The effect of insemination volume on pregnancy rates of pony mares

It has recently been reported that large insemination volumes might affect fertility of mares. The results from these studies are confounded by other factors, however, such as inadequate number of spermatozoa in the inseminate. We conducted a study to test whether volume alone affects fertility when sufficient numbers of spermatozoa are present. Semen from one stallion was collected, extended at 50 x 10(6) spermatozoa/ml, and stored in a commercial semen cooling device for 18 to 30 h before insemination. Ten pony mares were assigned during estrus in random pairs to be bred every other day with either 30 or 120 ml of extended cooled semen. Pregnancy was diagnosed by ultrasonography per rectum on Days 11, 12 and 13 after ovulation. On Day 13, the mares were given a luteolytic dose (5 mg) of PGF(2alpha). On the subsequent cycle, the mares were given the alternate treatment. The pregnancy rates in the 30- and 120-ml insemination volume groups were 7 9 and 10 10 , respectively; this difference was not significant (P=0.2). Embryonic growth from Day 11 to Day 13 was not different (P>0.05) between groups. We conclude that insemination volumes as large as 120 ml have no adverse effect on fertility.     

The effect of pregnancy and foaling on intravaginal pressure in pony mares

Pressures were recorded monthly at two sites in the vagina of each of five pregnant and five nonpregnant pony mares; pressures in five mares were also recorded weekly after foaling. The developing pregnancy did not influence pressure, and after foaling the integrity of the seal formed by the caudal reproductive tract was rapidly restored.     

The maternal recognition of pregnancy in mammals

Progesterone secreted by the corpora lutea stimulates uterine secretion of nutrients required for embryonic growth. But progesterone also blocks maturation of ovarian follicles and oestrous behaviour. When conception fails, therefore, the corpora lutea must lyse, in order to allow a second ovulation and oestrus, while in pregnancy the corpora lutea must be maintained. A number of different endocrine mechanisms exist to ensure the corpora lutea are controlled appropriately depending on pregnancy status; among the best studied of these are the secretion by the developing conceptus of interferons (in ungulates) or chorionic gonadotropins (in primates). These and other mechanisms are reviewed.     

Protein production by sheep embryos during the period of maternal recognition of pregnancy

An embryo must be present in the uterus 12-13 days after estrus to prevent regression of the ovine corpus luteum. The present experiments were designed to determine if embryo-specific secretory proteins could be detected in the maternal blood at the time of maternal recognition of pregnancy. In two experiments, 92 embryos were flushed from 47 ewes at 14-15 days after estrus. Embryos were incubated in vitro for 24 h and the proteins in the media were harvested. Antisera to proteins in both flushing and incubation medium were produced in rabbits. In experiment 1, crude fractions were used for antibody production and radioimmunoassays were established for protein peaks separated on a 1.1 X 75 cm G-100 Sephadex column. Two low molecular weight fractions (EPiv and EPv) appeared to be embryo specific but were not detectable in jugular vein sera of 14- to 15-day pregnant animals. In experiment 2, proteins derived from uterine flushes and from embryo incubations were chromatographed on a 2.5 X 85 cm column of G-100 Sephadex. The protein peaks were measured, pooled, lyophilized, and used for immunization of rabbits. As in experiment 1, antisera were generated, some of which seemed to be directed against embryo-specific proteins. However, we could not detect these fractions in the uterine vein blood of pregnant animals. Thus, embryo-specific proteins are either confined to the uterus or they appear in the blood in quantities that are undetectable with our assay system.     

The effect of the gonadotropin-releasing hormone analog, buserelin, on pregnancy rates in horse and pony mares

We conducted a series of trials over a four-year period on a total of 2,346 mares, to determine the effect of a single dose of the GnRH analog buserelin (20 to 40 microg i.m. or s.c.) on pregnancy rates when given between 8 and 12 days after service. Although there were some statistically significant improvements in pregnancy rates in individual trials, meta-analysis of the data overall showed significant improvements at all times examined, i.e. 13 to 16, 19 to 23, 28 to 31 and 38 to 42 days after service. These results indicate that treatment of mares with 20 to 40 microg buserelin between Days 8 and 12 significantly increases pregnancy rates by approximately 10 percentage points.     

