Estrogen and LH dynamics during the follicular phase of the estrous cycle in the Asian elephant

Pituitary and corpus luteum hormone patterns throughout the elephant estrous cycle have been well characterized. By contrast, analysis of follicular maturation by measurement of circulating estrogens has been uninformative. This study tested the ability of a urinary estradiol‐3‐glucuronide radioimmunoassay to noninvasively assess follicular development during the nonluteal phase of the elephant estrous cycle, and to determine the relationship between estrogen production and the “double LH surge.” Daily urine and serum samples were collected throughout seven estrous cycles from three Asian elephants, and urine was collected from an additional three females, for a total of 13 cycles. Serum was analyzed for luteinizing hormone (LH), and urine was analyzed for estrogens and progestins. Elephants exhibited a typical LH pattern, with an anovulatory LH (anLH) surge occurring approximately 21 days before the ovulatory LH (ovLH) surge. The urinary estrogen pattern indicated the presence of two follicular waves during the nonluteal phase. The first wave (anovulatory) began 5 days before the anLH surge and reached a maximum concentration the day before the peak. Thereafter, urinary estrogens declined to baseline for 2 weeks before increasing again to peak concentrations on the day of the ovLH surge. Urinary progestins were baseline throughout most of the follicular phase, increasing 2–3 days before the ovLH surge and continuing into the luteal phase. These results support previous ultrasound observations that two waves of follicular growth occur during the nonluteal phase of the elephant estrous cycle. Each wave is associated with an increase in estrogen production that stimulates an LH surge. Thus, in contrast to serum analyses, urinary estrogen monitoring appears to be a reliable method for characterizing follicular activity in the elephant. Zoo Biol 22:443–454, 2003. © 2003 Wiley‐Liss, Inc.     

Analysis of urinary immunoreactive steroid metabolites and gonadotropins for characterization of the estrous cycle,breeding period,and seasonal estrous activity of captive killer whales (Orcinus orca)

To increase the basic understanding of killer whale (Orcinus orca) reproductive physiology necessary for the development of artificial breeding programs, we utilized radioimmunoassays (RIA) to detect urinary immunoreactive steroid metabolites (pregnanediol-3α-glucuronide [PdG] and estrone-conjugates [EC]) and gonadotropins (luteinizing hormone [LH] and follicle-stimulating hormone [FSH]) in urine samples from six female killer whales. Urine samples were collected from the whales by voluntary presentation behavior over a 2- to 4-year period. All urinary hormone values were corrected for intersample urine concentration variations by indexing with creatinine. Daily urine samples from four whales were collected during two conceptions and 18 complete estrous cycles. LH, FSH, EC, and PdG immunoreactive levels were determined and combined with observed copulatory activity in five cycles, including two conceptive cycles from two whales. Mean luteal phase lengths ranged from 9.7 to 19.2 days. Mean follicular phase lengths ranged from 6.5 to 16.8 days. Mean estrous cycle lengths based on the first detectable PdG levels were 41.6 ± 6.72 S.E.M. days. After PdG nadir, immunoreactive FSH levels showed a bimodal pattern with the first peak being greater in size, and both preceding a follicular phase EC increase. LH levels > the 95% confidence interval of the mean were considered significant. Combined LH immunoreactive values from whales 2 and 6 during two and three estrous cycles, respectively, had significant LH peak concentrations on day minus 2. These significant LH peaks were assumed to represent the preovulatory LH surge. Eight copulations during two conceptive cycles were observed between whales 2 and 6 and a breeding male. Six of these copulations (3 with each female whale) occurred within 72 hours of the beginning or the end of the presumptive preovulatory LH surge. Estrous activity was seen throughout the year for the herd. However, individuals had varying periods of anestrus that could not be linked to environmental, social, or nutritional influences. The whales that were reproductively successful had anestrus intervals that were usually influenced by gestation, postparturient period, or lactation. The information obtained during this research enhances the foundation for future artificial reproductive management techniques. © 1993 Wiley-Liss, Inc.     

