Corpus Luteum Group Research Articles

Local Regulatory Changes of HSD11B1 and NR3C1 in the Follicular and Luteal Tissue During Experimentally Induced Ovulation in the Cow.

The objective of the study was to characterise the expression patterns of the two key components of cortisol action namely HSD11B1 (11-beta-hydroxysteroid dehydrogenase type 1) and NR3C1 (nuclear receptor subfamily 3, group C, member 1, also known as the glucocorticoid receptor) in superovulation induced bovine follicles during the periovulation and subsequent corpus luteum (CL) formation. Bovine ovaries containing preovulatory follicles or CL were timely defined during induced ovulation as follows: 0 h before GnRH (Gonadotropin-releasing hormone) application, and 4, 10, 20, 25 (follicles) and 60 h (early CL) after GnRH. The low mRNA expression of HSD11B1 and NR3C1 in the follicle group before the GnRH application increased significantly in the follicle group 20 h after GnRH and remained high afterward also in the early CL group. In contrast, the high NR3C1 mRNA decreased in follicles 25 h after GnRH (close to ovulation) and significantly increased again after ovulation (early CL). Our results indicated the involvement of HSD11B1 and NR3C1 as the two key components of cortisol action in the local mechanisms coordinating final follicle maturation, ovulation, follicular-luteal transition and CL development in the cow.

Lactating dairy cows managed for second and greater artificial insemination services with the Short-Resynch or Day 25 Resynch program had similar reproductive performance

The objective of this randomized controlled experiment was to evaluate reproductive performance and reproductive physiological outcomes of lactating Holstein cows managed for second and greater artificial insemination (AI) services with the Short-Resynch or Day 25 Resynch program. Cows from 2 commercial farms were randomly assigned after first service to the Short-Resynch (SR; n = 870) or Day 25 Resynch (D25R; n = 917) program in which they remained until 210 d after first service or left the herd. Cows in D25R received GnRH 25 ± 3 d after AI, whereas cows in SR did not. Cows not reinseminated at detected estrus (AIE) by 32 ± 3 d after AI underwent nonpregnancy diagnosis (NPD) through transrectal ultrasonography (TUS). Nonpregnant cows from both treatments with a corpus luteum (CL) ≥15 mm and an ovarian follicle ≥10 mm (hereafter, CL cows) received 2 PGF2α treatments 24 h apart, GnRH 32 h after the second PGF2α, and timed AI 16 to 18 h later. Cows that did not meet the criteria to be included in the CL group (NoCL cows) received a modified Ovsynch protocol with progesterone (P4) supplementation [P4-Ovsynch; GnRH and controlled internal drug-release device (CIDR) in, 7 d later CIDR removal and PGF2α, 24 h later PGF2α, 32 h later GnRH, and 16 to 18 h later timed AI]. In a subgroup of cows, blood samples were collected and TUS conducted at each treatment to evaluate ovarian responses to resynchronization. Binary data were analyzed with logistic regression, continuous data by ANOVA, and time-to-event data by Cox's proportional hazard regression. A greater proportion (mean; 95% CI) of cows were AIE before NPD in the SR (60.5%; 57.0-63.8; n = 3,416) than the D25R (50.1%; 46.5-53.7; n = 3,177) treatment, whereas pregnancy per AI (P/AI) at 32 d for AIE services before NPD was greater for the D25R (41.3%; 38.8-43.8; n = 1,560) than the SR (37.6%; 35.5-39.8; n = 1,961) treatment. At NPD, a greater proportion of cows in the D25R (84.3%; 82.2-86.2) than the SR (77.0%; 74.4-79.4) treatment were considered CL cows. Pregnancy per AI at 32 d was greater for the D25R than the SR treatment for all timed AI services (D25R = 43.0%; 40.2-45.9 vs. SR = 36.8%; 33.8-39.8) and for CL cows (D25R = 42.8%; 39.7-45.9 vs. SR = 33.8%; 30.6-37.2) but did not differ for NoCL cows (D25R = 39.4%; 32.1-47.3 vs. SR = 44.0%; 36.8-51.4). The hazard ratio for time to pregnancy (1.03; 0.93-1.14) and the proportion of cows not pregnant at the end of the observation period (D25R = 5.9%; 4.4-7.8 vs. SR = 6.7%; 5.0-8.7) did not differ between SR and D25R treatments. The GnRH treatment 25 d after AI resulted in more cows with P4 >1 ng/mL (D25R = 80.5%; 75.3-84.9 vs. SR = 63.6%; 57.3-69.4) and smaller follicle diameter at NPD 32 ± 3 d after AI for D25R (16.2 ± 0.4 mm) than for SR (17.5 ± 0.4 mm); however, it did not affect follicle diameter and luteal regression risk (CL cows only) before TAI. We concluded that the use of reproductive management programs including SR and D25R for CL cows and the P4-Ovsynch protocol for NoCL cows resulted in similar hazard of pregnancy and proportion of nonpregnant cows for up to 210 d after first service.

