Asymmetric expression of proteins in the granules of the placentomal Binucleate cells in Giraffa camelopardalis†.

Mature granulated trophoblast binucleate cells (BNC) have been found in all ruminant placentas examined histologically so far. BNC are normally fairly evenly distributed throughout the fetal villus and all their granules contain a similar variety of hormones and pregnancy associated glycoproteins (PAGs). Only the Giraffe is reported to show a different BNC protein expression, this paper is designed to investigate that. Gold labelled Lectin histochemistry and protein immunocytochemistry were used on deplasticised 1 µm sections of a wide variety of ruminant placentomes with a wide range of antibodies and lectins. In the Giraffe placentomes, even though the lectin histochemistry shows an even distribution of BNC throughout the trophoblast of the placental villi, the protein expression in the BNC granules is limited to the BNC either in the apex or the base of the villi. Placental lactogens and Prolactin (PRL) are present only in basally situated BNC: PAGs only in the apical BNC. PRL is only found in the Giraffe BNC which react with many fewer of the wide range of antibodies used here to investigate the uniformity of protein expression in ruminant BNC. The possible relevance of these differences to ruminant function and evolution is considered to provide a further example of the versatility of the BNC system.

Ovarian and placental morphology and endocrine functions in the pregnant giraffe (Giraffa camelopardalis).

Gross, histological and immunocytochemical examinations carried out on maternal and fetal reproductive tissues from two pregnant giraffes at an estimated 8 and 13.5 months of gestation (term=15 months) revealed a typically ruminant macrocotyledonary placenta with binucleate trophoblast cells scattered sparsely in the placentome where they stained intensely with a prolactin antiserum. Binucleate cells were present in greater numbers in the intercotyledonary allantochorion where they did not stain for prolactin whereas the uninucleate trophoblast still did. A single large corpus luteum of pregnancy and several small luteinised follicles were present in the maternal ovaries while the fetal ovaries at 13.5 months gestation showed an assortment of enlarging antral follicles and partially and completely lutenised follicles, the granulosa and luteal cells of which stained positively for 3ß-hydroxysteroid dehydrogenase (3ß-HSD), 17,20 lyase, prolactin, progesterone receptor and androgen receptor, but negatively for aromatase. The uninucleate trophoblast of the placentome and intercotyledonary allantochorion, the epithelium of the maternal endometrial glands, the seminiferous epithelium in the fetal testis at 8 months of gestation and the zonae fasciculata and reticularis of the fetal adrenal at 13.5 months also stained positively for 3ß-HSD and negatively for aromatase. Endocrinologically, it appears that the giraffe placenta is more similar to that of the sheep than the cow with a placental lactogen as the likely driver of the considerable degree of luteinisation seen in both the maternal and the fetal ovaries.

Intrauterine administration of plant oils inhibits luteolysis in the mare.

REASONS FOR PERFORMING THE STUDY: The maternal recognition of pregnancy (MRP) signal in the mare has not been determined, although oestrogens have been proposed as a potential candidate. OBJECTIVES: To determine effects of intrauterine administration of oestrogen and various oils on cyclic luteolysis in the mare. HYPOTHESIS: Intrauterine oestradiol or fatty acids may suppress luteolysis in the cycling mare when administered during late dioestrus. METHODS: A single 1 ml dose of slow-release oestradiol (10 mg/ml) in fractionated coconut oil was infused into the uterine lumen of cycling mares on Days 6, 8, 10, 12 or 14 post ovulation (n=12 in each group). Four further groups, each of 12 mares, received an intrauterine infusion of either 1 ml of fractionated coconut oil, peanut oil, mineral oil or a slow-release preparation of oestradiol (10 mg/ml) in mineral oil on Day 10 post ovulation. Serial blood samples were assayed for progesterone concentrations to monitor luteal function. RESULTS: Intrauterine administration of oestradiol in fractionated coconut oil showed peak efficacy at Day 10 when luteolysis was delayed in 11/12 (92%) mares. The ability of the treatment to delay luteolysis was not significantly different when administered on Days 8 (9/12; 75%), 12 (10/12; 83%) or 14 (6/12; 50%) of dioestrus, but declined significantly when given on Day 6 (3/12; 25%). Oestradiol was not needed to initiate luteostasis since fractionated coconut oil alone or peanut oil administered at Day 10 induced the same high rate of luteal persistence (11/12; 92% for both oils). In contrast, mineral oil did not prolong luteal lifespan, either when administered alone (2/12; 17%) or combined with oestradiol (3/12; 25%). CONCLUSION: These results do not unequivocally rule out a possible involvement of embryonic oestrogens in MRP in the mare but suggest it is unlikely. The results demonstrate that plant oils can postpone luteolysis, suggesting they may modulate synthesis or release of prostaglandins from the mare's endometrium. POTENTIAL RELEVANCE: Administration of fractionated coconut or peanut oil on Day 10 post ovulation provides an effective and practical method of prolonging luteal function ('pseudopregnancy') thereby suppressing unwanted oestrous behaviour. Further studies to elucidate the mechanism by which this is achieved may increase understanding of both luteostasis and MRP signal in the mare.

