Comparison of luteolytic effectiveness of several prostaglandin analogs in heifers and relative binding affinity for bovine luteal prostaglandin binding sites

The relative binding affinities for both the prostaglandin (PG)E₁ and PGF_2α specific bovine luteal binding sites were determined for five PGE and fourteen PGF derivatives and analogs. Relative binding affinity was determined using membranes prepared from bovine corpora luteal (CL) obtained from the slaughterhouse. The parent structure of the analog was a dominant feature in determining the affinity for the respective PG binding site. Luteolysis was determined in cattle following intramuscular injection of various doses of prostaglandin once between days 6 and 14 after estrus and measuring CL regression by ovarian palpation per rectum, interval between injection and return to estrus and duration of the subsequent estrous cycle. A dose which was luteolytic was established for each of eight PGF-type compounds, and a dose which was not luteolytic was also established. There appeared to be limited association between the relative affinity for the PGF_2α specific site and the estimated luteolytic dose range of these PGF analogs when tested in cattle. Differences in luteolytic potency for the compounds tested could not be explained by differences in binding affinity. Differences in metabolism and absorption may also be important in the determination of potency.     

Prostaglandin E1 and F2alpha specific binding in bovine corpora lutea: comparison with luteolytic effects.

Preliminary characterization indicated the presence of separate prostaglandin (PG)E1 and (PG)F2alpha binding sites in membrane fractions prepared from bovine corpora lutea. These differ in the rate and temperature dependence of the specific binding. Equilibrium binding data indicate the apparent dissociation constants as 1.32 x 10(-9)M and 1.1 x 10(-8)M for PGE1 and PGF2alpha, respectively. Competition of several natural prostaglandins for the PGE1 and PGF2alpha bovine luteal specific binding sites indicates specificity for the 9-keto or 9alpha-hydroxyl moiety, respectively. Differences in relative ability to inhibit 3H-PG binding were found due to sensitivity to the absence or presence of the 5, 6-cis-double bond as well. Bovine luteal function was affected following treatment of heifers with 25 mg PGF2alpha as measured by reduced estrous cycle length, decreased corpus luteum size and significantly decreased plasma progesterone levels. In contract, treatment with 25 mg PGE1 resulted in cycle lengths comparable to those of non-treated herdmates with no apparent modification in corpus luteum size. However, plasma progesterone levels were increased significantly following PGE1 treatment compared to pretreatment values. In so far as data obtained in vitro on PGF2alpha relative binding affinity to the bovine CL can be compared to data obtained independently in vitro on PGF2alpha induced luteolysis in the bovine, PGF2alpha relative binding to the CL and luteolysis appeared to be associated. By similar reasoning, there was no apparent relationship between PGE1 relative binding affinity in the luteal fractions and luteolysis in estrous cyclic cattle.     

Endothelin-1 mediates prostaglandin F(2alpha)-induced luteal regression in the ewe

A diversified series of experiments was conducted to determine the potential role of endothelin-1 (ET-1) in ovine luteal function. Endothelin-1 inhibited basal and LH-stimulated progesterone production by dispersed ovine luteal cells during a 2-h incubation. This inhibition was removed when cells were preincubated with cyclo-D-Asp-Pro-D-Val-Leu-D-Trp (BQ123), a highly specific endothelin ET(A) receptor antagonist. Administration of a luteolytic dose of prostaglandin F(2alpha) (PGF(2alpha)) rapidly stimulated gene expression for ET-1 in ovine corpora lutea (CL) collected at midcycle. Intraluteal administration of a single dose of BQ123 to ewes on Day 8 or 9 of the estrous cycle mitigated the luteolytic effect of PGF(2alpha). Intramuscular administration of 100 microg ET-1 to ewes at midcycle reduced plasma progesterone concentrations for the remainder of the estrous cycle. Following pretreatment with a subluteolytic dose of PGF(2alpha), i.m. administration of 100 microg ET-1 caused a rapid decline in plasma progesterone and shortened the length of the estrous cycle. These data complement and extend previously published reports in the bovine CL and are the strongest evidence presented to date in support of a role for ET-1 in PGF(2alpha)-mediated luteal function in domestic ruminants.     