Qualitative and quantitative assessment of bovine oocytes

The reproductive development of cattle is must to hasten the production of milk and meat. It relies mostly on the quality of oocytes. Thus the present study was carried out to sort out the superior ovary to be used in further research. Cow ovaries were collected from slaughter house, processed and categorized as right or left and ovaries with or without corpus luteum (CL). Morphology of ovaries were studied on the basis of length (cm), width (cm), weight (g), total number of follicles on the surface of each category ovaries, number of follicles aspirated, total number of cumulus-oocyte-complex (COCs) and then graded as normal and abnormal. Significantly higher mean weight (3.47±1.85 vs 2.44±1.76), length (1.94±0.34 vs 1.71±0.39) and width (1.36±0.34 vs 1.09±0.29) were found in right ovaries than left. Forty seven of right and 20% of left ovaries were found with CL. Numerically higher number of normal COCs was found in left ovaries than right ovaries whereas total and abnormal COCs were higher in right ovaries. On the other hand, in comparison between the ovaries with and without CL group, significantly (p<0.05) higher mean weight (3.81±1.68 vs 2.53±1.83), length (2.04±0.30 vs 1.72±0.38) and width (1.45±0.25 vs 1.11±0.32) were found in with CL ovaries than without CL. The average number of normal COCs of with CL ovary (0.75±0.85) was lower than that of without CL (0.85±0.83) but they are not significant. Finally, it can be concluded that, oocytes from left ovaries in comparison to right and without CL in comparison to with CL are superior in the context of selection of oocytes for embryo development.

Effect of Mode of Conception on Maternal Serum Relaxin, Creatinine, and Sodium Concentrations in an Infertile Population.

To investigate how the mode of conception affects maternal relaxin, creatinine, and electrolyte concentrations. Pregnancies achieved by fertility treatment often begin in a nonphysiologic endocrine milieu with no corpus luteum (CL) or with many corpora lutea. The CL produces not only estradiol and progesterone but is also the sole source of relaxin in early pregnancy, a hormone that may contribute to maternal systemic and renal vasodilation. There is limited data about maternal physiology in early pregnancy during fertility treatment, and studies have rarely considered the potential effect of the absence of the CL. To begin to address this gap in knowledge, we sought to investigate how the mode of conception affects maternal relaxin, creatinine, and electrolyte concentrations. One hundred eighty-four women who received care at an academic infertility practice provided serum samples. Levels of relaxin 2, creatinine, and electrolytes were compared between 4 groups defined on the basis of mode of conception which corresponded to categories of CL number: (1) absence of the CL, (2) single CL, (3) multiple CL from ovarian stimulation not including in vitro fertilization (IVF), and (4) multiple CL from IVF with fresh embryo transfer. Relaxin-2 levels were undetectable in patients lacking a CL. Creatinine, sodium, and total CO2 levels were significantly higher in the 0 CL group (relaxin absent) compared to all other groups (relaxin present). Compared to clomiphene, use of letrozole was associated with a lower relaxin level. Early creatinine and sodium concentrations are increased in the absence of relaxin. Given the increasing utilization of frozen embryo transfer, further studies comparing programmed with natural cycles are warranted.

Sex steroids modulate morphological and functional features of the bovine oviduct.

In cattle, the oviduct plays a major role in the reproductive process; however, molecular control of oviduct receptivity to the embryo is poorly understood. A model for receptivity based on size of the pre-ovulatory follicle (POF) was used to compare oviductal morphology, cellular proliferation, and candidate transcript abundance. Growth of the POF of Nelore (Bos indicus) cows was manipulated to produce two groups: a large POF-large corpus luteum (CL) group (LF-LCL; greater receptivity) and a small POF-small CL group (SF-SCL). Samples of the ampulla and isthmus ipsilateral and contralateral to CL were collected 4days after GnRH-induced ovulation. Tissues were either embedded in paraffin for Harris-Hematoxylin and Eosin and periodic acid-Schiff staining and KI67 immunostaining, followed by morphological analyses, or stored at -80°C for RNA extraction, cDNA synthesis, and qPCR analyses. The effects of group (LF-LCL and SF-SCL), region (ampulla and isthmus), and side (ipsilateral and contralateral) were analyzed using three-way nested ANOVA. The ipsilateral ampulla of the LF-LCL group presented more primary mucosal folds, a greater mucosal-folding grade and luminal perimeter, and more secretory cells and proliferating cells when compared with the ampulla of the SF-SCL group and with the contralateral ampulla of both groups. There were no morphological differences in the isthmus between groups and sides. Changes in transcript abundance are suggestive of LF-LCL-stimulated secretory activity. In summary, ovulation of a larger POF generates a periovulatory endocrine milieu that modulates morphological and functional features of the bovine oviduct which may support embryo survival and development.

