Plasma Progesterone Concentrations Research Articles

61 Low plasma progestin concentration during the early luteal phase impairs equine conceptus development until placentation

During the early luteal phase, low progesterone concentrations delay downregulation of endometrial progesterone receptors. This contributes to impaired histotroph production at Day 14 of pregnancy (Beyer et al. 2019 Theriogenology 125, 236-241). Until the beginning of placentation (i.e. Day 37 of pregnancy), nutritional supply of the equine conceptus depends on histotrophy alone. The aim of the present study was to analyse development of the equine conceptus under reduced plasma progesterone concentrations until shortly before placentation. Fertile Haflinger mares (n=11; 4-11 years old) were examined daily by transrectal ultrasonography, and when in oestrus, they were inseminated every 48h until spontaneous ovulation. Mares were randomly assigned to either the treatment group and received the prostaglandin F2α agonist cloprostenol (PGF2α; 125μg) once daily for 3 days after ovulation (Beyer et al. 2019) or the control group and left untreated. After conceptus collection on Day 34, mares were allowed one oestrous cycle for recovery and subsequently assigned to the opposite treatment, thus serving as their own controls. From Day 10 after ovulation, conceptus development including size, uterine fixation, and heartbeat detection was evaluated daily by transrectal ultrasonography. On Day 34, conceptus and fetal membranes were recovered transcervically. The recovered material was weighed, measured, and stored for further analysis. Conceptuses underwent microcomputed tomography that was evaluated by Amira (Thermo Fisher Scientific). Statistical comparison for differences between control and treatment pregnancies was performed by nonparametric Wilcoxon test or chi-square analysis. The day of first visualisation of the conceptus and of the embryo proper and heartbeat did not differ between treatments. Uterine fixation occurred on Day 19.3±0.5 in conceptuses from PGF2α-treated pregnancies but on Day 16.7±0.4 in controls (P<0.05). At 34 days of gestation, the conceptuses from PGF2α-treated mares were smaller (P<0.05) than control conceptuses when measured with ultrasound callipers in utero (maximal length: PGF2α 17.4±0.3mm, control 19.0±0.4mm). Conceptus weight determined after recovery was less in treated pregnancies (PGF2α 2.5±0.2 g; control 3.5±0.3 g; P<0.05). Microcomputed tomography analysis of selected inner organs showed some differences in development. Seven complete pairs of undestroyed conceptuses were available. Mean numbers of 33.4±3.2 and 18.8±7.5 bronchi were detected in control and PGF2α conceptuses, respectively (P<0.05). Heart volume did not differ, but communication between ventricles was detected in only 1/7 control but 5/7 PGF2α conceptuses (P<0.05). The footpad was present in 6/7 control and 0/7 PGF2α conceptuses (P<0.001). In conclusion, subphysiological progesterone concentration during the early luteal phase delays development of the equine conceptus before placentation. The condition may contribute to early conceptus loss in horses, which occurs in up to 20% of pregnancies.

Prostaglandin F<sub>2α</sub> induced luteolysis is delayed but not prevented by acute administration of luteotropic drugs in lactating nonpregnant Holstein cows

Inadvertent use of prostaglandin F2α (PGF2α) in pregnant cattle could result in luteolysis and pregnancy loss. Using a nonpregnant cow model, we determined if luteotropic agents could counteract luteolytic effects of PGF2α. Ovarian status of 20 lactating nonpregnant Holstein cows was synchronized using an Ovsynch protocol and ovulation confirmed by transrectal ultrasonography 48 hours after the second gonadotropin releasing hormone (GnRH). Eight days after ovulation, 25 mg of dinoprost (native PGF2α) was administered IM to induce luteolysis. Five minutes after PGF2α, cows were treated IM with gonadorelin (GnRH, 100 μg; n = 5), human chorionic gonadotropin (hCG, 1,000 IU; n = 6), porcine pituitary luteinizing hormone (pLH, 25 mg; n = 5), or saline (control, 2 ml; n = 4). Blood samples were collected before PGF2α (0 hour) and at 1, 6, 12, 18, 24, 30, 36, 42, 48, 60, 72, and 84 hours after PGF2α to monitor subsequent luteal activity by measuring plasma progesterone concentrations. Although none of the treatments counteracted the luteolytic action of PGF2α, the rate of decline in plasma progesterone within 1 hour after PGF2α was greater (p = 0.04) in control than in GnRH, hCG, and pLH (2.2 versus 0.2, 0.3, and 0.1 ng/ml/hour, respectively). In addition, pLH-treated cows tended (p = 0.07) to have greater overall mean plasma progesterone for up to 84 hours (1.6 ± 0.2 ng/ml) than other treatments combined (1.1 ± 0.1 ng/ml). In the present study, giving GnRH, hCG or pLH 5 minutes after PGF2α administration was not effective in countering luteolytic effects of PGF2α. Nevertheless, pLH treatment tended to delay PGF2α induced luteolysis, which warrants further investigation.

