Spontaneous Luteolysis Research Articles

Characteristics of Pulses of 13,14-Dihydro-15-Keto-Prostaglandin F2alpha Before, During, and after Spontaneous Luteolysis and Temporal Intrapulse Relationships with Progesterone Concentrations in Cattle1

Pulses of the prostaglandin F2alpha (PGF) metabolite 13,14-dihydro-15-keto-PGF (PGFM) were compared among heifers that were in the preluteolytic, luteolytic, and postluteolytic periods (n = 7 or 8 heifers/period). Hourly blood sampling was done in 18-h sessions 15, 16, or 17 days after ovulation. Hourly sampling and statistical identification of a PGFM pulse allowed novel comparisons of PGFM pulses among the three periods. Each period had a similar number of PGFM pulses (2.3 +/- 0.2). The pulses were more prominent during the luteolytic period than during the other periods, as indicated by significantly greater concentration for the peak and amplitude between nadir and peak. Significantly more fluctuations that did not meet the definition of a pulse occurred at the beginning of the preluteolytic period and end of the postluteolytic period than during the luteolytic period. The same nadir ended a pulse and began the next pulse in 85% of adjacent pulses. Seven heifers were selected objectively, based on a progesterone concentration >5 ng/ml at Hour -3 (Hour 0 = peak of PGFM pulse) and a progressive decrease in progesterone from Hours -3 to 0. Progesterone increased (P < 0.03) between Hours 0 and 1, remained at a mean plateau at Hours 1 and 2, and then decreased. Results support the hypothesis of a transient intrapulse rebound in progesterone during an individual PGFM pulse, but only during the first portion of luteolysis. These findings should be considered in future proposals on the mechanisms involved in the effects of PGF on progesterone concentrations.

Is interleukin-1α a luteotrophic or luteolytic agent in cattle?

Cytokines are thought to regulate prostaglandin (PG) secretion in the bovine endometrium. However, there is no consensus about the role of interleukin-1alpha (IL1A) on PG secretion. The objective of this study was to examine the influence of IL1A on basal and interferon-tau (IFNT)-regulated PG in vitro secretion, as well its effects on PG secretion, progesterone (P(4)) output, and corpus luteum (CL) in vivo lifespan. Explants of bovine endometrium (days 16-17 of the estrous cycle or early pregnancy) were stimulated with IL1A (10 ng/ml), IFNT (30 ng/ml), or IL1A combined with IFN. IL1A alone strongly stimulated luteotrophic PGE(2) secretion by endometrial tissues of both pregnant and nonpregnant cows. IL1A also stimulated luteolytic PGF(2alpha) output in the late luteal phase. IFNT augmented the stimulatory effect of IL1A on PGE(2) secretion. In an in vivo experiment, saline or IL1A at different doses (0.001-10 microg/per animal) was applied to the uterine lumen on day 16 of the cycle. Only the highest dose of IL1A caused a temporal increase in PGF(2alpha) secretion, while it had no effect on P(4) secretion or CL lifespan. Application of 0.1 and 1 microg IL1A stimulated P(4) and PGE(2) output and prolonged the CL lifespan. Although IL1A may stimulate in vitro luteolytic PGF(2alpha) secretion during the estrous cycle, it only acts as a luteotrophic factor in vivo. IL1A increased luteotrophic PGE(2) and P(4) output, inhibiting spontaneous luteolysis. These luteotrophic effects may result in appropriate luteal development and function in cows during the estrous cycle and early pregnancy.

Concentrations of circulating hormones normalized to pulses of a prostaglandin F 2α metabolite during spontaneous luteolysis in mares

The temporal relationships between a pulse of 13,14-dihydro-15-keto-PGF 2α (PGFM) and the concentrations of circulating hormones during the luteolytic period were studied for 11 pulses in 11 mares (Equus caballus) using samples collected hourly. Mean PGFM pulses encompassed 4 h before to 4 h after the peak, and hormone data were normalized to the PGFM peak (Hour 0). Concentration of progesterone decreased (P < 0.05) between Hours –4 and –3 and continued to decrease linearly throughout the PGFM pulse. The concentrations of cortisol and prolactin increased (P < 0.004) during Hours –4 to 0 and decreased (P < 0.002) during Hours 0 to 4. Estradiol concentration increased (P < 0.02) during Hours –4 to 0 but did not change significantly after Hour 0. Concentrations of follicle-stimulating hormone and luteinizing hormone did not change significantly during the PGFM pulse, and the oxytocin results were equivocal. Percentage of corpus luteum area with color-Doppler signals of blood flow did not change significantly between Hours –4 and 0 and first began to decrease (P < 0.004) between Hours 0 and 2. Results demonstrated that concentrations of progesterone decreased linearly during a PGFM pulse, and cortisol, prolactin, and estradiol increased during the ascending portion of the pulse. The progesterone and gonadotropin results supported the hypothesis that the initial progesterone and gonadotropin increases that have been reported to occur in response to a single bolus luteolytic treatment with prostaglandin F 2α do not occur in response to the natural secretion of prostaglandin F 2α.

