Peripheral inhibin levels during estrous cycle in buffalo (Bubalus bubalis)

A sensitive, specific RIA was validated and used for measurement of peripheral plasma immunoreactive inhibin (irinhibin) levels during the estrous cycle in Murrah buffalo. The RIA employed an 125-I iodinated inhibin as tracer and an antiserum against dimeric inhibin. The procedure had a sensitivity of 16 pg/tube, and the nonspecific effects of buffalo plasma were compensated for by including 200 ul bullock plasma in the standards. Separation of free and bound inhibin was affected by the use of a second antibody and precipitation with polyethylene glycol. Blood samples were collected once daily for 30 d from Murrah buffalo (n = 6) during the hot month of July. Cyclic activity and estrus were confirmed by plasma progesterone determination. Peripheral plasma concentrations of ir-inhibin fluctuated between 0.40 +/- 0.07 and 0.67 +/- 0.13 ng/ml during the estrous cycle in buffalo. During the same period, plasma progesterone levels increased from 0.21 +/- 0.01 ng/ml at Day 0 to a peak of 3.30 +/- 0.72 ng/ml on Day 13, declining sharply by Day -5. Ir-inhibin levels exhibited an increase during the follicular phase, with the maximum concentration of 0.65 +/- 0.01 ng/ml occuring on the day of estrus, a decline thereafter, and no pattern during the luteal phase. The differences, however, were not statistically significant throughout the estrous cycle.     

Amino acids in oviduct and uterine fluid and blood plasma during the estrous cycle in the bovine

Up to 40% of cattle embryos die within 3 weeks of fertilization while they are nutritionally dependent on the maternal environment provided by the oviduct and uterine fluids for their development and survival. Despite this dependence there is limited information on the composition of these fluids in cattle. Amino acids are essential for the normal growth and development of the early embryo, acting as precursors of proteins and nucleic acids and as energy sources, osmolytes and signaling molecules. The objective of this study was to measure and compare the amino acid concentrations of oviduct and uterine fluid and blood plasma on different days of the estrous cycle. Oviduct fluid was collected in situ from anaesthetised heifers on Days 0, 2, 3, 4 and 6 and uterine fluid on Days 6, 8 and 14 of the estrous cycle and the concentrations of 19 amino acids determined. Glycine was the most abundant amino acid in both oviduct and uterine fluid. However, the concentrations of many amino acids differed between oviduct and uterus and many were present at higher concentrations in oviduct and uterine fluid than in blood plasma. Oviduct fluid concentrations of amino acids were not affected by day of cycle in contrast to uterine fluid for which there was a day of cycle effect on most of the amino acids. These results provide novel information on the amino acid concentrations in the maternal environment of the early cattle embryo and could form the basis for devising improved media for the production of embryos in vitro.     

Changes in patterns of luteinizing hormone secretion before and after the first ovulation in the postpartum mare

To determine whether luteinizing hormone (LH) secretion during the first estrous cycle postpartum is characterized by pulsatile release, circulating LH concentrations were measured in 8 postpartum mares, 4 of which had been treated with 150 mg progesterone and 10 mg estradiol daily for 20 days after foaling to delay ovulation. Blood samples were collected every 15 min for 8 h on 4 occasions: 3 times during the follicular phase (Days 2-4, 5-7, and 8-11 after either foaling or end of steroid treatment), and once during the luteal phase (Days 5-8 after ovulation). Ovulation occurred in 4 mares 13.2 +/- 0.6 days postpartum and in 3 of 4 mares 12.0 +/- 1.1 days post-treatment. Before ovulation, low-amplitude LH pulses (approximately 1 ng/ml) were observed in 3 mares; such LH pulses occurred irregularly (1-2/8 h) and were unrelated to mean circulating LH levels, which gradually increased from less than 1 ng/ml at foaling or end of steroid treatment to maximum levels (12.3 ng/ml) within 48 h after ovulation. In contrast, 1-3 high-amplitude LH pulses (3.7 +/- 0.7 ng/ml) were observed in 6 of 7 mares during an 8-h period of the luteal phase. The results suggest that in postpartum mares LH release is pulsatile during the luteal phase of the estrous cycle, whereas before ovulation LH pulses cannot be readily identified.     

