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.
Miró J, Morató R, Vilagran I, Taberner E, Bonet S, Yeste M.Three separate experiments were conducted to improve preservation of stallion epididymal sperm. In the first one, two different cooling extenders (Kenney and Gent) were compared. Sperm viability and motility patterns were assessed in 10 different epididymal sperm samples after 0 hours, 24 hours, 48 hours, 72 hours, and 96 hours of preservation at 4°C. No significant differences were observed in any of the evaluated parameters either between extenders or throughout the storage period. The second set of experiments was designed to determine whether supplementing thawing medium (INRA Freeze...
Papa PM, Guasti PN, De Vita B, Nakazato NG, Maia L, Freitas Dell'Aqua CP, Scheeren VFDC, Segabinazzi LGTM, de Andrade Junior LRP, Silva LFMC....Although stem cell therapy is a promising alternative for treatment of degenerative diseases, there are just few reports on the use of stem cells therapy in horse's reproductive system. This study aims to evaluate the effect of intratesticular injection of bone marrow mesenchymal stromal/stem cells (MSCs) in healthy stallions, and its outcome on seminal parameters and fertility. In Experiment 1, 24 stallions were divided into treatment group (TG) and control group (CG). In the TG, an intratesticular application of MSC was performed, and in the CG, only PBS was used. Measurements of testicular ...
Höfner L, Luther AM, Waberski D.Ongoing progress in proteomic characterization of seminal plasma has stimulated research on the identification of biomarkers for male fertility and sperm preservability. So far, many studies have evaluated the benefits of reconstituting cryopreserved or sex-sorted semen with seminal plasma. Less information is available about the effect of remaining or added seminal plasma in liquid preserved semen. The interaction between seminal plasma and spermatozoa is species -specific, and within species often complex and ambiguous. This article aims to review the action of seminal plasma on sperm functi...
Kadivar A, Shams Esfandabadi N, Dehghani Nazhvani E, Shirazi A, Ahmadi E.Protamines substitute DNA-binding histones during late spermatogenesis in sperm nucleus. Stallion sperm contains all three variants of these arginine-rich and positively charged nuclear proteins (P1, P2 and P3). Two variants of protamine-2, that is P2 and P3, constitute approximately 15% of the entire protamine content. Also, the ratio of protamine-1 to protamine-2 varies among different mammalian species, and abnormal protamine ratios and protamine content are correlated with male infertility. In this study, changes in protamine mRNA abundance for all three protamines were investigated in sta...
Gacem S, Papas M, Catalan J, Miró J.The accessory sex glands play a major role in the production of seminal plasma, and testicular artery blood flow seems to strongly influence testicular function. However, very little ultrasound imaging of these organs has been undertaken in donkeys. The present work reports the results of such examinations in five jackasses along the year. The accessory glands were inspected by B-mode ultrasound while the testicular artery blood flow was assessed by colour pulsed-wave Doppler ultrasound. The testicular artery was examined at pampiniform plexus (PPT), supratesticular area (ST) and capsular arte...
Podico G, Ellerbrock RE, Curcio BR, Cheong SH, Lima FS, Canisso IF.Urospermia is a major ejaculatory dysfunction affecting stallions. It has been thought that urine-contaminated semen should not be cryopreserved; however, on select cases, urine contamination of semen cannot be avoided. A recent study suggested that urospermic semen can be cryopreserved after cushion centrifugation and extension. Thus, this study aimed to assess the use of single-layer colloid centrifugation (SLC) to process frozen-thawed urine-contaminated stallion semen. Raw ejaculates (n = 55) from eight stallions were split into three groups: no urine, low (20%), or high (50%) urine conta...
Cunha Dos Santos FC, Morrell JM, Nunes MM, Nogueira CE, Curcio BR, Malschitzky E.Epididymal sperm cryopreservation represents the ultimate option to preserve spermatozoa of valuable stallions. Objective: The study aims to evalute whether single layer centrifugation (SLC) prior to cryopreservation or after post-thawing improves the quality of stallion epididymal sperm. Methods: Epididymal sperms of stallions were harvested (N=20). Sperm samples were subjected to treatments: conventional centrifugation, SLC prior to cryopreservation (SLC-PC) or SLC post-thaw (SLC+). All samples were cryopreserved, thawed and evaluated. SLC+ were thawed, single layer cenrifuged and resuspende...
