Lack of complete regression of the Day 5 corpus luteum after one or two doses of PGF2α in nonlactating Holstein cows

The early corpus luteum (CL) (before Day 6) does not regress after a single PGF_2α treatment. We hypothesized that increasing PGF_2α dose or number of treatments would allow regression of the early CL (Day 5). Nonlactating Holstein cows (N = 22) were synchronized using the Ovsynch protocol. On Day 5 (Day 0 = second GnRH treatment), cows were assigned to: (1) control (N = 5): no further treatment; (2) 1PGF (N = 6): one dose of 25 mg PGF_2α; (3) 2PGF (N = 5): two doses of 25 mg PGF_2α (50 mg) given 8 hours apart (second PGF_2α on Day 5 at the same time as the other PGF_2α treatments); (4) DPGF (N = 6): double dose of 25 mg PGF_2α (50 mg) given on Day 5. Blood samples were collected to monitor progesterone (P4) profiles in two periods. In the first period (0 to 24 hours), there were effects of treatment (P = 0.01), time (P < 0.01), and an interaction of treatment and time (P = 0.02). Group 1PGF versus control was different only at 12 hours (P = 0.02). Cows treated with DPGF were different than control at 4 hours (P = 0.04), 12 hours (P < 0.01), and 24 hours (P < 0.01). Only cows treated with 2PGF had lower P4 than control during the entire period and low P4 (0.37 ± 0.17 ng/mL) at 24 hours, usually indicative of luteolysis. In the second period (Day 5 to 15 of the cycle), there were effects of treatment (P < 0.01), time (P < 0.01), and interaction of treatment and time (P = 0.002). Group 1PGF was not different than control from Day 5 to 13 and P4 was greater than control on Day 14 (P = 0.01) and 15 (P < 0.01). Circulating P4 in DPGF cows was lower than control from Day 7 (P = 0.05) through 12 (P < 0.01). Likewise, there were differences between control and 2PGF from Day 7 to 13, but not on Day 14 and 15. On Day 15, all PGF_2α-treated groups had circulating P4 consistent with an active CL. Ultrasound evaluation confirmed that no CL from any group completely regressed during the experiment and no new ovulations occurred to account for functional CL later in cycle. In summary, a double dose of PGF_2α (twice on Day 5 or 8 hours apart) can dramatically decrease P4, consistent with classical definitions of luteolysis; however, these CL recover and become fully functional. Thus, the Day 5 CL of mature Holstein cows do not regress even to two doses of PGF_2α.     

Inhibitory effects of aspirin and indomethacin on the biosynthesis of PGE2 and PGF2α

A gas-liquid chromatography system has been used to study the effects of indomethacin and aspirin on the biosynthesis of PGE₂ and PGF_2α by the prostaglandin synthetase system of bovine seminal vesicle. Both compounds were found to inhibit the production of PGE₂ and PGF_2α. However, based on statistical analyses, the inhibitory effect of indomethacin was found to be non-selective while aspirin produced statistically significant preferential inhibition of PGE₂ over PGF_2α.     

Effect of mifepristone and antiestrogens on uterine PGF2α and PGE2 concentrations in ovariectomized and pregnant rats

