Injection Of Prostaglandin F2 Alpha Research Articles

To Study the Efficacy of Intraamniotic Prostaglandin F2 Alpha versus Mifepristone and Misoprostol Combination for Mid-trimester Medical Termination of Pregnancy

Abstract Context: Aims: This study aims to compare the effectiveness, induction to abortion interval, blood loss, and side effect profile between administration of intraamniotic prostaglandin F2 alpha (PGF2α) with a combination of mifepristone and misoprostol for medical termination of pregnancy in mid-trimester. Prospective observational study. Subjects and Methods: The study was conducted by dividing the women with indicated mid-trimester abortions into two groups of 20 women each. Group-I received Tablet Mifepristone 200 mg orally on day 1 and then after 48 h, Tablet Misoprostol 400 μg given per vaginally every 4 h till abortion was completed or a maximum of 5 doses whichever was earlier. Group-II received a single injection of PGF2α (250 μg) intra-amniotically under ultrasonography guidance. Group comparisons were made using t-test/Mann–Whitney U-test for normally/nonnormally distributed continuous data, respectively, and Chi-square test for categorical variables. SPSS version 23 was used for analysis. P < 0.05 was taken as the cut-off for statistical significance. Results: The success rate was 100% in both the groups. There was a significant difference in the mean induction to abortion interval between Group-I, the mean being 12.92 ± 2.68 h compared to Group-II being 33.08 ± 6.35 h (P < 0.001). Group-I had a mean blood loss of 221 ± 55.15 mL, significantly more than Group-II 160 ± 25.13 mL (P < 0.001). Twenty-five percent of the Group-I woman had retained products of conception with none in Group-II (P = 0.029). The pain score was better in Group II (pain score 4.4 vs. 5.7, P < 0.001). Fewer women developed chills in Group II compared to Group I (0% vs. 50%, P < 0.001). Conclusions: PGF2α is equally effective as a combination of Mifepristone and Misoprostol in mid-trimester abortion. Although it has a longer induction to abortion interval, it appears to be better with respect to the completeness of evacuation, lesser blood loss, and better patient tolerance.

Efficacy of Second Prostaglandin F2 Alpha Treatment during Ovsynch Protocol on Pregnancy Rate Inseminated with Sex-sorted Semen in Buffaloes

Background: Ovsynch is one of the most adopted estrus synchronization protocols used for synchronization of ovulation and fixed-time artificial insemination however, new corpus luteum (CL) induced by the first GnRH treatment in Ovsynch is inappropriately regressed with a single treatment with prostaglandin F2α. Inadequate CL regression reduces fertility. Considering this rationale, present research was implemented with the effect of second prostaglandin F2 alpha treatment during Ovsynch protocol on pregnancy rate inseminated with sex-sorted semen in buffaloes. Methods: Post-partum true anestrus multiparous buffaloes (n=24) were categorized as a group I (Ovsynch; n=12) and group II (Double PG Ovsynch; n=12). Buffaloes from group I were treated with standard Ovsynch protocol (D=7; G-P-G), while additional second prostaglandin F2 alpha injection was administered on day eighth (D=7; G-P-P-G) in group II (Double PG Ovsynch). All the buffaloes were inseminated 16-20 hrs after the last GnRH injection with sex-sorted semen. Result: The estrus signs exhibited were numerically higher in group II than in group I. The first service pregnancy rate in double PG Ovsynch was higher than the Ovsynch group. The addition of a second prostaglandin F2 alpha injection in the Ovsynch protocol improves the estrus quality and increases pregnancy rate.

Progesterone profile of red Sokoto does treated with prostaglandin F2-alpha and progesterone sponges for clinical application

Progesterone profiles of Red Sokoto does were evaluated for clinical application. Fifty-one Red Sokoto goats does were assigned into three groups: (a) prostaglandin F2-alpha (n = 17), and given double injection of prostaglandin F2-alpha at 12-days interval; (b) progesterone sponges (n = 17), and administered progesterone sponges, inserted for 12-days; and (c) control (n = 17), no treatment. Blood samples were collected from all groups from day 0 to 6, day 9, day 12 to 15, day 19, and day 21 to 23 for progesterone profile. Group A had four profiles: 1) does in luteal phase at first and second injections; 2) does in luteal phase at first injection but insensitive at second; 3) does in follicular phase at first injection but luteal phase at second; 4) does, insensitive at first and second injections. Group B profile were: 1) does in luteal phase at sponge insertion; 2) does in luteal phase with decreased progesterone concentration; 3) does in follicular phase at sponge insertion; 4) does with insensitive corpus luteum at sponge insertion. It was concluded that progesterone profile assisted in describing exhibitions and non-exhibitions of behavioural oestrus in Red Sokoto does.