Sancler-Silva YFR, Monteiro GA, Ramires-Neto C, Freitas-Dell'aqua CP, Crespilho AM, Franco MMJ, Silva-Junior ER, Cavalero TMS, Scheeren VFC, Papa FO.Inflammation of the seminal vesicle interferes with fertility and is a persistent problem that is difficult to treat. The aim of this study was to evaluate the semen quality of 5 stallions with seminal vesiculitis before and after local treatment. All stallions were endoscopically treated for seminal vesiculitis during 10 consecutive days. The glandular lumen was accessed and flushed with a Ringer Lactate solution prior to antibiotic infusion. The antibiotic was selected based on the antibiogram from bacterial culture of samples previously collected from the seminal vesicles. The kinetic param...
Peña FJ, O'Flaherty C, Ortiz Rodríguez JM, Martín Cano FE, Gaitskell-Phillips GL, Gil MC, Ortega Ferrusola C.Redox regulation and oxidative stress have become areas of major interest in spermatology. Alteration of redox homeostasis is recognized as a significant cause of male factor infertility and is behind the damage that spermatozoa experience after freezing and thawing or conservation in a liquid state. While for a long time, oxidative stress was just considered an overproduction of reactive oxygen species, nowadays it is considered as a consequence of redox deregulation. Many essential aspects of spermatozoa functionality are redox regulated, with reversible oxidation of thiols in cysteine resid...
Len J, Beehan D, Eilts B, Ebrahimie E, Lyle S.The objective of the study was to investigate if reducing the seminal plasma of stallion extended semen by centrifugation once will suffice to maintain acceptable semen quality for insemination after 4 days of cool storage. Collected semen was extended to 25 × 106 sperm/mL and subjected to one of the following treatments: noncentrifuged (control), centrifuged for 10 minutes at 900 × g and 1800 × g. The supernatant was partially removed, and the sperm pellet, reconstituted and re-extended. It was then placed in a passive cooling device overnight and then transferred to a refrigerator fo...
Canisso IF, Panzani D, Miró J, Ellerbrock RE.Donkeys are nonseasonal, polyestrous, territorial, and nonharem breeders. Although there are many similarities between horses and donkeys, there are also reproductive features that differ, from the longer cervix in the jenny to spermatogenic efficiency in the jack. Mules display reproductive cyclic activity but are rarely fertile. Frozen donkey semen has high pregnancy rates in mares, but lower rates in jennies. This article reviews key aspects of donkey and mule reproductive physiology, reproductive medicine, and assisted reproductive techniques that are useful for practitioners offering assi...
Centromeres exert an inhibitory effect on meiotic recombination, but the possible contribution of satellite DNA to this "centromere effect" is under debate. In the horse, satellite DNA is present at all centromeres with the exception of the one from chromosome 11. This organization of centromeres allowed us to investigate the role of satellite DNA on recombination suppression in horse spermatocytes at the stage of pachytene. To this aim we analysed the distribution of the MLH1 protein, marker of recombination foci, relative to CENP-A, marker of centromeric function. We demonstrated that the sa...
Morris LH, Maclellan LJ.The increased commercialisation of intracytoplasmic sperm injection (ICSI) in horses creates more opportunities to incorporate advanced reproductive technologies, such as sex-sorted, refrozen and lyophilised spermatozoa, into a breeding program. This paper reviews the status of these semen-handling technologies in light of their use in equine ICSI programs. Pregnancies have been achieved from each of these advanced technologies when combined with ICSI in horses, but refinements in the semen-handling processes underpinning these technologies are currently being explored to produce more reliable...
Snoeck PPDN, Pessoa THO, Pereira MGS, Bastos ICL, de Melo MIV.The objective of this study was to compare the BotuCrio extender with the Merk - egg yolk and the INRA 82 modified by the inclusion of acetamide, methyl cellulose and trehalose in substitution of glycerol for freezing equine semen. The semen was diluted after centrifugation to obtain 100 x 10 of sperm/ml in: BotuCrio (control); Merk - egg yolk or INRA 82 modified (Experiment 1). The extended semen was packaged in 0.5 ml straws, cooled and frozen in a freezing machine. The control extender was superior in preserving the motility, VCL, VSL, VAP, LIN, STR and the BCF when compared to the Merk - e...