Abnormal umbilical cord attachment sites in the mare: a review illustrated by three case reports.

Abnormal cord attachment can be a manifestation of an inappropriate fixation position of the conceptus in the uterus, or it may result from disorientation of the conceptus post fixation. The potential for this resulting in fetal and neonatal compromise is reviewed in the light of previous reports and to which 3 cases within the authors' experience are added.

Post natal oestrogen administration stimulates precocious endometrial gland development in the horse.

REASONS FOR PERFORMING STUDY: Fillies completely devoid of endometrial glands (uterine gland knockout; UGKO) would make ideal experimental models in which to study the role of endometrial histotroph in embryogenesis and early fetal development in the mare. HYPOTHESIS: Administration of a synthetic progestagen plus oestrogen to newborn filly foals and, thereafter, at regular intervals to age 6 months, would permanently suppress endometrial gland development. METHODS: Nine half-sister Thoroughbred filly foals were treated, in 3 groups, with: A) the weakly active progestagen, norgestomet, administered from birth to age 6 months, in subcutaneous implant form plus oestradiol valerate and norgestomet i.m. at fortnightly intervals; B) the strongly active oral progestagen, altrenogest, administered daily from birth to age 6 months plus fortnightly injections of oestradiol valerate and norgestomet; C) nothing (untreated controls). Endometrial biopsies were recovered from all fillies at ages 6 months and 2 years to assess the degree of endometrial gland morphogenesis and to determine immunohistochemically the presence or absence of oestrogen and progesterone receptors in the endometrial tissues. RESULTS: Groups B and C showed no endometrial gland development, whereas Group A fillies showed a high degree of endometrial gland development, plus strong staining for both oestrogen and progesterone receptors at age 6 months. All 9 fillies showed full normal endometrial gland morphogenesis, development and function at age 2 years. CONCLUSIONS AND RELEVANCE: While the administration of a strongly active progestagen over-rode the actions of the concomitantly administered oestrogen and suppressed endometrial gland development during the period of administration, treatment with oestradiol valerate together with a weakly active progestagen, stimulated precocious endometrial gland development. Neither steroid was able to create the desired UGKO experimental model and all fillies showed normal endometrial gland development and fertility after puberty. Hence, ovarian oestrogen, not progesterone, appears to be the basic stimulus for endometrial gland morphogenesis in the horse.

Histological and immunohistochemical characterization of equine anovulatory haemorrhagic follicles (AHFs).

Anovulatory haemorrhagic follicles (AHFs) are often the reason for ovulation failure in the mare. As the underlying factors that lead to AHF development are not well understood, it was of interest to investigate the vascularization of AHFs compared with normal follicles and corpora lutea (controls). In the present study, the ovarian cell populations investigated immunohistochemically included granulosa and luteal cells as well as various vascular structures. None of these cell types showed differences in the expression of vascular endothelial growth factor A (VEGF-A) between control ovaries containing normal follicles and corpora lutea and ovaries with AHFs. In contrast, a considerable reduction in the proportion of Flk-1-expressing cells, together with a decreased intensity of staining, was apparent in the AHFs. This greatly reduced expression of Flk-1 in the luteinized cells and the vascular structures of AHFs may lead to a distinct decrease in the potential pro-angiogenic activity of VEGF-A in these structures compared with the situation in normal follicles and corpora lutea. Furthermore, the authors suspect that the distinct expression of angiopoietin2 and VEGF-A seen in the cells within the inner fibrous layers of the AHFs was caused by hypoxia resulting from deficient vascularization, as suggested by the irregularity of the capillaries present in the luteinized wall of the AHF. In addition, whereas LH-receptor (LH-R) expression occurred uniformly in all stages of development of the corpora lutea in normal control ovaries, there was highly variable labelling for LH-R in all the AHFs examined, thereby indicating a possible numerical deficiency of LH-receptors in AHFs. The authors concluded that, despite the apparent expression of sufficient VEGF-A in the AHFs allows ovulation and corpus luteum formation, a relative lack of receptor, Flk-1, effects the pro-angiogenic activity of VEGF-A which could be a reason for ovulation failure associated with AHF formation.