Regulatory changes of local produced prostaglandins in corpus luteum after experimentally induced luteolysis in the cow.

The objective of the study was to evaluate the expression patterns of prostaglandin F2alpha (PGF), prostaglandin E2 (PGE), PGF receptor (FP), PGE receptors (EP2 and EP4), prostaglandin-endoperoxide synthase 2 (PTGS2) and prostaglandin synthases (PGFS and PGES) in corpora lutea (CL) during experimentally induced luteolysis in cow. The Fleckvieh cows in the mid-luteal phase (days 8-12, control group) were injected with cloprostenol (PGF analogue), and CL were collected by transvaginal ovariectomy before (days 8-12, control group) and at 0.5, 2, 4, 12, 24, 48 and 64h after PGF application (n=5 per group). The mRNA expression was determined by RT-qPCR, the hormone concentrations by enzyme immunoassay and localization by immunohistochemistry. PTGS2gene expression increased significantly 2h after PGF application, followed by continuous and significant downregulation afterwards. The PGF tissue concentration increased significantly just after PGF injection and again during structural luteolysis (after 12h), whereas PGE concentration significantly decreased during structural luteolysis. The FP receptor mRNA decreased significantly at 2h and again at 12h after PGF. In contrast, EP4 receptor mRNA increased significantly just after the PGF application (0.5h). The immunostaining of PGES and PTGS2 on day 15-17shows numerous positive luteal cells, followed by lower activity afterwards on day 18 (luteolysis). In conclusion, the changes of examined prostaglandin family members in CL tissue after PGF application may be key components of the local mechanisms regulating the cascade of actions leading to functional and subsequent structural luteolysis in the bovine ovary.

Factors influencing multiple ovulation in dairy cows from Latvian brown, Latvian blue, and Danish red breeds at risk in Latvia

The aim of this study was to determine the general factors that influence multiple ovulation (MO) in the donor cows from the breed Latvian Brown (LB), Latvian Blue (LBl), and Danish Red (DR) using a follicle-stimulating hormone (FSH) or equine chorion gonadotropin (eCG). The 30 cows aged 2 to 15 years were selected for MO and embryo flushing (EF) from the database of the Agricultural Data Center of the Republic of Latvia, of which 13 were LB cows, 7 were LBl cows, and 10 were DR. Serum biochemistry parameters and serum progesterone (P4) concentrations were determined in the donor cows on the day of starting MO treatment (time point T1). Serum P4 detection was repeated at 4 additional time points: before injection of prostaglandin F2 alpha (PGF2α) (T2), on the days of the 1st and 2nd artificial insemination (AI) (T3 and T4), and the day of EF (T5). On the day of EF, ovaries were examined by ultrasonography, and the success of MO was assessed according to the number of corpora lutea (CL). On average, 7.7 (range 1 – 19) CL were found per cow. A successful response to the MO treatment (CL≥3) was observed in 24 cows (80%). The cows that failed to respond to MO treatment were equally distributed among cow breeds. The P4 concentration of the donor cows with a successful response to MO treatment was significantly higher than in cows without the response at T1, T2, and T5. The results of multivariable negative binomial regression showed that the number of CL was positively influenced by P4 level, phosphorus, and cholesterol concentration in the blood serum before MO treatment. It was concluded that cows from breeds LB, LBl, and DR at risk in Latvia effectively responded to MO treatment.

Effect of progesterone supplementation in a resynchronization protocol on follicular dynamics and pregnancy success

