Persistent breeding-induced endometritis after hysteroscopic insemination in the mare

Low-dose insemination has been proposed to reduce persistent breeding-induced endometritis (PBIE) in mares with delayed uterine clearance (DUC). Others proposed that hysteroscopic insemination induces an exaggerated inflammatory response and should be avoided in DUC mares. The objectives here were to evaluate presence and severity of PBIE in normal and DUC mares after hysteroscopic insemination with fresh semen, and to determine if hysteroscopy could be used in DUC mares without inducing excessive inflammation. Reproductively normal (n = 4) and DUC (n = 5) mares received four treatments in random order: uterine body insemination (UB, 1 × 10(9) spermatozoa, 20 ml), hysteroscopic insemination (HYST, 5 × 10(6) spermatozoa, 0.5 ml), sham hysteroscopic insemination (SHAM, semen extender, 0.5 ml) and hysteroscopic infusion of seminal plasma (SP, 0.5 ml). Significantly more DUC (50%) mares than normal (14%) mares accumulated intrauterine fluid 24 h post-treatment. The difference in fluid accumulation between DUC (40%) mares and normal (7%) mares was also significant 48 h post-treatment. Fluid scores were not significantly different between treatments in normal mares. However, treatments HYST and SHAM resulted in significantly higher fluid scores 24 h but not 48 h post-treatment in DUC mares. There was no effect of treatment or mare group on the percentage and total number of neutrophils in uterine fluid 48 h post-treatment. Percentage of neutrophils was correlated with duration of hysteroscopy in normal mares, with procedures lasting ≥ 9 min associated with PBIE. There was no effect of mare group, treatment or duration of hysteroscopy on pregnancy rate. Hysteroscopy induces a transient inflammation that is not more severe than that after conventional artificial insemination, suggesting no contraindication to its use in DUC mares.     

Termination of pseudopregnancy by administration of prostaglandin F2alpha and termination of early pregnancy by administration of prostaglandin F2alpha or colchicine or by removal of embryo in mares.

At day 24 of gestation, pregnant mares were allotted to 1 of 5 treatment groups (3 to 5 mares/group): group A--nontreated controls; group B--intraembryonic injection of 4 mg of colchicine on day 24; group C--removal of embryo on day 24; group D--subcutaneous injection of 1.25 mg of prostaglandin F2alpha (PGF2alpha) on day 32; and group E--removal of embryo on day 24 and subcutaneous injection of PGF2alpha on day 32. In all mares treated with colchicine (group B), the fetal bulge was absent within 2 days. The interval from injection of colchicine to onset of estrus was very short (mean, 4 days). These results indicated that treatment with colchicine was lethal to the 24-day embryo, and pseudopregnancy did not occur. Surgical removal of the embryo (group C) resulted in pseudopregnancy characterized by a prolonged interval from treatment to return to estrus (mean, greater than 31 days), prolonged production of progesterone, and prolonged maintenance of tense uterine and cervical tone. The interval from treatment to ovulatory estrus was longer (P less than 0.05) for group C mares than for group B mares. The mean interval from treatment to complete loss of tense tubular uterine tone was not significantly different between group A pregnant controls (28.3 days) and group C pseudopregnant mares (30 days). Treatment of pregnant mares (group D) with a single injection of PGF2alpha on day 32 resulted in loss of pregnancy in 4 of 4 mares within 2 to 5 days, and in all group D mares a large decrease in progesterone concentration occurred on day 33, 34, or 35. Although subsequent reproductive activity was variable, all group D mares rapidly lost the tense uterine and cervical tone characteristic of early pregnancy. These results indicated that a single subcutaneous injection of 1.25 mg of PGF2alpha caused loss of pregnancy, and pseudopregnancy did not occur. Treatment of group E mares, which had been made pseudopregnant by removal of embryo, with 1.25 mg of PGF2alpha resulted in termination of pseudopregnancy in 5 of 5 mares. All group E mares returned to estrus within 2 to 5 days after treatment, and progesterone concentration decreased (P less than 0.05) within 2 days after treatment. There was no significant difference in loss of tense tubular uterine or cervical tone between pregnant (group D) and pseudopregnant (group E) mares after PGF2alpha treatment.     