A predictive model to diagnose pregnancy in guanacos (Lama guanicoe) using non-invasive methods

Pregnancy status is usually not included in ecological studies because it is difficult to evaluate. The use of non-invasive methods to determine pregnancy, without physically restraining individuals, would enable pregnancy to be included in population studies. In this study, we evaluated sex steroid hormones in plasma and fecal samples from pregnant and non-pregnant females to develop a pregnancy predictive model for guanacos (Lama guanicoe (Müller, 1776)). Samples were obtained during live-shearing management (i.e., capture, shear, and release) of guanacos. Enzyme immunoassays were used to evaluate progesterone (P4) and estradiol (E2) concentrations in plasma and pregnanediol glucuronides (PdG) and conjugated estrogens (EC) in feces. Mean hormonal and fecal metabolite concentrations were significantly higher in pregnant females than in non-pregnant females. A linear relationship was found between each hormone and its fecal metabolite. Finally, hormonal data were combined with an independent source of pregnancy diagnosis such as abdominal ballottement to develop a logistic regression model to diagnose pregnancy in non-handled individuals. The use of predictive models and non-invasive methods might be suitable to incorporate pregnancy information in large-scale population studies on guanaco and other free-ranging ungulates.

Annual changes in plasma progesterone and estradiol-17β concentrations compared to pituitary-adrenal axis activity in the female goat reared under arid environment

ABSTRACT This study investigates the relationships between the pituitary-adrenal axis and the ovarian activity in the cyclic goat reared under arid environment. Plasma hormone levels are assessed on blood samples collected weekly at 9:00 a.m., during one year. The ovarian activity is characterized by an anestrous period from mid-March to mid-July and a breeding period during the rest of the year. Adrenal histomorphometry and immunolocalization of Estrogen Receptors α (ERα) and Progesterone Receptors (PR) were performed on adrenal glands collected during each season. During the anestrous period, all adrenal compartments appeared enlarged, as did cellular and nuclear areas. ERα and PR immunostaining were identified in all the adrenal gland zonae with more intensity for the ERα. ERα immunostaining is greater in zona fasciculata whereas PR immunostaining is more intense in zona fasciculata and zona reticularis during the breeding season. Plasma ACTH shows no difference between the two seasons, while cortisol rises slightly during the anestrous period. There is a negative but not significant correlation between progesterone levels with plasma ACTH and cortisol. The pituitary-adrenal axis is activated essentially at the anestrous onset and at the ovarian resumption which represents stressful events, but it varies slightly between these two seasons in this goat.

Effects of a single trace mineral injection at beginning of fixed-time AI treatment regimen on reproductive function and antioxidant response of grazing Nellore cows

Two experiments evaluated the effects of injectable trace minerals (ITM) administered 11 d before artificial insemination (AI) on body weight (BW), body condition score (BCS), ovarian structures, pregnancy rate, and antioxidant response of Nellore cows. In Experiment 1, 20 multiparous cows were assigned to one of two treatments: subcutaneous injection (6 mL/cow; 11 d before AI) of saline solution or ITM (60, 10, 5, and 15 mg/mL of Zn, Mn, Se and Cu, respectively) and BW, BCS, ovarian structures and blood were evaluated. In Experiment 2, 1,144 multiparous cows were assigned to same treatments described in Experiment 1 and pregnancy rate on d 30 was evaluated. In Experiment 1, ITM did not affect (P ≥ 0.23) BW, dominant follicle size, ovulation rate, and plasma concentrations of haptoglobin, ceruloplasmin and progesterone (P4). The ITM treatment tended to increase (P = 0.06) cow BCS and reduce (P ≤ 0.06) corpus luteum (CL) diameter and volume. Furthermore, ITM treatment tended to increase (P = 0.06) plasma concentrations of SOD and increased (P = 0.007) GSH-Px compared with saline injection. In Experiment 2, ITM treatment tended (P = 0.06) to increase pregnancy rate of cows with BCS ≤ 5.0 but not cows with BCS > 5.0 (P = 0.99). The ITM treatment did not alter BW, plasma P4, and acute phase response, but enhanced plasma concentrations of antioxidant enzymes, and tended to enhance BCS and pregnancy rates to AI of cows with BCS ≤ 5.0, even though there was a smaller corpus luteum size.