133 COMPARISON OF ENDOMETRIAL TRANSCRIPTOME CHANGES BETWEEN IPSI- AND CONTRALATERAL HORNS DURING DIESTROUS AND ITS RELATIONSHIP WITH THE ABILITY TO SUPPORT CONCEPTUS ELONGATION IN CATTLE

Data from embryo transfer studies indicate that transfer to the uterine horn contralateral to the ovary containing the corpus luteum (CL) reduces pregnancy rate. The aim of this study was to compare the ipsi- v. contralateral horns in terms of (1) endometrial gene expression and (2) the ability to support conceptus growth to Day 14. In Experiment 1, endometrial samples from the ipsi- and contralateral horns were collected from synchronized nonpregnant heifers slaughtered on Day 5, 7, 13 or 16 post-oestrus (n = 5 per time point) and snap frozen for subsequent analysis. In Experiment 2, fresh Day 7 in vitro-produced blastocysts were transferred to the uterine horn(s) ipsilateral (Group I, n = 9 recipients) or contralateral to the CL (Group C, n = 8) or to both horns (i.e. bilateral, Group B, n = 9) of synchronized recipients (10 embryos transferred per horn per recipient). All recipients were slaughtered on Day 14. Reproductive tracts were recovered and each horn was flushed independently. The number and dimensions (length and width) of recovered conceptuses were recorded independently for each horn. Embryo data were analysed using PROC MIXED in SAS (SAS Institute Inc., Cary, NC, USA). The model had treatment as a fixed effect and heifer within treatment as a random effect. Experiment 1, RNA-Seq analysis of endometrium, detected 217, 54, 14, and 18 differentially expressed genes (&gt;2-fold change, false discovery rate P &lt; 0.05) between ipsi- and contralateral horns on Day 5, 7, 13, and 16 of the oestrous cycle, respectively. In Experiment 2, no differences were observed in the recovery rate of conceptuses on Day 14 from the 3 groups [53.3% (48/90), 48.7% (39/80), and 45% (81/180)], for the I, C, and B transfer groups, respectively. Conceptus length from the ipsi- (2.24 ± 0.35 mm) and contralateral (2.91 ± 0.38 mm) horns was not different (P &gt; 0.05). Thus, significant differences in endometrial gene expression exist between the uterine horns ipsi- and contralateral to the CL in cattle, but those differences do not affect conceptus elongation on Day 14. Research was supported by Science Foundation Ireland (13/IA/1983) and the Irish Department of Agriculture, Food and The Marine (13S528).

Size of the Ovulatory Follicle Dictates Spatial Differences in the Oviductal Transcriptome in Cattle.

In cattle, molecular control of oviduct receptivity to the embryo is poorly understood. Here, we used a bovine model for receptivity based on size of the pre-ovulatory follicle to compare oviductal global and candidate gene transcript abundance on day 4 of the estrous cycle. Growth of the pre-ovulatory follicle (POF) of Nelore (Bos indicus) cows was manipulated to produce two groups: large POF large corpus luteum (CL) group (LF-LCL; greater receptivity) and small POF-small CL group (SF-SCL). Oviductal samples were collected four days after GnRH-induced ovulation. Ampulla and isthmus transcriptome was obtained by RNA-seq, regional gene expression was assessed by qPCR, and PGR and ERa protein distribution was evaluated by immunohistochemistry. There was a greater abundance of PGR and ERa in the oviduct of LF-LCL animals thus indicating a greater availability of receptors and possibly sex steroids stimulated signaling in both regions. Transcriptomic profiles indicated a series of genes associated with functional characteristics of the oviduct that are regulated by the periovulatory sex steroid milieu and that potentially affect oviductal receptivity and early embryo development. They include tissue morphology changes (extra cellular matrix remodeling), cellular changes (proliferation), and secretion changes (growth factors, ions and metal transporters), and were enriched for the genes with increased expression in the LF-LCL group. In conclusion, differences in the periovulatory sex steroid milieu lead to different oviductal gene expression profiles that could modify the oviductal environment to affect embryo survival and development.