Cannulation of the equine oviduct and chemical analysis of oviduct fluid

Siliconized rubber tubes were used to cannulate one oviduct in 7 mares, and secretions were collected in a polycarbonate container located externally, in the region of the left paralumbar fossa. Secretion rates were recorded daily during the estrous cycle. Concentrations of calcium, magnesium, sodium, potassium, inorganic phosphorus and glucose were determined in the oviduct fluids secreted throughout the estrous cycle. Secretion rates were greatest during estrus (days 1-9), with a significant decrease (P<.01) noted during nonestrus (days 10-21). Concentrations of all constitutents measured were low during estrus, with a marked elevation in concentration during the nonestrus period. Histologic examination of oviducts following long-term cannulation demonstrated infiltration of lymphocytes, plasma cells, and small areas of degenerated epithelium.     

Content of meiosis activating sterols in equine follicular fluids: correlation to follicular size and dominance

Meiosis activating sterols (MAS) are pre-cholesterol sterols that can be isolated from follicular fluid (FF-MAS) or testes (T-MAS). Meiosis activating sterols trigger the resumption of meiosis in cultured meiotically competent oocytes. In the present work MAS, cholesterol and progesterone were assayed by HPLC in follicular fluids collected from pony mares at fixed days after the last ovulation. Follicles were divided into two groups according to whether they were aspirated before or after Day 17 after the last ovulation. The latter group was further divided according to whether the follicle diameter was < or = 22 mm or > 27 mm. Both FF-MAS and T-MAS were detected in almost all samples. Overall, the total amount of MAS in the follicular fluids increased with the size of the follicles but was accompanied by a decrease in the amount of free cholesterol. The amounts of MAS and progesterone in > 27 mm follicles aspirated after Day 17 were significantly higher as compared to the other groups. A transversal cohort analysis showed that the largest follicle at the time of aspiration had the highest level of MAS after day 17 of the cycle, which was not always true for follicle samples aspirated before Day 17 of the cycle. The study demonstrates that the content of MAS in equine follicular fluids increased during follicular maturation concomitant with a decrease in the concentration of free cholesterol. Moreover, MAS concentration is higher in dominant follicles than in subordinate follicles. The MAS may therefore play an as yet unknown physiological role during pre-ovulatory maturation.     

Reproduction in high body condition mares with high versus low leptin concentrations

Previous results from our laboratory indicated that a majority of mares with high body condition scores (BCS) displayed estrous cycles or had considerable follicular activity during the winter. Among these high BCS mares, about 35% of them exhibited a persistent hyperleptinemia and hyperinsulinemia. The current experiment was designed to compare the reproductive characteristics of high BCS mares with hyperleptinemia to those with normal (low) plasma concentrations of leptin during the winter and the first estrous cycle (or the first full cycle encountered for those already cycling). Light horse mares with high BCS (6-8.5) were assigned to groups based on leptin concentrations (8/group): low (<5 ng/mL) and high (>10 ng/mL). Beginning 7 January, mares were assessed every 3d for follicular activity and then daily once a follicle >25 mm was detected. Mares were subsequently monitored through their first and second ovulations. Leptin concentrations remained higher (P<0.001) in mares in the high leptin group over the duration of the experiment. Also, high leptin mares had greater (P<0.0001) insulin response to glucose infusion and a faster (P<0.05) rate of glucose clearance. One mare with high leptin and three mares with low leptin had progesterone concentrations indicative of the presence of a corpus luteum at the onset of the experiment. Plasma concentrations of LH, FSH, and progesterone did not differ between groups (P>0.1) during the first estrous cycle occurring after 7 January. Date of first ovulation after 7 January and interovulatory interval were similar (P>0.1) for the two groups, as were estimates of follicular numbers on the ovaries (small, medium, and large; P>0.1). It is concluded that the perturbations in leptin and insulin secretion observed in some high BCS mares are not associated with alterations in ovarian activity or the estrous cycle during winter and into the period of vernal transition.     