Immunohistochemical localization of oestrogen receptors alpha and beta, progesterone receptor and aromatase in the equine placenta.

The functions of placental oestrogens during equine pregnancy are still unclear. Yet, they may act predominantly as local regulators of growth and differentiation in the microplacentomes. Thus, expression patterns of oestrogen receptors (ERs) alpha and beta were investigated in the microcotyledonary placenta from pregnant mares at 110, 121, 179, 199 and 309 days of gestation by immunohistochemistry. In microplacentomes, both the ER isoforms were detected in trophoblast (T) cells, chorionic villous stroma (FS), microcaruncular epithelium (ME) and microcaruncular stroma (MS). Proportions of positive cells were 38-91% (T), 11-41% (FS), 55-89% (ME), 17-51% (MS) for ERalpha and 66-76% (T), 21-37% (FS), 41-68% (ME) and 24-55% (MS) for ERbeta. Between days 110 and 199, proportions of cells positive for progesterone receptor (PR) varied between 19% and 62% (T), 3% and 50% (CS), 15% and 46% (ME), and 4% and 33% (MS). At day 309, PR was virtually absent in T, CS and ME (percentages < 0.1), whereas in MS 14.3% of cells were still positive. The expression of ERs and PR in equine microplacentomes gives evidence for a role of placental steroids as regulators of placental growth, differentiation and function. The detection of ERalpha, ERbeta and PR in foetal and maternal vascular tissue suggests that placental steroids are also involved in the control of placental angiogenesis and /or vascular functions. The co-localization of ERs with aromatase in T suggests auto- or intracrine functions of oestrogens in this cell type.

Vitrification of equine expanded blastocysts following puncture with or without aspiration of the blastocoele fluid.

BACKGROUND: Historically, cryopreservation of equine embryos >300 µm gave poor pregnancy rates until researchers collapsed the blastocoele cavity and aspirated the blastocoele fluid prior to vitrification. OBJECTIVE: To determine if aspiration of the blastocoele fluid prior to vitrification is essential for post warming survival. STUDY DESIGN: In vivo experiments. METHODS: Fifty embryos were recovered on day 7-8 and washed in holding medium (HM; M-199HEPES + 20% FBS + antibiotics). Embryos were punctured using a micromanipulator mounted 30 µm biopsy needle; following this 28 had >90% of their blastocoelic fluid actively aspirated while the remaining 22 were not-aspirated. Embryos were then vitrified using a two-step process with increasing concentrations of DMSO and ethylene glycol (7.5-15% v:v), and 0.5 mol/L sucrose in the final solution before being loaded onto a Cryolock device and plunged into liquid nitrogen. The embryos were warmed by plunging the Cryolock tip into HM with 1 mol/L sucrose at 37°C. After 1 min, the embryos were transferred to HM + 0.5 mol/L sucrose at RT for 4 min before transfer into HM for a further 4 min prior to transfer to a recipient mare. RESULTS: Mean (±s.e.) embryo diameter was not significantly different between the punctured and punctured plus aspirated group (646.4 ± 61.7 vs. 754.8 ± 59.1 µm, respectively; P = 0.215). Nonaspirated and aspirated embryos gave pregnancy rates of 10/22 (45%) and 21/28 (75%) respectively (P = 0.061). Sub-dividing embryos on the basis of size showed that vitrification of larger embryos (>550 µm) yielded a significantly higher pregnancy rate when they were aspirated vs. not-aspirated (13/18 [72%] vs. 1/10 [10%], respectively; P = 0.006), whereas there was no difference for smaller embryos (8/10 [80%] vs. 9/12 [75%], respectively; P = 0.8). MAIN LIMITATIONS: Group sizes are limited. CONCLUSION: Aspiration of blastocoele fluid from embryos ≤550 µm is not a pre-requisite for successful vitrification. The Summary is available in Spanish - see Supporting Information.