Negative effect of estradiol on luteinizing hormone throughout the ovulatory luteinizing hormone surge in mares

The negative effect of estradiol-17beta (E2) on LH, based on exogenous E2 treatments, and the reciprocal effect of LH on endogenous E2, based on hCG treatments, were studied throughout the ovulatory follicular wave during a total of 103 equine estrous cycles in seven experiments. An initial study developed E2 treatment protocols that approximated physiologic E2 concentrations during the estrous cycle. On Day 13 (ovulation = Day 0), when basal concentrations of E2 and LH precede the ovulatory surges, exogenous E2 significantly depressed LH concentrations to below basal levels. Ablation of all follicles > or = 10 mm when the largest was > or =20 mm resulted in an increase in percentage change in LH concentration within 8 h that was greater (P < 0.03) than for controls or E2-treated/follicle-ablated mares. Significant decreases in LH occurred when E2 was given when the largest follicle was either > or =25 mm, > or =28 mm, > or =35 mm, or near ovulation. Treatment with 200 or 2000 IU of hCG did not affect E2 concentrations during the initial portion of the LH surge (largest follicle, > or =25 mm), but 2000 IU significantly depressed E2 concentrations before ovulation (largest follicle, > or =35 mm). Results indicated a continuous negative effect of E2 on LH throughout the ovulatory follicular wave and may be related to the long LH surge and the long follicular phase in mares. Results also indicated that a reciprocal negative effect of LH on E2 does not develop until the E2 surge reaches a peak.     

Reproductive hormone profiles in mares during the autumn transition as determined by collection of jugular blood at 6 h intervals throughout ovulatory and anovulatory cycles

The aim was to define precisely the FSH secretion pattern in mares during the two ovulatory cycles before, and for 24 days after, the last ovulation of the season and to compare this with the profiles of other reproductive hormones and follicular growth to identify changes which may lead to the termination of follicular cycles. Jugular blood was collected every 6 h from ten light horse mares for 6 weeks in autumn. Samples were assayed for FSH, LH, prolactin, inhibin, oestrone conjugates and progesterone. Luteolysis occurred earlier and periovulatory oestrone, but not inhibin, concentrations were significantly lower in the last than in the second to last cycles. In ovulatory and anovulatory cycles, daily mean FSH concentrations were low at the expected time of ovulation and high between days 9 and 11 (day 0 = ovulation), which were usually after luteolysis. However, the periovulatory FSH nadir was prolonged in the last compared with the second to last cycles, and the difference between peak and trough values was not significant in anovulatory cycles. Between day 5 and day 8, the FSH interpulse interval was approximately 2 days, and did not vary in successive cycles. The LH profile also showed progressive changes as mares entered acyclicity; the surge terminated sooner in the last than in the second to last cycles, and failed to occur when expected in acyclicity. Sporadic prolactin pulses occurred at luteolysis in a similar proportion of ovulatory and anovulatory cycles. These results indicate that inadequate gonadotrophin stimulation in early dioestrus may be a critical event leading to suboptimal follicular and luteal development, and eventually acyclicity. Moreover, the time relationships amongst changes in pituitary and ovarian hormones and follicular growth become increasingly disrupted during the autumn transition, which may contribute to the cessation of cyclicity.     

Role of the double luteinizing hormone peak, luteinizing follicles, and the secretion of inhibin for dominant follicle selection in Asian elephants (Elephas maximus)

Elephants express two luteinizing hormone (LH) peaks timed 3 wk apart during the follicular phase. This is in marked contrast with the classic mammalian estrous cycle model with its single, ovulation-inducing LH peak. It is not clear why ovulation and a rise in progesterone only occur after the second LH peak in elephants. However, by combining ovarian ultrasound and hormone measurements in five Asian elephants (Elephas maximus), we have found a novel strategy for dominant follicle selection and luteal tissue accumulation. Two distinct waves of follicles develop during the follicular phase, each of which is terminated by an LH peak. At the first (anovulatory) LH surge, the largest follicles measure between 10 and 19.0 mm. At 7 ± 2.4 days before the second (ovulatory) LH surge, luteinization of these large follicles occurs. Simultaneously with luteinized follicle (LUF) formation, immunoreactive (ir) inhibin concentrations rise and stay elevated for 41.8 ± 5.8 days after ovulation and the subsequent rise in progesterone. We have found a significant relationship between LUF diameter and serum ir-inhibin level (r(2) = 0.82, P < 0.001). The results indicate that circulating ir-inhibin concentrations are derived from the luteinized granulosa cells of LUFs. Therefore, it appears that the development of LUFs is a precondition for inhibin secretion, which in turn impacts the selection of the ovulatory follicle. Only now, a single dominant follicle may deviate from the second follicular wave and ovulate after the second LH peak. Thus, elephants have evolved a different strategy for corpus luteum formation and selection of the ovulatory follicle as compared with other mammals.     