Effect of phenylbutazone (PBZ) on luteolysis in the mare induced by uterine biopsy

Uterine biopsy in the mare on day 4 post-ovulation causes an acute inflammatory reaction which results in premature luteolysis. In this study, seven mares (4 to 6 years of age) were used in a switchback experimental design to test the hypothesis that in the mare parenterally administered PBZ will block luteolysis induced by uterine biopsy on day 4 post-ovulation. All mares were allowed two normal estrous cycles (range 18 to 24 days). On the first day of estrus of the third estrous cycle each mare was intravenously given 2 grams PBZ (treatment) or 10 ml 0.9% saline (control) daily until signs of estrus were exhibited. The day of ovulation (day 0) was determined by rectal palpation and subsequently verified by peripheral plasma progesterone concentrations. On day 4 following ovulation all mares were subjected to uterine biopsy, and subsequent estrus detection was performed daily using an andro-genized gelding. A total of 19 estrous cycles (ten for PBZ treatment and nine for controls) were evaluated. Mean number of days (+/-SE) from uterine biopsy to induced estrus was 5.00+/-0.16 for control cycles and was significantly different (P<0.025) when compared with 9.20+/-0.34 days for treatment cycles. Results of this study suggest that PBZ can block luteolysis in the mare induced by uterine biopsy on day 4 post-ovulation, possibly as a result of accumulating PBZ in acutely inflamed uterine tissue and inhibiting prostaglandin synthesis.     

Plasma oxytocin concentrations in cyclic mares and sexually aroused stallions

An experiment was conducted to measure plasma oxytocin concentrations at 4 different stages of the estrous cycle in 11 pony mares. Plasma oxytocin concentrations (muU/ml +/- SE) were found to be higher (P<.01) on day 2 of estrous (39.8 +/- 12.5) and day 5 post-ovulation (33.1 +/- 12.0) than on day 10 (2.3 +/- 1.6) and day 15 post-ovulation (6.8 +/- 4.1). A second experiment was conducted to measure jugular plasma oxytocin concentrations before and after sexual arousal in six pony stallions. Oxytocin concentrations (muU/ml +/- SE) were higher (P<0.06) after sexual arousal (50.5 +/- 8.9) than before sexual arousal (23.8 +/- 2.9). These data suggest plasma oxytocin concentrations may be associated with ovarian function in mares and with sexual behavior in stallions.     

Effect of beta-carotene administration on reproductive function of horse and pony mares

The objective of this study was to determine whether supplemental beta-carotene would influence reproductive function in mares maintained on spring and summer pastures and to characterize plasma carotene concentrations during the estrous cycle. Carotene concentrations in plasma did not vary with day of estrous cycle (P = 0.7455). Mares receiving every other day injections of beta-carotene (400 mg; n = 4) or saline (10 ml; n = 4) during proestrus/estrus did not differ in plasma estradiol (E(2)) concentrations (P = 0.6313), follicle development (P = 0.8068), or plasma progesterone (P(4)) concentrations during the following diestrus (P = 0.4954). Moreover, no differences in plasma P(4) concentrations (P = 0.9047) were detected between mares receiving every other day injections of beta-carotene (400 mg; n = 4) or saline (10 ml; n = 4) during diestrus. However, administration of beta-carotene raised plasma carotene concentrations relative to controls when injected during proestrus/estrus (P = 0.0096) and diestrus (P = 0.0099). Pregnancy rates (P = 0.4900) and number of cycles required for pregnancy (P = 0.2880) were similar for mares administered injections of saline (10 ml; n = 37), beta-carotene (400 mg; n = 37), vitamin A (160,000 IU; n = 38), or vitamin A + beta-carotene (160,000 IU + 400 mg; n = 43), on the first or second day of estrus and on the day of breeding. Therefore, these results collectively suggest that supplemental beta-carotene does not affect the reproductive function of mares fed adequate dietary carotene. Whether supplemental beta-carotene would enhance reproductive function in mares on low carotene diets warrants further investigation.     

Effects of norgestomet, altrenogest, and/or estradiol on follicular and hormonal characteristics of late transitional mares

Two experiments were conducted to investigate the effect of norgestomet and altrenogest, alone or in combination with estradiol, on late transitional mares. In the first experiment, 32 mares were assigned to four treatment groups: controls (C), those treated with 1.5 mg of norgestomet (N1), 3.0 mg norgestomet (N2) or 26 mg altrenogest (AT). Treatments were initiated during the months of April and May and given daily for 15 d. During treatment, altrenogest suppressed estrous behavior and diameter of the largest follicle, whereas norgestomet had no effect at either dose. The rise in serum luteinizing hormone (LH) levels following the withdrawal of altrenogest treatment was significantly greater than that for the other three groups. In the second experiment, 24 late transitional mares were assigned to three treatments: controls (C), those receiving 26 mg altrenogest (AT) daily, or 26 mg altrenogest plus 10 mg estradiol (AE) daily for 16 d. Both altrenogest treatments suppressed estrous behavior and follicular growth compared with controls. However, suppression of follicular activity was significantly greater for the combined steroid treatment. Following treatment, the interval to ovulation and estrus was longer for the combined steroid group. We concluded that: 1) norgestomet at a dose up to 3.0 mg per day had no effect on follicular activity, estrous behavior or serum LH levels in late transitional mares, 2) estradiol combined with altrenogest had greater suppressive activity on follicular growth than altrenogest alone, and 3) the greater suppression by the combined steroid treatment had no advantage over altrenogest alone on induction of estrus and ovulation in late transitional mares.     