Low Concentration of Anti-Müllerian Hormone in Mares with Delayed Uterine Clearance

Luteal dysfunction has been observed in mares with defective uterine clearance. Association of low ovarian reserve with luteal dysfunction and abnormal endometrial thickness has been reported in bovine. Anti-Müllerian hormone (AMH) has been indicated as a marker for ovarian reserve in bovine and originates primarily from the ovary in equine. The present study evaluated serum AMH concentration in mares with delayed uterine clearance versus that in mares without delayed uterine clearance. Of 49 mares assigned to the study, 12 and 37 mares were diagnosed with and without delayed uterine clearance, respectively. Delayed uterine clearance was determined based on history and the observation of intrauterine fluid in ultrasonographic examination 24 hours after natural breeding. Serum AMH concentration was measured during estrus. Concentration of AMH was lower in mares with delayed uterine clearance (0.4 ± 0.1 ng/mL) than in those without delayed uterine clearance (1.1 ± 0.1 ng/mL). In conclusion, the present study indicated possible associations between ovary-lined mechanisms and uterine clearance failure in mares.     

Inhibitory effects of CIDR-based ovulation-synchronization protocols on uterine PGF2alpha secretion at the following luteal phase in early postpartum non-cycling beef cows

We investigated whether CIDR-based ovulation-synchronization protocols inhibit secretion of prostaglandin (PG) F2alpha from the uterus in the following luteal phase in non-cycling beef cows. Ten early (a month) postpartum non-cycling Japanese Black beef cows were treated with (1) Ovsynch (GnRH analogue on Day 0, PGF2alpha analogue on Day 7, and GnRH analogue on Day 9; n=3), (2) Ovsynch+CIDR (Ovsynch protocol plus a CIDR for 7 days from Day 0; n=4), or (3) estradiol benzoate (EB) Ovsynch+CIDR (EB on Day 0 in lieu of the first GnRH treatment followed by the Ovsynch+CIDR protocol; n=3). An oxytocin challenge was administered on Day 24 to examine uterine PGF2alpha secretion. Plasma concentrations of 13,14-dihydro-15-keto- PGF2alpha were lower at 30-120 min after oxytocin administration in the Ovsynch+CIDR group and 75 min after administration in the EB Ovsynch+CIDR group than in the Ovsynch group (P<0.05). Plasma progesterone concentrations were higher from Days 1 to 7 in the Ovsynch+CIDR group and from Days 1 to 5 in the EB Ovsynch+CIDR group than in the Ovsynch group (P<0.05). The progesterone concentrations were higher on Days 27 and 29 in both CIDR-treated groups than in the Ovsynch group (P<0.05). In conclusion, in non-cycling beef cows, CIDR-based ovulation-synchronization protocols inhibit uterine PGF2alpha secretion in the following luteal phase and prevent premature luteolysis as is seen with the Ovsynch protocol.     

Making sense of equine uterine infections: the many faces of physical clearance

Equine uterine infections inflict major losses on the equine industry. Persistent inflammation of the oviduct and uterus leads to loss of the conceptus and mares susceptible to infection have weakened uterine defences partly due to retention of inflammatory exudate. Bacteria may trigger inflammation, resist phagocytosis, or adhere to the endometrium and types of infection range from genital commensals in susceptible mares to reproductive pathogens in normal mares. Uterine infections are diagnosed by history, detection of uterine inflammation, and isolation of typical organisms and susceptible mares may be identified by detection of intrauterine fluid during oestrus, or at 6-48 h post-breeding. Therapy includes oxytocin, uterine lavage, antibiotics, and prostaglandin analogues and clinical studies indicate additive benefits of oxytocin and antibiotics. Improved conception rates have been associated with autologous, intrauterine plasma, despite controversy about its bactericidal efficacy. Because of the potential for endometrial damage, intrauterine antiseptics require caution.     

Equine post-breeding endometritis: A review

The deposition of semen, bacteria and debris in the uterus of the mare after breeding normally induces a self-limiting endometritis. The resultant fluid and inflammatory products are cleared by 48 hours post cover. Mares that are susceptible to persistent post-breeding endometritis (PPBEM) have impaired uterine defence and clearance mechanisms, making them unable to resolve this inflammation within the normal time. This persists beyond 48 hours post-breeding and causes persistent fluid accumulation within the uterus. Mares with PPBEM have an increased rate of embryonic loss and a lower overall pregnancy rate than those without the condition. To enhance conception rates, mares at high risk need optimal breeding management as well as early diagnosis, followed by the most appropriate treatment. This article reviews the pathogenesis, diagnosis and treatment of PPBEM and the management of affected mares.     