Álvarez C, González N, Luño V, Martínez F, Gil L.The aim of this study was to test and compare two new components in extenders for freezing donkey semen: mare colostrum and jenny colostrum. Colostrum was obtained from four mares and four jennies right after the foal's birth. Ejaculates were collected from five fertile donkeys. Sperm samples were pooled, diluted and cryopreserved in three different experimental extender groups: lactose supplemented with egg yolk extender (20%) as the control group, lactose supplemented with jenny colostrum extender (20%), and lactose supplemented with mare colostrum extender (20%). After thawing, we evaluated...
Gonzalez-Castro RA, Amoroso-Sanches F, Stokes JE, Graham JK, Carnevale EM.Oocyte activation is initiated when a fertilising spermatozoon delivers sperm-borne oocyte-activating factor(s) into the oocyte cytoplasm. Candidates for oocyte activation include two proteins, phospholipase Cζ1 (PLCZ1) and postacrosomal WW-binding protein (PAWP; also known as WBP2 N-terminal like (WBP2NL)). We localised PLCZ1 and WBP2NL/PAWP in stallion spermatozoa and investigated the PLCZ1 content and sperm parameters as well as cleavage after intracytoplasmic sperm injection (ICSI). PLCZ1 was identified as 71-kDa protein in the acrosomal and postacrosomal regions, midpiece and principal p...
Whitesell K, Stefanovski D, McDonnell S, Turner R.The stallion Breeding Soundness Examination (BSE), as proposed by the Society for Theriogenology, recommends that a stallion produce a minimum of one billion progressively motile, morphologically normal sperm (PMMNS) in the second of two ejaculates collected 1 h apart to be classified as a Satisfactory Prospective Breeder. With this in mind, the first objective of this study was to determine if the classification outcome of the traditional BSE differs depending on the methods used to evaluate sperm motility, morphology and concentration. We hypothesized that application of Computer Assisted ...
Greiser T, Sieme H, Martinsson G, Distl O.The objectives of the present study were to analyze systematic effects on semen quality traits from fresh and frozen-thawed semen collected in and outside season. A total of 4,681 reports on semen traits of 121 stallions representing Arabian, Thoroughbred, Quarter Horse and four warmblood breeds used for artificial insemination at the Lower Saxon National stud Celle were edited for analysis of gel-free volume, sperm concentration, total number of sperm and total number of motile sperm in fresh semen and in frozen-thawed semen progressive motility, DNA fragmentation index and non-viable sperm. ...
Kumar P, Kumar R, Mehta JS, Chaudhary AK, Ravi SK, Chandra Mehta S, Ansari MM, Legha RA, Tripathi BN, Talluri TR.The present study was designed to study the adverse effects of cryopreservation and evaluation of the cryoprotective effect of reduced glutathione (GSH) and ascorbic acid (AA) supplementation on exotic jack semen in combination or alone. For this, 24 semen samples from four adult and fertile jacks were collected via artificial vagina using an estrus jenny as dummy. After semen collection, the semen was evaluated for various qualitative and quantitative parameters in fresh, cooled, and frozen-thawed semen. The semen pellet was extended with the freezing extender containing either AA (0.9 g/L),...
Papas M, Arroyo L, Bassols A, Catalán J, Bonilla-Correal S, Gacem S, Yeste M, Miró J.This study compared the activities of four antioxidant enzymes (superoxide dismutase, SOD; catalase, CAT; glutathione peroxidase, GPX; and glutathione reductase, GSR) in the seminal plasma of stallions and jackasses. Eighteen stallion ejaculates and 24 jack ejaculates were collected through an artificial vagina. Seminal plasma was obtained by several centrifugations at 3000×g and 4 °C for 10 min, and activities of SOD, CAT, GPX and GSR were subsequently determined. We also evaluated whether the collecting season had any influence on the activities of these four enzymes in both stallions ...