Estrogen and progesterone concentrations in bovine milk during the estrous cycle

Estrogen and progesterone concentrations in milk during the estrous cycle were estimated in 18 normally cycling Holstein dairy cows. The estrogen and progesterone concentrations in milk during the estrous cycle followed the pattern described for them in blood in the corresponding period. During most of the estrous cycle, estrogen concentration remained at approximately 200 pg/ml and reached a proestrous peak of 360 +/- 127 pg/ml on day 19. The progesterone concentration in milk during the estrous cycle increased to a peak on day 13 (45.5 +/- 6.6 ng/ml) and thereafter declined towards estrus. Estrus detection/prediction based on milk progesterone concentrations appears feasible in view of the significant differences in milk progesterone concentrations between the early luteal (post-ovulatory), luteal and rapid follicular growth periods of the estrous cycle.     

The efficacy of recombinant equine follicle stimulating hormone (reFSH) to promote follicular growth in mares using a follicular suppression model

The efficacy of a recently engineered single chain recombinant equine follicle stimulating hormone (reFSH) was investigated in estrous cycling mares whose gonadotropins and follicular activity had been suppressed by concurrent treatment with progesterone and estradiol (P&E). Time of estrus was synchronized in 15 estrous cycling mares during the breeding season with prostaglandins F_2α (PGF_2α). The day after ovulation, mares were treated once daily with P&E for 14 days. Mares received a second injection of PGF_2α on day 6 of the synchronized estrous cycle to induce luteolysis. On day 8 post-ovulation mares were randomly assigned to three groups: small dose reFSH-treatment group (0.5 mg reFSH IV, twice daily); large dose reFSH-treatment group (0.85 mg reFSH IV twice daily); control group (saline IV, twice daily). reFSH treatment occurred concurrently with the last week of P&E treatment. After a follicle or cohort of follicles reached 35 mm in diameter, mares were injected with 0.75 mg of recombinant equine luteinizing hormone (reLH) to induce ovulation. Post-treatment ovulation was assessed. Daily blood samples were collected for analysis of FSH, LH, estradiol, progesterone, and inhibin by radioimmunoassay (RIA). On the first day of reFSH/saline treatment, blood samples were collected periodically from 1 h prior to treatment to 6 h post-injection via an indwelling jugular catheter to determine acute changes in FSH concentrations. Monitoring of follicular activity, estrus, and ovulation was performed daily by utilizing a stallion and transrectal ultrasonography.A difference (p ≤ 0.05) between the largest diameter follicle in the reFSH-treatment groups compared to controls occurred on day 14 post-ovulation, the day treatments ended, and the difference continued until day 21 post-ovulation. reFSH-treatment groups had larger (p ≤ 0.05) numbers of 20–29 mm follicles (days 13–18), 30–34 mm follicles (days 15–20) and ≥35 mm follicles (days 16–21) than controls. Mares treated with reFSH, at either dose, took less time (average: 2.95 ± 0.42 days) to develop 2–3 times more pre-ovulatory follicles than control mares (7.8 ± 0.51 days) (p ≤ 0.05). The number of ovulations between treated mares and controls were similar due to a greater incidence of ovulation failure in reFSH-treated mares. During reFSH treatment, concentrations of plasma FSH, inhibin and estradiol were greater (p ≤ 0.05) compared to control concentrations. Plasma LH concentrations in reFSH-treated mares were suppressed and did not exhibit the ovulatory surge of controls (p ≤ 0.05). Plasma progesterone concentrations were not different across groups.These findings demonstrate the specific effects of reFSH to increase number of total follicles including pre-ovulatory follicles in mares with endogenous pituitary gonadotropins and follicular growth suppressed by a regimen of P&E.     