The objective of this study was to evaluate the necessity of a controlled internal drug releasing device (CIDR) in a fixed-time AI resynchronization protocol as well as to compare a commercially available blood pregnancy test with transrectal ultrasonography for Day 28 pregnancy detection. Over a two-year period, beef cows and heifers from twelve herds were inseminated using the 7-day CO-Synch + CIDR protocol. On Day 21 following the first insemination, the protocol was repeated, with animals receiving either a CIDR or no CIDR. Pregnancy status (AI1) was determined on Day 28 by both transrectal ultrasonography and the IDEXX Rapid Visual Pregnancy Test. Non-pregnant animals by both methods (CIDR: n = 190 cows, n = 228 heifers; no CIDR: n = 185 cows, n = 223 heifers) received an injection of Prostaglandin F2alpha and were inseminated at the appropriate time or bred following detection of estrus. Corpora lutea (CL) number and largest follicle diameter were recorded on a subset of non-pregnant animals (CIDR: n = 66 cows, n = 46 heifers; no CIDR: n = 76 cows, n = 41 heifers) at time of pregnancy diagnosis on Day 28. Final pregnancy status was determined a minimum of 31 days following the second AI (AI2). The GLIMMIX procedure of SAS was utilized for estrus and pregnancy data; while the MIXED procedure was utilized for analyses of CL number and largest follicle diameter. There was no effect (P ≥ 0.55) of treatment on AI1 pregnancy, AI2 pregnancy, or overall pregnancy rates. The presence of a CIDR during the resynchronization increased (P < 0.001) estrus expression prior to AI2. There was an effect of treatment by age on AI2 pregnancy (P < 0.01); heifers that received a CIDR had greater AI2 pregnancy rates than heifers that did not receive a CIDR (P = 0.04), but there was no difference between cows with and without a CIDR. Treatment had no effect (P > 0.10) on embryonic loss (between the first and second pregnancy diagnosis), CL number, or follicle diameter. Although, there was a tendency for the interaction of treatment by age on follicle size (P = 0.07), with cows having larger follicles than heifers in the no CIDR group but not the CIDR group. In conclusion, use of a CIDR in this resynchronization protocol increased estrus expression, increased AI2 pregnancy for heifers, but did not improve pregnancies in cows, and did not influence overall pregnancy or embryonic loss.

Expression pattern of HIF1alpha and vasohibins during follicle maturation and corpus luteum function in the bovine ovary.

The aim of this study was to characterize expression patterns of hypoxia-inducible factor-1alpha (HIF1A) and vasohibin family members (VASH1 and VASH2) during different stages of ovarian function in cow. Experiment 1: Antral follicle classification occurred by follicle size and estradiol-17beta (E2) concentration in the follicular fluid into 5 groups (<0.5, 0.5-5, 5-40, 40-180 and >180 E2 ng/ml). Experiment 2: Corpora lutea (CL) were assigned to the following stages: days 1-2, 3-4, 5-7, 8-12, 13-16 and >18 (after regression) of oestrous cycle and of pregnancy (months 1-2, 3-4, 6-7, >8). Experiment 3: Cows on days 8-12 were injected with a prostaglandin F2alpha (PGF) analogue and CL were collected before and 0.5, 2, 4, 12, 24, 48 and 64hr after PGF injection. Expression of mRNA was measured by qPCR, steroid hormone concentration by EIA and localization by immunohistochemistry. HIF1A mRNA expression in our study increases significantly in follicles during final maturation. The highest HIF1A mRNA expression was detected during the early luteal phase, followed by a significant decrease afterwards. In contrast, the mRNA of vasohibins in small follicle was high, followed by a continuous and significant downregulation in preovulatory follicles. The obtained results show a remarkable inverse expression and localization pattern of HIF1A and vasohibins during different stages of ovarian function in cow. These results lead to the assumption that the examined factors are involved in the local mechanisms regulating angiogenesis and that the interactions between proangiogenic (HIF1A) and antiangiogenic (vasohibins) factors impact all stages of bovine ovary function.

Effect of exogenous progesterone and gonadotropin-releasing hormone application on maintenance of pregnancy in early pregnant ewes after prostaglandin F2 alpha injection

Effect of exogenous progesterone and gonadotropin-releasing hormone application on maintenance of pregnancy in early pregnant ewes after prostaglandin F2 alpha injection

In Vivo Expression of Cysteine-Rich 61 and Angiogenic Regulators in Bovine Ovarian Follicles.