Follicular Population during the Oestrous Cycle in Nili-Ravi Buffaloes Undergoing Spontaneous and PGF2a Induced Luteolysis

The objective of this study was to compare the follicular population during spontaneous and PGF 2α induced oestrous cycles in Nili-Ravi buffaloes. In Exp.1, (n = 13 oestrous cycles) follicular population was monitored using ultrasonography on alternate days. Buffaloes were monitored for ovarian follicles from day 0 (first oestrus) until next oestrus. These animals were observed for oestrus twice daily using a teaser bull. Of 12 oestrous cycles, 9 (75%) had two waves of follicular activity and only 3 (25%) had three waves during the oestrous cycle. The mean number of small, medium and large follicles among various days of the oestrous cycle between two and three waves of follicular development were not significantly different (p>0.05). In Exp. 2, follicular population 3 days before oestrus was compared in buffaloes undergoing spontaneous (n = 12 oestrous cycles) and PGF 2α induced (n = 6) luteolysis. The mean number of small and large follicles increased (p<0.05) and the number of medium follicles decreased (p<0.05) during the 3 days before oestrus in buffaloes undergoing induced luteolysis as compared to those with spontaneous luteolysis. These results showed that the mean number of small, medium and large follicles among various days of the oestrous cycle were similar between the two and three waves of follicular development, and three days before oestrous the number of small, medium and large follicles altered due to induced luteolysis on day 9, compared to those with spontaneous luteolysis.

Dynamic Changes in Gene Expression that Occur during the Period of Spontaneous Functional Regression in the Rhesus Macaque Corpus Luteum

Luteolysis of the corpus luteum (CL) during nonfertile cycles involves a cessation of progesterone (P4) synthesis (functional regression) and subsequent structural remodeling. The molecular processes responsible for initiation of luteal regression in the primate CL are poorly defined. Therefore, a genomic approach was used to systematically identify differentially expressed genes in the rhesus macaque CL during spontaneous luteolysis. CL were collected before [d 10-11 after LH surge, mid-late (ML) stage] or during (d 14-16, late stage) functional regression. Based on P4 levels, late-stage CL were subdivided into functional-late (serum P4 > 1.5 ng/ml) and functionally regressed late (FRL) (serum P4 < 0.5 ng/ml) groups (n = 4 CL per group). Total RNA was isolated, labeled, and hybridized to Affymetrix genome microarrays that contain elements representing the entire rhesus macaque transcriptome. With the ML stage serving as the baseline, there were 681 differentially expressed transcripts (>2-fold change; P < 0.05) that could be categorized into three primary patterns of expression: 1) increasing from ML through FRL; 2) decreasing from ML through FRL; and 3) increasing ML to functional late, followed by a decrease in FRL. Ontology analysis revealed potential mechanisms and pathways associated with functional and/or structural regression of the macaque CL. Quantitative real-time PCR was used to validate microarray expression patterns of 13 genes with the results being consistent between the two methodologies. Protein levels were found to parallel mRNA profiles in four of five differentially expressed genes analyzed by Western blot. Thus, this database will facilitate the identification of mechanisms involved in primate luteal regression.

Is Interleukin 1α a Luteotropic or Luteolytic Agent in Cattle?