Effects of melatonin implants in pony mares. 1. Acute effects

The effects of melatonin implant treatment over a four week period on LH, estradiol (E2) and progesterone (P4) secretion during the breeding season were studied in ovary-intact and ovariectomized pony mares. Mares with melatonin implants had significantly higher daytime melatonin concentrations than mares with sharm implants (P = 0.0065). In ovariectomized mares, LH secretion did not differ between mares with melatonin and sham implants. In ovary-intact mares, melatonin implants altered the pattern of LH secretion (P = 0.0023) in such a way that an increase in LH secretion was observed during the periovulatory period. Estradiol and P4 secretion were unaffected by melatonin implants. These results suggest that constant administration of melatonin may enhance the secretion of LH during the periovulatory surge but does not adversely affect E2, P4 or basal LH secretion in mares during the breeding season.     

Effects of melatonin implants in pony mares. 2. Long-term effects

The effects of melatonin implant treatment over a 4 wk period at the summer solstice on the transition into and out of the following anovulatory season were evaluated in ovary-intact and ovariectomized mares. Melatonin implants tended to delay the timing of the final ovulation of the breeding season (P = 0.0797) in the ovary-intact mares. Although the decline in LH secretion associated with the end of the breeding season was parallel between treatments and ovarian statuses, the rate of LH secretion, as expressed by its mathematical accumulation, was lower in ovariectomized, melatonin-treated mares than in ovariectomized, control mares suggesting that melatonin administration advanced the offset of the breeding season in ovariectomized mares (P = 0.0001). The first ovulation of the subsequent breeding season was significantly delayed in the melatonin-treated mares as compared with that of control mares (P = 0.0031). During reproductive recrudescence, the time of the onset of the increase in LH secretion was similar among all 4 groups but the patterns of LH secretion were different for each treatment and ovarian status combination (P = 0.0112). Mares with melatonin implants had a slower rate of increase in LH secretion than control mares (P = 0.0001), and ovariectomized mares had a faster rate of LH increase than intact mares (P = 0.0001). These results suggest that melatonin implants during the summer solstice can alter the annual reproductive rhythm in mares and support the concept that endocrine patterns of reproductive recrudescence are not entirely independent of the ovary.     

The effect of human chorionic gonadotrophin and exogenous progesterone on luteal function during early pregnancy in pony mares

Previous reports have indicated that repeated administrations of human chorionic gonadotrophin (hCG) to pony mares before day 38 of gestation caused pregnancy failure, but that this could be prevented by the prior or simultaneous administration of progesterone. The present paper describes the circulating progestagen concentrations during these experiments, and shows that pregnancy failure resulted from luteolysis after hCG administration. Most of the mares which received progesterone maintained their pregnancies after hCG injection, but progestagen concentrations fell, suggesting that partial lysis of the corpus luteum occurred. After day 38 of gestation repeated doses of hCG had no effect on plasma progestagen concentrations.     

Role of interferons in maternal recognition of pregnancy in ruminants.

It has recently become evident that a type I interferon (IFN) subtype signals the presence of a viable conceptus to the mother during early pregnancy in cattle, sheep, and related mammalian species. This IFN, which is a product of the epithelium (trophectoderm) of the expanding trophoblast, is expressed in extremely large quantities for a few days just prior to implantation. It appears to be involved in modulating the release of the luteolytic hormone, prostaglandin F2 alpha, from the uterine endometrium and, hence, preventing the destruction of the corpus luteum that normally occurs at the end of an estrous cycle if an egg has not been fertilized. These trophoblast IFN have antiviral, antiproliferative, and immunomodulatory properties quite similar to other type I IFN, such as IFN-alpha, -beta, and -omega. However, they constitute a structurally and serologically distinct subtype. In addition, they are poorly inducible by virus, and the promoter regions of their genes are organized differently than other type I IFN. The genes for these trophoblast IFN are confined to ruminant species in the Artiodactyla order and probably evolved from IFN-omega less than 55 million years ago. There is no evidence for comparable production of type I IFN by trophoblast and placental tissues of mammals outside this ruminant group. Recent experiments have indicated that IFN treatment may have value in improving reproductive performance of sheep when provided during the period of maternal recognition of pregnancy, when much embryonic loss is believed to occur.     