We have previously shown that the angiogenic inducer, cysteine-rich, angiogenic inducer, 61 (CYR61), is highly expressed in the day 4 bovine corpus luteum (CL). Along with some of the known angiogenic regulators, vascular endothelial growth factor A (VEGF), fibroblast growth factor 2 (FGF2) and matrix metalloproteinase 2 (MMP2), CYR61 may function to promote the high rate of angiogenesis that occurs during the early development of the CL. Since the folliculo-luteal transition results in the infiltration of blood vessels from the vascularized theca interna into the avascular granulosa compartment, we hypothesized that CYR61 may play a critical role in switching on this angiogenic process. However, because it is not known whether ovarian follicles or the surrounding stroma may also synthesize CYR61, our objectives were to determine the expression of CYR61 in bovine granulosa and theca, and relate its expression to know angiogenic regulators such as VEGF, FGF2 and MMP2. Ovaries were removed from midcycle Holstein cows at time 0 (control, no treatment) and at 24 hours or 48 hours following prostaglandin F2 alpha (PGFa) injection. Dissected follicles were measured (diameter; mm); follicular fluid was aspirated; and the granulosa cells and theca interna were separated before RNA was extracted from them. cDNAs generated from granulosa cells and theca interna were analyzed by quantitative PCR to determine the levels of known angiogenic factors, including CYR61, VEGF, FGF and MMP2. Additionally, follicular fluid was analyzed for expression and activity of MMP2 and MMP9 by zymography. Follicular fluid estradiol (E2) and progesterone (P4) concentrations were determined by radioimmunoassay to assess the health status of follicles. The E2 concentrations in the follicular fluid obtained from the largest follicles (diameter 18.9 mm to 22.4 mm) were high, indicating that they were healthy. In addition, plasma P4 concentrations decreased accordingly following PGF2a injection. Further, although there were no differences in mRNA expression of selected angiogenic factors between follicles of PGF2a treated and control cows for any of the endpoints, the mRNA expression of CYR61, FGF2 and MMP2 was always higher (p<0.05) in the thecal than in the granulosa compartment within controls and within each treatment group. In contrast, the mRNA expression of VEGF in the thecal compartment was always lower (p<0.05) than in the hypoxic, granulosa compartment within controls and each treatment group. In summary, CYR61 is reported, for the first time, to be expressed in vivo by granulosa cells and theca interna of bovine ovarian follicles. The function of CYR61 and other angiogenic regulators in the folliculo-luteal transition remains to be elucidated, awaiting analysis of follicles collected at earlier time points, e.g. at 1, 6 and 12 hrs following PGFa injection. This research was supported in part by: NIH grant P01 CA045548 to MAM and by the Multistate Northeast Regional Project NE-1027 to PCWT. (poster)

The effects of three methods of synchronization on estrus induction and hormonal profile in Kalkuhi ewes: A comparison study

Due to the importances of estrus synchronization in sheep reproduction and fertility, the present trial had been carried out to compare the three methods of estrus synchronization in ewes during breeding season. 60 cycling, fat-tailed Iranian Kalkuhi ewes were randomly allocated in equal numbers (n = 20) to 3 treatment groups. In two treatment groups, sponges (medroxyprogesterone) and controlled intravaginal drug-releasing devices (CIDR) were inserted for a period of 14 and 12 days respectively; also, another group received two injections of prostaglandin F2alpha (PGF2α) (with 11 days interval). At the time of CIDR/sponge removal and second PGF2α administration, all the animals received 400 IU of pregnant mare serum gonadotropin (PMSG). Blood samples were collected from all of the treated ewes, and progesterone (P4) and estradiol (E2) concentrations were measured by double-antibody enzyme immune-assay (ELA). Estrus synchronization by CIDR was as effective as Sponge and PGF2α treatments; also CIDR treated ewes exhibited overt signs of estrus earlier than those in other treatment groups. Following hormonal assays, P4 concentrations at first and second sampling and estradiol concentrations at second and third sampling were significantly different (P < 0.05). Key words: Ewe, reproduction, synchronization, estradiol, progesterone.

Evidence That Mammary Gland Infection/Injury During Pregnancy in Dairy Cows May Have a Negative Impact on Size of the Ovarian Reserve in Their Daughters.

The effect of disease on maternal environment, which has a crucial role in embryo development and subsequent health of offspring, has not been examined extensively in cattle. Therefore, the purpose of this study was to determine if a persistent mammary gland infection such as mastitis and correspondingly a chronically high (> 200,000 units) somatic cell count (SCC) in milk of pregnant dairy cows has a potential negative impact on size of the ovarian reserve (total number of healthy follicles and oocytes in ovaries) and potentially fertility of their offspring. This study was conducted at Green Meadow Farms Inc. in Elsie, MI and used 192, 12-mo-old Holstein heifers. Each animal was injected with 2 injections of prostaglandin F2 alpha (PG) spaced 11 d apart. To assess relative size of the ovarian reserve in each heifer, a single blood sample was removed 4 d after the 2nd PG injection and assayed for concentration of anti-Müllerian hormone (AMH), and ovarian ultrasonography was used to determine number of follicles ≥ 3 mm in diameter and ovary size. Results confirm previous observations that AMH concentrations were highly variable among animals but positively correlated (P < 0.001) with both ovary size and number of antral follicles. To determine if a chronic mammary gland infection during pregnancy of dairy cows was associated with alterations in AMH concentrations and correspondingly size of the ovarian reserve in their offspring, we next tested whether number of SCC measurements > 200,000 for each heifer's dam was associated with alterations in the serum AMH concentration in each heifer. Number of SCC measurements > 200,000 per individual cow is an accurate index of recurrent udder infections, injury and/or inflammation. A total of 5 to 7 measurements of SCC were made for each dairy cow beginning 2 mo before pregnancy and continuing throughout pregnancy. Number of SCC measurements > 200,000 ranged from 0 to 5 per individual cows, thus cows were separated into groups based on their number of SCC measurements > 200,000. Cows with 4 or 5 SCC measurements > 200,000 were ~2 years older (P < 0.01, 6.1 vs 4.4 yrs old) and tended (P < 0.09) to produce less milk compared with cows that had 0 to 3 SCC measurements > 200,000. Results showed that the daughters of cows with 4 and 5 SCC measurements > 200,000 also had markedly lower (P < 0.02) AMH concentrations as young adults compared with daughters of the cows with 0 to 3 SCC measurements > 200,000. Although mechanisms are unclear, our results imply that a chronic mammary infection or injury during pregnancy of older cows (as predicted by a high number of SCC measurements > 200,000) is associated not only with low milk production, but also with diminished size of the ovarian reserve and perhaps reduced reproductive performance (fertility and reproductive lifespan) of their daughters. Research supported by USDA-NRI 2004-01697, 2007-01289 to JJI. (poster)