Flavonoid Intake and Plasma Sex Steroid Hormones, Prolactin, and Sex Hormone-Binding Globulin in Premenopausal Women.

Background: Flavonoids potentially exert anti-cancer effects, as suggested by their chemical structures and supported by animal studies. In observational studies, however, the association between flavonoids and breast cancer, and potential underlying mechanisms, remain unclear. Objective: To examine the relationship between flavonoid intake and sex hormone levels using timed blood samples in follicular and luteal phases in the Nurses’ Health Study II among premenopausal women. Methods: Plasma concentrations of estrogens, androgens, progesterone, dehydroepiandrosterone (DHEA), DHEA sulfate (DHEAS), prolactin, and sex hormone-binding globulin (SHBG) were measured in samples collected between 1996 and 1999. Average flavonoid were calculated from semiquantitative food frequency questionnaires collected in 1995 and 1999. We used generalized linear models to calculate geometric mean hormone concentrations across categories of the intake of flavonoids and the subclasses. Results: Total flavonoid intake generally was not associated with the hormones of interest. The only significant association was with DHEAS (p-trend = 0.02), which was 11.1% (95% confidence interval (CI): −18.6%, −3.0%) lower comparing the highest vs. lowest quartile of flavonoid intake. In subclass analyses, the highest (vs. lowest) quartile of flavan-3-ol intake was associated with significantly lower DHEAS concentrations (−11.3% with 95% CI: −18.3%, −3.7%, p-trend = 0.01), and anthocyanin intake was associated with a significant inverse trend for DHEA (−18.0% with 95% CI: −27.9%, −6.7%, p-trend = 0.003). Conclusion: Flavonoid intake in this population had limited impact on most plasma sex hormones in premenopausal women. Anthocyanins and flavan-3-ols were associated with lower levels of DHEA and DHEAS.

Relationships between size, steroidogenesis and miRNA expression of the bovine corpus luteum

In a previous study, a subset of miRNAs were identified the expression of which increases substantially during the follicle-luteal transition in cattle. Here, we investigated the functional involvement of some of these miRNAs (miR-96, miR-182, miR-132, miR-21, miR-378) by determining whether there is an association in vivo between their expression in the corpus luteum (CL), CL size and progesterone production. The two largest and two smallest CL were collected from 12 donor beef heifers on Day 7 following ovarian super-stimulation (Day 0 = 28–32 h after first standing to be mounted). Additionally, the CL and a plasma sample were collected from 29 recipient heifers on Day 15. Luteal expression of miRNAs and mRNAs, and plasma progesterone concentrations were quantified by RT-qPCR and RIA, respectively. There were no differences in the mean expression of any miRNAs examined or the steroidogenic enzymes, STAR or CYP11A1, between the largest and smallest CL in donor heifers (P > 0.1). In addition, there were no significant correlations of luteal volume or weight with any miRNA, CYP11A1 or STAR in donor heifers. However, a correlation (r ≥ 0.5, P ≤ 0.001) existed between the transcript levels of CYP11A1 and STAR in the CL, as well as between each of those and miR-182 levels. In addition, CYP11A1 abundance was moderately correlated (r ≤ 0.4, P < 0.05) with each of miR-96 and miR-378. In recipient heifers, progesterone levels were moderately correlated with luteal weight (r = 0.41, P = 0.03) but not with the expression of any miRNA, CYP11A1 or STAR (P > 0.1). Moreover, luteal CYP11A1 and STAR were correlated (r = 0.6, P ≤ 0.001) with miR-182 as well as with each other, consistent with data in donor heifers. Finally, both CYP11A1 and STAR were moderately correlated (r ≤ 0.5) with miR-132 and, in the case of STAR, with miR-378. In summary, there was no association between either luteal weight/volume or plasma progesterone concentrations and any of the miRNAs analysed in donor and recipient heifers. However, CYP11A1 and STAR transcript levels were significantly correlated with several miRNAs, most notably miR-182, as well as with each other, in luteal tissues from both donor and recipient heifers. This finding confirms results of previous in vitro studies and, importantly, provides the first in vivo evidence of a role of the miR-183-96-182 cluster in regulating luteal steroidogenesis.

