Decrease of body temperature after aglepristone treatment in bitches

Body temperature responses and the timing of abortions were evaluated in pregnant bitches with the anti-progestin aglepristone. Fifteen purebred and crossbred, 25-45 days pregnant, were included in this study and seven untreated bitches at the same stage of pregnancy served as controls. Treated bitches were administered two applications of aglepristone (10 mg/kg SC) 24 h apart for pregnancy termination. Pregnancy termination was confirmed by ultrasonographic assessment. Body temperature was rectally measured three times a day for 6 days beginning 24 h before treatment or pregnancy diagnosis in the treated and control bitches, respectively. Additionally, serum progesterone concentrations were assessed at time points during the study in the treated bitches. Pregnancy was terminated in 14 treated bitches in a mean+/-S.E.M. of 4.3+/-0.7 days after treatment. Control bitches remained pregnant. In the treated bitches, but not in the controls, body temperature significantly decreased 24 h after the beginning of the treatments (P < 0.01) and then gradually returned to pre-treatment values. Correlation between the day of mean minimum body temperature and the day of pregnancy termination was low (0.07; > 0.05). Progesterone did not show significant change throughout the study. Body temperature does not seem to be a suitable variable to clinically monitor the aborting effect of aglepristone. Decrease of body temperature after aglepristone treatment could represent further evidence of its hypothalamic effects.     

In vitro effects of estradiol-17β, monobutyl phthalate and mono-(2-ethylhexyl) phthalate on the secretion of testosterone and insulin-like peptide 3 by interstitial cells of scrotal and retained testes in dogs

The objective was to determine the effects of estradiol-17β, monobutyl phthalate (MBP) and mono-(2-ethylhexyl) phthalate (MEHP) on testosterone and insulin-like peptide 3 (INSL3) secretions in cultured testicular interstitial cells isolated (enzymatic dispersion) from scrotal and retained testes of small-breed dogs. Suspension cultures were treated with estradiol-17β (0, 10, and 100 ng/mL), MBP (0, 0.8, and 8 mmol/L) or MEHP (0, 0.2, and 0.8 mmol/L) for 18 h, in the presence or absence of 0.1 IU/mL hCG. Testosterone (both basal and hCG-induced) and INSL3 (basal) concentrations were measured in spent medium. Effects of estradiol-17β, MBP, and MEHP on testosterone and INSL3 secretions were not affected (P > 0.15) by cell source (scrotal versus retained testis); therefore, data were combined and analyzed, and outcomes reported as percentage relative to the control. In testicular interstitial cells, basal testosterone secretion was increased (P < 0.01) by 100 ng/mL estradiol-17β (130.2 ± 10.6% of control). Among phthalates, 0.2 and 0.8 mmol/L MEHP stimulated (P < 0.01) basal testosterone secretion (135.5 ± 8.3% and 154.6 ± 12.9%, respectively). However, hCG-induced testosterone secretion was inhibited (P < 0.01) by 8 mmol/L MBP (67.7 ± 6.0%), and tended to be inhibited (P = 0.056) by 0.8 mmol/L MEHP (84.5 ± 5.6%). Basal INSL3 secretion was inhibited (P < 0.01) by 8 mmol/L MBP (73.6 ± 6.8%) and 0.8 mmol/L MEHP (76.9 ± 11.3%). In conclusion, we inferred that estradiol-17β and certain phthalate monoesters had direct effects on secretions of testosterone and INSL3 in canine testicular interstitial cells, with no significant difference between scrotal and retained testes.     

Pharmacokinetics of eCG and induction of fertile estrus in bitches using eCG followed by hCG