Sensitivity of bovine corpora lutea to prostaglandin F2alpha is dependent on progesterone, oxytocin, and prostaglandins.

Prostaglandin (PG) F2alpha that is released from the uterus is essential for spontaneous luteolysis in cattle. Although PGF2alpha and its analogues are extensively used to synchronize the estrous cycle by inducing luteolysis, corpora lutea (CL) at the early stage of the estrous cycle are resistant to the luteolytic effect of PGF2alpha. We examined the sensitivity of bovine CL to PGF2alpha treatment in vitro and determined whether the changes in the response of CL to PGF2alpha are dependent on progesterone (P4), oxytocin (OT), and PGs produced locally. Bovine luteal cells from early (Days 4-5 of the estrous cycle) and mid-cycle CL (Days 8-12 of the estrous cycle) were preexposed for 12 h to a P4 antagonist (onapristone: OP; 10(-4) M), an OT antagonist (atosiban: AT; 10(-6) M), or indomethacin (INDO; 10(-4) M) before stimulation with PGF2alpha. Although OP reduced P4 secretion (p < 0.001) only in early CL, it reduced OT secretion in the cells of both phases examined (p < 0.001). OP also reduced PGF2alpha and PGE2 secretion (p < 0.01) from early CL. However, it stimulated PGF2alpha secretion in mid-cycle luteal cells (p < 0.001). AT reduced P4 secretion in early and mid-cycle CL (p < 0.05). Moreover, PGF2alpha secretion was inhibited (p < 0.05) by AT in early CL. The OT secretion and the intracellular level of free Ca2+ ([Ca2+]i) were measured as indicators of CL sensitivity to PGF2alpha. PGF2alpha had no influence on OT secretion, although [Ca2+]i increased (p < 0.05) in the early CL. However, the effect of PGF2alpha was augmented (p < 0.01) in cells after pretreatment with OP, AT, and INDO in comparison with the controls. In mid-cycle luteal cells, PGF2alpha induced 2-fold increases in OT secretion and [Ca2+]i. However, in contrast to results in early CL, these increases were magnified only by preexposure of the cells to AT (p < 0.05). These results indicate that luteal P4, OT, and PGs are components of an autocrine/paracrine positive feedback cascade in bovine early to mid-cycle CL and may be responsible for the resistance of the early bovine CL to the exogenous PGF2alpha action.     

Production of leukotriene B4 by luteal tissues of sheep treated with prostaglandin F2 alpha

Sheep were injected on Day 10 of the estrous cycle with a luteolytic dose of prostaglandin (PG) F2 alpha or injection vehicle. Corpora lutea were recovered at laparatomy 2, 4, 8, and 16 h following treatment. Portions of tissue were processed for light microscopy and analyzed for concentrations of leukotriene (LT) B4 and progesterone. There was a significant increase in LTB4 in tissues collected before evidence of either functional or structural regression (2 h); such levels were sustained. Tissues isolated 2 h after administration of PGF2 alpha and placed in short-term incubation produced considerable quantities of LTB4. Although no direct in vitro effect of PGF2 alpha on luteal production of LTB4 was demonstrated, it seems possible that lipoxygenase products of arachidonate metabolism, such as LTB4, could act as intermediates in the luteolytic mechanism of action of PGF2 alpha in vivo.     

Is tumor necrosis factor alpha a trigger for the initiation of endometrial prostaglandin F(2alpha) release at luteolysis in cattle?