Recent Advances in the Synchronization of Estrus and Ovulation in Dairy Cows

Synchronization programs have become standard components in the current breeding management of cows in the dairy herds of most dairy industries. Many are based on protocols that allow timed inseminations (TAI) so as to circumvent the practical difficulties associated with estrus detection. These difficulties are exacerbated in modern herds of high producing cows either because of increasing herd size in which individual animal monitoring is difficult and often subjective, or because small intensively managed herds are milked in robotic systems that minimize animal: staff interactions. Additional reasons arise from high producing cows having less obvious symptoms of estrus, partly because of housing systems combined with intensive feeding and milking, partly because of higher metabolic clearance rates of reproductive hormones like estradiol and partly because of the increasing prevalence of prolonged post-partum anestrus and reproductive tract pathology. The most recently developed programs include protocols for resynchronization following first or subsequent inseminations. These re-synchronization protocols may involve selected forms of hormonal intervention during the diestrous and pro-estrous periods following TAI, or following pregnancy diagnosis by ultrasound from 28 days after TAI. The latter form of re-synchronization has become increasingly important with the recognition that late embryonic/early foetal death has become a major factor compromising the reproductive performance of high producing Holstein cows in many dairy industries. Although cows detected in estrus without any hormonal treatment before insemination have higher conception rates than those inseminated following synchronization and TAI, the low detection rates combined with embryonic death means that intervals from calving to conception (days open) are usually less when synchronization programs have been successfully implemented. One of the significant factors affecting a program's success is the compliance rate that may sometimes be less than 70%. Almost all programs involve strategically timed injections of prostaglandin F2alpha (PGF) and gonadotropin releasing hormone (GnRH). Injections of an estradiol ester and progesterone supplementation per vaginum may be included in some programs. The basic program is the "Ovsynch" regimen. Numerous variations have been tested and developed. Many involve increasingly complex protocols that increase the risk of non-compliance, none has consistently achieved conception rates that exceed 40% and few have reduced the incidence of embryonic death. These synchronization programs are the best that are currently available. They have not been able to overcome the consequences of lowered fertility associated with high levels of milk yield, forms of nutrition and environmental factors like heat stress that have profound effects on the physiology and metabolism of the high producing dairy cow.

Vascular Endothelial Growth Factor A 164b (Vegfa164b) Inhibitory Isoform is Up-regulated in Bovine Granulosa Cells Prior to the LH Surge May Be a Marker of Follicle Status.