The genes for interleukin (IL)1α and its functional receptors are present in the bovine female reproductive tract during the estrous cycle. IL1α stimulates prostaglandin (PG) production in cultured bovine endometrial cells. The target of IL1α for stimulating both luteotropic PGE2 and luteolytic PGF2α production is the stromal cells. The stimulatory effect of IL1α on PG production is mediated by differently modulating of phospholipase A2 and expression of respective PG synthases in the stromal cells. Moreover, IL1α stimulates PG secretion in the steroidogenic cells of bovine corpus luteum (CL) in vitro without any effect on progesterone (P4) production. However, physiological meaning of these findings is unclear. To determine physiological role of IL1α, it's influence on PG secretion by the cultured bovine endometrial tissues obtained from the cyclic or early pregnant cows, and the effect of IL1α on PG, progesterone (P4) output and the CL lifespan in vivo were examined. In Exp. 1, effects of IL1α with/without an embryonic signal (interferone-τ, INFτ) on PG secretion by cultured endometrial tissues were examined. Explants of endometrium (30-40 mg) obtained from the cows at Day 16/17 of the estrous cycle (Group 1; n=7) or early pregnancy (Day 16/17 after insemination, Group 2; n=6) were stimulated for 18-h with: saline (control), TNFα (10 ng/ml), IL1α (10 ng/ml), IFNα(30 ng/ml) or IL1α combination with IFNτ. Concentrations of PGE2 and PGF2α in the culture medium were measured by EIA. IL1α alone stimulated secretion of luteotropic PGE2 in endometrial tissues of both Group 1 and Group 2 (more than 200% of the control, P<0.01). IL1α also stimulated luteolytic PGF2α output from endometrial explants, but only during the late luteal phase (150%, P<0.05). This stimulatory effect of IL1α on PGF2α secretion was inhibited by INFτ (P<0.05). Moreover, IFNτ augmented the stimulatory effect of IL1a on PGE2 secretion (P<0.05). In Exp. 2, the effects of IL1α administrated into the uterine lumen on PGF2α, PGE2 and P4 secretions, and on the lifespan of the CL were determined in vivo. Saline (5 ml, control; n=5) or IL1α at different doses: 0,001 (n=3); 0.01 (n=5); 0.1 (n=5); 1 (n= 5) and 10 μg (n=3) in 5 ml of saline were applied at Day 16 of the estrous cycle. P4, PGE2 and PGFM (stable metabolite of PGF2α) concentrations in peripheral blood were measured by EIA. IL1α at doses 0.1 and 1 μg stimulated P4 and PGE2 output (P<0.01), and simultaneously inhibited spontaneous luteolysis with prolonging the lifespan of CL (P<0.01). Although only the lowest dose of IL1α (0.001 μg) caused a temporal increase in PGFM concentration during the experiment (until 12-h after application; P<0.05), any doses of IL1α had effect on the CL lifespan. Concluding, IL1α may play some important autocrine or paracrine roles in regulating PG production in the endometrium and CL during the estrous cycle and early pregnancy in cattle. IL1α increased P4 and luteotropic PGE2 concentrations in blood plasma that may result in appropriate development and function of the bovine CL during the estrous cycle. These luteotropic effects of IL1α during pregnancy may support the early embryo development and implantation, and may play a supporting role in the maternal recognition of pregnancy in cattle. Supported by Grant 2P06K02529.

Prostaglandin F2 increases endothelial nitric oxide synthase in the periphery of the bovine corpus luteum: the possible regulation of blood flow at an early stage of luteolysis

Prostaglandin F(2)(alpha) (PGF(2)(alpha)) released from the uterus causes alterations in luteal blood flow, reduces progesterone secretion, and induces luteolysis in the bovine corpus luteum (CL). We have recently discovered that luteal blood flow in the periphery of the mature CL acutely increases coincidently with pulsatile increases in a metabolite of PGF(2)(alpha) (PGFM). In this study, we characterized changes in regional luteal blood flow together with regional alterations in endothelial nitric oxide synthase (eNOS) expression during spontaneous luteolysis and in response to PGF(2)(alpha). Smooth muscle actin-positive blood vessels larger than 20 microm were observed mainly in the periphery of mature CL. PGF(2)(alpha) receptor was localized to luteal cells and large blood vessels in the periphery of mid-CL. PGF(2)(alpha) acutely stimulated eNOS expression in the periphery but not in the center of mature CL. Injection of the NO donor S-nitroso-N-acetylpenicillamine into CL induced an acute increase in luteal blood flow and shortened the estrous cycle. In contrast, injection of the NOS inhibitor l-NAME into CL completely suppressed the acute increase in luteal blood flow induced by PGF(2)(alpha) and delayed the onset of luteolysis. In conclusion, PGF(2)(alpha) has a site-restricted action depending on not only luteal phase but also the region in the CL. PGF(2)(alpha) stimulates eNOS expression, vasodilation of blood vessels, and increased luteal blood flow in periphery of mature CL. Furthermore, the increased blood flow is mediated by NO, suggesting that the acute increase in peripheral blood flow to CL is one of the first physiological indicators of NO action in response to PGF(2)(alpha).