To determine the physiological significance of tumor necrosis factor alpha (TNFalpha) in the regulation of luteolytic prostaglandin (PG) F(2alpha) release by the bovine endometrium, the effect of TNF-alpha on PGF(2alpha) output by the endometrial tissues in vitro was investigated and compared with the effect of oxytocin (OT). Furthermore, the presence of specific receptors for TNFalpha in the bovine endometrium during the estrous cycle was determined. Endometrial slices (20-30 mg) taken from six stages of the estrous cycle (estrus: Day 0; early I: Days 2-3; early II: Days 5-6; mid-: Days 8-12; late: Days 15-17; and follicular: Days 19-21), as determined by macroscopic examination of the ovaries and uterus, were exposed to TNFalpha (0.06-6 nM) and/or OT (100 nM). OT stimulated PGF(2alpha) output at the follicular stage and at estrus (P < 0.001), but not at the late luteal stage. On the other hand, the stimulatory effects of TNFalpha on PGF(2alpha) output were observed not only at the follicular stage but also at the late luteal stage (P < 0.001). When the endometrial tissues at late luteal stage were simultaneously exposed to TNFalpha (0.6 nM) and OT (100 nM), the stimulatory effect on PGF(2alpha) output was higher than the effect of TNFalpha or OT alone (P < 0.05). Specific binding of TNFalpha to the bovine endometrial membranes was observed throughout the estrous cycle. The concentration of TNF-alpha receptor at the early I luteal stage was less than the concentrations at other luteal stages (P < 0.01). The dissociation constant (K(d)) values of the endometrial membranes were constant during the estrous cycle. The overall results lead us to hypothesize that TNFalpha may be a trigger for the output of PGF(2alpha) by the endometrium at the initiation of luteolysis in cattle.     

Effects of in vivo and in vitro administration of prostaglandin F2 alpha on lipoprotein utilization in cultured bovine luteal cells

Two experiments were conducted to determine if a loss in the ability to utilize lipoprotein-cholesterol is one mechanism whereby prostaglandin F2 alpha (PGF2 alpha) decreases steroidogenesis in bovine luteal cells. In the first experiment, serum-free cultures of bovine luteal cells were treated with PGF2 alpha (100 ng/ml) for 5 days prior to addition of lipoproteins. Exposure to PGF2 alpha completely suppressed low-density lipoprotein (LDL)- and high-density lipoprotein (HDL)-stimulated progesterone production (p less than 0.01) compared to control (no PGF2 alpha) cultures. Luteal cells cultured in the presence of LDL + luteinizing hormone (LH, 10 ng/ml) + PGF2 alpha produced significantly less progesterone than luteal cells cultured with LDL + LH (p less than 0.05). Treatment with PGF2 alpha had no significant effect on HDL + LH-stimulated progesterone synthesis. In the second experiment, cows were injected with a luteolytic dose of PGF2 alpha (25 mg), and the corpora lutea were removed at 0 (no PG), 1, 4, or 12 h post-injection. Dissociated luteal cells were placed in culture for 7 days, either with or without LH (10 ng/ml), and lipoproteins were added on Days 5-7. LH stimulation of progesterone production was apparent in cultures obtained at 0 and 12 (p less than 0.05) but not 1 and 4 h post-PGF2 alpha. Addition of either LDL or HDL increased progesterone synthesis in all cultures, regardless of time following in vivo administration of PGF2 alpha. It is concluded that PGF2 alpha can inhibit bovine luteal cell utilization of either LDL or HDL in vitro. However, luteal cell utilization of lipoproteins in vitro is not adversely affected by in vivo exposure to PGF2 alpha, if collected within 12 h post-PGF2 alpha.     

Administration of a nitric oxide synthase inhibitor counteracts prostaglandin F2-induced luteolysis in cattle

The objective of this study was to determine whether nitric oxide (NO) is produced locally in the bovine corpus luteum (CL) and whether NO mediates prostaglandin F2alpha (PGF2alpha)-induced regression of the bovine CL in vivo. The local production of NO was determined in early I, early II, mid, late, and regressed stages of CL by determining NADPH-d activity and the presence of inducible and endothelial NO synthase immunolabeling. To determine whether inhibition of NO production counteracts the PGF2alpha-induced regression of the CL, saline (10 ml/h; n = 10) or a nonselective NOS inhibitor (Nomega-nitro-l-arginine methyl ester dihydrochloride [L-NAME]; 400 mg/h; n = 9) was infused for 2 h on Day 15 of the estrous cycle into the aorta abdominalis of Holstein/Polish Black and White heifers. After 30 min of infusion, saline or cloprostenol, an analogue of PGF2alpha (aPGF2alpha; 100 microg) was injected into the aorta abdominalis of animals infused with saline or L-NAME. NADPH-diaphorase activity was present in bovine CL, with the highest activity at mid and late luteal stages (P < 0.05). Inducible and endothelial NO synthases were observed with the strongest immunolabeling in the late CL (P < 0.05). Injection of aPGF2alpha increased nitrite/nitrate concentrations (P < 0.01) and inhibited P4 secretion (P < 0.05) in heifers that were infused with saline. Infusion of L-NAME stimulated P4 secretion (P < 0.05) and concomitantly inhibited plasma concentrations of nitrite/nitrate (P < 0.05). Concentrations of P4 in heifers infused with L-NAME and injected with aPGF2alpha were higher (P < 0.05) than in animals injected only with aPGF2alpha. The PGF2alpha analogue shortened the cycle length compared with that of saline (17.5 +/- 0.22 days vs. 21.5 +/- 0.65 days P < 0.05). L-NAME blocked the luteolytic action of the aPGF2alpha (22.6 +/- 1.07 days vs. 17.5 +/- 0.22 days, P < 0.05). These results suggest that NO is produced in the bovine CL. NO inhibits luteal steroidogenesis and it may be one of the components of an autocrine/paracrine luteolytic cascade induced by PGF2alpha.     