Seasonal effects on the endocrine pattern of semi-captive female Asian elephants (Elephas maximus): timing of the anovulatory luteinizing hormone surge determines the length of the estrous cycle

Better breeding strategies for captive Asian elephants in range countries are needed to increase populations; this requires a thorough understanding of their reproductive physiology and factors affecting ovarian activity. Weekly blood samples were collected for 3.9 years from 22 semi-captive female Asian elephants in Thai elephant camps to characterize LH and progestin patterns throughout the estrous cycle. The duration of the estrous cycle was 14.6+/-0.2 weeks (mean+/-S.E.M.; n=71), with follicular and luteal phases of 6.1+/-0.2 and 8.5+/-0.2 weeks, respectively. Season had no significant effect on the overall length of the estrous cycle. However, follicular and luteal phase lengths varied among seasons and were negatively correlated (r=-0.658; P<0.01). During the follicular phase, the interval between the decrease in progestin concentrations to baseline and the anovulatory LH (anLH) surge varied in duration (average 25.9+/-2.0 days, range 7-41, n=23), and was longer in the rainy season (33.4+/-1.8 days, n=10) than in both the winter (22.2+/-4.5 days, n=5; P<0.05) and summer (18.9+/-2.6 days, n=8; P<0.05). By contrast, the interval between the anLH and ovulatory LH (ovLH) surge was more consistent (19.0+/-0.1 days, range 18-20, n=14). Thus, seasonal variation in estrous cycle characteristics were mediated by endocrine events during the early follicular phase, specifically related to timing of the anLH surge. Overall reproductive hormone patterns in Thai camp elephants were not markedly different from those in western zoos. However, this study was the first to more closely examine how timing of the LH surges impacted estrous cycle length in Asian elephants. These findings, and the ability to monitor reproductive hormones in range countries (and potentially in the field), should improve breeding management of captive and semi-wild elephants.     

Deep intra-uterine artificial inseminations using cryopreserved spermatozoa in beluga (Delphinapterus leucas)

Artificial insemination (AI) with liquid-stored spermatozoa and sperm cryopreservation using directional freezing (DF) have been successful in the beluga. This study built on this foundation to develop a deep intra-uterine AI technique with frozen-thawed semen in beluga. Forty-two ejaculates from one male were cryopreserved using DF technology and subsequently used for 10 insemination attempts with seven females. Percentage pre- and post-thaw progressive motility and viability were (mean ± SD) 73.0 ± 12.2, 38.4 ± 8.8, 88.0 ± 0.1, and 59.3 ± 15.7%, respectively. A series of GnRH injections (3 x 250 μg, IV, 1.5 to 2 h apart) were used to induce ovulation, once a growing follicle >2.5 cm in diameter was visualized via trans-abdominal ultrasonography. Artificial insemination was performed at 30.1 ± 3.8 h post-initial GnRH injection with semen deposited in the uterine horn, 92.6 ± 16.2 cm beyond the genital opening using a flexible endoscope. The external cervical os (cEOS) was located beyond a series of 5 to 10 vaginal rings, 44.8 ± 9.3 cm from the external genital opening. The internal bifurcation of the uterus was 27 ± 6.8 cm beyond the cEOS. Ovulation occurred at 8.5 ± 7.6 h post-AI. Two of 10 inseminations (20%) resulted in pregnancy. The first pregnancy resulted in twins; both calves were born 442 d after AI, with one surviving. The second pregnancy is ongoing. These findings represent the first successful application of AI using frozen-thawed semen in beluga, and are important examples of how assisted reproductive technologies can provide tools for the global management of threatened species.     