Intrauterine infusion of BQ-610, an endothelin type A receptor antagonist, delays luteolysis in dairy heifers

Three separate in vivo experiments were conducted to evaluate the putative role of endothelin-1 (ET-1) during luteal regression in heifers. In Experiment 1, a single intraluteal injection of 500μg BQ-610 [(N,N-hexamethylene) carbamoyl-Leu-d-Trp (CHO)-d-Trp], a highly specific endothelin A (ETA) receptor antagonist, did not diminish the decline in plasma progesterone following a single exogenous injection of 25mg prostaglandin F2 alpha (PGF2α) administered at midcycle of the estrous cycle. In Experiment 2, six intrauterine infusions of 500μg BQ-610 given every 12h on days 16–18 delayed spontaneous luteolysis, as evidenced by an extended elevation (P=0.054) of plasma progesterone concentration. In Experiment 3, heifers were administered six intrauterine infusions of BQ-610 or saline on days 16–19, and peripheral blood samples were collected from day 11 to 16 (before infusion), hourly on days 16–19 (during infusion), and on days 20–25 (after infusion). BQ-610 treated heifers had markedly higher (P<0.0001) levels of plasma progesterone compared with saline controls, and this effect was most notable during the infusion period (treatment by period interaction; P≤0.05). Heifers infused with BQ-610 also had higher progesterone levels on day 21 (treatment by time interaction; P≤0.05). Mean plasma concentrations of 13,14-dihydro-15-keto-PGF2α (PGFM), the primary metabolite of PGF2α, were measured in the samples collected hourly and were not different (P≥0.05) between treatments. These results indicate that the in vivo antagonism of the ETA receptor can delay functional luteolysis, and supports the theory that ET-1 regulates luteal function in ruminants.

Positive Association, in Local Release, of Luteal Oxytocin with Endothelin 1 and Prostaglandin F2alpha During Spontaneous Luteolysis in the Cow: A Possible Intermediatory Role for Luteolytic Cascade Within the Corpus Luteum1

Luteolysis is caused by a pulsatile release of prostaglandin F(2alpha) (PGF(2alpha)) from the uterus in ruminants, and a positive feedback between endometrial PGF(2alpha) and luteal oxytocin (OXT) has a physiologic role in the promotion of luteolysis. The bovine corpus luteum (CL) produces vasoactive substances, such as endothelin 1 (EDN1) and angiotensin II (Ang II), that mediate and progress luteolysis. We hypothesized that luteal OXT has an additive function to ensure the CL regression with EDN1 and Ang II, and that it has an active role in the luteolytic cascade in the cow. Thus, the aim of the present study was to observe real-time changes in the local secretion of luteal OXT and to determine its relationship with other local mediators of luteolysis. Microdialysis system (MDS) capillary membranes were implanted surgically into each CL of six cyclic Holstein cows (18 lines total among the six cows) on Day 15 (estrus == Day 0) of the estrous cycle. Simultaneously, catheters were implanted to collect ovarian venous plasma ipsilateral to the CL. Although the basal secretion of OXT by luteal tissue was maintained during the experimental period, the intraluteal PGF(2alpha) secretion gradually increased up to 300% from 24 h after the onset of luteolysis (0 h; time in which progesterone started to decrease). In each MDS line (microenvironment) within the CL, the local releasing profiles of OXT were positively associated with PGF(2alpha) and EDN1 within the CL in all 18 MDS lines implanted in the six CLs (OXT vs. PGF(2alpha), 50.0%; OXT vs. EDN1, 72.2%; P < 0.05). On the other hand, the intraluteal OXT was weakly related to Ang II (OXT vs. Ang II, 27.7%). In the ovarian vein, the peak concentration of PGF(2alpha) increased significantly when the peak of PGF(2alpha) coincided with the peak of OXT after the onset of spontaneous luteolysis (P < 0.05). In conclusion, intraluteal OXT may locally modulate secretion of vasoactive substances, particularly EDN1 and PGF(2alpha) within the CL, and thus might be one of the luteal mediators of spontaneous luteolysis in the cow.

PREOVULATORY CHANGES AND OVULATION IN CATTLE UNDERGOING SPONTANEOUS OR CLOPROSTENOL-INDUCED LUTEOLYSIS

The follicular population, diameter of the ovulatory and subordinate follicles, corpus luteum (CL) size, concentration of progesterone and estradiol-17p were studied following spontaneous or cloprostenol-induced luteolysis. A total of 14 heifers received cloprostenol treatment on Day 9-11 of the cycle to synchronize their estrus. Subsequently, they were divided into two groups, one group which was allowed to undergo spontaneous luteolysis and the other group in which estrus was induced from days 9 to 12. In the induced-group, transrectal ultrasonography were performed daily started two days prior to injection until the onset of estrus. In the spontaneous-group, ultrasonography was done daily from day 15 until the onset of estrus. In both groups scanning were performed every 4 h from the onset of estrus until ovulation was ascertained. Small (SF, 2-4 mm), medium (MF.5-9 mm ) and large (LF,&gt;9mm) size follicles were recorded. The diameter of largest and subordinate follicles were measured and blood were drawn from jugular vein at approximately around scanning and the plasma were used for measurement of progesterone (P4) and estradiol-17p (E2) concentration. There was no different in term of number of SF, MF and LF (P&gt;0.05) between the two groups. Similarly, no effect of side (left vs. right ovary) and CL position (ipsi- vs contralateral to the ovary) was found (P&gt;0.05). However, it was demonstrated that mean number of ovulatory follicles was higher (PO.01) in the spontaneously ovulating group while the regressing-CL size was larger in the cloprostenol induced animals (P