Changes in steady-state concentrations of messenger ribonucleic acids in luteal tissue during prostaglandin F2alpha induced luteolysis in mares

Transvaginal ultrasound-guided luteal biopsy was used to evaluate the effects of prostaglandin (PG)F2alpha on steady-state concentrations of mRNA for specific genes that may be involved in regression of the corpus luteum (CL). Eight days after ovulation (Hour 0), mares (n=8/group) were randomized into three groups: control (no treatment or biopsy), saline+biopsy (saline treatment at Hour 0 and luteal biopsy at Hour 12), or PGF2alpha+biopsy (5mg PGF2alpha at Hour 0 and luteal biopsy at Hour 12). The effects of biopsy on CL were compared between the controls (no biopsy) and saline+biopsy group. At Hour 24 (12h after biopsy) there was a decrease in circulating progesterone in saline group to 56% of pre-biopsy values, indicating an effect of biopsy on luteal function. Mean plasma progesterone concentrations were lower (P<0.001) at Hour 12 in the PG group compared to the other two groups. The relative concentrations of mRNA for different genes in luteal tissue at Hour 12 was quantified by real time PCR. Compared to saline-treated mares, treatment with PGF2alpha increased mRNA for cyclooxygenase-2 (Cox-2, 310%, P<0.006), but decreased mRNA for LH receptor to 44% (P<0.05), steroidogenic acute regulatory protein to 22% (P<0.001), and aromatase to 43% (P<0.1) of controls. There was no difference in mRNA levels for PGF2alpha receptor between PG and saline-treated groups. Results indicated that luteal biopsy alters subsequent luteal function. However, the biopsy approach was effective for collecting CL tissue for demonstrating dynamic changes in steady-state levels of mRNAs during PGF2alpha-induced luteolysis. Increased Cox-2 mRNA concentrations suggested that exogenous PGF2alpha induced the synthesis of intraluteal PGF2alpha. Thus, the findings are consistent with the concept that an intraluteal autocrine loop augments the luteolytic effect of uterine PGF2alpha in mares.     

Reduction by human chorionic gonadotropin of the luteolytic effect of prostaglandin F2 alpha in ewes

The ability of human chorionic gonadotropin (HCG) to reduce the luteolytic effect of prostaglandin (PGF2 alpha) was demonstrated in cycling ewes. As expected, treatment with 10 mg of PGF2 alpha alone on Day 10 of the estrous cycle exerted a potent negative effect on the function and structure of corpus luteum (CL) as indicated by reduced plasma progesterone, CL progesterone, and CL weight. However, the identical PGF2 alpha treatment failed to significantly reduce either luteal function or luteal weight when administered to ewes that were also treated with HCG on Days 9 and 10 of the estrous cycle. Treatment with HCG alone had a positive effect on CL as indicated by increased plasma progesterone, CL progesterone, and CL weight. Treatment with HCG did not render the CL totally insensitive to the negative effects of PGF2 alpha because plasma progesterone was reduced when the dose of PGF2 alpha was doubled. Whether CL regressed or continued to function after treatment with both HCG and PGF2 alpha appeared to depend upon a balance between the positive and negative effects of the two hormones.     