Comparison of progesterone concentrations in blubber and plasma among female Antarctic minke whales of known reproductive status

The utility of progesterone concentration in blubber as a means of determining reproductive status in the Antarctic minke whale Balaenoptera bonaerensis was assessed through a comparative analysis of progesterone concentration in blubber and plasma among 230 female whales of known reproductive status (immature, resting, ovulating or pregnant). Whales were sampled during the austral summer in the Antarctic Ocean. The general pattern of progesterone concentration by reproductive category was well correlated between blubber and plasma samples, validating in principle the use of progesterone concentrations in blubber to determine the reproductive status of females. However, some differences were found for resting and ovulating females, which require further consideration. For blubber, overlap of progesterone concentrations was observed between reproductive categories with the exception of immature/ovulating and immature/pregnant. This result suggests that the method of using progesterone concentration in blubber cannot distinguish between pregnant and non-pregnant mature females. However, it can be used to distinguish between immature and mature females. Although a low overlap ratio in concentration was found between immature and resting females, the method is still useful for determining sexual maturity, because resting females of the Antarctic minke whale are seldom found in the Antarctic Ocean.

Ultrasonographic and hormonal characterization of reproductive health and disease in wild, semiwild, and aquarium-housed southern stingrays (Hypanus americanus).

To characterize physical examination, plasma biochemical, and ultrasonographic findings in aquarium-housed, managed semiwild, and wild southern stingrays (Hypanus americanus) with and without reproductive disease. Southern stingrays from aquarium (n = 48), lagoon (managed semiwild; 34), and wild (12) habitats. Limited, opportunistic prosections were performed of presumed anatomically normal wild southern stingrays and compared with findings for aquarium-housed stingrays with reproductive disease. Ultrasonographic video data from both groups were used to assign a score (1 to 5) indicating increasing severity of ovarian and uterine reproductive disease. Plasma total 17β-estradiol, estrone, progesterone, and testosterone concentrations were measured with enzyme immunoassays validated for use in southern stingrays. Ultrasonographic ovarian scores were significantly correlated with uterine scores. No reproductive disease was detected in semiwild or wild stingrays, but 65% (31/48) of aquarium-housed stingrays had developing or advanced reproductive disease (ie, ultrasonographic ovarian or uterine score of 4 or 5). Significant correlations were identified between ovarian and uterine disease status and plasma concentrations of all steroid hormones except testosterone. Findings suggested that ultrasonography and plasma hormone concentrations may be useful in the identification of reproductive disease and determination of disease severity in southern stingrays.

Effects of recombinant bovine somatotropin on pregnancy per artificial insemination, corpus luteum cellular composition and endometrial gland morphometry in beef cattle

The aim of this research was to evaluate the effect of recombinant bovine somatotropin (bST) on pregnancy per artificial insemination (P/AI), cellular composition of the corpus luteum (CL) and endometrial gland morphometry. In Experiment 1, Nelore cows (n = 587) received a fixed-time artificial insemination (FTAI) protocol and, at insemination, received 0, 250 or 500 mg of bST subcutaneously (SC). In Experiment 2, Nelore cows (n = 243) received 0 or 500 mg of bST, SC, on D7 (D0 = day of FTAI). Blood samples were collected on D7 and D16 to measure progesterone (P4) concentrations. In Experiments 1 and 2, pregnancy diagnosis was performed 30 days after FTAI. In Experiment 3, Nelore heifers (n = 20) received a FTAI protocol, but were not inseminated, and on D0 (ovulation day), they received 0 (bST 0; n = 9) or 500 mg of bST (bST 500; n = 11), SC. The heifers were slaughtered on D15 (D0 = ovulation day), at which time the CL was evaluated for diameter, weight, a percentage of large (LLC) and small (SLC) luteal cells, and the concentration of progesterone in plasma measured. The number, perimeter and area of superficial and deep endometrial glands were evaluated. There was no difference in P/AI when bST was applied on D0 and D7. In Experiment 1, P/AI did not differ among treatments, with 59.28% (115/194), 58.38% (115/197) and 65.82% (129/196) for the bST 0, 250 and 500 treatments, respectively. In Experiment 2, P/AI did not differ between treatments, with 57.3% (71/124) and 60.5% (62/119) for the bST 0 and 500 treatments, respectively. Plasma progesterone concentrations on D16 was greater in the bST 500 (11.63 ± 0.84 ng/mL) than bST 0 (9.83 ± 0.88 ng/mL). In Experiment 3, there was no difference in ovarian diameter and weight, CL diameter, percentage of SLC, P4 concentrations and endometrial gland morphology. Heifers in the bST 500 treatment had heavier CL (3.11 ± 0.32 vs. 2.25 ± 0.20 g); however, the bST 0 treatment heifers had a greater percentage of LLC than did the bST 500 treatment (13.72 ± 1.16% vs. 8.60 ± 1.52). It was concluded that the doses of bST used in this study do not increase P/AI; however, they do cause changes in P4 concentration and the cellular composition of the CL.