194 EFFECT OF CORPUS LUTEUM PRESENCE FOR THE DURATION OF FOLLICULAR GROWTH ON BOVINE OOCYTE DEVELOPMENTAL COMPETENCE

It has been reported that follicular wave control is effective to increase bovine oocyte (OC) quality of ovum pickup (Imai et al. 2008 Reprod. Fertil. Dev. 20, 182). However, the effect on the bovine OC developmental competence to the presence of corpus luteum (CL) for the duration of follicular growth (FG) is not known. This study was conducted to examine the developmental competence of OC after stimulation of FG with and without CL. Japanese Black cows (n = 4/session) were synchronised in oestrus (Day 0; D0) with prostaglandin F2α (PGF2α; 0.5 mg cloprostenol). On Day 9, cows were removed follicles (FL) ≥ 8 mm in diameter and CIDR were inserted. Cows in the PGF2α treatment group (PG) were administered PGF2α (0.75 mg cloprostenol) for regression of the CL; the PGF2α non-treatment group (non-PG) were not administered PGF2α for the presence of CL. The cows received 20 mg of FSH twice a day from the evening of Day 10 to the morning of Day 14 in decreasing doses. Oocytes were collected from FL ≥ 5 mm by OPU on Day 15. Collected OC were classified as follows: Grade 1, several layers of cumulus cells (CC) and homogeneous OC cytoplasm; Grade 2, 1 to 3 layers of CC; Grade 3, denuded OC; Grade 4, expanded CC or degenerated OC. The OC that were classified as Grade 1 to 3 were matured in vitro for 20–22 h and then inseminated with 3 × 106 sperm mL–1 for 6 h. Then presumptive zygotes were cultured in 5% newborn calf serum + CR1aa for 216 h in micro-well culture dishes (Dai, Nippon, Japan). Cows were assigned to groups of 4 with each pair placed into 1 of the treatments, and groups switched at ≥40 days of interval for crossover trials. To compare the quality of OC derived from CL ipsilateral FL and CL contralateral FL, the OC were subdivided in CL ipsilateral group (ipsi-CL) and CL contralateral group (contra-CL) in the non-PG. In addition, half of the data from PG were regarded as unilateral ovary (regression CL group; reg-CL). Statistical analysis was carried by Student’s t-test, Tukey’s test, Fisher’s exact probability test, and chi-squared test. Part of data was log-transformed. At OPU, all cows in the non-PG group had an observed CL, and the average diameter of the CL was 16 ± 0.1 mm in the long axis and short; all cows in the PG were observed to not have a CL. In PG and non-PG, there were no significant differences in the number of FL ≥ 5 mm, the rate of cleavage, or the rate of blastocyst (BL) formation (17 ± 2.0 v. 20.8 ± 2.6, 82.7 ± 5.6% v. 86.1 ± 3.9% and 53.4 ± 5.9% v. 62.1 ± 4.7%, respectively). The number of BL tended to increase in non-PG cows compared with PG cows (10.0 ± 1.2 v. 7.8 ± 0.9; P = 0.078). The number of BL rated as Code 1 or 2 (IETS code) was higher in non-PG cows than in PG cows (76 v. 46; P &lt; 0.01). In ipsi-CL, contra-CL, and reg-CL, statistical differences were observed between the rate of Grade 1 to 2 OC and groups; the rate of Grade 1 to 2 OC was higher in ipsi-CL (93.8 v. 79.0 v. 86.7; P &lt; 0.01). These results suggested that the presence of CL for the duration of FG did not affect the rate of cleavage and BL formation but may have improved efficiency of embryo production by increasing quality of OC and BL.

The periovulatory endocrine milieu affects the uterine redox environment in beef cows.