Prostaglandin f(2alpha) induces distinct physiological responses in porcine corpora lutea after acquisition of luteolytic capacity

This study examines differences in intracellular responses to cloprostenol, a prostaglandin (PG)F(2alpha) analog, in porcine corpora lutea (CL) before (Day 9 of estrous cycle) and after (Day 17 of pseudopregnancy) acquisition of luteolytic capacity. Pigs on Day 9 or Day 17 were treated with saline or 500 microgram cloprostenol, and CL were collected 10 h (experiment I) or 0.5 h (experiment III) after treatment. Some CL were cut into small pieces and cultured to measure progesterone and PGF(2alpha) secretion. In experiment I, progesterone remained high and PGF(2alpha) low in luteal incubations from either Day 9 or Day 17 saline-treated pigs. Cloprostenol increased PGF(2alpha) production 465% and decreased progesterone production 87% only from Day 17 luteal tissue. Cloprostenol induced prostaglandin G/H synthase (PGHS)-2 mRNA (0.5 h) and protein (10 h) in both groups. In cell culture, cloprostenol or phorbol 12, 13-didecanoate (PDD) (protein kinase C activator), induced PGHS-2 mRNA in luteal cells from both groups. However, acute cloprostenol treatment (10 min) decreased progesterone production and increased PGF(2alpha) production only from Day 17 luteal cells. Thus, PGF(2alpha) production is induced by cloprostenol in porcine CL with luteolytic capacity (Day 17) but not in CL without luteolytic capacity (Day 9). However, this change in PGF(2alpha) production is not explained by a difference in induction of PGHS-2 mRNA or protein.     

Local changes in blood flow within the early and midcycle corpus luteum after prostaglandin F(2 alpha) injection in the cow

One of the postulated main luteolytic actions of prostaglandin (PG) F(2 alpha) is to decrease ovarian blood flow. However, before Day 5 of the normal cycle, the corpus luteum (CL) is refractory to the luteolytic action of PGF(2 alpha). Therefore, we aimed to determine in detail the real-time changes in intraluteal blood flow after PGF(2 alpha) injection at the early and middle stages of the estrous cycle in the cow. Normally cycling cows at Day 4 (early CL, n = 5) or Days 10--12 (mid CL, n = 5) of the estrous cycle (estrus = Day 0) were examined by transrectal color and pulsed Doppler ultrasonography to determine the blood flow area, the time-averaged maximum velocity (TAMXV), and the volume of the CL after an i.m. injection of a PGF(2 alpha) analogue. Ultrasonographic examinations were carried out just before PG injection (0 h) and then at 0.5, 1, 2, 4, 8, 12, 24, and 48 h after the injection. Blood samples were collected at each of these times for progesterone (P) determination. The ratio of the colored area to a sectional plane at the maximum diameter of the CL was used as a quantitative index of the changes in blood flow within the luteal tissue. Blood flow within the midcycle CL initially increased (P < 0.05) at 0.5-2 h, decreased at 4 h to the same levels observed at 0 h, and then further decreased to a lower level from 8 h (P < 0.05) to 48 h (P < 0.001). Plasma P concentrations decreased (P < 0.05) from 4.7 +/- 0.5 ng/ml (0 h) to 0.6 +/- 0.2 ng/ml (24 h). The TAMXV and CL volume decreased at 8 h (P < 0.05) and further decreased (P < 0.001) from 12 to 24 h after PG injection, indicating structural luteolysis. These changes were not detected in the early CL, in which luteolysis did not occur. In the early CL, the blood flow gradually increased in parallel with the CL volume, plasma P concentration, and TAMXV from Day 4 to Day 6. The present results indicate that PGF(2 alpha) induces an acute blood flow increase followed by a decrease in the midcycle CL but not in the early CL. This transitory increase may trigger the luteolytic cascade. The lack of intraluteal vascular response to PG injection in the early CL appears to be directly correlated with the ability to be resistant to PG.     