Effects of flunixin meglumine on endotoxin-induced prostaglandin F2 alpha secretion during early pregnancy in mares

The role of prostaglandin F2 alpha (PGF2 alpha) in embryonic loss following induced endotoxemia was studied in mares that were 21 to 44 days pregnant. Thirteen pregnant mares were treated with a nonsteroidal anti-inflammatory drug, flunixin meglumine, to inhibit the synthesis of PGF2 alpha caused by Salmonella typhimurium endotoxin given IV. Flunixin meglumine was administered either before injection of the endotoxin (group 1, -10 min; n = 7), or after endotoxin injection into the mares (group 2, 1 hour, n = 3; group 3, 2 hours, n = 3); 12 pregnant mares (group 4) were given only S typhimurium endotoxin. In group 4, the secretion of PGF2 alpha, as determined by plasma 15-keto-13,14-dihydro-PGF2 alpha concentrations, was biphasic, initially peaking at 30 minutes followed by a second, larger peak approximately 105 minutes after the endotoxin was given IV. When flunixin meglumine was administered at -10 minutes, synthesis of PGF2 alpha was inhibited for several hours, after administration of flunixin meglumine at 1 hour, the second secretory surge of PGF2 alpha was blocked, and administration of the drug at 2 hours did not substantially modify the secretion of PGF2 alpha. Plasma progesterone concentrations were unchanged after endotoxin injections were given in group 1. In group 2, progesterone values decreased less than 2 ng/ml and remained low for several days. In group 3 and group 4, progesterone concentrations decreased to values less than 0.5 ng/ml by 48 hours after endotoxin injections were given.(ABSTRACT TRUNCATED AT 250 WORDS)     

Concentrations of uterine luminal prostaglandins in mares with acute and persistent endometritis

Intrauterine infusion of 1 per cent oyster glycogen solution was used to induce acute endometritis in four genitally normal mares. Numbers of viable neutrophils recovered in uterine washings had increased by 1 h after infusion and remained elevated for at least 72 h. There was a significant correlation between numbers of viable neutrophils and total protein concentrations and between prostaglandin (PG)F and PGE2 concentrations in washings. There was also a significant relationship between concentrations of 15-keto-13, 14-dihydro PGF2 alpha in plasma and PGF in washings. Intrauterine concentrations of PGF were influenced by cycle stage and in turn the induced acute endometritis interfered with normal ovarian function. Mares with persistent endometritis had significantly higher concentrations of PGF and total protein and percentage of neutrophils and mononuclear cells in washings than normal mares. White blood cells from mares were capable of producing PGF and PGE2 in vitro.     

Recording uterine motility in the nonanesthetized bitch

Spontaneous and drug-induced uterine motility (UM) was recorded in 5 nonanesthetized bitches for 2 to 4 days. Catheter-tip pressure transducers were surgically implanted in 1 uterine horn, tunneled subcutaneously to exit from the skin over the dorsal lumbar area, and protected by a bandage. On the day after implantation, spontaneous UM was recorded in the awake bitch. Effects of IV prostaglandin (PG) F2 alpha (5 micrograms/kg of body weight) and oxytocin (0.05 USP U/kg) and IM PGF2 alpha (25 micrograms/kg) were measured. Estradiol (1 to 25 micrograms/kg) was administered and the study was repeated 24 hours later. In awake bitches, spontaneous UM was 190% greater than UM in anesthetized bitches. Uterine motility was increased by more than 100% after IV PGF2 alpha or oxytocin and by 52% after IM PGF2 alpha. Estradiol abolished spontaneous UM, but did not affect drug-induced responses. Seemingly, spontaneous and drug-induced UM can be documented in the nonanesthetized bitch.     

Effectiveness of a two-dose regimen of prostaglandin administration in inducing luteolysis without adverse side effects in mares