Effects of injecting increased doses of vitamins C and E on reproductive parameters of Holstein dairy cattle

Vitamins C and E have been supplemented separately to improve fertility in cattle. The objective of this study was to evaluate the effect of combined injections of increased doses of vitamins C and E on reproductive parameters in dairy cattle. Lactating Holstein cows (n= 44) were randomly assigned to one of three treatments: 1) Control: n= 15, cows were not injected with vitamins; 2) VCE3: n= 15, cows received a single intramuscular injection of 3,000 IU of vitamin E before estrus and multiple subcutaneous injections of vitamin C with a total dose of 3,000 mg before and after estrus; 3) VCE6: n= 14, cows were treated as in VCE3, but doses of vitamins C and E were increased to 6,000 mg and 6,000 IU. The reproductive indicators measured were diameter of the preovulatory follicle, time to estrus, area of the corpus luteum, pregnancy rate 35 and 45 d after AI and plasma concentrations of estradiol and progesterone. There was no effect of treatment on any of the evaluated reproductive parameters (P˃0.05), except that the lowest dose of vitamins sustained similar pregnancy rates among treatments, even though they had lower progesterone concentrations (P≤0.05) (19.4 ± 2.66 vs 10.1 ± 2.55 vs 19.2 ± 0.44 ng mL-1 for Control, VCE3 and VCE6, respectively). In conclusion, the supplementation with the highest amount of vitamin C and E (6,000 mg and 6,000 IU versus 3,000 mg and 3,000 IU) does not significantly increase the reproductive parameters measured.

Plasma steroid hormone concentrations and their relationships in Suffolk ewes during gestation and parturition.

In this study, we measured plasma concentrations of progesterone, pregnenolone, estradiol, estrone, estrone sulfate, and cortisol and analyzed the correlations between these hormones during gestation in 13 Suffolk ewes, the main breed in Japan. Progesterone increased during gestation and decreased a few days before parturition; however, this pattern was different in samples with high progesterone concentrations (P4 spike samples). This P4 spike was associated with a high pregnenolone concentration. Apart from the P4 spike, the progesterone change was similar to that in other sheep breeds. Pregnenolone increased during gestation and decreased after parturition. A significant correlation between progesterone and pregnenolone was observed a few days before parturition. Estrone sulfate and estradiol concentrations increased during gestation, but estrone did not. They increased shortly before parturition, and then decreased immediately after parturition. At parturition, the correlation between estrone and estrone sulfate was significantly stronger. Moreover, a strong correlation between estrone sulfate and estradiol was observed after parturition. Cortisol did not change during gestation and increased shortly before parturition. The results showed steroid hormone dynamics in normal pregnant Suffolk ewes, which were mostly in line with those of other sheep breeds. It should be noted that high progesterone concentrations altered the typical patterns.

Evaluation of molt induction using cassava meal varying the length of feeding period in older (90 week) laying hens.

Physiological changes during molt and postmolt egg production variables were compared among hens induced to molt by using cassava meal varying the length of the feeding period. Hens were randomly assigned to 3 treatments of 72 birds each: 1) molted by full feeding with cassava meal for 4 wk (CAS4); 2) molted by full feeding with cassava meal for 5 wk (CAS5); and 3) molted by full feeding with cassava meal for 6 wk (CAS6). All groups were fed a pullet developer diet for 2 wk following treatment. During the molt period, the birds were exposed to an 8L:16D photoperiod and had access to drinking water at all times. Thereafter, all hens were fed a layer diet (17.71% CP), and exposed to a 16L:8D photoperiod, and production performance was measured for 16 wk. The molt treatments resulted in total cessation of egg production in all treatments within 2 wk following feeding of the molt diet. BW loss of birds in the CAS6 treatment was approximately 41.81%, which was significantly higher (P < 0.05) than those in the CAS5 (37.63%) and CAS4 (31.89%) treatments. The CAS5 and CAS6 hens had significantly lower (P < 0.05) ovarian and oviducal weights than the CAS4 hens. No consistent differences were observed for concentrations of plasma estradiol and progesterone, bone quality, or mortality among the treatments. Postmolt feed consumption, egg weight, and mortality were not different (P > 0.05) as a result of molt induction method. However, the CAS4 hens had significantly higher (P < 0.05) cumulative egg production and lower feed conversion ratio than hens in the CAS6 treatment. Significant differences were found among treatments for Haugh units and shell thickness in some periods of the postmolt. It was concluded that cassava meal could be used to induce molt in laying hens effectively, feeding the molt diet for 4 wk produced optimum body weight loss and postmolt performance.

