Passage of postovulatory follicular fluid into the peritoneal cavity and the effect on concentrations of circulating hormones in mares

Reported data were reviewed and reexamined to evaluate the concept that most of the follicular fluid enters the peritoneal cavity at ovulation in mares and transiently alters the circulating concentrations of LH, FSH, estradiol, and inhibin. A transrectal ultrasonographic study supported the hypothesis that the large volume (40-50 ml) of evacuated follicular fluid passes through the infundibular fimbriae into the peritoneal cavity. A spike in circulating inhibin and a decrease in the rate of reduction in circulatory estradiol occurs at ovulation. Simultaneously, a disruption occurs in the increasing concentrations of the ovulatory LH surge and in the FSH surge that begins before ovulation. The concept was further supported by the present finding that the estradiol content of follicular fluid within a few hours before ovulation is equivalent to the amount reported to be needed for a negative effect on LH and for a synergistic negative effect of estradiol and inhibin on FSH.     

Effect of passive immunization against inhibin on FSH secretion, folliculogenesis and ovulation rate during the follicular phase of the estrous cycle in mares

Physiological roles of inhibin in mares were investigated by means of passive immunization using an antiserum to inhibin that had been raised in a castrated goat. Eight mares were given an intravenous injection of either 100 mL (n = 4) or 200 mL (n = 4) of inhibin antiserum 4 d after a single intramuscular injection of PGF2 alpha on Day 8 after ovulation, 4 control mares were treated with 100 mL castrated goat serum in the same manner. Jugular vein blood samples were collected after treatment with the serum until 192 h post treatment. Follicular growth and ovulations were monitored by ultrasound examination at 24-h intervals. The ability of the inhibin antiserum to neutralize the bioactivity of equine inhibin was examined in vitro using a rat pituitary cell culture system. Suppression of secretion of FSH from cultured rat pituitary cells by equine follicular fluid was reversed by the addition of increasing doses of the inhibin antiserum, thereby indicating its bioactivity. Plasma levels of FSH and estradiol-17 beta were higher in mares treated with the inhibin antiserum. The ovulation rate was significantly higher in mares treated with antiserum (100 mL = 3.75 +/- 0.63; 200 mL = 4.50 +/- 0.65) than in control mares (1.25 +/- 0.25). These results demonstrate that inhibin is important in regulating FSH secretion and folliculogenesis in mares. They also show that neutralization of the bioactivity of inhibin may become a new method for the control of folliculogenesis and ovulation rate in mares.     

Treatment with recombinant equine follicle stimulating hormone (reFSH) followed by recombinant equine luteinizing hormone (reLH) increases embryo recovery in superovulated mares

The dynamics of ovarian follicular development depend on a timely interaction of gonadotropins and gonadal feedback in the mare. The development and efficacy of genetically cloned recombinant equine gonadotropins (reFSH and reLH) increase follicular activity and induce ovulation, respectively, but an optimum embryo recovery regimen in superovulated mares has not been established. The objective of this study was to determine if treatment with reFSH followed by reLH would increase the embryo per ovulation ratio and the number of embryos recovered after superovulation in mares. Sixteen estrous cycling mares of light horse breeds (4-12 years) were randomly assigned to one of two groups: Group 1; reFSH (0.65mg)/PBS (n=8) and Group 2; reFSH (0.65mg)/reLH (1.5mg) (n=8). On the day of a 22-25mm follicle post-ovulation mares were injected IV twice daily with reFSH for 3 days (PGF(2α) given IM on the second day of treatment) and once per day thereafter until a follicle or cohort of follicles reached 29mm after which either PBS or reLH was added and both groups injected IV twice daily until the presence of a 32mm follicles, when reFSH was discontinued. Thereafter, mares were injected three times daily IV with only PBS or reLH until a majority of follicles reached 35-38mm when treatment was discontinued. Mares were given hCG IV (2500IU) to induce ovulation and bred. Embryo recovery was performed on day 8 day post-treatment ovulation. Daily jugular blood samples were collected from the time of first ovulation until 8 days post-treatment ovulation. Blood samples were analyzed for LH, FSH, estradiol, progesterone and inhibin by validated RIA. Duration of treatment to a ≥35mm follicle(s) and number of ovulatory size follicles were similar between reFSH/reLH and reFSH/PBS treated mares. The number of ovulations was greater (P<0.01) in the reFSH/reLH group, while the number of anovulatory follicles was less (P<0.05) compared to the reFSH/PBS group. Number of total embryos recovered were greater in reFSH/reLH mares than in the reFSH/PBS mares (P≤0.01). The embryo per ovulation ratio tended to be greater (P=0.07) in the reFSH/reLH mares. Circulating concentrations of estradiol, inhibin, LH and progesterone were not statistically different between groups. Plasma concentrations of FSH were less (P<0.01) in the reFSH/reLH treated mares on days 0, 1, 4, 6, 7 and 8 post-treatment ovulation. In summary, reFSH with the addition of reLH, which is critical for final follicular and oocyte maturation, was effective in increasing the number of ovulations and embryos recovered, as well as reduce the number of anovulatory follicles, making this a more viable option than treatment with reFSH alone. Further evaluation is needed to determine the dose and regimen of reFSH/reLH to significantly increase the embryo per ovulation ratio.     