Four antiestrogens (anordiol, tamoxifen, RU 39411, ICI 182780) and the antiprogestin, mifepristone (RU 486), were administered to the following three animal models: (1) ovariectomized rats, (2) mated rats treated post-coitally; and (3) pregnant rats treated post-implantation. The antiestrogens were administered alone or in combination with mifepristone at doses effective in preventing and/or terminating pregnancy in rats. The objective of the study was to determine whether these drugs influenced uterine concentrations of prostaglandins (PGF_2α and PGE₂).Antiestrogens administered alone to ovariectomized rats did not effect uterine PGE₂ or PGF_2α concentrations; whereas the combination of anordiol/mifepristone increased uterine PGF_2α concentration, resulting in an increase in the PGF_2α/PGE₂ ratio.Mated rats were treated post-coitally for three consecutive days with anordiol, tamoxifen, estradiol and mifepristone alone and with the combination of anordiol/mifepristone and tamoxifen/mifepristone. An increase in uterine PGF_2α concentrations and in the PGF_2α/PGE₂ ratio occurred only in anordiol/mifepristone treated group. A decrease in uterine PGE₂ concentrations occurred in animals treated with anordiol, tamoxifen and estradiol, resulting in an increase in the PGF_2α/PGE₂ ratio.Anordiol (5.0 mg/kg/day) and mifepristone (4.0 mg/kg/day) alone and the combination of anordiol/mifepristone (2.5/1.0 mg/kg/day) administered to pregnant rats on days 7, 8 and 9 of pregnancy induced an increase in PGF_2α levels without affecting uterine PGE₂ concentration. The changes in uterine PGF_2α concentrations induced by anordiol and the combination of anordiol/mifepristone resulted in an increase in the PGF_2α/PGE₂ ratio.The antiestrogens tested except for ICI 182780 possessed agonist activity when assayed by measuring their capacity to increase the uterine weights in ovariectomized rats. Also, ICI 182789 was the only antiestrogen that did not influence uterine PG concentrations. It can be concluded that ICI 182780 is the only “pure” antiestrogen among those tested.The present results show that antiestrogens and the combination of mifepristone plus anordiol at doses preventing implantation and terminating pregnancy increase uterine PGF_2α and/or decrease PGE₂ concentrations, resulting in an alteration of PGF_2α/PGE₂ ratio. These findings suggest that there exists a critical balance of PGF_2α to PGE₂ concentrations in the uterus required for the normal passage of fertilized ova through the oviduct, initiating implantation of the blastocysts, development of embryos, and maintenance of pregnancy.     

Omega chain methylated analogs of PGF2α and PGE2

Our previously published prostaglandin (PG) synthesis route, in which the ω-chain is added in the penultimate step, provides facile access to a wide variety of ω-chain variant PG analogs. Each series requires only the synthesis of the appropriate methylated acylphosphonate for the Emmons' condensation. The syntheses of analogs bearing the following methylation pattern are detailed: 15-Me; 17, 17-(Me)₂; 17, 17, 20-(Me)₃; 18, 18, 20-(Me)₃; 15, 18, 18, 20-(Me)₄; and 15-Ome-18, 18, 20-(Me)₃. The well-known 16, 16-dimethyl prostaglandins have also been prepared by this sequence. The synthesis of 16, 16-tetramethylene-PG analogs is also described.     

Effects of PGF2α infusion on human cortisol biosynthesis

Intravenous administration of prostaglandin F_2α to normal volunteers induced a three-fold increase in urinary cortisol output. Dexamethasone administration significantly inhibited the cortisol response, indicating that prostaglandin does not directly stimulate adrenal cortisol biosynthesis. Prostaglandin infusion appears to increase cortisol biosynthesis through stimulation of pituitary ACTH release.     

Radioimmunoassay measurement of ACTH-facilitated PGE2 and PGF2α release from isolated cat adrenocortical cells

Prostaglandin (PG) biosynthesis by trypsin-dispersed cat adrenocortical cells was studied by radioimmunoassay (RIA). Parallel assays of incubation media using PGF_2α and PGF_1α antisera established that PGF_2α is the primary PGF released by feline cortical cells. Following the reduction of PGE to PGF with sodium borohydride (NaBH₄) these same two antisera were also used to identify PGE₂ as the primary PGE released. RIA using a PGE antiserum confirmed the presence of PGE in the incubation medium. Steroidogenic concentrations of ACTH (50–250μU) enhanced PGE and PGF release, and indomethacin suppressed the ACTH-facilitated release. These studies provide additional evidence for ACTH-induced PG synthesis by feline cortical cells, and support the hypothesis that PGs play some role in the steroidogenic action of ACTH.     

Luteal function in sheep injected with prostaglandin F2α directly into the corpus luteum

Groups of ewes received either saline or prostaglandin F_2α (PGF_2α) as an injection directly into the corpus luteum. Changes in circulating progesterone levels were measured as well as subsequent histological examination of the corpora lutea. Saline, or PGF_2α given at the two lower doses (60 and 120 μg respectively), failed to suppress progesterone levels permanently, or to induce degenerative changes in the corpora lutea. Treatment with a higher dose of PGF_2α (240 μg) was followed by a marked elevation in progesterone levels. These results are discussed in relation to reported effects of PGF_2α arriving at the ovary via the arterial circulation.     