Temporal Associations among Pulses of 13,14-Dihydro-15-keto-PGF2alpha, Luteal Blood Flow, and Luteolysis in Cattle1

Luteal blood flow was studied in heifers by transrectal color-Doppler ultrasound. Data were normalized to the decrease in plasma progesterone to <1 ng/ml (Day 0 or Hour 0). Blood flow in the corpus luteum (CL) was estimated by the percentage of CL area with color flow signals. Systemic prostaglandin F2alpha (PGF) treatment (25 mg; n=4) resulted in a transient increase in CL blood flow during the initial portion of the induced decrease in progesterone. Intrauterine treatment (1 or 2 mg) was done to preclude hypothetical secondary effects of systemic treatment. Heifers were grouped into responders (luteolysis; n=3) and nonresponders (n=5). Blood flow increased transiently in both groups; induction of increased blood flow did not assure the occurrence of luteolysis. A transient increase in CL blood flow was not detected in association with spontaneous luteolysis when examinations were done every 12 h (n=6) or 24 h (n=10). The role of PGF pulses was studied by examinations every hour during a 12-h window each day during expected spontaneous luteolysis. At least one pulse of 13,14-dihydro-15-keto-PGF2alpha (PGFM) was identified in each of six heifers during the luteolytic period (Hours -48 to -1). Blood flow increased (P<0.02) during the 3-h ascending portion of the PGFM pulse, remained elevated for 2 h after the PGFM peak, and then decreased (P<0.03) to baseline. Results supported the hypothesis that CL blood flow increased and decreased with individual PGFM pulses during spontaneous luteolysis.

Transgene Insertion on Mouse Chromosome 6 Impairs Function of the Uterine Cervix and Causes Failure of Parturition1

The molecular mechanisms controlling the initiation of parturition remain largely undefined. We report a new animal model in which parturition does not occur. A line of mice expressing a human apolipoprotein B (APOB) gene fail to deliver their young if the transgene is present in homozygous (Tg/Tg), but not in heterozygous (Tg/Wt), form. Cloning and mapping of the transgene insertion locus indicate that 10 copies of the 80-kilobase APOB genomic fragment inserted into mouse chromosome 6 result in a small, 390-base pair deletion of mouse genomic DNA. Nine other lines expressing the transgene have normal labor, suggesting that transgene insertion in this mutant line disrupted a mouse gene crucial for successful parturition. The pathophysiology of parturition failure in these animals was defined using physiological, endocrinological, and morphological techniques. Results indicate that luteolysis occurs in Tg/Tg mice but is delayed by 1 day. Delivery did not occur in mutant mice at term after spontaneous luteolysis or even after removal of progesterone action by ovariectomy or antiprogestin treatment. Biomechanical and functional studies of the uterus and cervix revealed that the primary cause of failed parturition in Tg/Tg mice was not inadequate uterine contractions of labor but, rather, a rigid, inelastic cervix at term that was abnormally rich in neutrophils and tissue monocytes. Characterization of the transgene insertional mutant, Tg/Tg, indicates that progesterone withdrawal is insufficient to complete parturition in the presence of inadequate cervical ripening at term.