Control of FSH, follicular development and estrus synchronization in the mare with steroid-free follicular fluid

Twenty-two pony mares were used in a project designed to determine the effectiveness of different treatments in controlling FSH, follicular development and synchronization of estrus and ovulation. Mares in Group 1 (n=8) received daily oral altrenogest (0.044 mg/kg); those in Group 2 (n=7) received daily altrenogest (0.044 g/kg) and, during the last 4 days of treatment they received steroid-free follicular fluid, (15 cc) intravenously (I.V.) two times a day; Mares in Group 3 (n=7) received daily intramuscular (I.M.) injections of progesterone (80 mg) and estradiol valerate (7 mg). All treatments lasted for 10 days, at the end of which prostaglandin (PgF(2)alpha, 10 mg) was administered. Sexual behavior, follicular development and FSH concentrations were monitor daily. Concentrations of FSH in Group 2 mares, were not significantly different (P>0.05) from those of Group 1 until the mares in Group 2 were treated with follicular fluid (P<0.05). Concentrations of FSH in Group 3 mares, were significantly lower than those of Groups 1 and 2 (P<0.05) until the mares in Group 2 were treated with steroid-free follicular fluid. At this point there was no significant difference between groups 2 and 3 (P>0.05). Steroid-free follicular fluid appears to induce atresia in larger follicles (>11 mm), and the initiation of new follicular wave. The combination of progesterone and estradiol valerate appears to delay follicular growth and not to induce atresia, since larger follicles (>11 mm) continued to grow after treatment. Both treatments (groups 2 and 3) resulted in ovulations within 5 days period. The treatment in Group 1 did not have any effect on FSH or follicular development and ovulations were dispersed through a 9-day period. We concluded that steroid-free follicular fluid offers a new possibility to synchronize ovulation in the mare by controlling FSH and follicular development.     

Steroid concentrations in follicular fluid aspirated repeatedly from transitional and cyclic mares

The objectives of the present study were to determine follicular progesterone (P4) and estradiol-17beta (E2) in transitional mares and to compare follicular steroid concentrations between transitional and cyclic mares. Follicles > 8 mm were aspirated under transvaginal ultrasound-guidance 4 times at 3 to 4 day intervals (T1-T4) in Norwegian pony mares during vernal transition. During the breeding season, follicular aspirations were conducted in each mare on Day 6, Day 14 and Day 18 after ovulation of 3 separate estrous cycles (Day of ovulation = Day 0). Plasma and follicular fluids were analyzed for P4 and E2 with ELISA and RIA, respectively. Plasma P4 concentrations remained below 1 ng/mL throughout T1-T4, while the follicular P4 concentrations increased significantly to cyclic levels after the first transitional aspiration. Plasma E2 concentrations similarly remained at low levels during the course of the transitional aspirations, while the follicular E2 concentrations increased gradually over the 4 aspirations to cyclic concentrations. The mares ovulated on average 9.8 +/- 1.6 (mean +/- SEM) days after the last transitional aspiration, and 16.6 +/- 0.2, 11.3 +/- 1.5 and 23.2 +/- 4.4 days after aspirations conducted on Day 6, 14 and 18, respectively. The present study demonstrates that in the transitional mare newly developing follicles exhibit biosynthesis of P4 and E2. Furthermore, an increase in follicular steroid concentrations is not necessarily reflected in the peripheral steroid concentrations.     