Moraes CR, Runcan EE, Blawut B, Coutinho da Silva MA.The iSperm is a newly released semen analysis tool from Aidmics Biotechnology Co. LTD, which allows an iPad Mini to be transformed into a handheld microscope with objective semen analysis software for equine available through the Apple Store (version 4.5.2). The aim of this study was to compare iSperm values for sperm motility and sperm concentration to current acceptable methods for semen analysis and to determine the agreement with these methods using statistical methods. Two ejaculates from each of five Standardbred stallions were used to compare sperm motility (computer-assisted semen anal...
Vitoria A, Romero A, Fuente S, Barrachina L, Vazquez FJ.This report describes a technique for standing laparoscopic vasectomy in stallions through a prospective descriptive study. A preliminary study was carried out with two experimental intact male horses and subsequently the procedure was performed in two clinical cases. These horse owners want to keep their animals in the most possible natural way, preserving its stallion behaviour in a herd without generating offspring. The horses were sedated and restrained in stocks and laparoscopic vasectomy was performed using three portal sites in both paralumbar fossae recording surgical times. A 4-cm seg...
Wilson M, Williams J, Montrose VT, Williams J.Most stallions within breeding programmes are expected to breed and compete concurrently. The exercising of stallions with regards to training regimes during the breeding season is a controversial subject. Daily exercise at low intensities is important for the mental and reproductive well-being of the stallion, however higher intensities of exercise, as seen in competing stallions, may have detrimental effects on seminal quality. To calculate if competition does affect semen quality, this study investigated the effect that equestrian discipline and timing of competition had on a range of stall...
Ruiz AJ, Castaneda C, Raudsepp T, Tibary A.This case report describes spermatogenic arrest and azoospermia in a stallion with a unique Y chromosome-autosome translocation. Clinical diagnosis of azoospermia was based on history of infertility and evaluation of ejaculates collected for artificial insemination. Clinical and ultrasonographic evaluation of the external and internal genitalia did not reveal any abnormalities except for smaller than normal testicular size. Azoospermia of testicular origin was confirmed by determining alkaline phosphatase concentration in semen. Histological evaluation of testicular tissue after castration con...
Sales FA, Ferreira-Silva JC, Vieira JI, Chaves MS, Caldas EL, Filho JP, Freitas VJ, Oliveira MA.Addition of extenders to thawed semen could improve fertility. Objective: To determine the efficiency of extenders to increase viability of thawed semen, measured by sperm parameters in vitro and pregnancy rates after artificial insemination (AI). Methods: Sperm motility and acrosin activity were measured during a thermoresistance test (TRT). Results: Progressive motility decreased (P<0.05) after 30 min in thawing semen treated with saline solution (SS) and only after 60 min with Tyrode's solution (TS) or freezing diluent (FD). The total motility decreased (P<0.05) after 60 min in thawed semen...
Tafuri S, Cocchia N, Carotenuto D, Vassetti A, Staropoli A, Mastellone V, Peretti V, Ciotola F, Albarella S, Del Prete C, Palumbo V, Esposito L....The present study was conducted to assess the chemical composition of Yellow Maca () and its biological activity on stallions following oral administration of hypocotyl powder. Maca was subjected to methanolic extraction and the chemical analysis was carried out by LC-MS-QTOF (liquid chromatography-mass spectrometry). Our results showed that Maca contains some effective antioxidants, a high percentage of glucosinolates, and other important components with a high antioxidant capacity. To evaluate the plant biological activity in stallions fed with Maca powder for 60 days, the redox status and s...
Griffin RA, Baker M, Aitken RJ, Swegen A, Gibb Z.Stallions experience lower per-cycle conception rates compared to other livestock species, largely because they are selected for breeding based on athletic prowess and not reproductive fitness. Mares are seasonal breeders, and pregnancies cannot be detected until 10-14 days post cover via transrectal ultrasonography. This means the detection of stallion fertility fluctuations is delayed by at least 2 weeks, which within the short breeding season employed by the thoroughbred horse breeding industry, can prove quite costly. For these reasons, there is increased demand for robust laboratory assay...