Immunolocalisation of the uterine secretory proteins uterocalin, uteroferrin and uteroglobin in the mare's uterus and placenta throughout pregnancy.

Previous studies have shown that the equine uterus produces many progesterone-dependent proteins throughout gestation. In particular, uterocalin and uteroferrin are detectable using electrophoresis or blot analyses but information regarding the immunohistochemical placental distribution of these two proteins is rare and information regarding uteroglobin is still lacking. The aim of the present study was to co-immunolocalise these three secretory proteins in the mare's uterus throughout gestation in an effort to understand their functional role in the maintenance of pregnancy. Therefore, endometrial biopsy samples were obtained from 20 pregnant mares between 16 and 309 days of gestation and labelled immunohistochemically for uteroglobin, uteroferrin and uterocalin. Uteroferrin remained detectable in almost every endometrial gland at all stages but with an increase in staining intensity as gestation advanced. The most progesterone-dependent protein, uterocalin, showed variable staining throughout gestation with the most intense labelling in early pregnancy and during the period of endometrial cup reaction. Uteroglobin secretion was only detectable in traces and only in individual glands throughout gestation. The results indicate that uterocalin and uteroferrin, but not uteroglobin, may play important roles in supplying nutrients for the conceptus, thereby contributing to the maintenance of pregnancy. However, further investigations are necessary to understand the role of uteroglobin during gestation.

The effects of maternal health and body condition on the endocrine responses of neonatal foals.

REASONS FOR PERFORMING THE STUDY: Chronic and acute alterations in maternal nutrient intake during pregnancy alter pancreatic and hypothalamo-pituitary-adrenal (HPA) axis function in the offspring, before and after birth. Little is known about these effects. OBJECTIVE: To determine whether maternal nutrient restriction caused by natural infection with Streptococcus equi altered endocrine function in neonatal foals born from mares fed a maintenance or high plane of nutrition throughout pregnancy. METHODS: Ten primiparous mares received either a diet to maintain moderate body condition score (Moderate, n = 5) or a near ad libitum feeding regime to maintain a high body condition score (High, n = 5) throughout pregnancy. All mares inadvertently became infected with Streptococcus equi in mid gestation and lost approximately 10% body mass. RESULTS: Maternal insulin and glucose concentrations decreased (P < 0.05) during, and one month following, the weight loss period. High mares weighed more (P < 0.05) at parturition than Moderate mares; all foals were healthy. Gestational age, foal bodyweights, placental and clinical parameters after birth were no different between the 2 groups. Foal plasma cortisol and glucose responses to exogenous adrenocorticotrophic hormone and insulin, respectively, were similar for both groups. Insulin concentrations during glucose tolerance test were significantly higher (P < 0.05) in foals from Moderate than High mares and compared with foals studied previously from healthy, well-fed mares, suggesting that the beta cell sensitivity to glucose was enhanced in Moderate. CONCLUSION: Acute nutrient restriction in mid gestation caused by maternal illness and inappetence, superimposed on a maintenance feed intake throughout pregnancy, enhanced insulin secretion to glucose in foals. Nutritional programming of pancreatic beta cells, but not the HPA axis, appeared to depend on the level of nutrition before and after the weight loss period. POTENTIAL RELEVANCE: Disturbances in neonatal pancreatic beta cell function programmed during pregnancy may predispose foals to metabolic problems in later life.

Half a century of equine reproduction research and application: A veterinary tour de force.

Over the past 50 years, per season pregnancy rate in Thoroughbred mares have risen from 70 to >90% and foaling rates from 55 to >80%. Some of the significant pharmacological treatments and diagnostic methods which have driven this marked improvement in breeding efficiency are reviewed. They include the application of artificial lighting to hasten the onset of ovulatory oestrous cyclicity in early Spring, rapid steroid hormone assays to aid in determining the stage and normality of the mare's cycle, prostaglandin analogues, synthetic progestagens and Gonadotrophin-releasing Hormone (GnRH) analogues to better control and manipulate her cycle to good effect, transrectal ultrasound scanning to monitor follicle growth, endometrial architecture and ovulation and to allow accurate, early pregnancy diagnosis thereby enabling successful ablation of one of twin conceptuses. Also, flexible videoendoscopy to monitor physiological and pathological changes in the uterine endometrium and rigid laparoscopy to apply prostaglandin to the oviducts to dislodge and clear suspected blockages of them to restore fertility. The outbreak of Contagious Equine Metritis in Newmarket in the spring of 1977 and the swabbing-related changes to mare and stallion management, plus the improved veterinary hygiene methods, which followed are also recounted. The past half century has witnessed many technical and therapeutic advances that have enhanced tremendously the diagnostic and treatment capabilities of studfarm veterinary surgeons. They, in turn, have improved greatly the efficiency of breeding Thoroughbreds and other types of horses.