Delayed follicular development and ovulation following inhibition of FSH with equine follicular fluid in the mare

In two experiments, PGF(2alpha) was given to all mares on Day 10 (ovulation = Day 0). In experiment 1, mares received either whole follicular fluid or saline i.v. every 12 hours on Days 10 to 14. Experiment 2 was similar to experiment 1, except the follicular fluid was extracted with charcoal to remove steroids. Analysis of the FSH data for Days 10 to 21 indicated an effect of treatment (P<0.08) with whole follicular fluid, but not with charcoal-extracted follicular fluid. However, there was an effect of day (P<0.05) and an interaction (P<0.01) of treatment with day for both experiments. The interaction of treatment with day seemed primarily due to a marked post-treatment increase in FSH concentrations between Days 15 and 17 for mares treated with either whole follicular fluid or charcoal-extracted follicular fluid. Analysis of the diameter of the largest follicle for Days 10 to 18 indicated a main effect of treatment (P<0.05) and day (P<0.05) and an interaction (P<0.05) of treatment with day for both experiments. The interaction of treatment with day was attributable to the inhibition of follicular growth by Day 14 for mares treated with whole follicular fluid and by Day 15 for mares treated with charcoal-extracted follicular fluid. The length of the interovulatory interval was longer (P<0.05) in the treated group than in controls for both experiments. Results indicated that equine follicular fluid contained a proteinaceous substance that suppressed circulating concentration of FSH. The inhibited follicular growth and the delay in ovulation were attributed to the reduced concentrations of circulating FSH.     

Disruption of periovulatory FSH and LH surges during induced anovulation by an inhibitor of prostaglandin synthesis in mares

The role of passage of follicular fluid into the peritoneal cavity during ovulation in the transient disruption in the periovulatory FSH and LH surges was studied in ovulatory mares (n=7) and in mares with blockage of ovulation by treatment with an inhibitor of prostaglandin synthesis (n=8). Mares were pretreated with hCG when the largest follicle was ≥32 mm (Hour 0). Ultrasonic scanning was done at Hours 24 and 30 and every 2h thereafter until ovulation or ultrasonic signs of anovulation. Blood samples were collected at Hours 24, 30, 32, 34, 36, 38, 48, and 60. Ovulation in the ovulatory group occurred at Hours 38 (five mares), 40, and 44. Until Hour 36, diameter of the follicle and concentrations of FSH, LH, and estradiol-17β (estradiol) were similar between groups. Between Hours 34 and 36, a novel transient increase in estradiol occurred in each group, and color-Doppler signals of blood flow in the follicular wall decreased in the ovulatory group and increased in the anovulatory group. In each group, FSH and LH periovulatory surges were disrupted by a decrease or plateau between Hours 38 and 48 and an increase between Hours 48 and 60. The discharge of hormone-laden follicular fluid into the peritoneal cavity at ovulation was not an adequate sole explanation for the temporally associated transient depression in FSH and LH. Other routes from follicle to circulation for gonadotropin inhibitors played a role, based on similar depression in the ovulatory and anovulatory groups.     

Repeatability of preovulatory follicular diameter and uterine edema pattern in two consecutive cycles in the mare and how they are influenced by ovulation inductors

Follicular diameter is used as a guiding tool to predict ovulation in the mare. However, the great range in preovulatory follicular diameter makes prediction of optimal breeding time based on follicular diameter unreliable. Uterine edema pattern is also useful to determine the best time to breed, since intensity of edema tends to dissipate as ovulation approaches, however, not every mare follows this pattern. The aims of this study were to assess the repeatability of preovulatory follicular diameter and uterine edema pattern in two consecutive spontaneous cycles and to determine how induction treatments (hCG, PGF(2)alpha and GnRH analogues) influence them. Fifty-three mares were followed during two consecutive cycles and scanned three times a day from 2 to 3 days before ovulation. During the first cycle, mares had a spontaneous ovulation and in the consecutive cycle mares received either: (a) no hormonal treatment; (b) 1500 IU hCG; (c) 125-250 microg Cloprostenol or (d) 2.1 mg Deslorelin implant. Mares ovulated consistently from similar follicular diameters in two consecutive spontaneous cycles (r=0.89; P<0.000). All three induction treatments had a significant effect on reducing the preovulatory follicular diameter (P<0.005). Mares showed fair correlation in uterine edema patterns in both consecutive non-induced cycles (r=0.71; P<0.005). In conclusion mares in consecutive cycles ovulated from consistent follicular diameters. Follicular diameters recorded from previous ovulations can be relied on to predict the optimal breeding time in successive cycles especially in mares that ovulate from unusually small follicles.     