The aim was to design a protocol combining eCG followed by hCG for estrus induction in the bitch. In Experiment 1, three ovariohysterectomized bitches received 10 000 IU of eCG iv, and 15 days later 10 000 IU of eCG im. Blood samples were taken up to 144 h after each injection to measure eCG concentrations. In Experiment 2, 25 healthy, intact late anestrous bitches were assigned to one of five doses of eCG (5, 10, 15, 20, 44, or 50 IU/kg eCG im; [TRT5-TRT50]). Sexual behavior (SB), clinical signs of estrus (CSE) and vaginal cytology (VC) samples were obtained and scored before eCG administration and every other day until onset of estrus, or for 14 days. In Experiment 3, intact late anestrous bitches were assigned to a treatment group (TRT; n = 16) and received eCG (50 IU/kg im) followed by hCG (500 IU im) 7 days later; or to a placebo group (PLA; n = 8) where they received 1 mL saline solution im. All bitches that were induced in estrus were mated or AI with fresh semen. In Experiment 1, maximum observed concentration (C_max) eCG were similar between im and iv routes (6.1 ± 0.9 vs. 8.6 ± 0.5 IU/mL, P > 0.08), whereas time for maximum observed concentration (T_max.) was longer for im compared to iv routes (17.5 ± 0.5 vs. 11.6 ± 0.3 h, P < 0.01). The area under the curve (AUC) was similar for im and iv routes (P > 0.48), and eCG was detectable in serum for at least 144 h for both routes. In Experiment 2, 3 days or 3 to 5 days after treatment, all bitches in TRT50 had higher scores compared to TRT5-44 animals (P < 0.01). In TRT50, the mean interval from treatment to estrus was 4.0 ± 0.4 days. In Experiment 3, the mean interval from treatment to estrus was shorter in the TRT group compared to the PLA group (4.1 ± 3.3 vs. 68.5 ± 4.4 days, P < 0.01). The previous interestrus interval was similar for TRT and PLA groups (199.6 ± 7.2 vs. 197.5 ± 10.2 days), but the new interestrus interval was shorter for the TRT compared to the PLA group (164.0 ± 7.2 vs. 212.2 ± 10.2 days; treatment by interval interaction, P < 0.007). Serum P₄ concentrations increased on the first day of cytologic diestrus after treatment in bitches in TRT (0.7 ± 0.3 vs. 22.8 ± 4.2 ng/mL; P < 0.01); but did not change in PLA (P > 0.84). Ninety-four percent of animals were bred (15/16; AI, n = 7; natural mating, n = 8), and 80% (12/15) became pregnant. None of the bitches had any side effects from the eCG and hCG therapy. We concluded that 50 IU/kg of eCG combined 7 days later with 500 IU of hCG was effective to induce normal and fertile estrus in bitches at 164 days post estrus, with an 80% pregnancy rate, with no side effects, and with a reduction of 48 days of the interestrus interval.     

In vitro PGF2alpha production by endometrium and corpus luteum explants from pregnant and nonpregnant diestrus bitches and placental explants from pregnant bitches

To better understand the process of slow luteal regression of the nonpregnant cycle in dogs and the acute luteolysis that occurs prepartum, the present study investigated in vitro PGF2alpha production by the endometrium, corpus luteum and placental explants obtained at known times of the cycle from pregnant bitches (days 63, 64 and immediately postpartum; day 0 = estimated day of the ovulatory LH surge) and from nonpregnant diestrus bitches (approximately days 65, 75 and 85). Both basal PGF2alpha production and its production in the presence of the protein kinase C (PKC) stimulator 12,13-phorbol dibutyrate (PDBu) were determined. For PDBu-supplemented incubations, mean PGF2alpha production (pg/mL/mg/6 h) by endometrium explants of the nonpregnant bitches in late diestrus was highest on day 65 (205 +/- 87) and reduced to low levels (38 +/- 17 and 11 +/- 11) on days 75 and 85, respectively. The production by corpus luteum explants from these bitches was significantly less on day 65 (46 +/- 14) than that of the day 65 endometrium explants, and was slightly increased on day 85 (103 +/- 52). The corresponding mean PGF2alpha production by the endometrium explants of pregnant bitches was on average much greater (i.e., two to three-fold) compared to nonpregnant bitches (P < 0.01) and involved high concentrations at day 64 (1523 +/- 467) and postpartum, compared to somewhat lower levels on day 63 (830+/-65); luteal PGF production (165 +/- 4) was also higher than in nonpregnant bitches around day 65. For pregnant bitches, PGF production per gram of tissue in the endometrium explants was greater than for the CL or placenta explants (180 +/- 37). Therefore, the endometrium of the pregnant bitch has an increased capability to produce PGF2alpha immediately prepartum, which on a tissue weight basis, exceeds that of either corpora lutea or the placenta. However, assuming a larger mass of placental tissue in vivo, we inferred that the placenta may contribute substantially to peripheral PGF concentrations.     