Reproductive efficiency of Flatrace and National Hunt Thoroughbred mares and stallions in England.

REASONS FOR PERFORMING STUDY: Previous surveys of reproductive efficiency in British Thoroughbreds included only mares and stallions standing on studfarms in and around Newmarket. The present study was widened to compare Flatrace (FR) (Group A) and National Hunt (NH) (Group B) mares and stallions on studfarms throughout England. OBJECTIVES: To assess the influences of mare type, status and age, and veterinary manipulations on reproductive efficiency parameters. To compare the inherent fertility of stallions, based on singleton and twin pregnancy rates and pregnancy loss rates, in Groups A and B Thoroughbred breeding stock. METHODS: Managers of 24 FR and 9 NH public studfarms were asked to complete a questionnaire for each mated oestrous cycle shown by 2321 Group A and 1052 Group B mares throughout the 2002 mating season. Parameters such as per cycle singleton and twin pregnancy rates, and pregnancy loss rates were noted, and the success of hormone treatments to induce oestrus and ovulation assessed. The number of matings per oestrus and per pregnancy were recorded, together with the incidence and effectiveness of uterine and other veterinary treatments. The inherent fertility of 84 Group A and 43 Group B stallions in the study, as measured by the singleton and twin early pregnancy rates and the pregnancy loss rates recorded in the mares they mated, was also estimated. RESULTS: Per cycle early pregnancy (Days 13-16) was 63.2% for Group A and 65.3% for Group B mares; and 10.3% and 13.1%, respectively, of those pregnancies were twins or triplets. Early, middle and late pregnancy loss rates were 7.2% vs. 8.0% (Days 15-42), 3.6% vs. 6.1% (Days 42-1st October) and 2.7% vs. 2.1% (October-foaling), respectively. Matings per oestrus and per early pregnancy were significantly higher in Group B vs. Group A mares. For stallions that mated > or = 30 mares, overall early pregnancy rates per cycle in mares mated ranged from 30-89% across the 2 groups. CONCLUSIONS: No major differences in reproductive efficiency were identified between FR and NH mares and stallions. Increasing mare age was the single biggest limiting factor to an otherwise high rate of fertility in well-managed English Thoroughbreds. POTENTIAL RELEVANCE: This study identified factors that influence reproductive efficiency in the Thoroughbred.

Structural and haemovascular aspects of placental growth throughout gestation in young and aged mares.

This investigation was carried out to study the equine placenta from early gestation to near term, with special reference to morphological changes associated with the development of the vasculature of the fetal component of the microcotyledons. Pregnant uteri were removed post mortem from five Thoroughbred mares between 110 and 309 days of gestation, two of which were aged, multiparous animals suffering age-related degenerative changes in their endometrium (endometrosis), while the other three were young, and had primigravid normal healthy uteri. Pieces of endometrium with placenta attached were fixed for light microscopy and fetal vascular casts were made by injecting the placental arteries with a mixture of Mercox and methylmethacrylate. The casts were examined under the scanning electron microscope. In an aged, endometrotic mare at 110 days of gestation, most of the microplacentomes were irregular in shape with a mean+/-sem diameter of 399+/-30.53 microm. Capillaries with variable diameters made up widely meshed network villi with pointed ends (Type 1 terminal villi), and narrow-meshed networks with finger-like ends (Type 2 terminal villi). In the "paired" young healthy mare at day 121 of gestation, most of the microplacentomes were globular in shape and appeared smaller in diameter than those in the 110-day "pair". The narrow-meshed capillary networks formed villi with stems that consisted of both intermediate and terminal parts, the latter of which represented more the Type 2 than the Type 1 terminal villus. In another aged endometrotic mare at 179 days of gestation, the microplacentomes were typically globular in shape and they showed a mean diameter of 534+/-36.07 microm. The villi were short and thick and they were distinctly differentiated into stem, intermediate and terminal parts. The density of the fetal capillaries had now greatly increased so that, three dimensionally, they constituted bulb-like capillary networks at the base of the stem of each villus. At 199 days in the young healthy "pair", the microplacentomes were again smaller in diameter (402+/-16.24 microm) than in the old mare at 179 days and the interhaemal distance had now reduced to 14.28+/-0.42 microm. The vascular density was lower than in the day 179 aged mare and the fetal villi were much longer and thinner. In the single late stage, healthy young mare at 309 days of gestation (term=336 days), the microplacentomes, each of around 2 mm diameter, exhibited maximal length villi. The capillaries were arranged simply, mostly in straight lines along the axis of the villus, and with communications visible at irregular intervals. Simple and slightly more complicated side capillary loops could be seen along the whole length of the villi and at the top of the terminal villi. Most of the capillaries were characterized by zones containing dilated sinusoids, which increased the surface area for materno-fetal exchange. Thus, the morphological development of the microplacentomes on the surface of the horse placenta during gestation was studied, with special reference to the growth and organisation of the fetal and maternal capillary beds within each microplacentome. The study also reinforced previous work showing the disadvantageous influence of age-related endometrial degenerative changes on microplacentome development and on both the extent and intimacy of physical and haematological contact at the fetomaternal interface, and hence upon fetal growth.