Time of ovarian follicular recruitment in cyclic pony mares

An experiment was carried out on pony mares to establish the time of the oestrous cycle at which ovarian follicles are recruited for ovulation. In one group (n=7), the cycle was interrupted at the preovulatory stage by removing the preovulatory follicle; in another group (n=13) the cycle was interrupted at day 6 of the luteal phase by inducing luteolysis with a prostaglandin injection (PG). In a subgroup (n=7) of those given PG, the ovary not bearing the corpus luteum was removed at the time of injection. A further group (n=6) served as surgical controls. The interval to the next ovulation and blood concentrations of FSH were observed. Anaesthesia alone induced in preovulatory mares was followed by normal ovulation 2.5+/-1 days later. Removal of the preovulatory follicle delayed the next ovulation (14.6+/-2.1 days; P < 0.01). Following PG injection, the interval to ovulation was similar regardless of whether an ovary was removed (12.8+/-4.3 days) or not (10+/-4.1 days). This similarity occurred despite a large and prolonged rise in plasma FSH levels that occurred only in the hemiovariectomized group. In addition, the intervals found after PG injection did not differ from those found after ablation of the preovulatory follicle. These observations indicate that 1) in the presence of the early active corpus luteum or dominant follicle, follicles grow to a similar stage of development; 2) recruitment of the follicle due to ovulation occurs 12 to 14 days before ovulation; 3) limiting new follicular growth to one ovary does not affect the time course to ovulation; and 4) prolonged high FSH levels do not alter the time course or ovulation rate.     

Responses of pony mares to the agent of contagious equine metritis 1977.

Reproduction of contagious equine metritis 1977 in Pony mares was achieved with cultures of an unclassified Gram-negative coccobacillus. Infected mares developed a vaginal discharge and associated inflammatory changes of the cervix and vagina. There was evidence of variation in pathogenicity between different strains of the organism. Although all infected mares made spontaneous clinical recoveries, the Gram-negative coccobacillus persisted in the genital tracts of a considerable proportion for a variable period after challenge. Recovery of the organism was not associated solely with the occurrence of oestrus. None of the mares has carried over infection into the following breeding season. There was no evidence of localization of the organism in the urinary tract. Cytological examination of smears of cervical and urethral swabs was of diagnostic value only during the clinical phase of the infection. A serological response was demonstrable in all mares that became infected after exposure to the Gram-negative coccobacillus. The complement-fixation test gave more specific and clear-cut results than either the agglutination or the antiglobulin test, with which there was a problem with non-specific reactions. The experimental findings indicate the value of the complement-fixation test for confirming recent cases of contagious equine metritis in the mare.     

Ovarian oxytocin and the maternal recognition of pregnancy

The secretion of oxytocin by the corpus luteum is thought to stimulate the episodic release of PGF-2 alpha from the uterus, thereby contributing to luteolysis. In pregnancy corpus luteum function is maintained, and secretion of oxytocin, or its actions on the uterus, appear to be inconsistent with the successful establishment of gestation. Protection against the effects of oxytocin is ensured by a number of mechanisms, including the cessation of luteal oxytocin secretion, which is evident by Day 20 after mating in sheep, and the maintenance of low levels of the oxytocin receptor in the uterus.     

Hemostatic profile during late pregnancy and early postpartum period in mares

Hemostasis is a physiological process that prevents excessive blood loss and represents a protective mechanism at the time of delivery. Peripartum hemorrhage is a recurring hazardous condition to mare's health; therefore, we aimed to study mares' hemostatic profile to investigate whether physiological adjustments occur during late pregnancy and early postpartum. Fifteen pregnant mares have been monitored from the 34th week of pregnancy until the third week after foaling. Fifteen nonpregnant mares were used as control group. Jugular blood samples were analyzed for platelet count (Plt), prothrombin time (PT), activated partial thromboplastin time (aPTT), and fibrinogen (Fb). Platelet count showed significant changes at foaling (P < 0.05) and a negative correlation (r = −0.968; P = 0.032) with postpartum. Prothrombin time changed (P < 0.05) showing a significant correlation (r = 0.675; P = 0.016) with late pregnancy. Fibrinogen concentrations changed throughout the experimental period (P < 0.0001). The linear regression model revealed a positive correlation (r = 0.9210; P < 0.0001) between Fb and late pregnancy and a negative correlation (r = −0.9583; P = 0.042) between Fb and early postpartum. The shortening in PT recorded in the imminence of parturition along with the increase in Plt and Fb at foaling might reflect a physiological hypercoagulable state that constrains excessive bleeding, enhancing mares' odds of surviving. Our research improves the knowledge about blood coagulation in periparturient mares providing specific information on routine coagulation tests that may support in monitoring mare's hemostatic profile during late pregnancy and early postpartum.     