Ghallab ARM, Abou-Ahmed MM, Fad AM, El-Badry DA, Shahat AM, Moawad AR.Cryopreservation of Arabian stallion semen is important in order to improve the function and fertility of frozen/thawed semen in this breed. Objective: To investigate the effects of centrifugation, type of semen extenders, and type of cryoprotectants on the quality of frozen/thawed Arabian stallion spermatozoa. Methods: Semen samples collected from four adult Arabian stallions (one ejaculate per week for 10 consecutive weeks) were either processed directly without centrifugation (no centrifugation; NC) or subjected to centrifugation on the gel-free fraction. Centrifugation protocols were divid...
Sales F, Ferreira-Silva J, Vieira J, Basto S, Filho P, Rocha J, Moura MT, Oliveira M.Post-diluents could potentially increase semen cryotolerance, but remain poorly explored in horses. The aim was to evaluate the efficiency of post-diluents on frozen-thawed semen viability of two stallions (S1-S2). The cryopreserved semen was thawed at 50°C for 40 seconds. Semen motility and acrosin activity (AA) were determined during the thermo-resistance test (TRT). Progressive motility of S2 semen decreased after 60 and 90 minutes of TRT (TRT60 and TRT90) on the control compared to both post-diluents. The total motility of both S1 and S2 decreased on TRT60 and TRT90 semen control versus b...
Sipahutar H, Sourdaine P, Moslemi S, Plainfossé B, Séralini GE.High levels of plasma estrogens constitute an endocrine peculiarity of the adult stallion. This is mostly due to testicular cytochrome p450 aromatase, the only irreversible enzyme responsible for the bioconversion of androgens into estrogens. To identify more precisely the testicular aromatase synthesis sites in the stallion, testes from nine horses (2-5 years) were obtained during winter or spring. Paraplast-embedded sections were processed using rabbit anti-equine aromatase, followed by biotinylated goat anti-rabbit antibodies, and amplified with a streptavidin-peroxidase complex. Immunoreac...
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 ...
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...
Prasinou P, De Amicis I, Fusaro I, Bucci R, Cavallini D, Parrillo S, Caputo M, Gramenzi A, Carluccio A.Fatty acid-based lipidomic analysis has been widely used to evaluate health status in human medicine as well as in the veterinary field. In equine species, there has been a developing interest in fertility and sperm quality. Fatty acids, being the principal components of the membranes, play an active role in the regulation of the metabolic activities, and their role on spermiogenesis seems to be of great importance for the resulting quality of the sperm and, thus, fertility. With the application of widely used lipidomic techniques, the aim of this study was to evaluate: (a) the fatty acid cont...
Gacem S, Catalán J, Yánez-Ortiz I, Soler C, Miró J.The evaluation of the male fertility potential is based on the analysis of the basic spermatic characteristics of concentration, motility and morphology. Thus, the study of sperm morphology is a fundamental element in the seminal analysis, but its real meaning has been biased by the techniques used for its evaluation. These techniques involve dehydration phases and subsequent staining, which involves the production of artifacts. The aim of the study is to compare two methods for equid semen morphology evaluation, Trumorph using living sperm vs. eosin-nigrosine stain. A total of 49 ejaculates f...
Zong E, Fan G.Generally speaking, the majority of F1 hybrids between the horse and donkey are sterile and do conform to the hypothesis of gametogenic breakdown. The results of our own research, nevertheless, lead us to conclude that there is variability in degree of sterility and fertility for both F1 and B1 individuals, with a gradual advance from sterility to fertility being seen. This is the subject we will discuss from the point of view of oestrus, ovulation, mating, pregnancy, spermatogenesis, endocrine activity, isozymes and karyotypic analysis in this paper.
Stout TA, Colenbrander B.There are a number of situations in which it is desirable to suppress part or all of the reproductive endocrine system in a horse, notably the competing animal whose tractability during training, or performance during competition, is compromised by the expression of sexual or aggressive behavior. The current therapeutic approaches to reproductive endocrine suppression include gonadectomy and progestagen administration, where the former carries surgical risks and entails irreversible loss of breeding potential, and effective progestagen therapy requires frequent administration for extended peri...
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...