Effects of a Streptococcus equi infection--mediated nutritional insult during mid-gestation in primiparous Thoroughbred fillies. Part 1: placental and fetal development.

REASONS FOR PERFORMING STUDY: There is a paucity of information on factors that influence placental development in the horse. HYPOTHESIS: Changes in nutrition, particularly around the time of proliferative placental growth, can affect development of the placenta and fetal growth. OBJECTIVE: To investigate the effects of 2 planes of nutrition and an unforeseen infection-mediated nutritional insult on placental and fetal development in the mare. METHODS: Twenty maiden Thoroughbred fillies, age 3 or 4 years, mated to one Thoroughbred stallion, were maintained on either High or Moderate food intake throughout pregnancy. In mid-gestation all the mares unexpectedly became infected with Streptococcus equi and suffered varying degrees of weight loss as a consequence. Gross and stereological measurements of the placenta and measurements of foal birthweight, crown-rump length, ponderal index and plasma IGF-1 concentration were made at term. RESULTS: Gains in bodyweight during gestation were significantly higher in the High vs. the Moderate nutrition groups. Placental and fetal growth parameters were not influenced by the plane of nutrition. However, transient weight loss in mid-gestation from acutely reduced food intake resulted in morphological changes to the allantochorion and decreased foal birthweight. CONCLUSIONS: Excessive weight gain during gestation in maiden Thoroughbred fillies does not result in nutrient partitioning to the dam at the expense of the fetus. However, sudden weight loss in mid-gestation has detrimental effects on placental development which results in reduced fetal growth. POTENTIAL RELEVANCE: Clinicians should appreciate the likely effects of maternal infection and resulting weight loss on pregnancy outcome.

Factors associated with failure of thoroughbred horses to train and race.

REASONS FOR PERFORMING STUDY: The low productivity associated with training and racing of young Thoroughbreds (TBs) in the UK due to athletic inability, injury and disease requires further study. OBJECTIVES: To identify the time points and causes of losses during growth and training phases of a cohort of 1022 TB foals born in 1999 up to the end of their third year. METHODS: Movement and fate of 1022 Thoroughbred foals conceived in 1998 and born live in 1999 were monitored from birth to age 3 years. Those (n = 562) that entered training age 2 and/or 3 years with one of 161 registered trainers in Britain or Ireland in 2001/2 were analysed as to their subsequent performance and the incidence of 9 common injuries or infective or metabolic illnesses. RESULTS: Of the 1022 foals identified, 537 (52%) entered training at age 2 years, 289 (28%) were exported, 60 (6%) were kept as 'stores' for National Hunt (NH) racing, 58 (6%) died or were destroyed, 25 (2%) were waiting to enter training at age 3 years, 17 (2%) were never intended for racing and 36 (4%) were untraceable. Race records showed that 327 (61%) of the 2-year-olds in training competed one or more times, 95 (18%) won and 165 (31%) were placed. Only 28 (5%) earned enough prize money to cover their training fees. Sore shins and inflammatory airway disease (IAD) were the 2 ailments most commonly encountered. In 2002, 431 (80%) of the previous year's 2-year-olds remained in training aged 3 years and 25 entered training having not raced previously. Of the 456 3-year-olds in training, 347 (76%) raced one or more times 138 (30%) won, 218 (48%) were placed and 78 (17%) recouped their training fees. Joint problems and sore shins were the ailments most commonly suffered and, with the exception of rhabdomyolysis ('tying up'), colts and geldings suffered a higher rate of muscloskeletal injuries than fillies. CONCLUSION: Previous reports of high nonrun and nonplaced rates, high incidence of injury and cost-ineffectiveness of 2-year-olds in flat-race training were confirmed. POTENTIAL RELEVANCE: This study lays the basis for further studies of the facets involved in wastage.