Prediction of the estrous cycle and optimal insemination time by monitoring vaginal electrical resistance (VER) in order to improve the reproductive efficiency of the Okinawan native Agu pig

The present study was conducted to determine whether vaginal electrical resistance (VER) can be exploited to improve the low reproductive efficiency of the rare Okinawan native Agu pig, in which estrous signs are difficult to ascertain by visual observation. The lowest VER (272.0+/-12.4 units, n=5) and the preovulatory LH surge were detected at 57.6+/-5.3 and 36.8+/-9.6h before the onset of estrus, respectively. The initiation of gradual increase in VER was found after 9.6+/-4.7h following the peak LH, and the higher levels of VER were plateaued during the luteal phase. These VER fluctuations were correlated with changes in plasma LH (P<0.05) and progesterone (P<0.001), but not estrogen. Moreover, the conception rate (41%, n=32) was dramatically improved by artificial insemination at 24 and 34 h after the beginning of the VER increase when compared with insemination at the conventional time (12 and 24h after detection of estrus, 20%, n=45), widely used in commercial pigs (P<0.05). These data suggest that VER fluctuation can be used to estimate the stage of the estrous cycle, and the scheduling artificial insemination according to increase in VER as an index for the preovulatory LH surge could improve Agu reproductive efficacy.     

In vitro fertilization and subsequent development of porcine oocytes using cryopreserved and liquid-stored spermatozoa from various boars

We had previously developed a porcine IVF system using a chemically defined medium, i.e., porcine gamete medium supplemented with theophylline, adenosine, and cysteine (PGMtac). In the present study, we investigated the utility of this IVF system using different types of semen: (1) cryopreserved ejaculated (n = 8); (2) cryopreserved epididymal (n = 4); and (3) liquid-stored ejaculated (n = 5). Cryopreserved spermatozoa were prepared by three methods. In vitro-matured porcine oocytes were fertilized for 20 h in PGMtac using each type of semen, and the presumptive zygotes were cultured in porcine zygote medium (PZM)-4 for 5 days. In the case of frozen-thawed spermatozoa, the number of spermatozoa per penetrated oocyte (1.1-1.7), rate of blastocyst formation (26-56%), and total number of cells per blastocyst (34-49) differed (P < 0.05) among freezing methods. However, blastocysts were produced using all types of cryopreserved spermatozoa (14-75%). When spermatozoa were liquid-stored for 1-14 days after semen collection, the rate of sperm penetration (P < 0.05) decreased as storage time increased, although there was no significant reduction in sperm motility during storage. In all groups, semen that had been stored within 10 days after collection enabled blastocyst production in vitro (20-48%). In conclusion, this IVF system, which uses a chemically defined medium, had widespread utility with both frozen-thawed and liquid-stored spermatozoa.     

Reproductive cycle of the elephant

The combination of a few factors, including poor captive reproduction, secession of importation from the wild and advances in hormone detection and ultrasonography, has contributed to the current knowledge on the elephant reproductive cycle. Several reproductive features in elephants differ markedly from other mammals. These include the urogenital tract anatomy, length and structure of the reproductive cycle, the formation of multiple corpora lutea and the type and secretion pattern of reproductive hormones. Being 13-18 weeks in length, the elephant estrous cycle is the longest amongst all studied non-seasonal mammals to date. Progesterone increases 1-3 days after ovulation, indicating the start of the luteal phase, which lasts 6-12 weeks. This is followed by a 4- to 6-week follicular phase that is concluded by two, precisely spaced and timed, LH surges. In general, the first, anovulatory LH surge occurs exactly 19-21 days before the second, ovulatory surge. Normally, a single follicle is ovulated. However, beside a corpus luteum (CL) forming on the site of ovulation, multiple accessory CLs can be found on the ovaries. Unlike many other species, the predominant progestagen secreted by luteal tissues is not progesterone, but rather its 5-alpha-reduced metabolites. The currently known aspects of the unique estrous cycle in Asian and African elephants, covering estrous behavior, circulating hormones, ultrasonography and anatomy of the reproductive organs as well as hormonal manipulation treatment possibilities, will be reviewed here.     