Pukazhenthi BS, Johnson A, Guthrie HD, Songsasen N, Padilla LR, Wolfe BA, Coutinho da Silva M, Alvarenga MA, Wildt DE.Two studies were conducted to understand sperm cryosensitivity in an endangered equid, the Przewalski's horse (Equus ferus przewalski), while testing the cryoprotectant ability of formamides. The first assessed the toxicity of permeating cryoprotectants (glycerol, methylformamide [MF] and dimethylformamide [DMF]) to Przewalski's horse spermatozoa during liquid storage at 4°C. The second examined the comparative influence of three diluents (with or without formamides) on cryosurvival of sperm from the Przewalski's versus domestic horse. When Przewalski's horse spermatozoa were incubated at 4°...
Li B, He X, Zhao Y, Bai D, Li D, Zhou Z, Manglai D.Numerous studies have shown that microRNAs (miRNAs) are essential for testicular development and spermatogenesis. In order to further characterise these physiological processes, three immature and three mature testes of the Mongolian horse were collected and six libraries were established. Using small RNA sequencing technology, 531 mature miRNAs were identified, including 46 novel miRNAs without previously ascribed functions. Among the 531 miRNAs, 421 were expressed in both immature and mature libraries, 65 miRNAs were found solely in immature testis libraries and 45 miRNAs were found solely i...
Pozor MA, Macpherson ML, McDonnell SM, Nollin M, Roser JF, Love C, Runyon S, Thomas BF, Troedsson MH.The objective of this study was to determine whether an indenopyridine derivative RTI-4587-073(l) was a good candidate for male contraception in horses. We hypothesized that a single administration of RTI-4587-073(l) causes significant suppression of testicular function in stallions without affecting sexual behavior. Three Miniature horse stallions received a single dose of 12.5 mg/kg RTI-4587-073(l) orally (group "treated"), whereas three other Miniature horse stallions received placebo only (group "control"). Semen was collected and evaluated from all stallions twice a week for three baselin...
Gadella BM, Rathi R, Brouwers JF, Stout TA, Colenbrander B.During sexual reproduction, the sperm and oocyte must fuse before the production of a diploid zygote can proceed. In mammals such as equids, fusion depends critically on complex changes in the plasma membrane of the sperm and, not surprisingly, this membrane differs markedly from that of somatic cells. After leaving the testes, sperm cease to synthesize plasma membrane lipids or proteins, and vesicle-mediated transport stops. When the sperm reaches the female reproductive tract, it is activated by so-called capacitation factors that initiate a delicate reorientation and modification of molecul...
Jiménez-Trejo F, Coronado-Mares I, Boeta M, González-Santoyo I, Vigueras-Villaseñor R, Arriaga-Canon C, Herrera LA, Tapia-Rodríguez M.Assisted reproductive techniques have been used on several domestic animals to preserve desirable traits in strains of high genetic and commercial value; however in equines its efficiency rate is relatively low. To increase the conception ratio in stallions, some research groups have used pharmacological treatments which promote sperm hyperactivation in order to increase male's fertility rates. In this way, our previous work suggests that serotonin (5-HT) could be a good pharmacological candidate that facilitates conception rate in domestic horses. 5-HT is a neurohormone involved in several re...
Foote RH.Several inherited conditions associated with testicular defects, abnormal spermatogenesis and morphologically abnormal sperm have been found. These usually are controlled by single gene pairs. A notable exception is testicular size, with heritability in young bulls ranging from .42 to .88. Testicular size directly affects sperm output potential. The major contributor to variation in semen quality is the environment. Environmental effects may be temporary or permanent. Permanent effects occurring during prenatal and prepubertal periods and temporary or permanent factors acting after spermatogen...
Pirhonen A, Linnala-Kankkunen A, Mäenpää PH.Protamines isolated from ejaculated human, stallion, bull, boar, and ram spermatozoa were subjected to phosphoserine conversion reaction and protein sequencing. Phosphoserines were detected as S-ethylcysteines. Endogenously phosphorylated protamines have previously been found only in ejaculated human sperm. In this study, we demonstrate that ejaculated sperm from other species also contain protamines phosphorylated at serine residues. In P1-protamines, the endogenously phosphorylated serines were located at the N-terminal region in all species studied, whereas in major forms of human and stall...