Meclofenamic acid extends donor-recipient asynchrony in equine embryo transfer.

REASONS FOR PERFORMING STUDY: A level of synchrony between embryo and uterine environment is essential for the establishment of pregnancy when performing embryo transfer. The ability to extend the acceptable degree of asynchrony would allow more efficient use of recipient mares. OBJECTIVES: To establish if administration of the prostaglandin synthetase inhibitor, meclofenamic acid, to asynchronous recipient mares could widen the acceptable window of asynchrony for embryo transfer. HYPOTHESIS: The prostaglandin synthetase inhibitory action of meclofenamic acid may act to suppress luteolysis and thereby allow for a greater degree of asynchrony between donor and recipient mares. METHODS: A total of 72 Grade 1 horse embryos were transferred nonsurgically into the uteri of recipient mares that had ovulated 2 (n = 20), 3 (n = 20), 4 (n = 16) or 5 (n = 16) days before the donor. Half of the mares in each group were treated orally with 1 g meclofenamic acid, beginning on Day 9 after ovulation and continuing for 7 days after embryo transfer. RESULTS: Comparison of recipient:donor asynchrony between treated and untreated mares was: +2 days, 9/10 pregnancies vs. 8/10 (P= 1.00); by +3 days, 8/10 vs. 2/10 (P= 0.025); by +4 days, 5/8 vs. 1/8 (P = 0.121); and by +5 days 3/8 vs. 0/8 (P = 0.20). In 10/11 meclofenamic acid-treated and 23/25 untreated recipient mares that failed to become pregnant, luteolysis occurred at the normal time (14-19 days) after ovulation. CONCLUSION: Treatment with meclofenamic acid supported the establishment of pregnancy in recipient mares that ovulated before the donors. However, meclofenamic acid appeared to provide this support in a manner other than by suppression of luteolysis. POTENTIAL RELEVANCE: Pregnancy rates can be increased in recipient mares that ovulate 3 days before the donor by administration of meclofenamic acid.

Laparoscopic application of PGE2 to re-establish oviducal patency and fertility in infertile mares: a preliminary study.

REASONS FOR PERFORMING STUDY: Mares are occasionally encountered that consistently fail to conceive when inseminated, naturally or artificially, with fertile stallion semen in the absence of any identifiable pathology of either the structure or function of their reproductive tract. HYPOTHESIS: Temporary blockage of the oviducts by accumulations of naturally occurring oviducal masses may be preventing oviducal transport of the embryo to the uterus. METHODS: Mares, with known reproductive histories, that had exhibited inexplicable failure of conception were treated by laparoscopically guided administration of PGE2-laced triacetin gel directly onto the surface of their oviducts. RESULTS: Fifteen mares age 10-21 years that had exhibited inexplicable failure of conception during 1-4 years were treated, of which 14 (93%) conceived within the same or subsequent breeding season. CONCLUSIONS: The high success rate of this treatment supports the tentative diagnosis of oviducal obstruction in these mares and indicates that blockage of the mare's oviducts may occur in the form of a moveable accumulation of debris rather than from permanent fibrous adhesions resulting from salpingitis. POTENTIAL RELEVANCE: This laparoscopic application of PGE2 to the oviducts constitutes a sound and practical method of restoring fertility in mares suffering oviducal obstruction and further studies involving the procedure are warranted.

Immunocytochemistry of the placentas of giraffe (Giraffa cameleopardalis giraffa) and okapi (Okapi johnstoni): comparison with other ruminants.

