either at the Lower Saxon National Stud Celle or the North Rhine-Westphalian National Stud Warendorf. A total of 71,078 fresh semen reports of the years 2001 to 2014 were edited for analysis of gel-free volume, sperm concentration, total number of sperm, progressive motility, and total number of progressively motile sperm. Breed differences were studied for warmblood and light horse breeds of both national studs (model I) and for warmblood b...
Naden J, Amann RP, Squires EL.Puberty was studied using 15 colts of Quarter Horse phenotype. Total scrotal width was measured every 8 weeks from 48 to 96 weeks. Blood samples were taken from 8 colts at 8, 16 and 24 weeks and then every 4 weeks until 100 weeks to measure changes in LH, FSH and testosterone concentrations. Seminal collections were attempted monthly from 48 to 64 weeks and every 2 weeks thereafter until puberty resumed every 3rd day from 96 weeks for 15 ejaculates. For all collections, times to erection, mount and ejaculation and seminal characteristics were recorded. Age at puberty was defined as the first e...