The variability in the interval between estrus and ovulation in cattle and its determinants

Fertility of Holstein cows has been decreasing for years and, to a lesser extent, the fertility of heifers too but more recently. A hypothesis to explain this phenomenon may be that the chronology of events leading to ovulation is different for those animals bred nowadays when compared to what was reported previously; this would result in an inappropriate time of insemination. Therefore, two experiments were designed to investigate the relationships among estrus behavior, follicular growth, hormonal events and time of ovulation in Holstein cows and heifers. In the first experiment, the onset of estrus, follicular growth, patterns of estradiol-17beta, progesterone and LH, and the time of ovulation were studied in 12 cyclic Holstein heifers that had their estrus synchronized using the Crestar method; this was done twice, 3 weeks apart. The intervals between estrus and ovulation, estrus and the LH peak, and between the LH peak and ovulation were, respectively, 38.5 h +/-3.0, 9.1 +/- 2.0 and 29.4 h +/-1.5 (mean+/- S.E.M). The variation in the interval between estrus and the LH peak explained 80.6% of the variation in the interval between estrus and ovulation. The intervals between estrus and the LH peak, and estrus and ovulation were correlated with estradiol-17beta peak value (r=-0.423, P <0.04 and r=-0.467, P<0.02, respectively). Positive correlation coefficients for the number of follicle larger than 5 mm, and negative correlation coefficients for the size of the preovulatory follicle with the intervals between estrus and LH peak, LH peak and ovulation, and estrus and ovulation suggest an ovarian control of these intervals. In respect to its role to explain the variation in the interval between estrus and ovulation, the variation in the interval between estrus and the LH peak was evaluated further in a second set of experiments utilizing 12 pubertal Holstein heifers and 35 Holstein cows. The duration of the interval between the beginning of estrus and the LH peak was longer in heifers than in cows (4.15 h versus -1.0 h; P <0.002); the variation for this interval was higher in cows than in heifers (S.E.M.= 1.2 h versus 0.8 h; P=0.01). According to the results of these studies it can be proposed that estradiol and other product(s) of ovarian origin regulate not only the duration of intervals between the onset of estrus and the LH surge but also between the LH surge and ovulation. From the results obtained in the first experiment, it may be postulated that differences observed between cows and heifers for the duration of the interval between onset of estrus and the LH surge as well as for the variation of this interval would be observed also for the interval between the onset of estrus and ovulation. Therefore, on a practical point of view, the long interval between the onset of estrus and ovulation and the high variation of this interval, especially in cows, may be a source of low fertility and should be considered when analysing reproductive disorders.     

LH surge in Nelore cows (Bos indicus), after induced estrus or after ovarian superestimulation

Considering that there is limited information about the preovulatory LH surge in Zebu cattle (Bos indicus), the purpose of the present work was to assess the LH surge in Nelore cows during the estrous cycle and after ovarian superestimulation of ovarian follicular development with FSH. This information is particularly important to improve superovulatory protocols associated with fixed-time artificial insemination. Nelore cows (n=12) had their estrus synchronized with an intravaginal device containing progesterone (CIDR-B) associated with estradiol benzoate administration (EB, 2.5 mg, i.m., Day 0). Eight days later all animals were treated with PGF2alpha (Day 8) in the morning (8:00 h) and at night, when CIDR devices were removed (20:00 h). Starting 38h after the first PGF2alpha injection, blood sampling and ovarian ultrasonography took place every 4h, during 37 consecutive hours. Frequent handling may have resulted in a stress-induced suppression of LH secretion resulting in only 3 of 12 cows having ovulations at 46.7+/-4.9 and 72.3+/-3.8 h, respectively, after removal of CIDR-B. Thirty days later, the same animals received the described hormonal treatment associated with FSH (Folltropin), total dose=200 mg) administered twice a day, during 4 consecutive days, starting on Day 5. Thirty-six hours after the first injection of PGF2alpha, to minimize stress, only seven blood samples were collected at 4h interval each, and ultrasonography was performed every 12 h until ovulation. In 11 of 12 cows (92%) the LH surge and ovulation were observed 34.6+/-1.6 and 59.5+/-1.9 h, respectively, after removal of progesterone source. The maximum values for LH in those animals were 19.0+/-2.6 ng/ml (mean+/-S.E.M.). It is concluded that, in Nelore cows submitted to a ovarian superstimulation protocol, the LH surge occurs approximately 35 h after removal of intravaginal device containing progesterone, and approximately 12h before the LH surge observed after an induced estrus without ovarian superstimulation.     