Corpus luteum regression induced by ultra-low pulses of prostaglandin F2α

In view of the pulsatile nature of PGF_2α secretion from the ovine uterus at the time of luteolysis, experiments were designed to examine the effect of pulsed infusions of PGF_2α on luteal function and to re-examine the minimal effective levels of PGF_2α required to induce luteolysis. To mimic physiological conditions, hour-long infusions of PGF_2α in increasing concentrations were given either 4 times in 19 h or 5 times in 25 h into the arterial supply of the autotransplanted ovary in conscious sheep on day 12 of an induced cycle. Blood flow and progesterone secretion rate from the ovary were used to monitor directly the luteolytic effect of administered PGF_2α. The concentration of LH in peripheral plasma was measured throughout each infusion experiment and the presence of a preovulatory peak of LH was used as an indicator of the permanence of luteal regression. Four pulses of PGF_2α in 19 h caused complete corpus luteum regression in only 1 of 4 animals whereas the addition of a fifth pulse (5 pulses in 25 h) caused permanent regression in 4 out of 4 animals. Infusion of 5 hour-long pulses of saline or PGF_2α at a rate of <0.04 μg/h did not induce permanent suppression of progesterone secretion. The average total effective dose of PGF_2α required to induced luteal regression when given as 5 pulses was 1/40th of the amount currently regarded as the minimal effective one when given by constant infusion into the ovarian artery. In another series of experiments the luteolytic effect of a single hour-long pulse of 0.1 μg/h PGF_2α given daily for either 3 or 4 days was investigated. A significant fall (ANOVA, F_0.01) in progesterone secretion rate, which reached a nadir at $\text{5.3 ± 2.2}\text{h (}\text{x}\text{±}\text{S.D.}\text{, n=15)}$, was followed by a recovery of progesterone secretion rate. Permanent luteal regression did not occur with this protracted regimen, suggesting that a relatively short pulse frequency of PGF_2α over a minimal period of 24 h is a necessary condition for physiological regression of the corpus luteum in sheep.     

Effect of increasing GnRH and PGF2α dose during Double-Ovsynch on ovulatory response,luteal regression,and fertility of lactating dairy cows

Ovsynch-type synchronization of ovulation protocols have suboptimal synchronization rates due to reduced ovulation to the first GnRH treatment and inadequate luteolysis to the prostaglandin F_2α (PGF_2α) treatment before timed artificial insemination (TAI). Our objective was to determine whether increasing the dose of the first GnRH or the PGF_2α treatment during the Breeding-Ovsynch portion of Double-Ovsynch could improve the rates of ovulation and luteolysis and therefore increase pregnancies per artificial insemination (P/AI). In experiment 1, cows were randomly assigned to a two-by-two factorial design to receive either a low (L) or high (H) doses of GnRH (Gonadorelin; 100 vs. 200 μg) and a PGF_2α analogue (cloprostenol; 500 vs. 750 μg) resulting in the following treatments: LL (n = 263), HL (n = 277), LH (n = 270), and HH (n = 274). Transrectal ultrasonography and serum progesterone (P4) were used to assess ovulation to GnRH1, GnRH2, and luteal regression after PGF_2α during Breeding-Ovsynch in a subgroup of cows (n = 651 at each evaluation). Pregnancy status was assessed 29, 39, and 74 days after TAI. In experiment 2, cows were randomly assigned to LL (n = 220) or HH (n = 226) treatment as described for experiment 1. For experiment 1, ovulation to GnRH1 was greater (P = 0.01) for cows receiving H versus L GnRH (66.6% [217/326] vs. 57.5% [187/325]) treatment, but only for cows with elevated P4 at GnRH1. Cows that ovulated to GnRH1 had increased (P < 0.001) fertility compared with cows that did not ovulate (52.2% vs. 38.5%); however, no effect of higher dose of GnRH on fertility was detected. The greater PGF_2α dose increased luteal regression primarily in multiparous cows (P = 0.03) and tended to increase fertility (P = 0.05) only at the pregnancy diagnosis 39 days after TAI. Overall, P/AI was 47.0% at 29 days and 39.7% at 74 days after TAI; P/AI did not differ (P = 0.10) among treatments at 74 days (LL, 34.6%; HL, 40.8%; LH, 42.2%; HH, 40.9%) and was greater (P < 0.001) for primiparous cows than for multiparous cows (46.1% vs. 33.8%). For experiment 2, P/AI did not differ (P = 0.21) between H versus L treatments (44.2% [100/226] vs. 40.5% [89/220]). Thus, despite an increase in ovulatory response to GnRH1 and luteal regression to PGF_2α, there were only marginal effects of increasing dose of GnRH or PGF_2α on fertility to TAI after Double-Ovsynch.     