Growth hormone, but not luteinizing hormone, acts with luteal peptides on prostaglandin F2alpha and progesterone secretion by bovine corpora lutea in vitro*

Prostaglandin F2alpha (PGF2alpha) is a major physiological luteolysin in the cow. However, injection of PGF2alpha before day 5 (day 0 = estrus) of the estrous cycle dose not induce luteolysis. On the other hand, the early corpus luteum (CL) actively produces PGF2alpha. This indicates that luteal PGF2alpha may play a key role in the refractoriness to PGF2alpha injected during the early luteal phase when angiogenesis is active in the CL. Thus, this study aimed to investigate the possible interaction between pituitary hormones and local factors (luteal peptides) on secretion of PGF2alpha and progesterone (P) by the early bovine CL, and to evaluate the effect of growth hormone (GH) as well as its interactions on production of PGF2alpha in the developing CL. A RT-PCR analysis revealed that mRNA for GH receptor in CL was fully expressed from early in the luteal phase throughout the estrous cycle, while luteinizing hormone (LH) receptor mRNA was expressed less by the early and regressing CL than those at mid or late luteal phases (P < 0.05). For the stimulation test, an in vitro microdialysis system (MDS) was used as a model. Each bovine early CL (days 3-4) was implanted with the MDS, and maintained in an organ culture chamber. The infusion of GH, insulin-like growth factor-1 (IGF-1) and oxytocin (OT) increased (P < 0.05) PGF2alpha and P release. In contrast, LH had no effect (P > 0.05) on PGF2alpha secretion and little effect on P release. Unexpectedly, there was no distinct interaction between pituitary hormones and luteal peptides on secretion of PGF2alpha and P. These results indicate that GH is a more powerful stimulator of PGF2alpha and P production in the early bovine CL than LH and suggest that GH and luteal peptides, IGF-1 and OT, contribute to maintenance of elevated PGF2alpha production in the developing bovine CL.     

Specific PGF-2 alpha binding by the corpus luteum of the pregnant and non-pregnant mare.

The binding of prostaglandin (PG) F-2 alpha to corpora lutea (CL) from pregnant and non-pregnant Pony mares was examined. Studies of the rates of association and dissociation indicated that [3H]PGF was bound specifically and reversibly to a luteal cell membrane preparation (MP) that was isolated by high speed (100,000 g) ultracentrifugation. Various PGs and PG metabolites displaced [3H]PGF from the receptors in the following decreasing order: PGF-2 alpha greater than 13, 14-dihydro-PGF-2 alpha = 13,14-dihydro-15-keto PGF-2 alpha greater than PGD-2 greater than PGF-1 alpha = PGE-2 greater than PGE-2 beta greater than PGE-1. These data implicate the 9 alpha-OH and 5,6 cis double bond as major contributors to PGF receptor recognition. The membrane preparation appeared to contain at least two receptor populations, a high affinity, low capacity and a low affinity, high capacity receptor. The binding of PGF (pg/mg MP protein +/- s.e.m. (n)) to CL of the non-pregnant mare increased from 4.09 +/- 11.6 (4), on Day 4 after ovulation, to reach maximal levels by Day 12, 15.01 +/- 2.5 (4), and declined thereafter. In pregnancy the binding of PGF continued to increase until Day 18, reaching 27.47 +/- 1.7 (3), before it declined on Day 20. The reduction in binding by Day 16 in the non-pregnant mare may reflect the process of luteolysis, while high PGF binding capacity of CL between Days 16 and 18 of pregnancy indicated that luteal maintenance during pregnancy is not associated with a reduction of PGF binding capabilities.     

Growth hormone, but not luteinizing hormone, acts with luteal peptides on prostaglandin F2alpha and progesterone secretion by bovine corpora lutea in vitro