Pregnancy in the domestic cat after vaginal or transcervical insemination with fresh and frozen semen

The objective was to compare pregnancy rates in domestic cats using fresh semen for intravaginal artificial insemination (IVI), either at the time of hCG treatment for induction of ovulation, or 28 h later, and to compare pregnancy rates following IVI or transcervical intrauterine insemination (IUI) of frozen-thawed semen. Eighteen queens were inseminated during 39 estrus cycles. Fresh semen with 13.5+/-5.4 x 10(6) sperm (range, 6.8-22 x 10(6)) collected by electroejaculation from four male cats was used in Experiment 1, and cryopreserved semen (20 x 10(6) sperm, with 70+/-5% post-thaw motility) from one male cat was used in Experiment 2. Serum concentrations of estradiol-17beta and progesterone were determined in most queens on the day of AI and again 30-40 days later. Treatment with 100 IU of hCG 3 days after the onset of estrus induced ovulation in 95% of treated queens. Pregnancy rates to IVI with fresh semen at the time of hCG administration versus 28 h later were not different (P=0.58); overall 33% (5/15) of the queens became pregnant. For frozen-thawed semen, AI was consistently done 28h after hCG administration; IUI and IVI resulted in pregnancy rates of 41.7% (5/12), whereas no queen (0/12) became pregnant by IVI (P=0.0083). In conclusion, an acceptable pregnancy rate was obtained with frozen-thawed semen in the domestic cat by non-surgical transcervical IUI; this method might also be useful in other small felids.     

Follicular dynamics and temporal relationships among body temperature, oestrus, the surge of luteinizing hormone and ovulation in Holstein heifers treated with norgestomet

The effects of chronic treatment with norgestomet on follicular dynamics, corpus luteum growth and function as well as the temporal relationships among body temperature, oestrous behaviour, the luteinizing hormone (LH) surge and ovulation following implant removal were studied in 16 Holstein heifers. Oestrous cycles of the heifers were initially synchronized using 2 injections of prostaglandin F-2 alpha (PGF-2 alpha) 12 days apart. The heifers were then implanted with a norgestomet ear implant for 9 days, beginning either at the middle of the synchronized cycle (dioestrus) or at the end of the synchronized cycle (pro-oestrus). Follicular dynamics, corpus luteum growth and regression, and plasma progesterone were not affected by norgestomet treatment at dioestrus. The dominant follicle present at the time of norgestomet implantation in the pro-oestrus group was maintained during the 9-day implant period of 6 of 8 heifers and ovulated after implant removal. Time from implant removal to onset of standing oestrus and time to LH peak following implant removal were highly correlated with the time of ovulation (r = 0.92 and 0.96, respectively). Onset of standing oestrus and the LH peak and the onset of standing oestrus and peak vaginal and rectal temperatures were also highly correlated (r = 0.96, 0.82 and 0.81, respectively). It is concluded that any decrease in pregnancy rates following treatment with norgestomet is not due to asynchrony among oestrus, the LH surge and ovulation.     