Effect of PGD2, PGE2, PGF2α and PGI2 on blood pressure,heart rate and plasma catecholamine responses to spinal cord stimulation in the rat

The following experiments were designed in order to examine the inter-relationships of various prostaglandins (PG's) and the adrenergic nervous system, in conjunction with blood pressure and heart rate responses, $\text{in vivo}$. Stimulation of the entire spinal cord (50v, 0.3–3 Hz, 1.0 msec) of the pithed rat increased blood pressure, heart rate and plasma epinephrine (EPI) and norepinephrine (NE) concentration (radioenzymatic-thin layer chromatographic assay). Infusion of PGE₂(10–30 μg/kg. min, i.v.) suppressed blood pressure and heart rate responses to spinal cord stimulation while plasma EPI (but not NE) was augmented over levels found in control animals. PGI₂ (0.03–3.0 μg/kg. min, i.v.) suppressed the blood pressure response to spinal cord stimulation without any effect on heart rate or the plasma catecholamine levels. PGE₂ and PGF_2α(10–30 μg/kg. min, i.v.) did not change the blood pressure, heart rate or plasma EPI and NE responses to the spinal cord stimulation although PGF_2α disclosed an overall vasopressor effect during the pre-stimulation period. At the pre-stimulation period it was also observed that PGE₂, PGF_2α and PGI₂, had a positive chronotropic effect on the heart rate, the cardiac accelerating effect of PGE₂ was not abolished by propanolol. These $\text{in vivo}$ studies suggest that in the rat, PGE₂ and PGI₂ modulate sympathetic responses, primarily by interaction with the post-synaptic elements — PGE₂ on both blood vessels and the heart and PGI₂ by acting principally on blood vessels.     

The induction of labour by the intravenous infusion of prostaglandin F2α

labour was induced by the intravenous infusion of prostaglandin F_2α in 106 patients at 36–44 weeks of pregnancy. The induction was successful in 80% of the women. The total dose needed ranged from 0.1 to 14.2 mg of PGF_2α. The uterine activity and fetal heart rate were recorded by cardiotocography. The contraction pattern and induction-delivery time were the same as reported for induction with oxytocin. In one case uterine hyperactivity occurred after rupture of the membranes. No serious adverse effects were seen, but in a few cases local irritation was noted at the site of infusion. The condition of the infants was generally good.It might be concluded that PGF_2α seems valuable for the induction of labour, but due to the risk for overstimulation careful supervision is needed.     

Luteolytic activity of a synthetic prostaglandin and PGF2α in heifers

Two experiments involving 44 cycling heifers were conducted to evaluate the luteolytic activity of a synthetic prostaglandin, AY 24366, and PGF_2α. Activity was assessed by the decline in progesterone level of peripheral blood and occurrence of estrus. Progesterone concentrations of jugular blood plasma were quantified by radioimmunoassay. In the first experiment, 36 heifers were treated during diestrus with AY 24366 (A - 10mg intrauterine, B - 30mg intramuscular and C - 60mg im) or with PGF_2α (D - 5mg, iu, E - 15mg im and F - 30mg im). Mean progesterone 0, 24 and 48 hours after treatment were A - 6.33, 5.55 and 5.06; B - 6.35, 2.79 and 3.92; C - 5.23, 2.69 and 3.91; D - 5.19, 1.50 and 1.51; E - 4.69, 0.85 and 0.61; F - 6.66, 0.80 and 0.48 ng/ml. Standing estrus was observed in 1, 1, 1, 4, 5 and 6 females in groups A, B, C, D, E and F respectively within 72 hours of treatment. PGF_2α resulted in significantly (P<0.01) lower progesterone at 24 and 48 hours than AY 24366. However, im administration of the latter did significantly (P<0.05) lower progesterone at 24 hours. In the second trial six heifers were treated with either 120 or 180mg of AY 24366 im on day 12 of the cycle. Mean progesterone declined from 3.84 to 2.12 ng/ml (P<0.01) by 6 hours and to 1.59 ng/ml by 12 hours. Thereafter the decline was gradual and reached a level of 0.65 ng/ml at 72 hours. All six heifers showed standing estrus at 78±2 hours and were inseminated. Two in each group conceived. Doses of 15mg PGF_2α and 120mg AY 24366 were effective in causing luteal regression, however, the latter caused respiratory discomfort for 5 to 10 minutes post treatment.     