Systemic treatment with high dose of flunixin-meglumine is able to block ovulation in mares by inducing hemorrhage and luteinisation of follicles

Prostaglandins play an obligatory role during the process of ovulation in mammals. Ovulation can be blocked by intrafollicular administration of non-steroidal anti-inflammatory drugs (NSAIDs) in several domestic species including the mare as well as by systemic administration of these drugs in women. In the mare, the effect of systemic NSAIDs treatment on ovulation has not been critically studied. The objectives of this study were: a) to determine whether high dose of flunixin-meglumine (FM) administered systemically to mares during the periovulatory period was able to block ovulation; and b) to study the follicular ultrasound characteristics of FM treated mares. Six mares were used in the study during two consecutive estrous cycles. Each mare received 2 mg FM/kg i.v. twice a day starting at the time of treatment with hCG when the follicle reached a diameter of ≥ 32 mm and continuing until ovulation. During the consecutive control cycle (CON) the mares received the same dose of hCG but were not administered FM. During the FM cycles five of six mares failed to ovulate and collapse the preovulatory follicle; but echoic specks were observed within the follicles, which continued to grow until a mean diameter of 55 mm. Eventually, the follicular contents were organised and luteinised. All CON mares ovulated normally. In conclusion, when mares were treated with FM, they had a higher incidence of ovulatory failure and development of luteinised unruptured follicles (83%, P = 0.015) compared with untreated mares.     

Elevated plasma testosterone concentrations during stallion-like sexual behavior in mares (Equus caballus)

Mounting interactions in mares isolated from stallions and the relationship to stage of the estrous cycle and level of circulating hormones were studied for 3 years in a herd averaging 105 mares. Mares were assigned to mounting, standing, and control groups. A control mare was selected by being within 1 day of the number of days after ovulation in a mounting mare. A total of 15 mounting interactions were detected by chance observation during the 3 years. A blood sample was collected immediately after the mounting interaction from each mare in the three groups, and a transrectal ultrasonographic examination of the reproductive tract was done. Two mounting interactions occurred during the early luteal phase and 13 during the follicular phase. The interactions that occurred during the follicular phase were used for comparisons among groups. The interval between mounting and the next ovulation, diameter of the two largest follicles, and the number of follicles larger and smaller than 20 mm were not different significantly among the mounting, standing, and control groups. Testosterone concentrations were higher (P<0.01) in the mounting group (17.7+/-2.3 pg/ml) than in standing group (10.9+/-0.5 pg/ml), and the difference between the mounting group and the control group (12.8+/-0.6 pg/ml) approached significance (P<0.08). Concentrations of androstenedione, estradiol, estrone, and progesterone did not differ significantly among groups. Results indicated that mounting behavior between mares is rare, usually occurs during the follicular phase, and is related to high circulating concentrations of testosterone.     

The mature mid-cycle follicle in the mare.

Oestrogen and progesterone concentrations in blood and follicular fluid and blood levels of LH were determined in 426 mares at different stages of the oestrous cycle. Mature follicles occur at all stages of the cycle; they ovulate readily in early metoestrus, occasionally in late metoestrus and very rarely in dioestrus. Maturation of a mid-cycle follicle is associated with intermediate levels of LH, which are less than those found during oestrus. This lower level of LH together with a high level of progesterone are probably responsible for the failure of ovulation and regression of most of the mid-cycle mature follicles found in the mare.     

Changes in concentrations of follicular fluid factors during follicle selection in mares