BackgroundIn cattle, recent studies have shown positive associations between pre-ovulatory concentrations of estradiol (E2), progesterone (P4) at early diestrus and fertility. However, information on cellular and molecular mechanisms through which sex steroids regulate uterine function to support early pregnancy is lacking. Based on endometrial transcriptome data, objective was to compare function of the redox system in the bovine uterus in response to different periovulatory endocrine milieus.MethodsWe employed an animal model to control growth of the pre-ovulatory follicle and subsequent corpus luteum (CL). The large follicle-large CL group (LF-LCL, N = 42) presented greater levels of E2 on the day of GnRH treatment (D0; 2.94 vs. 1.27 pg/mL; P = 0.0007) and P4 at slaughter on D7 (3.71 vs. 2.62 ng/mL, P = 0.01), compared with the small follicle-small CL group (SF-SCL, N = 41). Endometrium and uterine washings (N = 9, per group) were collected for analyses of variables associated with the uterine redox system.ResultsThe SF-SCL group had lower endometrial catalase (0.5 vs. 0.79 U/mg protein, P < 0.001) and glutathione peroxidase (GPx; 2.0 vs. 2.43 nmol β-nicotinamide adenine dinucleotide phosphate reduced/min/mg protein, P = 0.04) activity, as well as higher lipid peroxidation (28.5 vs. 17.43 nmol malondialdehyde/mg of protein, P < 0.001) and superoxide dismutase (SOD) activity (44.77 vs. 37.76 U; P = 0.04). There were no differences in the endometrial reactive species (RS) or glutathione (GSH) concentrations between the groups. The uterine washing samples showed no differences in the concentrations of RS or GSH or in total SOD activity (P > 0.1). Additionally, catalase, GPx4, SOD1 and SOD2 gene expression was lower in the SF-SCL group than in the LF-LCL group.ConclusionsWe concluded that the intrauterine environment of cows from the LF-LCL group exhibited higher antioxidant activity than that of the cows from the SF-SCL group. We speculate that uterine receptivity and fertility are associated with an optimal redox environment, such as that present in the animals in the LF-LCL group.

Gonadotropin‐Releasing Hormone Administration to Dairy Cows without a Corpus Luteum 4 Weeks after Calving Increases Reproductive Performance

This field study investigated whether the administration of a single dose of gonadotropin-releasing hormone (GnRH) to dairy cows without a corpus luteum (CL) 4weeks after calving can improve reproductive performance. Holstein dairy cows underwent ultrasonography to assess the presence of ovarian structures at 29.2±5.2days post-partum, and cows were divided into two main groups based on the presence (CL group, n=230) or absence (non-CL group, n=460) of a CL. The non-CL group was further randomly divided into two subgroups based on the administration of GnRH (non-CL GnRH group, n=230) or no GnRH (non-CL control group, n=230). Subsets of cows from non-CL control (n=166) and non-CL GnRH (n=175) groups received a second ultrasonography at 44.5±5.4days post-partum to assess CL formation. The percentage of cows with CL at the second ultrasonography was greater in the non-CL GnRH group (70.9%) than in the non-CL control group (53.0%, p=0.0006). The hazard of the first post-partum insemination by 150days in milk (DIM) was higher in the CL group than in the non-CL control group (hazard ratio [HR]: 1.36, p=0.001). The probability of a pregnancy to the first insemination was higher in non-CL GnRH (odds ratio [OR]: 1.50, p=0.04) and CL groups (OR: 1.55, p=0.03) compared to the non-CL control group. Furthermore, the hazard of pregnancy by 210 DIM was higher in non-CL GnRH (HR: 1.30, p=0.01) and CL (HR: 1.51, p=0.0001) groups than in the non-CL control group. In conclusion, administration of GnRH to dairy cows without a CL 4weeks after calving was associated with an increase in ovulation and improved reproductive performance.

13 THE RELATIONSHIP BETWEEN EARLIER POSTPARTUM CYCLICITY AND SUBSEQUENT REPRODUCTIVE PERFORMANCE IN DAIRY COWS: A FIELD TRIAL