Clinical evaluation of the use of aglepristone, with or without cloprostenol, to treat cystic endometrial hyperplasia-pyometra complex in bitches

The aim of the study was to evaluate the efficacy of aglepristone (10 mg/kg on days 1, 2 and 8) for the treatment of metritis or pyometra in bitches (n = 67) either alone for cases of metritis (n = 15), or in cases of pyometra (n = 52) with (n = 32) or without (n = 20) the addition of low doses (1 microg/kg) of cloprostenol for 5 days (days 3-7). Examinations performed on day 90, in addition to days 8, 14 and 28, determined that treatments had been curative in the long term in 54/67 bitches (80.6%). Bitches in whom pyometra did not resolve, were given additional aglepristone on day 14 (n = 38) and day 28 (n = 20). Aglepristone alone was curative in 15/15 bitches with metritis. In 17/17 bitches with closed pyometra, cervical opening occurred within 48 h of aglepristone administration. Amongst the 52 bitches with open (n = 35) or closed (n = 17) pyometra, the additional treatment with cloprostenol from days 3 to 7, significantly improved the overall success rate at day 90, which was 27/32 (84.4%), compared to 12/20 (60.0%) in bitches without cloprostenol (P < 0.05). The leucocyte count and plasma progesterone concentrations significantly decreased over the course of treatment. Thirteen of 15 bitches in whom plasma progesterone concentrations were initially low (< 3.18 nmol/L) were cured. The recurrence rate after 12 and 24 months was 13.0% (3/23) and 19.0% (4/21), respectively.     

Luteotrophic effect of ovulation-inducing factor/nerve growth factor present in the seminal plasma of llamas

The hypothesis that ovulation-inducing factor/nerve growth factor (OIF/NGF) isolated from llama seminal plasma exerts a luteotrophic effect was tested by examining changes in circulating concentrations of LH and progesterone, and the vascular perfusion of the ovulatory follicle and developing CL. Female llamas with a growing follicle of 8 mm or greater in diameter were assigned randomly to one of three groups (n = 10 llamas per group) and given a single intramuscular dose of PBS (1 mL), GnRH (50 μg), or purified OIF/NGF (1.0 mg). Cineloops of ultrasonographic images of the ovary containing the dominant follicle were recorded in brightness and power Doppler modalities. Llamas were examined every 4 hours from the day of treatment (Day 0) until ovulation, and every other day thereafter to Day 16. Still frames were extracted from cineloops for computer-assisted analysis of the vascular area of the preovulatory follicle from treatment to ovulation and of the growing and regressing phases of subsequent CL development. Blood samples were collected for the measurement of plasma LH and progesterone concentrations. The diameter of the dominant follicle at the time of treatment did not differ among groups (P = 0.48). No ovulations were detected in the PBS group but were detected in all llamas given GnRH or OIF/NGF (0/10, 10/10, and 10/10, respectively; P < 0.0001). No difference was detected between the GnRH and OIF/NGF groups in the interval from treatment to ovulation (32.0 ± 1.9 and 30.4 ± 5.7 hours, respectively; P = 0.41) or in maximum CL diameter (13.1 ± 0.4 and 13.5 ± 0.3 mm, respectively; P = 0.44). The preovulatory follicle of llamas treated with OIF/NGF had a greater vascular area at 4 hours after treatment than that of the GnRH group (P < 0.001). Similarly, the luteal tissue of llamas treated with purified OIF/NGF had a greater vascular area than that of the GnRH group on Day 6 after treatment (P < 0.001). The preovulatory surge in plasma LH concentration began, and peaked 1 to 2 hours later in the OIF/NGF group than in the GnRH group (P < 0.05). Plasma progesterone concentration was higher on Day 6 in the OIF/NGF group than in the GnRH group (P < 0.001). Results support the hypothesis that OIF/NGF exerts a luteotrophic effect by altering the secretion pattern of LH and enhancing tissue vascularization during the periovulatory period and early stages of CL development.     

Changes in Ovaries and Uterus after Aglepristone Administration in the Third Week of Luteal Phase of Non-Pregnant Bitches

Objective

The mechanism of aglepristone action in the placentation time in the bitch remains unclear. The aim of this study was to describe the mechanism by which aglepristone influences ovaries and uterus and to measure the levels of steroid sex hormones in non-pregnant bitches.

Materials and Methods

Fourteen bitches assigned to a study (n=9) and control (n=5) group were given aglepristone and saline solution, respectively, on the 19th and 20th day after LH peak. On the 26th day after LH peak an ovariohysterectomy was performed. Blood samples were screened for estradiol and progesterone concentrations. Ovaries and uterine horns and bodies were isolated for histological and morphometrical diagnosis and immunohistochemistry analysis of α-estrogen and progesterone receptor expression.

Results

A decrease of progesterone (p<0.01) and no differences in total estrogen level in the study group were observed. There were no significant differences either in the histomorphometry or α-estrogen and progesterone receptors expression in ovaries. Increase in expression of progesterone receptors in endometrium without surface epithelium of horns (p<0.05), endometrial surface epithelium (p<0.05), myometrium of uterine body (p<0.01) and estrogen receptors in endometrium without surface epithelium of horns (p<0.05) was observed. Elevated estrogen receptors probably increased sensitivity of tissues to estrogens in the bloodstream and led to notable inflammation, haemorrhages, and hyperplasia in endometrium with mononuclear immune cell infiltration. The myometrium of horns and endometrium of uterine body of study bitches were significantly thicker than in the control group (p<0.05 and p<0.01). Furthermore myometrium of uterine body was thicker than myometrium of horns (p<0.001) and expression of progesterone receptors was higher in uterine body (p<0.01). No differences were observed between endometrium of horns and body within groups.