The temporal relationships in the changes in concentrations of follicular fluid factors during follicle selection were characterized in mares. All follicles > or =5 mm were ablated 10 days after ovulation, followed by follicular fluid collection from the three largest follicles (F1, F2, and F3) when F1 of the new wave reached a diameter of 8.0-11.9, 12.0-15.9, 16.0-19.9, 20.0-23.9, 24.0-27.9, or 28.0-31.9 mm (n = 4-8 mares/range). Diameter deviation between F1 and F2 began during the 20.0- to 23.9-mm range, as indicated by a greater difference in diameter between the two follicles at the 24.0- to 27.9-mm range than at the 20.0- to 23.9-mm range. Androstenedione concentrations increased in F1, F2, and F3 between the 16.0- to 19.9- and 20.0- to 23.9-mm ranges. In contrast, estradiol, free insulin-like growth factor (IGF)-1, activin-A, and inhibin-A concentrations increased only in F1 beginning at the 16.0- to 19.9-mm range. As a result, the concentrations of all four factors were higher in F1 than in F2 and F3 at all the later ranges, including the 20.0- to 23.9-mm range (beginning of diameter deviation). Concentrations of progesterone differentially increased in F1, concentrations of androstenedione and IGF-binding protein (IGFBP)-2 increased only in F2 and F3, and concentrations of inhibin-B differentially decreased in F2 and F3 simultaneous with the beginning of deviation. Concentrations of FSH, LH, pro-alphaC inhibin, and total inhibin did not change differentially among follicles. Results indicated that, on a temporal basis, estradiol, free IGF-1, activin-A, and inhibin-A may have played a role in the initiation of follicle deviation. In addition, these four factors as well as progesterone, androstenedione, IGFBP-2, and inhibin-B may have been involved in the subsequent differential development of the follicles.     

Response of estradiol and inhibin to experimentally reduced luteinizing hormone during follicle deviation in mares

The increase in LH concentrations at the time of the decrease in FSH concentrations during follicle deviation in mares was studied to determine the role of LH in the production of estradiol and immunoreactive inhibin (ir-inhibin). Ten days after ovulation, all follicles > or =6 mm were ablated, prostaglandin F(2 alpha) was given, and either 0 mg (control group, n = 15) or 100 mg of progesterone in safflower oil (treated group, n = 16) was given daily for 14 days, encompassing the day of diameter deviation. The follicular and hormonal data were normalized to the expected day of the beginning of diameter deviation when the largest follicle first reached > or =20 mm (Day 0). The experimentally induced decrease in LH concentrations during follicle deviation beginning on Day -4 delayed and stunted the increase in circulating concentrations of ir-inhibin and estradiol beginning on Days -3 and -1, respectively, but did not alter the predeviation FSH surge and the initiation of diameter deviation between the two largest follicles. Combined for both groups, the interval to the expected day of deviation was 16.6 days after ovulation when the largest follicle was a mean of 21.6 mm. After deviation, the largest follicle started to regress in the treated group beginning on Day 1 and was associated with decreased concentrations of ir-inhibin and estradiol, and increased concentrations of FSH. The negative influence of the dominant follicle on the postdeviation decrease in FSH observed in the control group was alleviated and concentrations resurged in the treated group. Apparently this is the first in vivo evidence that the increase in LH that precedes follicle deviation has a positive effect in supporting the production of inhibin during diameter deviation. It was concluded that the increase in LH concentrations before diameter deviation played a role in the production of estradiol and inhibin by the largest follicle during deviation.     

eFSH in clinical equine practice

Equine follicle stimulating hormone (eFSH) has been used to induce follicular development in transitional mares and problem acyclic mares, as well as superovulate cycling mares. The most efficacious protocol is to administer 12.5 mg eFSH, intramuscularly, twice daily beginning 5 to 7 days after ovulation when the diameter of the largest follicle is 20 to 25 mm. Prostaglandins are to be administered on the second day of eFSH therapy. Treatment with eFSH is continued for 3 to 5 days until follicle(s) are >or=35 mm in diameter. The mare is subsequently allowed to 'coast' for 36 h, after which human chorionic gonadotropin is administered to induce ovulation.     

Pre-ovulatory changes in cyclic AMP and prostaglandin concentrations in follicular fluid of gilts

The concentrations of cyclic adenosine 3′,5′-monophosphate (cyclic AMP) and prostaglandins E and F (PGE and PGF) were determined in follicular fluid collected from follicles of prepubertal gilts at various times after treatment with pregnant mare serum gonadotropin (PMSG) and human chorionic gonadotropin (hCG) to induce ovulation. The concentrations of cyclic AMP, PGE and PGF in the follicular fluid after PMSG treatment but prior to hCG injection were about 1 pmol/ml, 1 ng/ml and 0.2 ng/ml, respectively. After hCG administration, the follicular fluid levels of cyclic AMP increased markedly, reaching a peak (400-fold increase) about 4 h after injection and then declined gradually to pre-hCG levels. A second rise (2.5- to 5-fold increase) occurred about 30 h after hCG with the levels being sustained up to the expected time of ovulation. In contrast, the levels of PGE and PGF remained relatively constant until 28–30 h after hCG treatment. Thereafter, the concentrations of both prostaglandins began to rise with the increases becoming more pronounced and reaching maximal values as the expected time of ovulation approached. These data provide further evidence for a physiological role of follicular prostaglandins in the process of ovulation but do not support an obligatory role for prostaglandins in the acute gonadotropin stimulation of cyclic AMP formation.     