Our objectives were to determine whether repeated administration of prostaglandin F2alpha (PGF2alpha) to simulate the endogenous mode of secretion would be more effective than a single injection in inducing luteolysis and enable use of smaller doses less likely to cause adverse side effects. The main study comprised 43 dioestrous mares, who were given im. either a single 10 mg dose of natural PGF2alpha (n = 22) or 2 doses of 0.5 mg PGF2, 24 h apart (n = 21). The intensity of side effects was assessed in 8 dioestrous mares given 5, 1.5, 0.5 or 0 mg PGF2alpha in consecutive cycles. Two doses of 0.5 mg PGF2alpha 24 h apart caused lysis of the corpus luteum in all mares, whether this was determined from a fall in plasma progesterone concentrations or reproductive tract/behavioural changes; and when 10 mg PGF2, was given, the corpus luteum was lysed in 17 of 22 mares i.e. a lower proportion (P = 0.0485). A single dose of 0.5 mg PGF2a was no more effective than saline in inducing luteolysis.The intensity of side effects of PGF2alpha increased with dose. Although the 0.5 mg dose was no more likely than saline to cause sweating or muscle spasms, it raised plasma cortisol concentrations and prevented the decline in heart rate seen after saline. We conclude that a 2 dose regimen of administration increases the luteolytic efficacy of PGF2alpha and thereby provides a way to minimise adverse side effects.     

Effect of constant infusion of oxytocin on luteal lifespan and oxytocin-induced release of prostaglandin F2α in heifers

Two experiments were conducted to determine whether constant infusion of oxytocin would prolong the luteal phase and inhibit uterine prostaglandin F2 alpha (PGF2 alpha) secretion in heifers. In Experiment 1, twelve heifers, treated with saline (SAL) or oxytocin (OXY) via jugular cannulae infusions (INF) or osmotic minipumps (OMP), were allotted at estrus into four treatment groups (n = 3). Treatments were: SAL-INF, SAL-OMP, OXY-INF and OXY-OMP. Physiological saline or oxytocin was given from Days 10 to 23 (Day 0 = estrus) of the estrous cycle. Method of treatment (jugular cannula infusion or osmotic minipump) had no effect (P greater than 0.05) on estrous cycle length or pattern of secretion of progesterone; therefore, data were pooled. Estrous cycle lengths were extended (P less than 0.01) for heifers which received oxytocin (25.3 +/- 0.4 d) compared to saline (20.5 +/- 0.4 d). Luteolysis did not occur in oxytocin-treated heifers until after treatment ceased. Experiment 2 was designed and conducted identically to Experiment 1 with the addition of a "challenge" injection of oxytocin (100 IU oxytocin, i.v.) given on Day 16 of the estrous cycle. Treatment of heifers with oxytocin extended (P less than 0.05) estrous cycle length by an average of 3 d compared to heifers treated with saline. The "challenge" injection induced (P less than 0.05) secretion of PGF2 alpha (as measured by the stable PGF2 alpha metabolite, 15-keto-13,14-dihydro-PGF2 alpha) in saline-treated but not oxytocin-treated heifers. In both Experiment 1 and 2, serum concentrations of FSH were elevated (P less than 0.05) in oxytocin-treated heifers. No increase was observed for LH or prolactin. The rise in estradiol-17 beta at luteolysis was not affected (P greater than 0.10) by treatment. In summary, constant infusion of oxytocin extended luteal lifespan, prolonged secretion of progesterone, and inhibited oxytocin-induced secretion of PGF2 alpha. Constant infusion of oxytocin did not affect serum concentrations of estradiol-17 beta, LH or prolactin; however, serum concentrations of FSH were elevated during the oxytocin treatment period.     

The effect of two different routes of administration of oxytocin on peripheral plasma prostaglandin F(2α) metabolite levels in early post-partum dairy cows

Various parenteral treatment forms of oxytocin, as often used under praxis circumstances, may act differently on contractility of the uterus during the first days of the puerperium. Various patterns of such induced uterotonic responses may lead to alterations in the emptying characteristics of the uterine lumen, thus influencing, as a late consequence, the process of involution. Therefore, this study was designed to test whether two different parenteral administration forms of oxytocin induce changes in peripheral plasma concentrations of 15-ketodihydro-prostaglandin F(2α) (PGF(2α) metabolite) in early post-partum cows. Between 13 and 15 h after uncomplicated calving, healthy dairy cows without retained foetal membranes were treated with 50 IU oxytocin, either intramuscularly (OT-IM group; n = 15) or intravenously (OT-IV group; n = 16). Saline solution was administered intramuscularly as controls (CON group; n = 15). Jugular blood samples were taken at 10-min intervals from 1 h before to 2 h after treatment. Plasma PGF(2α) metabolite levels were measured by radioimmunoassay. No significant differences in peripheral plasma PGF(2α) metabolite concentrations occurred in the OT-IM and CON groups, but mean values significantly increased in the OT-IV group, peaking at 20 min after treatment and reaching pre-treatment baseline values again at 120 min. Although the source of prostaglandins was not investigated in this study, our results suggest that exogenous oxytocin may enhance secretion of prostaglandins by the uterus during the first day after normal calving. These prostaglandins might contribute, by an endocrine or paracrine route, to the stimulation of myometrial contractility when exogenous oxytocin is given during this early post-partum stage.     