The effect of exogenous acth and pen-confinement on estrous cycle length, plasma steroid levels and ovarian function of beef heifers

Each of 24 pasture-reared crossbred beef heifers were herded from a large pasture on day 14 of the estrous cycle and assigned randomly to one of four treatment groups as follows: Field Control (FC), Field ACTH (FA), Pen Control (PC) and Pen ACTH (PA). Field groups were maintained in a small field, and pen groups were confined in a pen in a pole barn. ACTH groups received 200 IU of ACTH IM daily for days 17 through 21 of the cycle and control groups received only the gelatin carrier IM on the same cycle days. Average cycle lengths for FA and PA heifers were 25 and 24.3 days with an average period from plasma progesterone decline below 1 ng/ml to estrus of 5.5 days. During the ACTH injection period, follicular growth was suppressed and the proestrus plasma estrogen rise was delayed. Average cycle lengths for FC and PC heifers were 20.8 and 22.8 days respectively. All control group heifers exhibited estrus within 2 days of the plasma progesterone decline below 1 ng/ml. In addition, pen confinement heifers showed a trend for extended luteal function and consequent extended estrous cycle length.     

Amino acid concentrations in fluids from the bovine oviduct and uterus and in KSOM-based culture media.

Amino acids in bovine oviductal and uterine fluids were measured and compared with those in modified simplex optimized medium (KSOM) supplemented with either fetal calf serum or Minimum Essential Medium amino acids in addition to bovine serum albumin, fetal calf serum or polyvinyl alcohol. Concentrations of cysteine, threonine, tryptophan, alanine, aspartate, glycine, glutamate, proline, beta-alanine, and citrulline were higher in oviductal fluids than in KSOM-based culture media. Nonessential and essential amino acids were present in ratios of 5:1 and 2:1 in oviductal and uterine fluids, respectively. Concentrations of alanine (3.7 mM), glycine (14.1 mM) and glutamate (5.5 mM) were high in oviductal fluids, comprising 73% of the free amino acid pool. Of the amino acids measured in uterine fluids, alanine (3.1 mM), glycine (12.0 mM), glutamate (4.2 mM), and serine (2.7 mM) were highest in concentration, and the first three comprised 43% of the free amino acid pool. In conclusion, amino acid concentrations in the bovine reproductive tract were substantially higher than those in embryo culture media. Certain amino acids, particularly alanine, glutamate, glycine and taurine, are present in strikingly high concentrations in both oviductal and uterine fluids, suggesting that they might play important roles in early embryo development. The particular pattern of amino acid concentrations may be an important factor to be considered for the improvement of embryo culture media.     

Influence of immunization against GnRH on reproductive cyclicity and estrous behavior in the mare

The aim of this study was to evaluate the effect of active immunization against GnRH on ovarian activity, plasma progesterone and estradiol concentrations and on estrous behavior in adult mares. Eighteen cyclic mares were randomly divided into a treatment and control group. Nine mares were immunized twice with 2 mL (400 microg GnRH-protein conjugate) of a GnRH-vaccine (Improvac, CSL Limited, Australia) administered intramuscularly, 4 weeks apart. Control mares received the same amount of saline solution. Ovaries and uterus of all mares were examined weekly by ultrasonography from 3 weeks before to 60 weeks after first immunization. Thereafter, vaccinated mares were evaluated monthly until 100 weeks after first vaccination. In addition, mares were teased with a stallion for assessment of estrous behavior and blood was collected for progesterone, estradiol-17beta and GnRH antibody titer determination. Results demonstrate that vaccination against GnRH significantly (P<0.05) influenced all parameters, except estradiol-17beta concentration. All vaccinated mares ceased reproductive cyclicity (plasma progesterone <1 ng/mL, follicles <3 cm) within 8 weeks after the first injection and ovarian activity remained suppressed for a minimum of 23 weeks. Five mares resumed cyclicity (follicles >3 cm, progesterone >1 ng/mL) while three mares showed only follicular activity (follicles >3 cm) and one mare remained completely suppressed for the entire duration of the study. In spite of ovarian suppression, four mares expressed sporadic and one mare continuous estrous behavior. In conclusion, reproductive cyclicity in adult mares can be successfully suppressed by immunization against GnRH but the timing of resumption of cyclicity is highly variable and estrous behavior may occur in spite of ovarian suppression.     