Pre-ovulatory changes in cyclic AMP and prostaglandin concentrations in follicular fluid of gilts.

The concentrations of cyclic adenosine 3', 5'-monophosphate (cyclic AMP) and prostaglandins E and F (PGE and PGF) were determined in follicular fluid collected from follicles of prepubertal gilts at various times after treatment with pregnant mare serum gonadotropin (PMSG) and human chorionic gonadotropin (hCG) to induced ovulation. The concentrations of cyclic AMP, PGE and PGF in the follicular fluid after PMSG treatment but prior to hCG injection were about 1 pmol/ml, 1 ng/ml and 0.2 ng/ml, respectively. After hCG administration, the follicular fluid levels of cyclic AMP increased markedly, reaching a peak (400-fold increase) about 4 h after injection and then declined gradually to pre-hCG levels. A second rise (2.5- to 5-fold increase) occurred about 30 h after hCG with the levels being sustained up to the expected time of ovulation. In contrast, the levels of PGE and PGF remained relatively constant until 28-30 h after hCG treatment. Thereafter, the concentrations of both prostaglandins began to rise with the increases becoming more pronounced and reaching maximal values as the expected time of ovulation approached. These data provide further evidence for a physiological role of follicular prostaglandins in the process of ovulation but do not support an obligatory role for prostaglandins in the acute gonadotropin stimulation of cyclic AMP formation.     

Removal of deslorelin (Ovuplant) implant 48 h after administration results in normal interovulatory intervals in mares

Deslorelin implants, approved for use in inducing ovulation in mares, have been associated with prolonged interovulatory intervals in some mares. Administration of prostaglandins in the diestrous period, following a deslorelin-induced ovulation, has been reported to increase the incidence of delayed ovulations. The goals of the present study were: (1) to determine the percentage of mares given deslorelin that experience delayed ovulations with or without subsequent prostaglandin treatment, and (2) to determine if removal of the implant 48 h after administration would effect the interval to subsequent ovulation. We considered interovulatory intervals to be prolonged if they were greater than the mean +/- 2 standard deviation (S.D.) of the control group in study 1 and the hCG group in study 2. In study 1, we retrospectively reviewed reproduction records for 278 mares. We either allowed the mare to ovulate spontaneously or induced ovulation using deslorelin acetate implants or hCG. We administered prostaglandin intramuscularly, 5-9 days after ovulation in selected mares in each group. A higher percentage of mares which were induced to ovulate with deslorelin and given prostaglandins had a prolonged interovulatory interval (23.5%; n = 16), as compared to deslorelin-treated mares that did not receive prostaglandins (11.1%; n = 5). In study 2, we induced ovulation in mares with hCG (n = 47), a subcutaneous deslorelin implant via an implanting device provided by the manufacturer (n = 28), or a deslorelin implant via an incision in the neck (n = 43) and we removed the implant 48 h after administration. We administered prostaglandin to all mares 5-9 days after ovulation. In study 2, mares from which the implant was removed had a normal ovulation rate and none had a prolonged interval to ovulation. Administration of prostaglandin after deslorelin treatment was associated with a longer interval from luteolysis to ovulation than that found in mares not treated with deslorelin. Prostaglandin administration during diestrus may have exacerbated the increased interval to ovulation in deslorelin-treated mares. We hypothesize that prolonged secretion of deslorelin from the implant was responsible for the extended interovulatory intervals.     

The effect of treatment with flunixin meglumine at different times relative to hCG administration on ovulation failure and luteal function in mares