Effect of exogenous ovarian steroids on the uterine luminal prostaglandins in ovariectomised mares with experimental endometritis

Prostaglandins (PGs) F and E2 were measured in lavage fluid from the uterus of ovariectomised mares after experimental induction of uterine inflammation. Treatment with progesterone alone, or progesterone followed by oestradiol, significantly increased the concentrations of these PGs in the lavage compared with mares treated with oestradiol or control mares. Ovarian steroids, therefore, influenced uterine PG synthesis in response to an inflammatory stimulus. To determine whether the uterine lavage procedure might contribute to the concentrations of prostaglandins in the lavage, the procedure was also performed on six intact mares. With the exception of washings obtained at luteolysis, uterine concentrations of PGF (measured as the plasma metabolite 15-keto-13,14-dihydro PGF2 alpha) had returned to prewashing levels within 30 minutes of the start of uterine lavage. Lavage was therefore unlikely to have influenced the concentrations of prostaglandins in the lavage fluid.     

Effect of prostaglandin F2alpha on ovarian, adrenal, and pituitary hormones and on luteal blood flow in mares

The effect of a single injection of prostaglandin F2alpha (PGF) during mid-diestrus on systemic concentrations of progesterone, LH, FSH, estradiol, and cortisol and on blood flow to the corpus luteum was studied in 10 controls and 10 PGF-treated mares. Blood flow was assessed by estimating the percentage of corpus luteum with color-Doppler signals of blood flow during real-time scanning of the entire structure and by the diameter of the vascular pedicle near its attachment to the ovary. Treatment was done 8 days after ovulation and 0 h was immediately before the treatment. Examinations and collection of blood samples were done at 0 h, every 5 min until 1h, and then at 1.5, 2, 4, 8, 12, 24, 48, and 72 h. The concentrations of estradiol did not change, but progesterone, LH, FSH, and cortisol increased significantly within 5 min. Concentrations of LH and FSH in the PGF group remained elevated until a temporarily lower concentration at 8 or 4h, respectively, rebounded to 12h, and then slowly decreased. Cortisol remained elevated, until a decrease between 1 and 4h. Progesterone in the PGF group increased significantly until 10 min after 0 h and then decreased by 40 min to below the concentrations in controls. Within the PGF group, progesterone decreased significantly by 45 min to below the concentrations at 0 h. The values for each of the two indicators of blood flow did not differ significantly between the PGF and control groups until a decrease at 24h in the PGF group. Results did not support the hypothesis that the immediate transient post-PGF increase in progesterone was associated with an increase in luteal blood flow. Luteolysis, as indicated by decreasing progesterone, began well before the beginning of a decrease in luteal blood flow.     

Effect of prostaglandin F2 alpha, phenylephrine and ergonovine on uterine contractions in the ewe

Two experiments were conducted to determine the effect of prostaglandin F2 alpha (PGF2 alpha), phenylephrine and ergonovine on uterine contractions. In the first experiment, ewes were bilaterally ovariectomized, and a strain gauge force transducer was sutured to the serosa of one uterine horn. Each ewe was treated sc with 2 micrograms of estradiol-17 beta daily to prevent regression of the uterus. Beginning at least 5 d after ovariectomy, four dose levels of PGF2 alpha, phenylephrine and methoxamine were given by im injection and ergonovine was given by im or iv injection. Phenylephrine, methoxamine and ergonovine are alpha-adrenoceptor agonists. Uterine activity was recorded by physiograph for 30 min before and 90 min after treatment. Tracing were analyzed for 20-min periods before treatment and 4 to 24 min and 50 to 70 min after treatment. In Exp. 2, transducers were attached to uteri of intact ewes at d 10 to 12 of an estrous cycle. During subsequent estrus, one or two dose levels of PGF2 alpha, phenylephrine and ergonovine were given by im injection and uterine activity recorded. In Exp. 1, PGF2 alpha and phenylephrine increased (P less than .05 or .01) the number of amplitude of contractions at both 4 to 24 and 50 to 70 min. Ergonovine given im increased the number of contractions. In intact estrous ewes, PGF2 alpha increased the number and amplitude of contractions at 4 to 24 min, phenylephrine increased the number and amplitude at both 4 to 24 and 50 to 70 min, and ergonovine increased the number slightly but significantly at 4 to 24 min.(ABSTRACT TRUNCATED AT 250 WORDS)     