The Vegfa gene has 8 different exons that can be alternatively spliced to form several different angiogenic isoforms: Vegfa120, Vegfa164, Vegfa188, Vegfa205. Recently, human VEGFA inhibitory (b) isoforms were discovered that contain exon 8b instead of 8a (present in angiogenic isoforms). Furthermore, for every angiogenic isoform there is a similar size inhibitory or b isoform (ie. Vegfa120b, Vegfa164b, etc) that blocks its actions. Our laboratory has determined that VEGFA inhibitory isoforms regulate rat follicle progression in vitro. Thus, the objective of the current study was to amplify and sequence the bovine Vegfa164b isoform and determine its expression in relation to Vegfa164 prior to the LH surge in bovine granulosa cell aspirates. Furthermore, differences in these isoforms were determined in estrogenic verses non-estrogenic follicles. Cows were administered two injections of prostaglandin F2alpha (PG; 25 mg/ head) 14 days apart and the largest follicle was aspirated at 12, 18, 24, 30, 36 and 48 hr after the second injection of PG (n= 10 at each aspiration). Estrogen (E2) and progesterone (P4) assays were conducted on follicular fluid to determine whether the follicle was estrogenic (E2/P4 > 1) or non-estrogenic (E2/P4 < 1). Granulosa cells were extracted for RNA and quantitative real-time PCR (QRT-PCR) was conducted using reverse primers specific for: 1) Vegfa164 (exon 8a); 2) Vegfa164b (exon 8b); and 3) both Vegfa164 and Vegfa164b (reverse primer in exon 7; to compare with previous data). The Vegfa164b inhibitory isoform was amplified through RT-PCR and demonstrated to be 97% homologous with its human Vegfa165b counterpart. Furthermore, the predicted bovine protein sequence contained one amino acid less than the human. Prior to the LH surge, Vegfa164b in estrogenic follicles was greater at 18 hr (P=0.06) after the second injection of PG; however, there were no differences in amount of Vegfa164. The reverse primer that amplified both Vegfa164 and 164b (exon 7) had peak expression at 18 hr (P < 0.08) in estrogenic follicles similar to the results from the Vegfa164b primer (exon 8b). To determine if there were differences in isoforms based on follicle status, the area under the curve for Vegfa164 and 164b was determined for granulosa cells from estrogenic and non-estrogenic follicles aspirated at 12, 18, 24 and 48 hr after PG. In estrogenic follicles there was 2 fold greater Vegfa164b and 1.6 fold greater Vegfa164 isoforms than non-estrogenic follicles. However, in the reverse primers detecting both isoforms there was only 1.24 fold greater Vegfa164a&b isoform detected in estrogenic versus non-estrogenic follicles. Thus, inhibitory Vegfa164b isoform but not angiogenic Vegfa164 isoform is up-regulated in the granulosa cell prior to the LH surge and may be involved in follicle development or oocyte maturation. Furthermore, amount of Vegfa164b isoform in bovine granulosa cells may be utilized in the future to detect oocyte quality and developmental competence due to estrogenic or non-estrogenic status of the follicle.

Developmental Programming: Deficits in Reproductive Hormone Dynamics and Ovulatory Outcomes in Prenatal, Testosterone-Treated Sheep1

Prenatal testosterone excess leads to neuroendocrine, ovarian, and metabolic disruptions, culminating in reproductive phenotypes mimicking that of women with polycystic ovary syndrome (PCOS). The objective of this study was to determine the consequences of prenatal testosterone treatment on periovulatory hormonal dynamics and ovulatory outcomes. To generate prenatal testosterone-treated females, pregnant sheep were injected intramuscularly (days 30-90 of gestation, term=147 days) with 100 mg of testosterone-propionate in cottonseed oil semi-weekly. Female offspring born to untreated control females and prenatal testosterone-treated females were then studied during their first two breeding seasons. Sheep were given two injections of prostaglandin F2alpha 11 days apart, and blood samples were collected at 2-h intervals for 120 h, 10-min intervals for 8 h during the luteal phase (first breeding season only), and daily for an additional 15 days to characterize changes in reproductive hormonal dynamics. During the first breeding season, prenatal testosterone-treated females manifested disruptions in the timing and magnitude of primary gonadotropin surges, luteal defects, and reduced responsiveness to progesterone negative feedback. Disruptions in the periovulatory sequence of events during the second breeding season included: 1) delayed but increased preovulatory estradiol rise, 2) delayed and severely reduced primary gonadotropin surge in prenatal testosterone-treated females having an LH surge, 3) tendency for an amplified secondary FSH surge and a shift in the relative balance of FSH regulatory proteins, and 4) luteal responses that ranged from normal to anovulatory. These outcomes are likely to be of relevance to developmental origin of infertility disorders and suggest that differences in fetal exposure or fetal susceptibility to testosterone may account for the variability in reproductive phenotypes.

Induction of Estrus by Prostaglandin F2.ALPHA. Administration in the Vaginal Vestibules of Miniature Pigs

Estrus induction is an important step in embryo production. It has been difficult to induce estrus in miniature pigs by intramascular (i.m.) injection of prostaglandin F2alpha (PGF2(2alpha)) in the early luteal stage of the estrous cycle. In the present study, we injected two different doses of PGF2(2alpha) i.m. and into the submucosa of the vaginal vestibule (i.ves.) of miniature pigs, and examined the effect of these treatments on estrus induction. Fifteen miniature pigs were divided into five experimental groups (control, saline injected i.m.; PGF2alpha treated, 1.0 or 1.5 mg of PGF2alpha injected twice i.m. or i.ves.), and the estrus length and concentrations of 17beta-estradiol (E2) and progesterone (P) in the blood were examined. Estrus length was significantly shortened by a large amount of PGF2alpha injected i.ves. In addition, the concentration of P in the blood significantly decreased after two injections of PGF2alpha (i.m. or i.ves.). These results suggest that in miniature pigs, administration of at least 3.0 mg of PGF2alpha is required for the induction of luteolysis and injection of PGF2alpha into the vaginal vestibule is a useful method of estrus induction.