Amino acids in cat fallopian tube and follicular fluids

Aminograms of tubal and follicular fluids were obtained using fluids collected by aspiratory puncture from six cats. The amino acids were separated and quantified by high-performance liquid chromatography analysis. The serum of the cats was used as control. The three most prevalent amino acids quantified in cat tubal fluid were glycine, glutamic acid, and taurine. Their mean concentrations were 840 μmol/l (μm), 808 μm and 596 μm, respectively. The three most prevalent amino acids quantified in cat follicular fluid were alanine, glutamine, and taurine. Their mean concentrations were 359 μm, 351 μm, and 258 μm, respectively. This result is consistent with aminograms of tubal fluid previously determined in other mammals. As previously observed in other species and humans, glycine was quantitatively the most abundant and most prevalent free amino acid in cat tubal fluid. The total quantity of amino acids in tubal fluid was similar in cats and other species. However, in contrast with other species studied, hypotaurine was not detected in tubal and follicular fluids of female cats.     

Is one-wave follicular growth during the estrous cycle a usual phenomenon in water buffaloes (Bubalus bubalis)?

The pattern of growth and regression of ovarian follicles was monitored once daily for one complete estrous cycle in eight individual water buffaloes by ultrasonographic scanning of the ovaries for an entire interovulatory interval of normal cycle length. One-wave follicular growth was observed in five animals and two-wave follicular growth in three buffaloes during the estrous cycle. The first follicular wave of a two-wave cycle emerged significantly earlier (P < 0.05) than the emergence of the solitary wave of a one-wave cycle. One- and two-wave cycles differed significantly (P < 0.05) with respect to the mean interovulatory interval (21.0 +/- 0.54 days versus 22.7 +/- 0.33 days) and the mean interestrus interval (20.8 +/- 0.58 days versus 22.3 +/- 0.66 days). The overall linear growth rate of the ovulatory follicle was significantly greater (P < 0.01) in a two-wave cycle compared to that of a one-wave cycle (1.17 +/- 0.33 mm/day versus 0.32 +/- 0.01 mm/day). In a one-wave pattern, the growth profile of the solitary dominant follicle was atypical, showing three distinct phases, i.e. growth phase, regression phase and regrowth phase culminating in ovulation. The level of plasma progesterone steadily increased from day 0 of estrous cycle, attained peak level on day 14 and declined thereafter. A slower growth rate of the dominant follicle was observed in the presence of higher plasma progesterone concentration. The present study shows that one-wave follicular growth is a normal phenomenon in suckled water buffaloes.     

Changes in circulating plasma levels of cortisol in lactating and non-lactating dairy cattle during the estrous cycle

Plasma cortisol levels were determined by radioimmunoassay for five lactating cows and five open heifers from blood samples collected daily during the course of a complete estrous cycle. Each animal was fitted with an indwelling jugular catheter to minimize stress from bleeding. Mean plasma cortisol levels were 5.67 ng/ml for the lactating cows and 5.87 ng/ml for the open heifers. Mean values for individuals ranged from 3.79 ng/ml to 6.94 ng/ml in the lactating group, and 3.37 ng/ml to 11.69 ng/ml in the open group. These differences, both between and within groups, were not significant. Mean cortisol values during the estrous cycle ranged from 2.26 +/- 0.93 ng/ml on day one to 9.49 +/- 2.11 ng/ml on day six for the lactating group, and 2.74 +/- 0.52 ng/ml on day six to 14.66 +/- 10.78 ng/ml on day twelve for the open group. Group by day interactions were not significant. Attempts to correlate plasma cortisol with lactation or day of estrus were not significant.     

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.     

Studies on the effect of manual massage of the ovaries on the reproductive activity of the mare

The aim of this study was to determine the influence of hand massage of mares' ovaries on breeding activity and hormonal changes in the winter anestrous period and during the luteal phase of the oestrus cycle. The experiment was conducted on 5 experimental and 5 control mares. In winter, (January) the experimental mares underwent 30-sec daily massage of both ovaries, for 30 d, and in summer (August) from the 6th day of the cycle to the occurrence of estrus. The sexual behavior of all mares was determined each day by individual teasing by a vigorous stallion, and the ovaries were checked by palpation per rectum and with an USG. Every second day blood samples were drawn from each mare to determine progesterone and estradiol in the plasma. Ovarian massage during deep winter anestrus had no significant effect on acceleration of the mares' active breeding season. Nevertheless, a higher concentration of estradiol was observed in the experimental group. These differences occurring on the 11th, 17th and 20th days were found to be significant (P ≤ 0.05). It was shown that during the summer period, in the luteal phase of the cycle, ovarian massage shortened the length of the estrous cycle, and ovulation was brought on somewhat earlier.