Oxytocin precipitation of prostaglandin-induced farrowing in swine: determination of the optimal dose of oxytocin and optimal interval between prostaglandin F2 alpha and oxytocin

The influence of dose of oxytocin and the interval between prostaglandin (PG) F2 alpha and oxytocin administration on the synchrony of farrowing, the prevalence of intrapartum complications, and the number of pigs dying perinatally was investigated. In study 1, sows were given 10 mg of PGF2 alpha IM on day 112, 113, or 114 of gestation or were not treated. Twenty hours after PGF2 alpha administration, sows were given 0, 5, 10, 20, or 30 USP U of oxytocin IM. Sows treated with PGF2 alpha or PGF2 alpha plus oxytocin had a shorter interval to farrowing than sows not treated or treated with oxytocin alone. Treatment with PGF2 alpha plus 30 U of oxytocin induced the most rapid onset and the greatest synchrony of farrowing, with the mean onset occurring 2.1 +/- 0.4 hours after oxytocin vs greater than 8 hours for all other treatments. Sows treated with 5 or 10 U of oxytocin had a delayed onset and a less synchronous farrowing, compared with sows treated with 0 or 20 U. Day of PGF2 alpha treatment influenced (P less than 0.05) the interval from oxytocin to onset of farrowing. As day during gestation decreased, there was a corresponding decrease in the interval between oxytocin administration and farrowing. Number of interventions to remove retained pigs was not influenced (P greater than 0.05) by day of PGF2 alpha administration. All sows treated with PGF2 alpha followed by oxytocin had a higher rate of manual interventions, compared with that in sows given PGF2 alpha but not oxytocin.(ABSTRACT TRUNCATED AT 250 WORDS)     

Endometrial Oxytocin Receptor Concentration and Activity in Prepubertal Ewe Lambs

We have measured endometrial oxytocin receptor concentrations during prepuberty in ewe lambs, and have investigated the effect of progesterone on the activity of these receptors. In the first study, oxytocin receptor concentrations were undetectable in 2-week prenatal lambs but had increased immediately following birth and were then maintained throughout prepubertal life. Despite the presence of oxytocin receptors animals showed no prostaglandin F(2alpha) (PGF(2alpha)) release in response to exogenous oxytocin challenge at either 3 or 5 months of age. In a second study in 4-month-old ewe lambs treatment with exogenous progesterone resulted in the appearance of PGF(2alpha) release in response to oxytocin after 10 days of treatment. Thus, during the prepubertal life, ewe lambs possess the prerequisites of a luteolytic mechanism in that they have a dormant population of oxytocin receptors in which progesterone can induce oxytocin-stimulated PGF(2alpha) release.     

Plasma prolactin concentrations in mares and their neonates after oxytocin induction of parturition

Studies were undertaken to investigate the effects of oxytocin induction on prolactin release in term (Group II) and preterm (Group III) mares and to compare these effects to spontaneously foaling mares (Group I). Since physiological concentrations of prolactin in blood have not been measured in the neonatal foal, experiments were designed to monitor prolactin in the cord artery and jugular blood of the foals from all groups of mares. Although prolactin levels varied in term mares (Group I and II) during the last 11 days of pregnancy, an increase was observed between Day -6 and Day 0 (2.7 and 11.9 ng/ml respectively; P less than 0.1). The average concentration of prolactin over the last 4 days (Days -3 to 0) had increased by 40% when compared to the average concentration on Days -6, -5, and -4. These findings indicate a rising trend which appears to occur concomitantly with changes in concentrations of 2 mammary components tested, sodium and potassium. Prolactin concentrations did not significantly increase in term mares after oxytocin treatment or in spontaneously foaling mares. However, the preterm induced mares had higher prolactin concentrations during the first stage of labor (19.3 +/- 7.2 ng/ml) than prior to treatment with oxytocin (4.7 +/- 2.0 ng/ml; P less than 0.01). Levels of prolactin in all groups significantly declined by 20-min post-placental expulsion. For the first 30 min after birth, prolactin concentrations in foals from oxytocin-induced mares appeared to be 2-fold higher than those from spontaneously foaling mares. Thereafter, prolactin values declined to baseline values by 48 hrs. When comparing cord arterial plasma with cord venous plasma in each group, prolactin concentrations were similar. However, the average prolactin levels in both the cord artery and vein appeared higher (ave: 1.1 ng/ml) in Group II and III than in Group I (less than 0.5 ng/ml). From these results, the authors suggest that 1) prolactin may have a role in regulating mammary secretory products in mares just prior to parturition; 2) oxytocin may increase prolactin secretion in preterm induced mares; 3) oxytocin induction may have a short term effect to increase circulatory prolactin concentrations in neonates in utero regardless whether their dams were treated preterm or term.     