Prostaglandin F2alpha (PGF2alpha) is a major physiological luteolysin in the cow. However, injection of PGF2alpha before day 5 (day 0 = estrus) of the estrous cycle dose not induce luteolysis. On the other hand, the early corpus luteum (CL) actively produces PGF2alpha. This indicates that luteal PGF2alpha may play a key role in the refractoriness to PGF2alpha injected during the early luteal phase when angiogenesis is active in the CL. Thus, this study aimed to investigate the possible interaction between pituitary hormones and local factors (luteal peptides) on secretion of PGF2alpha and progesterone (P) by the early bovine CL, and to evaluate the effect of growth hormone (GH) as well as its interactions on production of PGF2alpha in the developing CL. A RT-PCR analysis revealed that mRNA for GH receptor in CL was fully expressed from early in the luteal phase throughout the estrous cycle, while luteinizing hormone (LH) receptor mRNA was expressed less by the early and regressing CL than those at mid or late luteal phases (P < 0.05). For the stimulation test, an in vitro microdialysis system (MDS) was used as a model. Each bovine early CL (days 3-4) was implanted with the MDS, and maintained in an organ culture chamber. The infusion of GH, insulin-like growth factor-I (IGF-I) and oxytocin (OT) increased (P < 0.05) PGF2alpha and P release. In contrast, LH had no effect (P > 0.05) on PGF2alpha secretion and little effect on P release. Unexpectedly, there was no distinct interaction between pituitary hormones and luteal peptides on secretion of PGF2alpha and P. These results indicate that GH is a more powerful stimulator of PGF2alpha and P production in the early bovine CL than LH and suggest that GH and luteal peptides, IGF-1 and OT, contribute to maintenance of elevated PGF2alpha production in the developing bovine CL.     

Interactions between prostaglandin analogues and a receptor in bovine Corpora lutea. Correlation of dissociation constants with luteolytic potencies in hamsters.

The dissociation constants for the interactions between some prostaglandin analogues and a prostaglandin F2 receptor in bovine corpora lutea were determined. These values were compared to the antifertility potencies of these compounds in hamsters and the rates of metabolism by 15-hydro-syprostaglandin dehydrogenase. The most active analogues with regard to both affinity for the receptor and luteolytic potency were 17-phenyl-18, 19, 20-trinorprostaglandin F2alpha and 15-methylprostaglandin F2alpha. The alkyl side chain of prostaglandins could be modified considerably without altering the affinity for the receptor. In this way metabolism by 15-hydroxyprostaglandin dehydrogenase could be blocked. Some of these compounds -ad greatly increased luteolytic effects. Substitution of a phenyl group for the 3 terminal carbon units of the alkyl side chain of prostaglandins increased both the affinity for the receptor and the luteolytic activity in vivo. 7-oxa-13-prostynoic acid, an antagonist of the luteolytic effect of prostaglandin F2alpha in vivo was a weak competitive inhibitor of the interation between prostaglandin F2alpha and the receptor.     

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 luteum

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.     

Administration of prostaglandin f(2 alpha) during the early bovine luteal phase does not alter the expression of ET-1 and of its type A receptor: a possible cause for corpus luteum refractoriness

Luteal regression is initiated by prostaglandin F(2 alpha) (PGF(2 alpha)). In domestic species and primates, demise of the corpus luteum (CL) enables development of a new preovulatory follicle. However, during early stages of the cycle, which are characterized by massive neovascularization, the CL is refractory to PGF(2 alpha). Our previous studies showed that endothelin-1 (ET-1), which is produced by the endothelial cells lining these blood vessels, plays a crucial role during PGF(2 alpha)-induced luteolysis. Therefore, in this study, we compared the effects of PGF(2 alpha) administered at the early and mid luteal phases on ET-1 and its type A receptors (ETA-R) along with plasma ET-1 and progesterone concentrations, and the mRNA levels of PGF(2 alpha) receptors (PGF(2 alpha)-R) and steroidogenic genes. As expected, ET-1 and ETA-R mRNA levels were markedly induced in midcycle CL exposed to luteolytic dose of PGF(2 alpha) analogue (Cloprostenol). In contrast, neither ET-1 mRNA nor its receptors were elevated when the same dose of PGF(2 alpha) analogue was administered on Day 4 of the cycle. In accordance with ET-1 expression within the CL, plasma ET-1 concentrations were significantly elevated 24 h after PGF(2 alpha) injection only on Day 10 of the cycle. The steroidogenic capacity of the CL (plasma progesterone as well as the mRNA levels of steroidogenic acute regulatory protein and cytochrome P450(scc)) was only affected when PGF(2 alpha) was administered during midcycle. Nevertheless, PGF(2 alpha) elicited certain responses in the early CL: progesterone and oxytocin secretion were elevated, and PGF(2 alpha)-R was transiently affected. Such effects probably result from PGF(2 alpha) acting on luteal steroidogenic cells. These findings may suggest, however, that the cell type mediating the luteolytic actions of PGF(2 alpha), possibly the endothelium, could yet be nonresponsive during the early luteal phase.