Heninger NL, Staub C, Blanchard TL, Johnson L, Varner DD, Forrest DW.Apoptosis in testicular germ cells has been demonstrated in many mammalian species. However, little is known about the stallion (Equus caballus) and rates of apoptosis during spermatogenesis. Morphological and biochemical features of apoptosis reported in other species were used to confirm that the TdT-mediated dUTP Nick end labeling (TUNEL) assay is an acceptable method for identification and quantification of apoptotic germ cells in histological tissue sections from stallion testis. Seminiferous tubules from eight stallions with normal testis size and semen quality were evaluated according t...
Tardif S, Brady HA, Breazeale KR, Bi M, Thompson LD, Bruemmer JE, Bailey LB, Hardy DM.Zonadhesin is a rapidly evolving protein in the sperm acrosome that confers species specificity to sperm-zona pellucida adhesion. Though structural variation in zonadhesin likely contributes to its species-specific function, the protein has not previously been characterized in organisms capable of interbreeding. Here we compared properties of zonadhesin in several animals, including the horse (Equus caballus), donkey (E. asinus), and Grevy's zebra (E. grevyi) to determine if variation in zonadhesin correlates with ability of gametes to cross-fertilize. Zonadhesin localized to the apical acroso...
Bogle OA, Carrasco RA, Ratto MH, Singh J, Adams GP.The objectives of the study were to compare the presence and localization of ovulation-inducing factor (OIF)/nerve growth factor (NGF) in male reproductive organs and determine the abundance in ejaculates of species representative of both spontaneous and induced ovulators. We hypothesized that the protein is a widely conserved component of semen among mammals, but is most abundant in camelids. Immunohistochemical analysis was performed on tissues from the male reproductive system of llamas, rats, cattle, bison, elk, and white-tailed deer (n = 2 males/species), and the abundance of OIF/NGF in...
Gross MK, Toscano DG, Toscano WA.Calmodulin (CaM), the calcium binding protein that modulates the activity of a number of key regulatory enzymes, is present at high levels in sperm. To determine whether CaM regulates adenylate cyclase in mammalian sperm, the actions of EGTA and selected CaM antagonists on a solubilized adenylate cyclase from mature equine sperm were examined. The activity of equine sperm adenylate cyclase was inhibited by EGTA in a concentration-dependent manner with a half-maximal inhibitory concentration (IC50) of 2 mM. Equine sperm adenylate cyclase was also inhibited in a concentration-dependent manner by...
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...
Calvete JJ, Mann K, Schäfer W, Sanz L, Reinert M, Nessau S, Raida M, Töpfer-Petersen E.We report the complete amino acid sequence of HSP-1, a major protein isolated from stallion seminal plasma or acid extracts of ejaculated spermatozoa. The protein consists of 121 amino acids organized in two types of homologous repeats arranged in the pattern AA'BB'. Each of the 13-15-residue A-type repeats contains two O-linked oligosaccharide chains. The B-type repeats span 44-47 amino acids each, are not glycosylated, and have the consensus pattern of the gelatin-binding fibronectin type-II module. This domain also occurs in the major bovine seminal plasma heparin-binding proteins PDC-109 (...
Nagata S, Kurosawa M, Mima K, Nambo Y, Fujii Y, Watanabe G, Taya K.The aim of this study was to clarify the effect of anabolic steroids on the testicular endocrine function of mature stallions. Mature thoroughbred stallions were treated with 800 mg nandrolone decanoate every 3 weeks for 3 months. After the first treatment, plasma concentrations of LH, immunoreactive inhibin and testosterone decreased rapidly to the nadir. These hormones were maintained at significantly lower concentrations compared with concentrations in intact stallions. Histology of the testicular tissue indicated the arrest of advanced spermatogenesis in the seminiferous tubules and a seve...
Lanneau M, Loir M.Using standardized methods for protein extraction and analysis, the testes of rams, bulls, goats, boars, stallions, rats, cats, hedgehogs, European mink and ferrets were examined for basic spermatid nucleoproteins by electrophoresis. The results suggest that differences exist in the total number of these proteins as well as in the number and amount of the cross-linked cystein-containing proteins. These differences appear to be more family-specific than species-specific.