Conclusion

To the knowledge of the authors this is the first study, which describes the inflammatory effect developing in uterus in response to aglepristone administration, and attempts to elucidate its mechanisms.     

A retrospective study of bitches with pyometra,medically treated with aglepristone

Pyometra is a common and life-threatening disease of intact bitches caused by hormonal influence of the uterus in combination with bacterial infection. The treatment of choice is ovariohysterectomy, but several purely medical options are available. Common drugs used for medical treatment in combination with antimicrobials are progesterone receptor blockers, prostaglandins, and dopamine agonists. The aim of this study was to evaluate long-term recovery and fertility after treatment with the progesterone receptor blocker aglepristone in bitches with pyometra. Data from 28 bitches with pyometra, admitted to the University Animal Hospital, Swedish University of Agricultural Sciences, during a 9-year period, were studied retrospectively and followed up by telephone interviews with the owners. The bitches had been treated with aglepristone at a dose of 10 mg/kg on a median of four occasions. All bitches had also been treated with antimicrobial drugs for a mean duration of 23 days, the most frequently used being enrofloxacin. Escherichia coli were the most commonly isolated bacteria from cranial vagina. The outcome was assessed for up to 6 years after treatment. The success rate, determined as restored clinically healthy status, was 75% (21/28 bitches), and the recurrence rate of disease was 48% (10/21 bitches). The mean time until recurrence was 10.5 months after the end of treatment. After treatment, 69% (9/13) of the mated bitches produced puppies. Of the seven bitches that did not have a successful treatment, six were ovariohysterectomized and one was euthanized. In conclusion, medical treatment with aglepristone in combination with antimicrobial therapy was successful in 75% of the bitches studied and the recurrence rate was 48%.     

Prevention of pregnancy in cats using aglepristone on days 5 and 6 after mating

The aim of this study was to test for the efficacy of aglepristone treatment for prevention of early pregnancy in the cat. Eleven cats (Gr. 1) were treated with 10 mg/kg aglepristone on days 5 and 6 after mating, 17 cats (Gr. 2) were used as untreated controls. Blood samples for progesterone (P4) determination were collected from 6 cats of Gr. 1 and 9 cats of Gr. 2, respectively. Ultrasound examination on day 25 revealed no pregnancy in any of the treated cats. In both groups, P4 concentrations increased from day 5 (before treatment) to day 20 (P < 0.01). In Gr. 1, the interval between aglepristone treatment and the subsequent estrus ranged from 5-299 d [18.5 (5.2) d]. Mean interestrus interval was 205 ± 72 d in Gr. 2. Mean duration of subsequent estrus was not different to duration of estrus before treatment in Gr. 1 and in Gr. 2, respectively. Mean time between treatment and next pregnancy was 56.4 (4.7) d, ranging from 5-325 d. Pregnancy rates after the next estrus following treatment were 64 and 82% after the first and second estrus, respectively. No major treatment-related side effects were observed. In conclusion, treatment was found to be highly effective for prevention of early pregnancy.     

Dynamics of follicular growth and progesterone concentrations in cyclic and anestrous suckling Nelore cows (Bos indicus) treated with progesterone,equine chorionic gonadotropin,or temporary calf removal