Synchronization of Ovulation and Fixed Time Intrauterine Insemination in Ewes

A novel method for oestrus-ovulation synchronization in sheep followed by fixed time insemination is presented herewith. Mature dry ewes (n = 28) of Karagouniko breed being at an unknown stage of the oestrous cycle, were used during the middle of breeding season. The treatment protocol consisted of an initial administration of a GnRH analogue followed 5 days later by a prostaglandin F2alpha injection. Thirty-six hours later a second GnRH injection was administered to synchronize ovulation, and laparoscopic intrauterine insemination was performed 12-14 h later. Three days after insemination, fertile rams were introduced into the flock twice daily and oestrus-mating detection was carried out. For progesterone (P(4)) determination, blood samples were collected on alternate days, starting 2 days before the first GnRH injection and continuing for 17 days after insemination. An additional sample was taken on the day of insemination. Pregnancy diagnosis was carried out by trans-abdominal ultrasonography. Fourteen ewes (50%) conceived at insemination and maintained pregnancy; from the remainder 14 ewes 10 became pregnant at natural service, while four, although they mated at least two to three times, failed to conceive. In response to the first GnRH, P(4) concentration increased at higher levels in ewes that conceived at AI compared with those that failed to conceive (47.54 and 22.44%, respectively; p < 0.05). Significant differences (p < 0.05) in mean P(4) concentration between pregnant and non-pregnant animals were detected 1 day before AI (0.17 +/- 0.06 and 0.26 +/- 0.14 ng/ml, respectively) on the day of AI (0.15 +/- 0.04 and 0.24 +/- 0.08 ng/ml, respectively) as well as 9 and 11 days thereafter (0.48 +/- 0.12 and 0.38 +/- 0.12 ng/ml; 0.68 +/- 0.14 and 0.50 +/- 0.18 ng/ml, respectively). These results indicate that using the proposed protocol, an acceptable conception rate can be achieved which could be further improved by modifying the time intervals between interventions.

In Vivo Evidence that Local Cortisol Production Increases in the Preovulatory Follicle of the Cow

The aim of the present in vivo study was to monitor real-time fluctuations of cortisol (Cr) in the wall of preovulatory follicles using a microdialysis system (MDS) implanted in the theca layer as well as changes in ovarian venous plasma (OVP) and jugular venous plasma (JVP). Seven cows were superovulated using FSH and prostaglandin F2alpha injections. Dialysis capillary membranes were surgically implanted into the theca layer of mature follicles and connected to a microdialysis system. Fractions of the perfusates were collected from Day -1 (Day 0=LH surge) to Day 3. No difference in the concentrations of Cr between JVP and OVP was detected throughout the experiment. Circulating concentrations of Cr ranged from 20 to 35 ng/ml 8 h after surgery in ovulatory and anovulatory cows. In five ovulatory cows, the Cr concentration decreased to basal levels (<10 ng/ml) between 12 and 24 h after surgery, however, two anovulatory cows retained high Cr levels (>10 ng/ml) up to 42 h after surgery. There was a clear increase in the local concentration of Cr from 13.3+/-2.1 pg/ml at -24 h to 27.5+/-1.7 pg/ml at 0 h (peak of the LH surge) within the wall of ovulatory follicles. This increase was not detected in anovulatory follicles. This transient increase in Cr occurred only in the follicle wall, but not in the OVP or JVP, indicating that the presence of a local regulatory mechanism for Cr production/conversion in ovulatory follicles, and this mechanism may modulate the inflammatory-like reaction induced by LH surge in the follicle wall. The present results demonstrate that the glucocorticoid environment in the follicular wall adjusts at the local level in bovine ovulatory follicles. This mechanism may protect follicles from the adverse effects of glucocorticoid, and it may prevent excess inflammatory reactions associated with ovulation by temporarily increasing local concentrations of glucocorticoid, thus forming an integral part of the regulatory mechanism in ovarian physiology.