Retrospective study on the incidence of postinsemination uterine fluid in mares inseminated with frozen/thawed semen

Uterine fluid accumulation has been reported after insemination or natural breeding of mares. This retrospective study examined the factors affecting the incidence of uterine fluid after insemination of frozen semen. Specifically, this study determined the association between mare age, reproductive status, fluid accumulation, and pregnancy rates in mares. Records were available from 283 warmblood mares throughout 496 cycles. Mares were divided into maiden, foaling, and barren and age groups of 3 to 9, 10 to 16, and more than 16 years. Mares were inseminated only once with frozen semen within 4 to 8 hours before or after ovulation. Ultrasound examinations were performed 12 to 18 hours after insemination. A depth of at least 20 mm of fluid was considered significant. Mares with less than 20 mm were treated with oxytocin, and those with more than 20mm of fluid were given oxytocin and uterine lavage. Pregnancy determination was performed at 14 to 16 and 30 to 50 days after ovulation. Fluid level of more than 20 mm was recorded in 25% of the cycles. Barren mares and aged mares (10-16 and > 16 years) had a higher incidence of uterine fluid accumulations. Per-cycle pregnancy rate was lower (45%) in mares with uterine fluid than in mares without uterine fluid (51%). This difference was primarily due to the reduction in fertility of mares who were older than 16 years and retained fluid after insemination. Apparently, oxytocin and lavage treatments provided acceptable fertility in the other groups of mares that had uterine fluid.

Introduction

Use of equine frozen semen is accepted by the majority of horse registries. According to several field studies,[1, 2, 3, 4 and 5] insemination of frozen semen has resulted in acceptable pregnancy rates. Postbreeding fluid accumulation is a physiologic inflammation that clears the uterus of foreign material such as excess spermatozoa, seminal plasma, bacteria, and extenders. [6, 7, 8, 9 and 10] Uterine fluid can be easily diagnosed with ultrasonography. [10, 11 and 12] Persistent postbreeding uterine fluid has been associated with a decrease in fertility after natural mating or artificial insemination (AI) of fresh semen. [11, 12 and 13] Predisposing factors to persistent fluid accumulations are reduced myometrial contractions, poor lymphatic drainage, large overstretched uterus, and cervical incompetence. [7, 14 and 15] Normal mares are able to expel uterine fluid quickly after inseminations, whereas susceptible mares accumulate fluid in their uterine lumen for more than 12 hours after breeding or insemination. [10]It is commonly stated that insemination with frozen semen leads to greater post-AI fluid accumulation than insemination with fresh or cooled semen or after natural mating. Apparently, there is only 1 controlled study on this comparison.[7] The authors reported that infusion of frozen semen resulted in a greater inflammatory response than natural breeding. In a field study, [16] 16% of mares naturally mated had persistent postbreeding fluid accumulations compared with a 30% rate reported for mares inseminated with frozen semen. [1 and 2] More recently, Watson et al. [17] reported a postbreeding fluid accumulation rate of 16%, which is identical to that reported for natural mating. [16] It is difficult to compare studies because details of mare selection and insemination or breeding frequencies are not always reported. Obviously, a higher proportion of barren and aged mares in a study would increase the incidence of postbreeding fluid accumulation. [1 and 2]The study presented herein was a retrospective study designed to determine the incidence of postbreeding fluid accumulation in a large number of mares inseminated with frozen semen. Associations were determined between mare age, reproductive status and fluid accumulation, and pregnancy rate in mares with and without uterine fluid accumulation.