Earlier postpartum cyclicity based on hormonal analysis was related to the subsequent fertility, although it has not been clarified under field conditions. This field study examined the relationship between the detection of corpus luteum (CL) at the first post partum month and subsequent reproductive performance in dairy cows. Holstein dairy cows underwent ultrasonography (Sonoace 600 with 7.5 MHz linear-array transducer; Medison Co. Ltd., Korea) at 30 (±7) days postpartum to determine the existence of CL in ovaries and then divided into 2 groups based on the existence of CL; the CL group (n = 156) or the non-CL group (n = 281). At the same time of ultrasonography, all cows were scored for body condition. Cows received the normal herd reproductive management program including oestrous detection or synchronization, or synchronization of ovulation in the breeding period. Pregnancy was determined per rectum 60 days after artificial insemination by both ultrasonography and palpation. Reproductive performance data were collected for a minimum of 210 days postpartum. Reproduction data between the CL and non-CL groups were evaluated by t-test, chi-square test, or logistic regression using an SAS program (Version 9.1: SAS Institute, Inc., Cary, NC, USA). The proportion of cows with CL in ovaries at 30 (±7) days postpartum was 35.7% of all cows. The number of days to first insemination after calving was more delayed (P &lt; 0.0001) in the non-CL group (100.1 ± 2.7 days) than in the CL group (80.9 ± 2.9 days), while the pregnancy rate to first insemination did not differ (P &gt; 0.05) between the CL group (44.2%) and the non-CL group (45.9%). The CL group had higher pregnancy proportions (PP) within 60 (odds ratio, OR = 2.3; P = 0.0063), 90 (OR = 1.6; P = 0.04), and 120 days (OR = 1.5; P = 0.05) postpartum than the non-CL group; however, the PP within 150 or 210 days postpartum did not differ between the 2 groups (P &gt; 0.05). Besides, logistic analysis revealed that cows with body condition score (BCS) ≥3.00 were more likely to have CL (OR = 1.8; P = 0.015) compared with cows with BCS ≤ 2.75, while primiparous cows were less likely to have CL (OR = 0.6; P = 0.0085) compared with multiparous cows. In addition, cows that underwent ultrasonography later than 30 days postpartum were more likely to have CL (OR = 1.6; P = 0.0192) compared with cows that underwent ultrasonography earlier. In conclusion, the earlier cyclicity based on the detection of CL using ultrasonography at the first postpartum month, which might be in part due to a higher BCS, predicts an enhanced reproductive performance in dairy cows, particularly during the early and early-mid lactations. This work was supported by the research grant of the Chungbuk National University in 2010.

Ovarian stimulation with follicle-stimulating hormone under increasing or minimal concentration of progesterone in dairy cows

The objective of this study was to investigate the effect of the presence or absence of Corpus luteum (CL) on the follicular population during superstimulation in dairy cows (Holstein-Friesian cattle). Animals were divided into two groups as follows: (1) Growing CL group (G1): Cows (n = 7) received a total dose of 28 Armour units (AU) follicle-stimulating hormone (FSH) through the first 4 d (twice daily) after spontaneous ovulation (Day 0). (2) CL Absence group (G2): Cows (n = 10) received prostaglandin F 2α (PGF 2α) at 9 or 10 d after ovulation. After 36 h, all the follicles (larger than 5 mm) were aspirated (Day 0). The FSH treatment started 24 h after aspiration and continued for 4 d. The number of small (3 to <5 mm), medium (5 to <8 mm), and large (≥8 mm) follicles was examined on Days 1, 3, and 5 in all groups. Blood samples were collected daily for 5 d, and progesterone (P 4), estradiol (E 2), insulin-like growth factor-1 (IGF-1), and growth hormone (GH) in plasma were measured by enzyme immunoassays. The results showed that in G1, the P 4 level increased gradually from 0.5 ng/mL at Day 1 to 2 ng/mL at Day 5, whereas in G2, the P 4 level was completely below 0.5 ng/mL. All cows of the G2 group showed an increase of E 2 at Day 3 or Day 4 followed by an increase of IGF-1 within 24 h, while GH increased concomitantly with the E 2 increase in 8 of 10 trials. On the other hand, cows of the G1 group showed neither E 2 nor IGF-1 increase. Moreover, at the end of the treatment, the number of follicles in the G2 group was significantly increased compared with that of the G1 group (22.8 ± 2.0 vs. 11.6 ± 2.0). In conclusion, low P 4 level during FSH treatment enhanced multiple follicular growth and E 2 secretion, which was followed by increase of IGF-1 and GH. Therefore, the absence of the CL may play a critical role in the superovulation response by controlling the number of growing follicles.

Association between evaluation of the reproductive tract by various diagnostic tests and restoration of ovarian cyclicity in high-producing dairy cows

The uterine condition of clinically normal postpartum Holstein-Friesian dairy cows (n = 45) was evaluated once weekly (Weeks 3 to 7) by endometrial cytology, vaginal mucus collection device (VMCD), vaginoscopy, and ultrasonography to establish a relationship with postpartum resumption of ovulatory cycles. The time of first detection of the corpus luteum (CL) by ultrasonography and plasma progesterone concentration ≥1 ng/mL was recorded. By 49 d postpartum, 78% of the cows (n = 35) had resumed ovarian function (CL group), whereas the remainder (n = 10) had no CL (NCL group). There was a positive correlation between VMCD score and presence of fluid in the uterus in cows with a CL (P < 0.01) during Week 3 postpartum but no significant correlation in cows without a CL. Percentage of polymorphonuclear leukocytes (PMN%) was higher in the NCL group (mean ± SEM, 24.6 ± 9.4%) than in the CL group (11.7 ± 2.2%) during Week 5 postpartum (P < 0.05). The PMN% (4.5 ± 6.5%) and VMCD (0.5 ± 0.5) scores during Week 5 in cows ovulating by Day 28 were lower (P < 0.01) than the PMN% (15.0 ± 14.3%) and VMCD (1.1 ± 0.9) scores in those ovulating by Day 49. In conclusion, higher PMN% at 5 wk postpartum was associated with delayed resumption of ovarian cyclicity in high-producing dairy cows.