Flunixin meglumine (FM), a prostaglandin synthetase inhibitor, causes ovulatory failure in the mare. However, the effect of the FM treatment relative to the time of hCG administration on the ovulation failure has not been determined nor has its effect on the luteal function of treated mares. Estrous mares with a follicle ≥32 mm (range of 32-38 mm) were treated with 1.7 mg/kg b.w. of FM iv at zero, 12, 24 and 36 h (n=6), at 24 and 36 h (n=6), at 28 and 36 h (n=6), at 24h (n=6) or at 30 h (n=6) after treatment with 1500 IU hCG. One group received no FM (control, n=6). Progesterone concentrations were determined using RIA. Mares treated with FM 0-36 h and 24-36 h had higher (P<0.05) incidence of ovulatory failure (83 and 80%, respectively) than mares treated twice at 28 and 36 h, or once at 24 or at 30 h after hCG (16.7, 0 and 0%, respectively). The anovulatory follicles of FM treated mares luteinized and produced progesterone (>2 ng/ml). The progesterone concentration was lower in mares treated with FM at zero to 36 h and at 24-36 h after hCG than in the other groups. In conclusion, the FM administration was effective in blocking ovulation only when the treatment began ≤24 h after hCG and was continued every 12 h until ≥36 h. In addition, the FM-induced anovulatory follicles underwent luteinization of follicular cells with active production of progesterone.     

Gonadotropin-releasing hormone treatment induces follicular growth and ovulation in seasonally anestrous mares

A study was conducted to evaluate the effectiveness of gonadotropin-releasing hormone (GnRH) pulse infusion to stimulate follicular development and induce ovulation in seasonally anestrous standardbred mares. Seventeen mares were selected for use in this experiment, on the basis of a previous normal reproductive history, and were housed under a photoperiod of 8L:16D beginning one week prior to the start of the experiment (second week in January). Mares were infused with 20 micrograms (n = 7) or 2 micrograms (n = 6) GnRH/h, or were subjected to photoperiod treatment only (controls, n = 4). Serum concentrations of luteinizing hormone (LH), follicle-stimulating hormone (FSH), and progesterone did not vary, and neither significant follicular development nor ovulation was observed in any control mare throughout the experimental period (greater than 60 days). By contrast, both groups of GnRH-treated mares showed elevated serum concentrations of LH and FSH within one day after the start of infusion. Mares infused with 20 micrograms GnRH/h had at least one follicle greater than or equal to 25 mm in 7.4 +/- 1.3 (mean +/- SEM) days following the start of infusion, and ovulated in 12.0 +/- 0.7 days. In the 2-microgram-GnRH/h treatment group, a 25-mm follicle was detected in 5.7 +/- 0.7 days, and ovulation occurred after 10.0 +/- 0.3 days of infusion. Ovulation in every instance was followed by a functional luteal phase, as indicated by the profiles of progesterone secretion.(ABSTRACT TRUNCATED AT 250 WORDS)     

Management and fertility of mares bred with frozen semen.

Semen quality, mare status and mare management during estrus will have the greatest impact on pregnancy rates when breeding mares with frozen semen. If semen quality is not optimal, mare selection and reproductive management are crucial in determining the outcome. In addition to mare selection, client communication is a key factor in a frozen semen program. Old maiden mares and problem mares should be monitored for normal cyclicity and all, except young maidens, should have at least a uterine culture and cytology performed. Mares with positive bacterial cultures and cytologies should be treated at least three consecutive days when in estrus with the proper antibiotic. With frozen semen, timing the ovulation is highly desirable in order to reduce the interval between breeding and ovulation. The use of ovulation inducing agents such as human chorionic gonadotropin (hCG) or the GnRH analogue, deslorelin, are critical components to accurately time the insemination with frozen semen. Most hCG treated mares ovulate 48h post-treatment (12-72h) while most deslorelin (Ovuplant) treated mares ovulate 36-42h post-treatment. However, mares bred more than once during the breeding cycle appear to have a slight but consistent increase in pregnancy rate compared to mares bred only once pre- or post-ovulation. In addition, the "capacitation-like" changes inflicted on the sperm during the process of freezing and thawing appear to be responsible for the shorter longevity of cryopreserved sperm. Therefore, breeding closer to ovulation should increase the fertility for most stallions with frozen semen. Recent evidence would suggest that breeding close to the uterotubal junction increases the sperm numbers in the oviduct increasing the chances of pregnancy. Post-breeding examinations aid in determining ovulation and uterine fluid accumulations so that post-breeding therapies can be instituted if needed. Average pregnancy rates per cycle of mares bred with frozen semen are between 30 and 40% with a wide range between sires. Stallion and mare status are major factors in determining the success of frozen semen inseminations. Pregnancy rates are lower for barren and old maiden mares as well as those mares treated for uterine infections during the same cycle of the insemination. To maximize fertility with frozen semen, a careful selection of the stallions and mares, with proper client communication is critical. Dedication and commitment of mare owner and inseminator will have the most significant impact on the pregnancy rates.