Puromycin-sensitive aminopeptidase is essential for the maternal recognition of pregnancy in mice

Maternal recognition of pregnancy in rodents requires semicircadian surges of hypophyseal PRL secretion during early gestation, which are required for the formation of the corpus luteum of pregnancy (CLP). Here we show that puromycin-sensitive aminopeptidase (Psa)-deficient mice display female infertility that results from impaired formation of CLP. Transplantation of mutant ovaries into normal females restored fertility but not vice versa. Psa-deficient females revealed no semicircadian surges of PRL induced after mating stimuli. Pregnancy in the mutant females was restored by grafting intact pituitaries to elevate circulating levels of PRL. Psa is thus required for the appearance of the semicircadian surges of PRL secretion that are crucial for maintaining pregnancy in rodents.     

Conceptus interferons and maternal recognition of pregnancy

Ovine trophoblast protein-1 (oTP-1) and bovine trophoblast protein-1 (bTP-1) are secreted by ovine and bovine conceptuses for a restricted period when the conceptus acts to block regression of the corpus luteum. Both are considered to be antiluteolytic, acting locally on the endometrium to prevent the production of the uterine luteolysin prostaglandin F2 alpha. Molecular cloning of cDNAs for oTP-1 and bTP-1 has shown that both are structurally related to alpha interferons (IFN-alpha s). In particular, they resemble members of the 172-residue long IFN-alpha II (or IFN-omega) subfamily. Both proteins possess the antiviral and antiproliferative properties of IFN-alpha s and appear to bind IFN-alpha receptors. A novel and previously unsuspected role for IFNs in early pregnancy is apparent.     

Response of plasma LH and FSH to gonadotropin-releasing hormone in pony foals and ovariectomized pony mares

Plasma FSH and LH response to a synthetic GnRH analog was measured in adult ovariectomized pony mares (OVX) and in pony foals (<70 days of age) during late spring (May-June). FSH and LH responded in a similar fashion (200% increase) in the OVX mare, which is different from other reports for intact mares. There was a greater mean response to a comparable dose of GnRH in the prepubertal foal for both FSH (500%) and LH (900%) than in the OVX mare. There was a positive correlation between age and the maximum FSH response to GnRH in male and female foals. The LH response was positively correlated with age in male foals, but not in females. The response to GnRH in the prepubertal foals was consistent with the previously observed patterns of gonadotropin secretion during this age period.     

An investigation of the uterine luminal environment of non-pregnant and pregnant pony mares.

Uterine flushings were collected from 30 non-pregnant Pony mares on Days 8, 12, 14, 16, 18 or 20 after oculation. Mares were allowed a recovery period of one oestrous cycle and were mated at the next oestrus. They were then ovario-hysterectomized on days which corresponded to the day of the oestrous cycle to which they were assigned. Uterine flushings were analysed for total recoverable protein and acid phosphatase activity. Least squares analysis indicated a status X day interaction for total protein (P less than 0.10) and acid phosphatase activity (P less than 0.005) in which the latter was higher in uterine flushings during pregnancy. Peripheral plasma oestrone and oestradiol concentrations were measured by radioimmunoassay and results indicated that plasma oestrone concentrations in pregnant and non-pregnant mares were not different, and oestradiol was lower (P less then 0.005) in the peripheral plasma during pregnancy. conceptus membranes were incubated in vitro for 120 min in a chemically defined medium. Incubation medium was then assayed to assess oestrone and oestradiol production capacilities at Days 8, 12, 14, 16, 18 and 20 of pregnancy. Conceptus membrane production of oestradios (pg/5 ml/h) increased (P less than 0.05) from Day 8 (243 pg/5 ml) to Day 20 (108 763 pg/5 ml). A similar trend, but of lower magnitude, existed for oestrone production.