Exogenous kisspeptin (kp-10) resumes cyclicity in postpartum anestrus mithun cows

Present study was conducted to assess the effect of exogenous kisspeptin administration on estrus induction, Kiss1gene expression, endocrinological profiles and follicular development in post partum (PP) anestrus mithun cows. Animals who failed to resume cyclicity after day 90–100 of parturition were selected. Experimental cows were examined thoroughly through per-rectal as well as ultrasonograpic examination to confirm the anestrous status. PP anestrous healthy mithun cows (12), aged 5–6 years (body condition score: 5–6 of 10) were selected with uniform anestrus status and randomly divided into 2 groups namely treatment (Gr I) and control (Gr II), each group consist 6 cows. Anestrus cows in Gr 1 were injected with kisspeptin @ 1.30 μg per kg body weight at 3 day interval for 21 days after 90–100 days of post parturition whereas in control group, normal saline was injected as placebo. Blood samples were collected on the days of injection and at estrus in both the groups. Trans-rectal ultrasonography was done at 3 days interval till onset of estrus and ovulation to study the follicular development. Exogenous kisspeptin administration caused significantly early resumption of cyclicity in treatment as compared to control (24.64±10.43 vs 66.56±14.66 days) and significantly increased kiss1 and GPR54 mRNA expression in treatment as compared to control on the day of estrus (1.943±0.29 vs 0.424±0.062 and 1.84±0.31 vs 0.416±0.082, respectively). Similarly, circulating level of estradiol and follicle stimulating hormone (FSH) increased gradually after exogenous administration of kisspeptin, which reached peak on the day of estrus in treatment group (25.36±1.27 pg/ml and 15.65±1.22 ng/ml, respectively) whereas no significant difference was observed between days of treatment in control group except on the day of estrus (11.29±1.76 pg/ml and 9.86±1.06 ng/ml, respectively). Level of estradiol and FSH on the day of estrus was significantly higher in treatment as compared to control whereas non-significant difference was observed in plasma progesterone concentration. Number of medium and large follicles increased in treatment whereas only small follicles were observed in control group. Improved endocrinological profiles, follicle development and kiss gene profiles in post partum anestrous mithun cows following exogenous kisspeptin indicates that kisspeptin inducted or resumed the cyclicity in early.

Monitoring follicular dynamics using ultrasonography in captive brown bears (Ursus arctos) during the breeding season

Artificial insemination (AI) may be a useful tool in the reproductive management of endangered animals, including bears. To establish an AI program for bears, we investigated follicular dynamics using weekly transrectal ultrasonography in six captive brown bears. Along with ultrasonography, we monitored plasma progesterone (P4) and estradiol-17β (E2) concentrations. Furthermore, two bears were administered a gonadotropin releasing hormone (GnRH) agonist to induce ovulation on the first day on which the largest follicle reached more than 10.0 mm in diameter. Brown bears showed two patterns of follicular development in the early and late periods of the breeding season. In the early period (May to mid-June), multiple follicular waves were observed; namely, many follicles developed, and the largest follicles grew to less than 6.0 mm in diameter then regressed. In the late period (mid-June to July), one or two follicles grew to greater than 6.0 mm in diameter and developed as dominant follicles. Moreover, the growth rate of the largest follicle in the late period was faster than that in the early period of the breeding season. One bear with a follicle of 13.1 mm ovulated spontaneously, and one bear ovulated when the follicle was 10.2 mm in diameter after GnRH agonist treatment. Plasma E2 concentrations increased and showed peaks five to seven days before the largest follicles reached their maximum size. Plasma P4 concentrations increased on the day the corpus luteum could be detected using ultrasonography. This is the first study that showed there are two patterns of follicular development in brown bears. Furthermore, the largest follicle reaching greater than 10.0 mm in diameter could be an indicator of the appearance of ovulatory follicles.

Quantitative expression of mRNA encoding BMP/SMAD signalling genes in the ovaries of Booroola carrier and non-carrier GMM sheep.