Pregnancies following long luteal phases in southern white rhinoceros (Ceratotherium simum simum)

All extant species in the Rhinocerotidae family are experiencing escalating threats in the wild, making self-sustaining captive populations essential genetic reservoirs for species survival. Assisted reproductive technologies (ARTs) will become increasingly important for achieving and maintaining ex situ population sustainability and genetic diversity. Previous reports have shown that a large proportion of captive southern white rhinoceros (SWR) females are irregularly cyclic or acyclic, and that cycling females display two different estrous cycle lengths of approximately 30 or 70 days. It has been suggested that the longer estrous cycle length is infertile or subfertile, as no term pregnancies have been observed following long cycles. Here we report the achievement of two pregnancies following long luteal phases, using ovulation induction and artificial insemination with either fresh or frozen-thawed semen. One female SWR conceived on the first insemination attempt and gave birth to a live offspring. A second female conceived twice in consecutive long cycles although the first embryo was resorbed by 33 days post-insemination. A pregnancy from this female's second insemination is ongoing with expected parturition in November 2019. Whether prolonged estrous cycles in SWR are subfertile or infertile in natural breeding situations remains unclear. However, our findings demonstrate that the application of ARTs following prolonged cycles can result the successful establishment of pregnancies in SWR. Therefore, with ARTs, female SWR otherwise considered nonreproductive due to long estrous cycles may still have the potential for representation and contribution to the ex situ population.     

Behavioral, follicular and gonadotropin changes during the estrous cycle in donkeys

Sexual behavior, follicular development and ovulation, and concentrations of circulating gonadotropins during the estrous cycle were studied during the summer in 7 jennies. Mean behavioral estrous length was 6.4 +/- 0.6 days (mean +/- SEM, n=19; 5.6 +/- 0.5 days preovulatory and 0.8 +/- 0.2 days post-ovulatory). Mean diestrous length was 19.3 +/- 0.6 days (n=14). Females in estrus typically showed posturing, mouth clapping, clitoral winking, urinating and tail raising. Mouth clapping began approximately one day sooner and lasted approximately one day longer than winking and tail raising, so that the total duration of clapping was significantly greater than for the other two signs. Follicular changes and concentrations of gonadotropins were determined for 14 estrous cycles (2 per jenny). The follicular end points [diameter of the largest follicle and number of large (>25 mm), medium (20-24 mm), and small follicles (<20 mm)] showed a significant day effect. The diameter of the largest follicle and the number of large follicles began to increase significantly 7 days prior to ovulation with a maximum value the day before ovulation. Medium follicles reached a maximum number 4 days prior to ovulation, and small follicles decreased significantly prior to ovulation. After ovulation, all follicular end points, except the number of small follicles, remained low for the next 12 days. Mean values of FSH were low during estrus and high during diestrus with 2 significant peaks, one 3 days and one 9 days after ovulation. In contrast, mean levels of LH were low during diestrus and high during estrus with a maximum value the day after ovulation. The LH profile showed a more prolonged gradual increase prior to ovulation, than that which has been reported for ponies and horses.     

Papanicolaou staining of exfoliated vaginal epithelial cells facilitates the prediction of ovulation in the giant panda

The giant panda is seasonally monoestrus, experiencing a single estrous with spontaneous ovulation in the spring. Therefore, accurate monitoring of the estrous cycle to pinpoint the time of ovulation is critical for the success of timed mating or artificial insemination. Analysis of exfoliated vaginal epithelial cells is a simple technique that rapidly yields information about the estrous status of a panda. Vaginal swabs were obtained during five estrous cycles of two nulliparous females. Cells were stained with the trichrome Papanicolaou and classified as basophils, intermediates or superficials. The color of stained cells, basophilic, acidophilic or keratinized, was recorded as a characteristic independent of the three standard cell types. The day urinary conjugates of estrogen fell from peak levels was considered the day of ovulation. A chromic shift occurred 8-9 days before ovulation when the majority of exfoliated vaginal cells changed from basophilic (blue) to acidophilic (pink) without accompanying nuclear or cytoplasmic changes. A second chromic shift was consistently observed 2 days prior to ovulation when keratinized (orange) cells replaced acidophils as the majority of vaginal cells. Monochrome staining of vaginal cells is sufficient to quantify superficial cells, which is a useful adjunct to behavioral and endocrinological data in determining estrous in the giant panda. However, the timing and duration of superficial cell elevations are substantially different between and within individual females, which limits the accuracy of timing ovulation for artificial insemination. The predictive value of vaginal cytology was greatly enhanced with the trichrome stain and evaluation of cell color.     