Blood flow: A key regulatory component of corpus luteum function in the cow

Prostaglandin F2α (PGF 2α) is the primary luteolysin in the cow. During the early luteal phase, the corpus luteum (CL) is resistant to the luteolytic effect of PGF 2α. Once mature, the CL becomes responsive to PGF 2α and undergoes luteal regression. These actions of PGF 2α coincide with changes in luteal blood flow (BF): PGF 2α has no effect on BF in the early CL, but acutely increases BF in the peripheral vasculature of the mature CL within 30 min of PGF 2α injection. During spontaneous luteolysis, luteal BF increases on Days 17–18 of the estrous cycle, prior to any decrease in plasma progesterone (P). The increase in luteal BF is synchronous with an increase in plasma PGFM levels, suggesting that pulsatile release of PGF 2α from uterus stimulates the increase in luteal BF. Serial biopsies of these CL showed that mRNA expression for endothelial nitric oxide synthase (eNOS) together with endothelin-1 (ET-1) and angiotensin converting enzyme (ACE) increases on Days 17–18 when the luteal BF is elevated. On Day 19 when plasma P level firstly decreases, eNOS mRNA returns to the basal level whereas ET-1 and ACE mRNA remains elevated. Cyclooxygenase-2 (COX-2) mRNA expression increases on Day 19. In support of these data, an in vivo microdialysis study revealed that luteal ET-1 and angiotensin II (Ang II) secretion increases and precedes PGF 2α secretion during spontaneous luteolysis. In conclusion, we show for the first time that an acute increase of BF occurs in the peripheral vasculature of the mature CL together with increases in eNOS expression and ET-1 and Ang II secretion in the CL during the early stages of luteolysis in the cow. We propose that the increase in luteal BF may be induced by NO from large arterioles surrounding the CL, and simultaneously uterine or exogenous PGF 2α directly increases ET-1 and Ang II secretion from endothelial cells of microcapillary vessels within the CL, thereby suppressing P secretion by luteal cells. Taken together, our results indicate that an acute increase in luteal BF occurs as a first step of luteolysis in response to PGF 2α. Therefore, local BF plays a key role to initiate luteal regression in the cow.

212. Follistatin secretion by the ovary is not directly related to PGF2α induced luteolysis in the ewe

Follistatin is a monomeric glycoprotein, expressed ubiquitously, and found in greatest levels in ovarian tissue. Our previous studies have demonstrated that ovarian venous follistatin levels are elevated at the same time as spontaneous luteolysis in merino ewes, but remain constant in circulating and uterine plasma across oestrous. It was hypothesized that the follistatin elevation may be part of functional luteolysis in the ovary. A cross-sectional study of ovarian venous follistatin during luteolysis was performed using merino ewes, which were synchronized with intravaginal sponges. Luteolysis was induced 10 days after oestrus using a PGF2α (IM) injection. Circulating (jugular and arterial), uterine venous and ovarian venous (luteinized and contralateral) blood samples were taken under general anaesthesia at times 0, 6, 12, 24, 36, 48 and &gt;48 h (54–71 h) after the PGF2α injection. All samples were assayed for progesterone by RIA and follistatin using a specific follistatin antiserum (#204) in a competitive ELISA with rhFS-288 used as standard. Progesterone concentrations in the venous plasma from the luteinized ovaries plasma were significantly higher than those in the contralateral ovarian and circulating plasma. As expected progesterone concentrations began declining within 6 h and continued until 24 h after PGF2α. Mean circulating plasma follistatin concentrations were similar throughout the experiment with overall means of 9.9 ± 0.5 ng/mL. Both luteinized and contralateral ovarian venous plasma follistatin concentrations (11.8 ± 3.4 ng/mL) were not significantly different from circulating concentrations. However at 48 h ovarian venous follistatin concentrations were significantly elevated (22.5 ± 2.3 ng/mL) and remained elevated for at least 71 h (29.5 ± 7.9 ng/mL). This experiment confirms our previous work that both luteinized and contralateral ovarian venous follistatin concentrations do elevate concurrently with each other but that the elevation in follistatin is not directly related to functional luteolysis as induced by PGF2α.

Endothelin-1 within the Corpus Luteum during Spontaneous Luteolysis in the Cow: Local Interaction with Prostaglandin F2?? and Angiotensin II

It is generally accepted that endothelin-1 (ET-1) is involved in the regression of the corpus luteum (CL) in cows, but there are few in vivo data available on the local release of vasoconstrictors, including ET-1. Thus, we aimed to determine in detail the local secretion of ET-1, angiotensin II (Ang II) and prostaglandin F2alpha (PGF2alpha) within the CL during spontaneous luteolysis in the cow. To observe real-time dynamics of the releasing profile of CL-derived factors, a microdialysis system was surgically implanted in the CL on day 15 of the estrous cycle and continuously perfused with Ringer's solution. Local secretion of ET-1, Ang II and PGF2alpha increased immediately after the onset of luteolysis (the time point when progesterone release started to decrease within the CL) and was maintained at high levels. A positive relationship was observed in intra-luteal changes among ET-1, Ang II and PGF2alpha release. This is the first real-time and in vivo evidence that the secretion of ET-1 together with Ang II and PGF2alpha immediately increases within the CL after the onset of spontaneous luteolysis. Consequently, we suggest that the activation of a local positive feedback mechanism among ET-1, Ang II and PGF2alpha might play a functional role in the paracrine modulation of luteolytic cascade and, simultaneously, the elevated ET-1 with Ang II and PGF2alpha should induce a strong vasoconstriction, thereby reducing the blood supplying the CL during spontaneous luteolysis.