The objective of this study was to investigate the effects of eCG and temporary calf removal (TCR) associated with progesterone (P4) treatment on the dynamics of follicular growth, CL size, and P4 concentrations in cyclic (n = 36) and anestrous (n = 30) Nelore cows. Cyclic (C) and anestrous (A) cows were divided into three groups. The control group received 2 mg of estradiol benzoate via intramuscular (IM) injection and an intravaginal device containing 1.9 g of P4 on Day 0. On Day 8, the device was removed, and the animals received 12.5 mg of dinoprost tromethamine IM. After 24 hours, the animals received 1 mg of estradiol benzoate IM. In the eCG group, cows received the same treatment described for the control group but also received 400 UI of eCG at the time of device removal. In the TCR group, calves were separated from the cows for 56 hours after device removal. Ultrasound exams were performed every 24 hours after device removal until the time of ovulation and 12 days after ovulation to measure the size of the CL. On the same day as the CL measurement, blood was collected to determine the plasma P4 level. Statistical analyses were performed with a significance level of P ≤ 0.05. In cyclic cows, the presence of the CL at the beginning of protocol resulted in a smaller follicle diameter at the time of device removal (7.4 ± 0.3 mm in cows with CL vs. 8.9 ± 0.4 mm in cows without CL; P = 0.03). All cows ovulated within 72 hours after device removal. Anestrous cows treated with eCG or TCR showed follicle diameter at fixed-timed artificial insemination (A-eCG 10.2 ± 0.3 and A-TCR 10.3 ± 0.5 mm) and follicular growth rate (A-eCG 1.5 ± 0.2 and A-TCR 1.3 ± 0.1 mm/day) similar to cyclic cows (C-eCG 11.0 ± 0.6 and C-TCR 12.0 ± 0.5 mm) and (C-eCG 1.4 ± 0.2 and C-TCR 1.6 ± 0.2 mm/day, respectively; P ≤ 0.05). Despite the similarities in CL size, the average P4 concentration was higher in the A-TCR (9.6 ± 1.4 ng/mL) than in the A-control (4.0 ± 1.0 ng/mL) and C-TCR (4.4 ± 1.0 ng/mL) groups (P < 0.05). From these results, we conclude that eCG treatment and TCR improved the fertility of anestrous cows by providing follicular growth rates and size of dominant follicles similar to cyclic cows. Additionally, TCR increases the plasma concentrations of P4 in anestrous cows.     

Effects of systemic and intrafollicular injections of LH,prostaglandins, and indomethacin on the luteinization of rabbit Graafian follicles

The possibility that initiation of luteinization in ovarian follicles by luteinizing hormone (LH) is mediated by prostaglandins (PG's) was investigated in rabbits. Estrous rabbits, given an ovulatory dose of LH (50 μg) intravenously, were administered indomethacin (IM), an inhibitor of PG biosynthesis, by various routes. Progesterone levels in the serum and in the induced corpora lutea (CL) were subsequently measured by radioimmunoassay. Continued daily subcutaneous injections of IM from 2 days before through 2 days after LH treatment reduced the corpus luteal level, measured at 72 hours post-LH, of PGF from 208 ± 43 to 98 ± 20 pg/CL (P < 0.025) and that of PGE from 272 ± 31 to 115 ± 9 pg/CL (P < 0.005). At the same time, progesterone levels were 72 ± 12 and 93 ± 10 ng/CL (P > 0.05) in the oil-treated and IM-treated rabbits, respectively. Serum progesterone continued to rise in a linear fashion during the period from 24 to 72 hours following LH treatment, whether IM was injected or not. Intrafollicular treatment with LH (100 ng/follicle) raised the progesterone content in the treated follicles 72 hours later from 1.1 ± 0.5 to 50.1 ± 13.5 ng. (P < 0.01). This progesterone content reached 21.5 ± 15.8 ng (P < 0.05) in follicles similarly treated with PGE₂ (5 μg/follicle), but remained meagre at lower doses of PGE₂ (100 ng/follicle and 2 ng/follicle). Serum progesterone increased from 0.5 ± 0.1 to 1.2 ± 0.1 ng/ml (P < 0.005) within 72 hours in rabbits treated intrafollicularly with LH, but remained unaltered in those similarly treated with PGE₂ (P > 0.1). Intrafollicular injections with PGF_2α failed to induce changes in either level of progesterone. It is concluded that prostaglandins probably do not mediate the luteinizing action of LH in rabbit Graafian follicles, although some degree of luteinization can be induced by high levels of exogenous PGE₂.     

Variables associated with peripartum traits in dairy cows. V. Hormonal profiles associated with retained fetal membranes

Profiles of certain hormones measured by radioimmunoassay in 41 Holstein cows and heifers with retained fetal membranes (RFM; >12 hr postpartum) were compared to 41 peers without RFM (NRFM). Peers were matched by parity, season of calving, gestation length, dystocia and parturient paresis within prepartum diet group. Linear covariates of natural photoperiod, mean daily temperature, calf birth weight, length of gestation, and age and body weight of cow were included in the leastsquares analyses of data. Plasma profiles of prolactin and estrone were nearly identical from day 8 prepartum to day 2.5 postpartum. Plasma estradiol-17α was approximately one-third higher each day in group RFM (P<.05 across days) but estradiol-17β (Eβ) tended to be lower until day 2 prepartum (not significant). Also, plasma progesterone (P) was higher in group RFM between days 8 to 3 prepartum (p<.05 across days).Relationships between plasma P and Eβ were indicative of subsequent RFM (>24 hr rather than >12 hr), but only on day 6 prepartum. Three of four cases of 12 to 24 hr RFM had P and Eß profiles similar to NRFM. Either a combination of low P (<3.0 ng/ml) and low Eß (<100 pg/ml) or only high P (>7.9 ng/ml) were associated with a ten-fold higher rate of RFM (>24 hr) than when P was intermediate (4 to 8 ng/ml) and Eß exceeded 99 pg/ml. These results are in agreement with prior data wherein RFM were induced at premature births either by ovariectomy during pregnancy or by glucocorticoids.     