360 THE EFFECTS OF PROGESTERONE AND PRESENCE OF CORPUS LUTEUM ON SUPEROVULATION IN BEEF CATTLE (BOS TAURUS)

Bovine embryo transfer is a multi-million dollar industry that relies on hyperstimulation of cattle ovaries with exogenous follicle stimulating hormone (FSH). This process is variable among donors, breeds, and embryo transfer organizations. Historically, donor cattle are administered FSH (IM; &lt;15 to 50 mg) for 4 days and a luteolytic dose of prostaglandin F2� (PGF2�; IM; 25 mg twice daily) on Day 3. The presence of a functional corpus luteum (CL) is crucial for this protocol; however, there are embryo donor cattle that do not exhibit progesterone indicative of a functional CL. Experiment 1 was performed to determine whether the number of embryos and ovulations that could be generated from donors without a functional CL might be increased by progesterone supplementation via a vaginal insert (CIDR). Experiment 2 was performed to determine an optimal FSH (Sioux Biochemical, Inc., Sioux Center, IA, USA) dosage (high dose = 30 units, n = 9; low dose = 20 units, n = 7) for cattle without a functional CL. Embryos were collected nonsurgically by uterine lavage and were evaluated morphologically according to the IETS system. Progesterone concentrations were determined using an RIA on venous blood samples (10 mL). Ovulations (as evident by resulting CL) were counted using transrectal ultrasonography. Embryo, ovulation, and progesterone data were analyzed using ANOVA with SAS (SAS Institute, Inc., Cary, NC, USA), to evaluate the main effects of group and time (progesterone concentrations only). In Experiment 1, there was no difference in the number of high-quality embryos that were recovered from hyperstimulated (IM; 35 units) Angus-cross cattle that either had a functional CL (n = 12) or a CIDR insert (n = 12) following PGF2� injection. Progesterone concentrations were similar until Day 3 (Day 0 = injection of PGF2� and CIDR insertion). On Day 3, the CL group had higher (P &lt; 0.05) progesterone concentrations (3.5 � 0.4 ng/mL) than the CIDR group (1.5 � 0.1 ng/mL). Further, there was no difference between the average number of ovulations for the cows with a CL (22.5 � 2.4) or cows without a CL (22.8 � 1.5). In Experiment 2, there was no difference in the number of high-quality embryos recovered from cattle in the high-dose group (8.1 � 0.4) compared to cattle in the low dose group (5.7 � 1.3). There were no differences during the treatment period in progesterone concentrations between high- and low-dose groups. Although there was no statistical difference in the number of ovulations between cattle in the high dose group (20.2 � 2.5) compared to cattle in the low dose group (15.6 � 0.9), the higher FSH dose did yield over 30% more ovulations. Embryo collection from cattle that do not show signs of a CL can increase the number of offspring and embryos, and the genetic progress of otherwise unavailable cattle, thus benefiting the beef and beef cattle industries.

Retention of a functional corpus luteum and peripheral concentrations of 13,14-dihydro-15-keto-prostaglandin F2alpha following metestrus administration of Syncro-Mate-B.