Effect of dexcloprostenol on endogenous prostaglandin F 2α release in dairy heifers

Cloprostenol was previously believed to be unable to release endogenous prostaglandin F 2α (PGF 2α) when administered during early bovine diestrus. A prostaglandin release is, however, seen in late diestrus. The aim of this study is to find out whether dexcloprostoenol (containing the only biologically active isomer, d-isomer, of cloprostenol) induces endogenous PGF 2α release during early and late diestrus. Twelve heifers of the Finnish Ayrshire breed were allocated into two equal groups. Their estrous cycles were synchronized with dexcloprostenol. A further luteolysis was induced with 0.15 mg of dexcloprostenol either on Day 7 (group D7 or early diestrus) or on Day 14 (group D14 or late diestrus) after ovulation. Blood for progesterone and the PGF 2α metabolite 15-ketodihydro-PGF 2α determinations was collected immediately before dexcloprostenol treatment and thereafter every second hour for 48 h. Five of the six heifers in both groups showed significantly increased blood levels of 15-ketodihydro-PGF 2α at some time during the 48-h experimental period. The intervals from treatment to the first significant increases of the PGF 2α metabolite were 32.8 ± 2.3 h (min. 30 h, max. 36 h) and 20.0 ± 4.2 h (min. 14 h, max. 24 h) in groups D7 and D14, respectively ( P < 0.01). We have concluded that dexcloprostenol induced endogenous PGF 2α release in most cases, regardless the time of its administration (early or late diestrus). This release, however, differs from that observed during spontaneous luteolysis.

Real-time dynamics of prostaglandin F2alpha release from uterus and corpus luteum during spontaneous luteolysis in the cow.

Prostaglandin (PG) F2alpha released from the uterus in a pulsatile fashion is essential to induce regression of the corpus luteum (CL) in the cow. In addition to the uterus, the CL has also been recognized as a site of PGF(2alpha) production. Therefore, this study aimed to determine the detailed dynamics of the releasing profile of CL-derived PGF2alpha together with uterus-derived PGF2alpha during spontaneous luteolysis in the cow. Non-lactating Holstein cows (n = 6) were surgically implanted with a microdialysis system (MDS) on day 15 (oestrus = day 0) of the oestrous cycle. Simultaneously, catheters were implanted to collect ovarian venous plasma ipsilateral to the CL as well as jugular venous plasma. The concentrations of PGF2alpha, 13,14-dihydro-15-keto-PGF2alpha (PGFM) and progesterone in the MDS and plasma samples were determined by enzyme immunoassays. The intra-luteal PGF2alpha secretion slightly increased after the onset of luteolysis (0 h) and drastically increased from 24 h, and was maintained at high levels towards the following oestrus. Furthermore, PGF2alpha was released from the CL into the ovarian vein in a pulsatile manner during spontaneous luteolysis. Also, the fact that intra-luteal secretion of PGF2alpha and PGFM showed a positive correlation indicates the existence of a local metabolic pathway for PGF2alpha in the CL. In conclusion, the present study clarified the real-time dynamics of uterus-derived PGF2alpha and CL-derived PGF2alpha during spontaneous luteolysis in the cow, and gives the first in vivo evidence that the CL releases PGF2alpha during spontaneous luteolysis in the cow. Although the physiological relevance of CL-derived PGF2alpha appears to be restricted to a local role as an autocrine/paracrine factor in the CL, overall results support the concept that the local release of PGF2alpha within the regressing CL amplifies the luteolytic action of PGF2alpha from the uterus.

Real-time relationships in intraluteal release among Prostaglandin F2alpha, endothelin-1, and angiotensin II during spontaneous luteolysis in the cow.