Effect of repeated laparoscopic surgery on the bovine estrous cycle

The effects of repeated laparoscopic surgery on the length of the bovine estrous cycle, estrus, ovulation and corpus luteum function were determined after one estrous cycle of normal duration (18 to 24 days). Five, Angus x Hereford cows were subjected to laparoscopy on days 5, 13, 18 and 20 (estrus = day 0) of the subsequent cycle. Blood was collected daily during the cycle in which laparoscopy was performed (surgical cycle) and during the next cycle (postsurgical cycle). Lengths of the surgical and postsurgical cycles (22.3 +/- .5 days and 21.5 +/- .6 days, respectively) did not differ (P>.05) from that of the presurgical cycle (21.8 +/- .2 days). Average concentrations (ng/ml) of LH and progesterone in serum were similar during the surgical and postsurgical cycles (1.2 +/- .1, 2.2 +/- .2 vs 1.3 +/- .2 and 2.3 +/- .1). Progesterone concentrations remained above 1 ng/ml for 17 and 16 days during the surgical and postsurgical cycles, respectively. A pre-ovulatory rise in LH, along with estrus and ovulation was confirmed in all animals. Follicular development, characterized by follicular volume, increased progressively from days 5 to 20, with the largest increase occurring between days 13 and 18. These results indicate that laparoscopy, used at the times and frequency specified, does not alter reproductive function of cyclic cows and can provide information on ovarian activity.     

Male stimulation of luteinizing hormone surge, progesterone secretion and ovulation in spontaneously persistent-estrous, aging rats

In aging, persistently estrous (PE) female rats, there are no estrous cycles or cyclic increases in luteinizing hormone (LH) secretion, but the sexual receptivity to the male is consistently maintained. We recently reported that caging and mating with fertile males elicits an LH surge followed by ovulation in aging PE rats. The present study examined the relationship between the LH surge, the increase in progesterone (P) secretion and ovulation in PE females exposed to males, and assessed whether intromission was essential for the male-induced pre-ovulatory LH surge. PE rats were implanted with intra-atrial cannulae. Six to eight days later, these females were individually caged with a fertile male and repeatedly sampled (once every 30 or 60 min) between 1400 and 1900 h for assays of plasma LH and P. Sexual behavior of the female was recorded and correlated with the changes in plasma LH and P values. Similar experiments were also performed on cannulated PE rats with their vaginal orifice blocked with adhesive tape during the caging and sampling session. In both experiments, over 90% of the PE females displayed a high degree of lordosis response to mounting by the male, and over 60% of those sexually receptive PE females exhibited an LH surge followed by ovulation. The male-induced preovulatory LH surge occurred in PE females without actual intromission. Caging with fertile males also elicited a marked increase in plasma P concentrations in PE rats and in PE females prevented from experiencing intromission.(ABSTRACT TRUNCATED AT 250 WORDS)     

Sustained but not repeated acute elevation of cortisol impaired the luteinizing hormone surge, estrus, and ovulation in gilts.

We tested the hypothesis that sustained and repeated acute elevation of cortisol would impair the LH surge, estrus, and ovulation in gilts. Cortisol was injected intramuscularly, to achieve a sustained elevation of plasma concentrations of cortisol, or intravenously, to achieve an acute elevation of plasma concentrations of cortisol. Control gilts received i.m. injections of oil and i.v. injections of saline. These treatments were administered to gilts (n = 6 per treatment) at 12-h intervals from Days 7 to 11 of the estrous cycle until after estrus ceased or until Day 27 or 28 of the estrous cycle, whichever came first. The repeated acute elevation of cortisol had no effect on the LH surge, estrus, or ovulation. In contrast, when the elevation of cortisol was sustained, the LH surge, estrus, and ovulation were inhibited. We conclude that cortisol is capable of direct actions to impair reproductive processes in female pigs but that plasma concentrations of cortisol need to be elevated for a substantial period for this to occur.