Johnson L.Spermatocytogenesis plays a pivotal role in regulation of spermatogenesis; however, its details remain relatively obscure in nonrodent species. The equine testis contains approximately 100% more spermatogonia in summer than in winter and appears to be a good model to identify the flexible components of spermatocytogenesis that cause seasonal changes in daily sperm production. Testes were taken from horses in the winter (n = 47) and in summer (n = 43). Tissues were fixed by glutaraldehyde-perfusion and submission in osmium, embedded in Epon or methacrylate, sectioned at 0.5 micron or 5 microns,...
Jeannerat E, Marti E, Berney C, Janett F, Bollwein H, Sieme H, Burger D, Wedekind C.The major histocompatibility complex (MHC) has repeatedly been found to influence mate choice of vertebrates, with MHC-dissimilar mates typically being preferred over MHC-similar mates. We used horses (Equus caballus) to test whether MHC matching also affects male investment into ejaculates after short exposure to a female. Semen characteristics varied much among stallions. Controlling for this variance with a full-factorial within-subject experimental design, we found that a short exposure to an MHC-dissimilar mare enhanced male plasma testosterone and led to ejaculates with elevated sperm nu...
Peña FJ, Ortega Ferrusola C, Martín Muñoz P.Flow cytometry is currently recognized as a robust tool for the evaluation of sperm quality and function. However, within equine reproduction, this technique has not reached the sophistication of other areas of biology and medicine. In recent years, more sophisticated flow cytometers have been introduced in andrology laboratories, and the number of tests that can be potentially used in the evaluation of sperm physiology has increased accordingly. In this review, recent advances in the evaluation of stallion spermatozoa will be discussed. These new techniques in flow cytometry are able to simul...
Kikuchi M, Takao Y, Tokuda N, Ohnami Y, Orino K, Watanabe K.Total 78 semen samples were obtained from 27 Thoroughbred stallions (aged 6 to 27 years), and were subjected to quantification of lactoferrin (Lf) in seminal plasma and examination of the seminal properties. The seminal plasma Lf concentration varied from 21 to 689 microg/ml, with a mean value of 244 +/- 151 microg/ml (S.D.). The seminal plasma Lf concentration and total seminal plasma Lf positively correlated with the sperm concentration (r=0.5938, P<0.001) and with the total sperm number (r=0.6959, P<0.001), respectively. There was no correlation between seminal plasma Lf and sperm mot...
Giesecke K, Hamann H, Stock KF, Klewitz J, Martinsson G, Distl O, Sieme H.The research of fertility in humans and other mammals has strongly advanced in the recent years. The examination of molecular mechanisms influencing horse fertility is relatively recent. We chose the angiotensin converting enzyme (ACE), the sperm autoantigenic protein 17 (SP17) and the follicle stimulating hormone (FSHB) as candidates for determining stallion fertility and to analyze associations of intragenic single nucleotide polymorphisms (SNPs), flanking microsatellites and candidate-gene linked haplotypes with the pregnancy rate per oestrus (PRO) in 179 Hanoverian stallions. Fertility tra...
Thompson DL, Pickett BW, Squires EL, Amann RP.Factors affecting testicular measurements in situ and the relationships among the measurements and various reproductive characteristics were studied using data from 48 stallions. Mean values during the breeding season are provided for scrotal width, widths and lengths of individual testes, combined weight of testicular parenchyma, daily sperm production and daily sperm output. Testicular measurements were highly repeatable from day to day and for repeated measurements on a given day; technician provided the largest source of variation in the measurements of a given stallion. Age significantly ...
Grignard E, Morin J, Vernet P, Drevet JR.We report here on the cloning of cDNAs coding bovine and equine orthologs of mouse epididymis-restricted and sperm-bound glutathione peroxidase 5 (GPX5), a selenium-independent member of the multigenic GPX family in mammals. The complete sequence of bovine GPX5 as well as a partial sequence of the equine GPX5 were characterized, conceptually translated and aligned with other known mammalian GPX5 proteins. Using Northern blotting assays, we show that the level of expression of GPX5 is high in bovine but low in equine and that in both species the regionalization of GPX5 expression in epididymis ...