Endocrine profiles during doe-litter separation and the subsequent pregnancy in rabbits

This study was carried out to determine the effects of a transient doe-litter separation on plasma prolactin, LH, FSH, estradiol-17β and progesterone concentrations before artificial insemination and during the subsequent pregnancy. Control does (n=12) had free access to nursing, whereas separated does (n=12) were kept away from their litters for 48 hours before artificial insemination. Both groups were inseminated on day 11 after parturition. Teat stimulation by suckling caused a high increase in prolactin concentrations in separated does (p<0.0001). Basal prolactin concentrations were observed in both groups on days 8 and 18 of pregnancy. No effect of the treatment was detected on LH and FSH concentrations during the sampling period. A rise of estradiol-17β concentrations was observed 48 hours after doe-litter separation, compared to control does and to previous values (p<0.003). Both groups showed low progesterone concentrations before artificial insemination. Pregnant rabbits in both groups showed increased progesterone concentrations on days 8 and 18 of pregnancy. Lower estradiol-17β concentrations were observed in control does on day 18 of pregnancy compared with separated rabbits (p<0.003). The results suggest that a transient separation of nursing does from their litters before artificial insemination may promote high follicular steroidogenesis activity leading to increased estradiol-l7β concentrations. This hormonal change could be a result of several stimulatory actions probably triggered by the absence of suckling episodes and may affect the luteotrophic function during the subsequent pregnancy.     

Differential regulation of estrous behavior and luteinizing hormone secretion by estradiol-17β in ovariectomized dairy cows

Holstein cows (n = 9) were used in an experiment to characterize the behavioral and endocrine responses to estradiol-17β when administered at rates designed to maintain peripheral concentrations within a physiological range. Cows were pretreated with progesterone for 3 d. Three days after progesterone treatment was completed, each cow was assigned to one of five estradiol-17β treatment groups (Doses 0 to 4), calculated to produce and maintain 0, 3, 6, 9, or 12 pg/mL in peripheral blood for 8 h. The experiment was conducted in eight replicates (with 3 to 7 cows each), with no dose repeated in any replicate. In each replicate, at least one additional cow was given an injection of estradiol-17β (500 μg im, in a corn oil vehicle) to facilitate estrus detection. Estrus was detected by visual observation for 30 min at 4 h intervals. Estrus was defined as a cow that stood to be mounted at least twice during the 50 h interval over which estrus was observed. Jugular venous blood samples were collected at 2 h intervals throughout the infusion and observation periods for quantification of luteinizing hormone (LH). Cows that received the highest dose (Dose 4, n = 7) all showed estrus, whereas those that received the two lowest doses (Dose 0, n = 5; Dose 1, n = 6) did not. Over the course of the experiment, five cows received each dose at least once. Of these, three showed estrus at Doses 2, 3, and 4, whereas the other two showed estrus only at Dose 4. Therefore, individual cows differed in the amount of estradiol-17β needed to induce estrus. There was a linear effect of dose on duration of estrus (P < 0.01). Estrus was shorter for Dose 2 (8.0 h) than for Dose 4 (18.4 h). The onset of estrus (after start of infusion) tended to be later for Dose 2 (20.0 h) than for Doses 3 and 4 (14.0 and 13.4 h, respectively; P = 0.15). Preovulatory-like surges of LH were induced in all cows at Doses 2, 3, and 4. Surges also were detected in 3 of 5 cows receiving Dose 1. The magnitude of the LH surge was less for Doses 1, 2, and 3 than for Dose 4 (P = 0.06). In contrast to the timing of estrus, the timing of the LH surge (after start of infusion) was not different among doses (P = 0.88). Thus, the hypothalamic centers responsible for regulating expression of estrus and secretion of LH responded differently to estradiol-17β.     