This study was conducted to examine the effects of metestrus administration of SyncroMate-B (SMB) on PGF2alpha secretion and corpus luteum (CL) development. In a study replicated over 2 yr, cows were observed for spontaneous estrus in yr 1, and cows received an injection of 25 mg of PGF2alpha and were observed for subsequent estrus in yr 2. At standing estrus (estrus = d 1), cows were randomly allotted to receive either the standard SMB regimen (n = 40) on d 3 of the estrous cycle or no treatment (n = 8). Fifty percent (n = 20) of SMB-treated cows were administered PGF2alpha on d 10 of the estrous cycle 48 h prior to implant removal. Twice-daily blood samples were collected in the morning (AM) and evening (PM) from d 2 AM through d 14 AM of the treated estrous cycle and subsequently analyzed for progesterone (P4) and PGF2alpha metabolite (PGFM). Prior to statistical analysis, SMB- and SMB/PGF2alpha-treated cows were sorted according to P4 concentration at d 10 of the treated estrous cycle to either a CL functional group (P4 > or = 1 ng/mL; n = 20) or a CL nonfunctional group (P4 < 1 ng/mL; n = 17). Following d 10 AM administration of PGF2alpha, functional and nonfunctional groups were further subdivided based on treatment. The groups were as follows: untreated control cows (n = 8); SMB-treated cows retaining a functional CL (SMB-F; n = 8); SMB-treated cows with a nonfunctional CL (SMB-N; n = 11); SMB/PGF2alpha-treated cows retaining a functional CL (SMB/PG-F; n = 12); and SMB/PGF2alpha-treated cows with a nonfunctional CL (SMB/PG-N; n = 6). Of all SMB-treated cows, 54% retained a functional CL through d 10 AM of the treated estrous cycle. Mean serum P4 concentrations increased for cows in all groups until d 7, after which P4 concentrations increased for cows in SMB/PG-F, SMB-F, and control groups and decreased for cows in SMB/PG-N and SMB-N groups. Following PGF2alpha administration on d 10, mean serum P4 concentrations remained < 1 ng/mL for cows in SMB/PG-N and SMB-N groups, decreased to < 1 ng/mL for cows in the SMB/ PG-F group, and remained > 1 ng/mL for cows in SMB-F and control groups. Mean serum PGFM concentrations tended (P = .06) to increase in cows with nonfunctional CL compared with control cows on d 8 AM and were greater (P < .05) in cows with functional CL on d 8 PM through d 9 PM. These results indicate that retention of a functional rather than a nonfunctional CL following metestrus administration of SMB is dependent on a premature release of uterine PGF2alpha.

Control of inhibin production by dispersed human luteal cells in vitro.

The production of inhibin in vitro by dispersed cells from early to mid (Days 16-19) and late stage (Day 23) human corpus luteum (CL) was examined, and the effects of human chorionic gonadotrophin (hCG), follicle stimulating hormone (FSH), oestradiol and testosterone on inhibin production were determined. Corpora lutea from five subjects in the early to mid luteal stage and three subjects in late luteal stage were dispersed with enzymes and the luteal cells cultured in medium supplemented with 5% calf serum and either FSH (1, 10 or 100 ng mL-1), oestradiol-17 beta (2.5, 5 or 10 micrograms mL-1) or testosterone (0.25, 1 or 5 micrograms mL-1) with or without hCG (1 I.U. mL-1). Cells were cultured for 1 to 3 days without changes of medium, and the concentrations of progesterone, oestradiol and immunoreactive inhibin in the medium were measured by radioimmunoassay. Cells from both types of CL produced inhibin in vitro under basal conditions, but only cells from early to mid CLs responded to hCG with a significant increase in inhibin production. Both progesterone and oestradiol production were stimulated by hCG in both groups of CL. Inhibin concentrations in the cell cultures declined with time in culture, particularly in the late CL group, whereas the concentration of steroids increased. Neither FSH, oestradiol nor testosterone significantly changed inhibin production in either CL group. It was concluded that inhibin production by human luteal cells in vitro is influenced by the age of the CL, and is dependent on LH (hCG) but not on FSH or sex steroids.

Pregnancy response after estrus synchronization of cyclic cows with or without a corpus luteum prior to breeding

Twenty-nine dairy heifers and 27 postpartum beef cows were utilized to determine if pregnancy rate following synchronization and artificially insemination (AI) was influenced by the presence of a functional corpus luteum (CL) prior to the induced estrus. Cyclic females were grouped for treatment by administering two injections of prostaglandin F 2 α (PGF 2 α; 25 mg; i.m.) 11 d apart. Heifers and cows were randomly assigned to receive a norgestomet ear implant either immediately following the second PGF 2 α injection (non-CL group) or 3 d after the induced estrus (CL group). Implants were removed from both groups 9 d after they were inserted. At that time, the CL group received an injection of PGF 2 α (25 mg; i.m.) to induce luteolysis. Females in both groups were artificially inseminated 12 h after the detection of estrus. Estrous response after implant removal was 96% (23/24) for non-CL cows and 84% (27/32) for CL cows. Time from implant removal to initiation of estrus was reduced (P <0.01) for the non-CL cows (54 ± 6 h) and heifers (38 ± 6 h) when compared with CL cows (74 ± 7 h) and heifers (83 ± 6 h). Pregnancy rate, as determined by palpation per rectum 45 d after AI, was not influenced by treatment. Although the presence of a CL prior to induction of estrus with norgestomet and PGF 2 α affected the interval to estrus, the presence of a CL prior to breeding did not affect pregnancy rates significantly.