INTRODUCTION: The trophoblast binucleate cell [BNC] is central to the structure and function of all ruminant placentas so far investigated. The Giraffidae are considered to form a separate family within the ruminant suborder. METHODS: The structure and function of two [mid and late pregnant] giraffe placentas and two term okapi placentas have been investigated immunocytochemically. RESULTS: Their major characteristics: polycotyledonary epitheliochorial structure, sequential glucose transport using two transporter isoforms, expression of water transporters in the interplacentomal [IP] and placentomal [P] trophoblast and restriction of calcium transport to the IP trophoblast are similar to those of the ruminant family Bovidae. . Giraffe and okapi also show characteristic ruminant trophoblast binucleate cells (BNC) which migrate and fuse with individual uterine epithelial cells as in the cow. However, there are many fewer BNC, of limited distribution, when compared with other ruminants so far investigated. The giraffe and okapi BNC also show a different range of proteins, Pregnancy Associated Glycoproteins (PAGs) and glycans which clearly distinguish the Giraffidae from the Bovidae. CONCLUSIONS: The results support a separate giraffid family derived from a common ancestry, possessing subpopulations of BNC with potentially different functions.

The influence of maternal size on placental, fetal and postnatal growth in the horse. II. Endocrinology of pregnancy.

Within-breed artificial insemination and between-breed embryo transfer were carried out in small pony (P) and large Thoroughbred (Tb) mares to create 4 types of horse pregnancy in which the fetus experienced spatial and nutritional deprivation (Tb-in-P; n=8), luxury (P-in-Tb; n=7) or normality (Tb-in-Tb; n=7 and P-in-P; n=7) in utero. Measurement of equine chorionic gonadotrophin (eCG), total conjugated oestrogens and progestagen concentrations in serial peripheral serum samples recovered from all the mares throughout gestation showed that the amount of eCG produced during the first half of gestation was dependent upon the breed of the mare rather than the breed of the fetus being carried. In contrast, the mean total amounts of oestrogens produced, as measured by area under the curve, were significantly greater (P=0.003) in the two types of pregnancy in which a Thoroughbred fetus was being carried (Tb-in-Tb and Tb-in-P) than those in which a pony fetus was gestated (P-in-P and P-in-Tb); the evidence suggests that the Tb fetus may have larger gonads than the P fetus and thereby secrete more C-19 precursor steroids for aromatisation to oestrogens by the placenta. In the final weeks of pregnancy mean plasma progestagen concentrations rose much earlier, and to significantly higher levels (P<0.001), in the Tb-in-P than in the P-in-Tb pregnancies, thereby reflecting the increased fetal stress in the former causing premature maturation of the fetal adrenal gland. This, in turn, resulted in increased secretion of pregnenolone by the adrenal cortex for conversion to progestagens by the placenta.

The effects of maternal age and parity on placental and fetal development in the mare.

REASONS FOR PERFORMING STUDY: The normality of equine placentation is essential for fetal health and development. Substantial information exists on the gross morphological status of the placenta but few studies have addressed the problem of degenerative lesions that interfere with placental morphology and placental efficiency. HYPOTHESIS: Degenerative changes in the endometrium with increasing age and parity are reflected in the morphology and density of the placental microcotyledons. OBJECTIVE: To assess placental efficiency on the basis of foal birthweight as a function of total microscopic area of fetomaternal contact. METHODS: Stereology was used to examine the placentae of 84 Thoroughbred mares grouped on the basis of age and parity. Placental efficiency, assessed by expressing foal birthweight as a function of total microscopic area of fetomaternal contact, was also determined. RESULTS: Mare age and parity influenced the development of the microcotyledons and microcotyledon surface density (Sv) was lowest in aged multiparous mares, presumably due to degenerative changes in their endometrium. However, primiparous mares also showed significantly lower Sv values than young multiparous mares, despite the virginal endometrium of the former group. This apparent 'priming' effect of a first pregnancy on microcotyledonary Sv was illustrated further by 11 maiden mares followed in 2 successive pregnancies. They all showed significant increases in Sv values in their second parity, with equivalent improvements in foal birthweight. CONCLUSIONS: Foal birthweight is a reflection of the balance between fetomaternal contact and placental efficiency. Increases in fetomaternal contact are correlated to reductions in placental efficiency, which may reflect the ability of the placenta to modify its exchange capabilities. POTENTIAL RELEVANCE: Further work is needed to elucidate how such changes in microcotyledon morphology and efficiency are brought about.