It is well known that prostaglandin F(2alpha) (PGF(2alpha)) is a physiological luteolysine, and that its pulsatile release from the endometrium is a luteolytic signal in many species. There is now clear evidence that the vasoactive peptides endothelin-1 (ET-1) and angiotensin II (Ang II) interact with PGF(2alpha) in the luteolytic cascade during PGF(2alpha)-induced luteolysis in the cow. Thus, we investigated the local secretion of PGF(2alpha), ET-1, and Ang II in the corpus luteum (CL) and their real-time relationships during spontaneous luteolysis in the cow. For this purpose, an in vivo microdialysis system (MDS) implanted in the CL was utilized to observe local secretion changes within the CL microenvironment. Each CL of cyclic Holstein cows (n = 6) was surgically implanted with MDS capillary membranes (18 lines/6 cows) on Day 15 (estrus = Day 0) of the estrous cycle. The concentrations of PGF(2alpha), ET-1, Ang II, and progesterone (P) in the MDS samples were determined by enzyme immunoassays. The intraluteal PGF(2alpha) secretion slightly increased from 12 h after the onset of luteolysis (0 h) and drastically increased (by about 300%) from 24 h. Intraluteal ET-1 secretion increased from 12 h. Intraluteal Ang II secretion was elevated from 0 h and was maintained at high levels (about 180%) toward estrus. In each MDS lines (in the same microenvironment) within the regressing CL, the local releasing profiles of PGF(2alpha), ET-1, and Ang II CL positively correlated with each other (P < 0.05) at high proportions in 18 MDS lines (PGF(2alpha) vs. ET-1, 44.4%; PGF(2alpha) vs. Ang II, 55.6%; ET-1 vs. Ang II, 38.9%). In contrast, there was no clear relationship among these substances released into different MDS lines implanted in the same CL (with different microenvironments). In conclusion, we propose that the increase of PGF(2alpha), ET-1, and Ang II within the CL during luteolysis is a common phenomenon for both PGF(2alpha)-induced and spontaneous luteolysis. Moreover, this study illustrated the in vivo relationships in intraluteal release among PGF(2alpha), ET-1, and Ang II during spontaneous luteolysis in the cow. The data suggest that these vasoactive substances may interact with each other in a local positive feedback manner to activate their secretion in the regressing CL, thus accelerating and completing luteolysis.

Expression patterns of cytokines, p53 and nitric oxide synthase isoenzymes in corpora lutea of pseudopregnant rabbits during spontaneous luteolysis.

The gene expressions for macrophage chemoattractant protein-1 (MCP-1), interleukin (IL)-1 beta, IL-2 and p53 were examined by semi-quantitative RT-PCR in corpora lutea (CL) of rabbits during spontaneous luteolysis at days 13, 15, 18 and 22 of pseudopregnancy. In the same luteal tissue, total activity of nitric oxide (NO) synthase (NOS) and genes for both endothelial (eNOS) and inducible (iNOS) isoforms were also analysed. From day 13 to 15, MCP-1 and IL-1 beta mRNA levels rose (P < or = 0.01) almost 2-fold, and the transcript for p53 almost 8-fold, but then all dropped (P < or = 0.05) from day 18 onward. IL-2 mRNA abundance was higher (P < or = 0.01) on day 13 and then gradually declined. During luteolysis, eNOS mRNA decreased 40% (P < or = 0.05) by day 15, but thereafter remained unchanged, while iNOS mRNA was barely detectable and did not show any clear age-related pattern throughout the late luteal stages. Total NOS activity progressively increased (P < or = 0.01) from day 13 to 18 of pseudopregnancy and then dropped to the lowest (P < or = 0.01) levels on day 22. Luteal progesterone content also declined during CL regression from 411 to 17 pg/mg found on days 13 and 22 respectively, in parallel with the decrease in blood progesterone concentrations. These data further support a physiological role of NO as modulator of luteal demise in rabbits. Locally, luteal cytokines may be involved in the up-regulation of NOS activity, while downstream NO may inhibit steroroidogenesis and induce expression of p53 gene after removal of the protective action of progesterone.

Intervalos do início e do final do estro à ovulação em vacas das raças Gir e Guzerá após luteólise natural ou induzida por prostaglandina

A duração do estro e os intervalos início do estro-ovulação e fim do estro-ovulação após luteólise induzida ou espontânea, e os intervalos administração de prostaglandina-início do estro e administração de prostaglandina-ovulação foram avaliados em 35 vacas da raça Gir e 12 da raça Guzerá, no inverno e no verão de dois anos consecutivos. A duração do estro não diferiu (P&gt;0,05) entre as raças Gir (12,13±0,47h) e Guzerá (12,30±0,63h) e não foi influenciada (P&gt;0,05) pelo tipo de luteólise e pela estação do ano. Os intervalos do início e final do estro à ovulação não diferiram (P&gt;0,05) entre as raças Gir (29,30± 0,96h e 17,28± 0,93h, respectivamente) e Guzerá (28,33± 1,26h e 16,04± 1,21h, respectivamente) e foram similares (P&gt;0,05) entre os estros induzido e natural, e entre estações. O intervalo aplicação de prostaglandina-início do estro foi menor (P&lt;0,05) na raça Gir (64,68± 2,41h) do que na Guzerá (73,47± 3,45h) e não diferiu (P&gt;0,05) entre as estações. O intervalo administração de prostaglandina-ovulação foi similar (P&gt;0,05) entre as raças Gir e Guzerá (96,62± 4,25h e 104,01± 6,51h, respectivamente). Observou-se ampla variação no intervalo prostaglandina-início do estro e alta correlação entre este e o momento de ovulação, sugerindo que as sincronizações do estro e da ovulação não foram suficientemente precisas para permitir a adoção de IA programada.