The GMM sheep is a carrier of Booroola fecundity (FecB) gene, which produces the twins and triplets in one lambing. The homozygous carrier GMM (FecBBB ), non-carrier GMM and Malpura (FecB++ ) ewes were synchronized by progesterone sponges, and the plasma progesterone concentration was measured by RIA. The results showed that the progesterone concentration did not differ significantly (p>.05) in homozygous carrier GMM (5.74±1.2ng/ml), non-carrier GMM (5.42±1.4ng/ml) and non-carrier Malpura ewes (5.67±1.5ng/ml). Further, quantitative expression of BMP factors/receptors and SMAD signalling genes were analysed in the ovaries of sheep by qRT-PCR. The study showed that the expression of BMP2 was slightly higher (p>.05) in carrier GMM than that of non-carrier GMM, but it was almost similar to Malpura ewes. Expression of BMP4 and BMP7 was significantly higher (p<.001; p<.05) in carrier GMM than that of non-carrier GMM and Malpura ewes. Although BMP6 expression was higher (p>.05) in carrier GMM than that of non-carrier GMM, but lower (p>.05) than the Malpura ewes. Expression of BMP15 (p<.05), GDF5 (p<.01) and GDF9 (p<.05) was significantly higher in carrier GMM than non-carrier GMM ewes. Surprisingly, BMPR1B expression was significantly higher (p<.001) in non-carrier GMM and Malpura than the carrier GMM ewes, while TGFβRI did not differ significantly (p>.05) among both GMM genotypes. On the other hand, expression of BMPR1A (p>.05) and BMPRII (p<.05) was higher in carrier GMM than the non-carrier GMM, but significantly lower (p<.001) than the Malpura ewes. It was interesting to note that the expression of SMAD1 (p>.05), SMAD2 (p<.001), SMAD3 (p<.05), SMAD4 (p<.001), SMAD5 (p<.001) and SMAD8 (p<.001) was lower in the carrier GMM than that of non-carrier GMM ewes. It is concluded that the FecB mutation alters the expression of BMPR1B and SMAD signalling genes in the ovaries of homozygous carrier GMM ewes.

Cortisol, progesterone, 17α-hydroxyprogesterone, and TSH responses in dogs injected with low-dose lipopolysaccharide.

BackgroundStress and diseases such as endotoxemia induce cortisol synthesis through a complex biosynthetic pathway involving intermediates (progesterone, and 17α-hydroxyprogesterone (17α-OHP)) and suppression of the hypothalamus-pituitary-thyroid axis.ObjectiveTo measure plasma concentrations of cortisol, progesterone, 17α-OHP, and thyroid stimulating hormone (TSH) in dogs experimentally injected with intravenous low-dose lipopolysaccharide (LPS). Our hypothesis was that LPS treatment would elicit a significant increase in cortisol and its precursors, and a significant decrease in TSH concentration.MethodsHormone measurements were performed on blood samples left over from a previous investigation (2011) on the effect of low-dose LPS on hematological measurands. Five sexually intact female dogs, none in estrous at the time of the study, were administered saline treatment two weeks prior to LPS treatment. LPS was administered intravenously at a dose of 0.1 µg/kg. Blood was collected before (baseline, time -24 hours) and 3-, 6- and 24-hours post-injection. Mixed model analysis for repeated measures was used, with both treatment and time as the repeated factors. Ranked transformation were applied when diagnostic analysis exhibited violation of normality and equal variance assumptions. Post hoc multiple comparisons were performed with Tukey’s adjustment. Statistical significance was defined as p < 0.05.ResultsSignificant differences relative to baseline values were detected following both treatments. Compared to baseline, dogs had significantly higher cortisol and 17α-OHP at 3-hours, and significantly lower TSH at 3- and 6-hours following LPS treatment. Dogs had significantly lower TSH at 6- and 24- following saline treatment. Though not statistically significant, the trend in progesterone concentrations was similar to cortisol and 17α-OHP, with an increase at 3-hours post-injection followed by a decrease close to baseline following both LPS and saline. Cortisol and 17α-OHP concentrations were higher after LPS treatment than after saline treatment at 3- and 6-hours post-injection, but differences were not statistically significant, and no significant differences between treatments were detected for any other hormone or timepoint.Discussion and conclusionCortisol and its adrenal precursors are released in the bloodstream following a low dose of LPS, while TSH appears to decrease. Similar changes occurred following saline treatment, suggesting that even routine handling and saline injection in conditioned dogs can elicit alterations in the internal equilibrium with subsequent modification of both hypothalamus-pituitary-adrenal and thyroid axes. Changes to adrenal and thyroid hormone concentrations must be interpreted in light of clinical information. Further studies are needed to elucidate mechanisms of adrenal steroidal hormone synthesis and secretion in response to various stressful stimuli in both neutered and intact animals.