Temporal relations between plasma concentrations of luteinizing hormone, follicle-stimulating hormone, estradiol-17beta, progesterone, prolactin, and alpha-melanocyte-stimulating hormone during the follicular, ovulatory, and early luteal phase in the bitch

Compared with other domestic animals, relatively little is known about the changes in, and temporal relations between, reproductive hormones around the time of ovulation in the domestic bitch. Therefore, plasma concentrations of luteinizing hormone (LH), follicle-stimulating hormone (FSH), estradiol-17beta, progesterone, prolactin (PRL), and alpha-melanocyte-stimulating hormone (alpha-MSH) were determined one to six times daily from the start of the follicular phase until 5 days after the estimated day of ovulation in six Beagle bitches. In all bitches, the pre-ovulatory LH surge was accompanied by a pre-ovulatory FSH surge. A pre-ovulatory PRL or alpha-MSH surge was not observed. The pre-ovulatory FSH and LH surges started concomitantly in four bitches, but in two bitches the FSH surge started 12 h earlier than the LH surge. The FSH surge (110+/-8 h) lasted significantly longer than the LH surge (36+/-5 h). In contrast with the pre-ovulatory FSH surge, the pre-ovulatory LH surge was bifurcated in four of six bitches. The mean plasma LH concentrations before (1.9+/-0.4 microg/L) and after (1.9+/-0.3 microg/L) the LH surge were similar, but the mean plasma FSH concentration before the FSH surge (1.6+/-0.3 U/L) was significantly lower than that after the FSH surge (3.1+/-0.2 U/L). In most bitches the highest plasma estradiol-17beta concentration coincided with or followed the start of the pre-ovulatory LH surge. In five of the six bitches the plasma progesterone concentration started to rise just before or concurrently with the start of the LH surge. In conclusion, the results of this study provide evidence for the differential regulation of the secretion of LH and FSH in the bitch. In addition, the interrelationship of the plasma profiles of estradiol-17beta and LH suggests a positive feedback effect of estradiol-17beta on LH surge release. The start of the pre-ovulatory LH surge is associated with an increase in the plasma progesterone concentration in this species.     

The roles of PGF2α and PGE2 in regression of the corpus luteum after intrauterine infusion of Arcanobacterium pyogenes in cows

To study the effect of bacteria in the uterus on the fate of the corpus luteum (CL), Arcanobacterium pyogenes was inoculated into the uteri of cows on Day 3 (Day 0 = day of spontaneous ovulation). Plasma concentrations of 13,14-dihydro-15-keto-PGF_2α (PGFM), 13,14-dihydro-15-keto-PGE₂ (PGEM) and progesterone (P₄) were determined. In five cows, the developing CL regressed and first-wave dominant follicles, which normally become atretic, ovulated (Group OV) after bacterial inoculation. In another five cows (Group NOV) and five control cows, the developing CL did not regress and first-wave dominant follicles did not ovulate. In Group OV, PGFM concentrations increased by 126.2 pg/mL (from 36.8 ± 7.8 pg/mL on Day 3 to 163 ± 37.2 pg/mL on Day 6), with an increase ratio of 5.8-fold. Conversely, in Group NOV, PGFM had a greater increase of 198.4 pg/mL (from 128.2 ± 27.8 pg/mL on Day 3 to 326.6 ± 115.1 pg/mL on Day 5), but the increase ratio was only 2.3-fold. Although PGEM tended to increase in both groups, raw increases and increase ratios were small. Bacterial inoculation into the uterus stimulated the release of prostaglandins and affected the fate of the CL; in that regard, the CL was affected more by PGF_2α than by PGE₂, and the increase ratio of PGF_2α was more important than the raw increase.     

Diurnal variation in LH and temporal relationships between oscillations in LH and progesterone during the luteal phase in heifers

Diurnal variation in progesterone and LH during the luteal phase and the temporal relationships between oscillations of the two hormones were studied in 10 heifers by collection of blood samples at 0100, 0700, 1300, and 1900 h each day, beginning on Day 1 (Day 0 = ovulation). Concentration of LH on Days 5-9, but not on Days 10-14, was lower (P < 0.05) at 0700 h (0.25 ± 0.02 ng/mL) than at each of the other three hours (combined, 0.32 ± 0.02 ng/mL). An oscillation was defined as an uninterrupted increase and decrease in concentrations. The number of LH oscillations/heifer with the peak at 1900 h (6.1 ± 0.7) throughout the luteal phase was greater (P < 0.01) than for each of the other hours (combined, 4.0 ± 0.2). Diurnal variation in progesterone was not detected. Only statistically defined LH oscillations were used to determine the temporal association between the peak of an LH oscillation and various components of a progesterone oscillation. On Days 5-14, the frequency of the peak of an LH oscillation occurring at the same hour as the peak of a progesterone oscillation (26/48, 54%) was greater (P < 0.0001) than at the progesterone nadir (3/48, 6%). The frequency of the LH peak occurring during increasing (11/34, 32%) and decreasing (8/25, 32%) progesterone concentrations was intermediate (P < 0.05). Results indicated the following: 1) diurnal variation occurred in LH as determined by concentration and by the hour of the peak of an oscillation; and 2) LH oscillations were temporally and positively related to progesterone oscillations.     