Second trimester abortion by laminaria followed by vaginal misoprostol or intrauterine prostaglandin F2α: a randomized trial

Intra-amniotic injection, as well as intravaginal application of prostaglandins, have been used to terminate second trimester pregnancies. There is as yet no consensus as to the most efficient protocol of such late abortions. Our goal was to compare the efficacy of intra-amniotic injection of prostaglandin F2 alpha (PGF2α) and intravaginal application of misoprostol in terminating second trimester pregnancies after pretreatment with intracervical laminaria. Women with live fetuses and requesting second trimester abortions were randomized into two groups. Eighteen hours following the insertion of intracervical laminaria, women were treated with either intra-amniotic injection of 40 mg PGF2α, or 12 hourly doses (to a maximum of 4 doses) of 200 mcg misoprostol. Fifty women were randomly assigned to each group. Failure to abort within 24 h of initiation of treatment occurred in 6 patients (12%) in the misoprostol group and 14 (28%) of the PGF2α group (p = 0.04). Mean time of induction of pharmacologic treatment to abortion was 13.6 h in the misoprostol group and 10.7 h in the PGF2α group (p = 0.03). The mean number of analgesic injections given were 0.8 in the misoprostol group and 1.6 in the PGF2α group (p = 0.0001). Only the method of abortion was predictive of abortion success and not other variables such as patient age, gestational age, gravidity, or parity. Following intracervical laminaria, vaginal misoprostol has been found to be more effective and less painful, compared with intra-amniotic PGF2α, for the termination of second trimester pregnancies with live fetuses.

Melengestrol acetate blocks the preovulatory surge of luteinizing hormone, the expression of behavioral estrus, and ovulation in beef heifers.

We tested the hypothesis that melengestrol acetate (MGA), an orally active progestin, blocks estrus and the preovulatory surge of luteinizing hormone (LH) in beef heifers. Cycling yearling Angus heifers were divided randomly into two groups: MGA-treated (n = 6) and control (n = 5). All heifers received injections of prostaglandin F2alpha (PGF) on d -25, -11, and 0 to synchronize estrus. Following the last PGF injection on d 0, heifers were fed either 0.5 mg MGA in a carrier or the MGA carrier each day for 8 d. At 4-h intervals on d 1 through 6, all heifers were observed for expression of estrous behavior, and blood samples were collected and assayed for LH. Daily blood samples were collected at 0800 on d 1 through 10 and assayed for circulating progesterone concentrations. All control heifers exhibited estrus and a preovulatory surge of LH. In each case, this was followed by increases in circulating concentrations of progesterone indicative of ovulation and normal luteal function. In contrast, none of the MGA-treated heifers exhibited estrus, LH surges, or evidence of ovulation. The results of this experiment show that MGA prevents ovulation in cattle by inhibiting the preovulatory surge of LH.

Indomethacin blocks pre-partum nest building behaviour in the pig (Sus scrofa): effects on plasma prostaglandin F metabolite, oxytocin, cortisol and progesterone.

In the pig, nest building occurs in the day preceding parturition (gestation=114--116 days). Nest building behaviour can be induced in pregnant, pseudopregnant and cyclic female pigs following injection of prostaglandin F2alpha. Here we investigated behaviour and endocrine changes after the administration of indomethacin, which inhibits cyclo-oxygenase enzymes and thus prostaglandin synthesis. In experiment 1, pregnant primiparous pigs (gilts) were blood sampled through jugular vein catheters every 20 min from 1000 h on day 113 of pregnancy and behaviour was recorded until birth. Two hours after pre-partum nest building began, animals received 4 mg/kg indomethacin (n=7) or control vehicle (n=8) intramuscularly. Indomethacin-treated animals showed less nest building than controls between 1 and 5 h after injection (P<0.05), during which time they were mostly inactive and lay down for longer than controls. From 5 h before birth until birth there was no significant treatment difference in nest building behaviour. There was a tendency for the start of birth to be delayed in indomethacin-treated animals. Plasma 13,14-dihydro-15-keto-prostaglandin F2 alpha (a major metabolite of prostaglandin F2 alpha) rose during pre-injection nest building and then fell following indomethacin treatment, but was not significantly different between groups when behaviour differed. Plasma oxytocin, cortisol and progesterone were not significantly affected by treatment. In experiment 2, indomethacin-treated non-pregnant gilts (n=7) did not show any changes in activity or posture compared with vehicle-treated controls (n=6) between 90 and 150 min after treatment. These results suggested that indomethacin treatment reversibly and specifically inhibits porcine pre-partum nest building by a mechanism that may involve endogenous prostaglandin F2 alpha synthesis inhibition but is independent of circulating oxytocin, cortisol and progesterone concentrations.