Are preconceptional stressful experiences crucial elements for the aetiology of autism spectrum disorder? Insights from an animal model

Autism spectrum disorders (ASD) are a group of neurodevelopmental disorders characterized by changes in social interactions, impaired language and communication, fear responses and presence of repetitive behaviours. Although the genetic bases of ASD are well documented, the recent increase in clinical cases of idiopathic ASD indicates that several environmental risk factors could play a role in ASD aetiology. Among these, maternal exposure to psychosocial stressors during pregnancy has been hypothesized to affect the risk for ASD in offspring. Here, we tested the hypothesis that preconceptional stressful experiences might also represent crucial elements in the aetiology of ASD. We previously showed that social isolation stress during adolescence results in a marked decrease in the brain and plasma concentrations of progesterone and in the quality of maternal care that these female rats later provide to their young. Here we report that male offspring of socially isolated parents showed decreased agonistic behaviour and social transmission of flavour preference, impairment in reversal learning, increased seizure susceptibility, reduced plasma oxytocin levels, and increased plasma and brain levels of BDNF, all features resembling an ASD-like phenotype. These alterations came with no change in spatial learning, aggression, anxiety and testosterone plasma levels, and were sex-dependent. Altogether, the results suggest that preconceptional stressful experiences should be considered as crucial elements for the aetiology of ASD, and indicate that male offspring of socially isolated parents may be a useful animal model to further study the neurobiological bases of ASD, avoiding the adaptations that may occur in other genetic or pharmacologic experimental models of these disorders.

Plasma steroid profiles and ovarian response in llamas treated with eCG for superovulation combined with exogenous progesterone during early luteal phase

The objective of this study was to evaluate the effects of plasma progesterone (P4) concentrations during eCG-ovarian follicular superstimulatory treatment performed in early luteal phase and estradiol concentrations during peri-ovulatory period on ovarian response, number and embryo quality. On Day −2, females (n = 75) having a follicle ≥7 mm were treated with GnRH to induce ovulation. On Day 0, females that had ovulations (n = 54) were treated with 1000 IU eCG and were assigned to one of two treatments: (1) intravaginal device (ID) containing 0.5 g P4 (P4 group) and (2) no ID (Control group). On Day 5, females were administered PGF2α and the ID was removed. On Day 7 and 8, females were mated and embryo recovery was performed 7 or 8 days later. Blood samples were collected from Day 0 to 9. Number (± SD) of follicles ≥7 mm on day of mating was greater (P = 0.04) in the control (9.7 ± 4.2) than P4-treated (6.7 ± 4.9) group; number of corpora lutea did not differ (5.5 ± 3.1 and 5.2 ± 3.4 respectively). Ovulation rate was greater (P < 0.01) in the P4-group (77.4%; 130/168) than control group (53.3%; 135/253). Number of embryos with an excellent grade (grade 1) tended to be greater (P = 0.07) in the P4-group (82.4%; 42/51) than control group (65.4%; 36/55). It was concluded that supplementation with exogenous P4 during eCG treatment in early luteal phase inhibits excessive follicular growth, increases ovulation rate and improves embryo quality.

Effects of central orexin A on gonadotropins and progesterone secretion in ewes during the luteal phase of the estrous cycle and during anestrus

Prior studies, mainly on rodents demonstrated that orexin A (OXA) plays an important role in regulating the hypothalamo-pituitary-gonadal (HPG) axis. However, the results of those works have indicated that the central impact of OXA on the HPG axis may vary depending on the species or sex of the animal, the phase of the estrous cycle, or the experimental model system (in vivo vs. in vitro). The aim of the present study was to further investigate the role of OXA in the regulation of reproduction in seasonally breeding, intact, non-steroid-primed ewes during the luteal phase of the estrous cycle and during anestrus. We examined the effect of fourth-ventricle injections of OXA (0.3 μg/kg body weight) on plasma luteinizing hormone (LH), follicle-stimulating hormone (FSH) and progesterone concentrations and on the level of gonadotropin-releasing hormone receptor (GnRH-R) protein in the anterior pituitary (AP). Radioimmunoassays revealed that OXA treatments inhibited (P < 0.001) LH and FSH secretion during both examined periods and progesterone (P < 0.001) during the luteal phase. The influence of OXA on LH and FSH release may be partially attributable to the additional response of the pituitary to OXA, as, under the influence of OXA, the concentration of GnRH-R protein in the AP was downregulated during the luteal phase (P < 0.01) and during anestrus (P < 0.05). We conclude that OXA may be a negative regulator of gonadotropin secretion in intact, non-steroid-primed ewes in periods when the HPG axis is suppressed by negative feedback through steroid sex hormones.