Materials and methods

Mares

Records were available from 283 warmblood mares inseminated with frozen semen at the Cristella Veterinary Clinic in Italy during 1998 to 2001. Mares ranging in age from 3 to 20 years were inseminated with semen that was frozen in 10 centers and was from 34 stallions. The broodmare population was subdivided into 3 reproductive groups: 89 maiden mares (mean age, 7.2 years), 106 foaling mares (mean age, 9.4 years), and 87 barren mares (mean age, 11.9 years). Maiden mares older than 7 years were selected with biopsy scores of 1 or 2 only. Barren mares were open for no more than 2 consecutive seasons and had negative cytology and bacteriology scores. Age groups were divided as follows: 3 to 9 years (n = 132), 10 to 16 years (n = 137) and older than 16 years (n = 14). Data from 496 cycles were used. Distribution of the estrous cycles was 172, 157, and 167 in the maiden, foaling, and barren groups, respectively; and 224, 244, and 28 in the youngest, intermediate, and oldest groups, respectively.

Mare reproductive management and artificial insemination protocol

During estrus, all mares underwent a daily ultrasound examination with a 5-mHz transrectal probe (SA 600 Vet; Medison Inc., Seoul, South Korea) until 1 or more 35-mm ovarian follicles were detected. Ovulation was then induced by the intravenous administration of 2000 IU of human chorionic gonadotropin (hCG). Ultrasound examination was performed 12 hours after hCG treatment and then every 4 to 8 hours until ovulation occurred. Mares were inseminated only once within a period of 4 to 8 hours before or after ovulation. The semen used was thawed according to the distribution center's instructions and had the following minimum post-thaw quality requirements: not less than 200 × 10⁶ progressively motile spermatozoa per dose and a minimum of 30% progressive spermatozoal motility. Foaling mares were not inseminated at their first postpartum (“foal heat”) estrous period, because pregnancy rates are recognized to be lower than during the subsequent estrous periods.[18] During the first postpartum estrus, ovarian ultrasound scan examinations were performed every 2 to 3 days until an ovulation was detected. A prostaglandin F₂α injection was given 5 days later to short-cycle the mare.

Postinsemination monitoring

An ultrasound examination of the reproductive tract was performed 12 to 18 hours after insemination to detect any intrauterine fluid accumulation. The presence and depth of intrauterine fluid was recorded. Twenty millimeters or more of grade II or III intrauterine fluid[19] was recorded as a significant amount of fluid. Mares with less than 20 mm of fluid were treated with an intravenous injection of 20 IU oxytocin. For mares with more than 20 mm of fluid, oxytocin was administered, and the uterus was flushed daily with buffered saline solution: 1-L aliquots were infused and recovered until the recovered fluid was clear. In these mares, oxytocin treatment was repeated up to 3 times daily. Post insemination treatments were performed for no more than 4 days after ovulation had occurred.Pregnancy diagnosis was performed with ultrasound at 14 to 16 days after ovulation. Scans were then repeated at 30 and 50 days of gestation to confirm the presence in the uterus of an apparently healthy developing conceptus.

Statistical analysis

χ² Analysis was used to determine the effect of reproductive status and age on the incidence of fluid accumulation. In addition, the influence of persistent uterine fluid accumulation on pregnancy rates per cycle was determined for each reproductive class and age by using χ² analysis.

Results

The per-cycle pregnancy rate at 14-16 days after ovulation was 49.3% (245/496 cycles). By the end of the season, 245 of 283 mares (86.5%) were confirmed pregnant. Fluid level of at least 20 mm (grade II or III) was recorded in 126 of the 496 cycles (25.4%). Barren mares had a higher (P < .05) incidence of postbreeding fluid accumulation (64/167; 38.3%) than maiden (34/172; 19.7%) and foaling (28/157, 17.8%; Table 1) mares. The incidence of fluid accumulation was also higher in mares older than 16 years (19/28; 67.8%) than those aged 10 to 16 years (69/244; 28.2%) and 3 to 9 years (38/224; 17%). The incidence of uterine fluid was also higher (P < .05) for mares aged 10 to 16 years than those aged 3 to 9 years (Table 2). Overall, the per-cycle pregnancy rate was lower (P < .05) for mares with post-AI fluid accumulations than for those with no uterine fluid or only a small quantity of fluid (57/126, 41.9% vs 188/360, 56.2%). Pregnancy rates were similar (P > .05) for mares with or without uterine fluid when comparisons were made within maiden and barren mare groups. However, more foaling mares became pregnant when no fluid was detected after insemination. Pregnancy rate for this group (68.1%) was higher than that for maiden (44.2%) and barren (44.6%) mares (Table 3). Older mares with uterine fluid accumulations had a lower per-cycle pregnancy rate (36.8%) than mares in the same group but without fluid. Surprisingly, if no fluid was detected, the highest pregnancy rates were in mares older than 16 years ( Table 4).