Repeated induction of abortion in bitches and the effect on plasma concentrations of relaxin, progesterone and estradiol-17beta

The aim of the present study was to investigate the effects of two medications on two subsequent abortions and plasma hormone concentrations of dogs. For this purpose, two groups of bitches (n=5 each), received the antiprogesterone aglepristone (Alizine) at 10mg/kg body weight on two subsequent days around day 30 after mating. In group II, the antiprolactin cabergoline (Galastop) was additionally administered po at 5 microg/kg body weight until the start of abortion. The plasma concentrations of relaxin, progesterone (P4) and estradiol-17beta (E2) were measured before, during and after each abortion. During the next cycle after the abortion, the same bitches were mated again and in pregnant animals, induction of abortion was performed as before. During the third cycle, pregnant bitches were allowed to whelp. Termination of first pregnancy occurred significantly earlier after the combined treatment (6.8 versus 10.6 days, p<0.05). In both groups and during both abortions, relaxin varied between individuals; however, there was a continuous decrease after the abortions and no significant differences between groups (p>0.05). In one bitch with high relaxin concentrations before treatment (11.6 ng/ml), a cystic endometrial hyperplasia was diagnosed. In the aglepristone only group, P4 concentrations increased significantly after the first application (p<0.05), then decreased continuously until day 45 after the beginning of abortion. In the combined group, there was a continuous decrease until day 45 (p>0.05). At this time, P4 concentrations between 0.47 and 84.9 nmol/l were measured in both groups. The level of E2 over time was not influenced by any medication. We therefore note that the two medications mainly influenced plasma concentrations of P4 in different ways, probably due to specific treatment-hormone interactions. However, all measurements fell within the range considered normal.     

The pituitary-ovarian axis in dogs with remnant ovarian tissue

It can be difficult to confirm the presence of remnant ovarian tissue (ROT) in bitches that are presumed to be ovariohysterectomised. A GnRH stimulation test can be used to distinguish ovariectomised bitches from those in anoestrus, but it is uncertain whether the GnRH-induced changes in plasma LH and oestradiol concentrations that occur in intact bitches also occur in ROT-bitches. We report here eighteen ROT-bitches and compare the results of GnRH stimulation tests with those of six ovariectomised and six bitches in anoestrus.The basal (n = 17) and/or GnRH-stimulated (n = 18) plasma oestradiol concentration was above the detection limit of the assay, i.e., < 7 pmol/l, in all ROT-bitches but below the detection limit in all ovariectomised bitches. Basal plasma LH concentration was significantly higher in ROT-bitches (4.1 ± 0.7 μg/L) than those in anoestrus (0.64 ± 0.04 μg/L), and significantly lower than in ovariectomised bitches (20.2 ± 3.6 μg/L). Basal plasma LH concentration was relatively high in bitches in which there was a long interval between ovariectomy and appearance of oestrus. GnRH administration resulted in a significant increase in plasma LH and oestradiol concentrations in ROT-bitches. The GnRH-induced increase and subsequent decline in plasma LH concentration were significantly less in ROT-bitches than in either ovariectomised bitches or those in anoestrus. The GnRH-induced increase in plasma oestradiol concentration was significantly smaller in ROT-bitches than in those in anoestrus.In conclusion, the results of this study demonstrate that in dogs ROT is associated with noticeable changes in the pituitary-ovarian axis and suggest that a GnRH stimulation test may be used to distinguish between completely ovariectomised bitches and those with ROT.     

Steroidogenic capabilities of the early mouse embryo

Preimplantation embryos from ICR albino mice were used to determine progesterone and estradiol-17β production during incubation in BMOC-2. Following culture of 40 embryos/culture at either the morula, early blastocyst or late blastocyst stages, progesterone and estradiol-17β contents were 192±27 and 82±22 pg, 289±50 and 147±46 pg and 157±28 and 88±23 pg, respectively, for incubated samples and 306±68 and 89±40 pg, 404±63 and 125±44 pg, and 241±54 and 86±39 pg, respectively for control samples. Although, there were significant stage of development and treatments effects (P<0.05) for progesterone, production of this steroid was not evident. These data suggest that the early preimplantation mouse embryo does not produce progesterone or estradiol-17β in a defined culture system.