Comparative study of the concentrations of peripheral progesterone before and after PGF2α injection between Bos taurus (Brown Swiss) and Bos indicus (Indobrazil) in the tropics

With the object of studying the changes in progesterone concentration during the oestrous cycle and of verifying prostaglandin F_2α (PGF_2α) response in the luteal phase, 10 Indobrazil and 6 Brown Swiss cows, all non-lactating, were bled three times a week during the months of March and April in the Mexican tropics.Progesterone levels in both groups followed a similar pattern, maximum mean levels being reached at day 13 of the cycle (Indobrazil 2.2 ng/ml and Brown Swiss 2.8 ng/ml). No significant differences were found in the progesterone levels throughout the cycle. Nevertheless, a highly significant difference (P < 0.001) was established between breeds with respect to progesterone levels before and after PGF_2α injection. This was possibly due to a seasonal effect on progesterone production in the two types of cow.     

Contractile effects of PGD2 and PGF2α on mechanical and electrical activities in longitudinal and circular muscles of the guinea-pig colon

Contractile effects of PGD₂ and PGF_2α on mechanical and electrical activities were investigated in longitudinal and circular muscles of the guinea-pig isolated proximal colon.PGD₂ and PGF_2α (1 μM)-induced contractions were reduced with a decrease in external Ca ion concentration and were prevented at zero Ca (EGTA 1mM) in both muscles.In electrical activities recorded by the single sucrose gap method, PGD₂ and PGF_2α (1 μM) increased the frequency of spike generation on the slow rates in both muscles.Verapamil (10 μM), a Ca antagonist, blocked spontaneous and PG-induced spike activities with little effect on the slow waves in both muscles. Verapamil markedly reduced the contractions to PGs in the longitudinal muscle, but not in the circular muscle.The results suggest that PGD₂ and PGF_2α produced contractions which may depend largely on an increase in the inward Ca current of the spikes in longitudinal muscle, and may depend both on a release of Ca from the Ca sequestration in the cell, and on a spike-dependent Ca influx in circular muscle of the guinea-pig colon.     

Decreased blood flow through rat testis after intratesticular injection of PGF2α

The blood flow through the testes was calculated for 26 rats after measurements of the clearance rate of intratesticularly injected ¹³³Xenon dissolved in saline.The testicular blood flow in the rats was measured immediately after injection of 1 or 10 μg PGF_2α into each testis, as well as one and four weeks later. The range of group mean flows in the contral groups was 27.5–37.0 ml/100 g/min. The injection of 1 or 10 μg PGF_2α immediately decreased (p < 0.025 and p < 0.0005) the flow to 12.0 and 4.3 ml/100 g/min, respectively. One and four weeks later, no difference was found between the control and experimental groups. At autopsy during the fourth week, no difference was found between the control and PGF_2α - treated groups with regard to the weight of the adrenals, testes, prostate gland and seminal vesicles, or in the histological appearance of the testes.Unilateral injection of 10 μg PGF_2α decreased the flow through the injected testis and tended to decrease the flow through the non-injected testis.     

Effect of exogenous estradiol and progesterone on the uterine tissue levels of prostaglandin F2α (PGF2α) in ovariectomized mice

Mice ovariectomized for 14 days were treated for 6 days with estradiol and/or progesterone. Both the steroids were effective in increasing the levels of PGF_2α in the uterine tissue, but the treatment with progesterone for 3 days followed by 3 days of estrogen resulted in a highly significant production of PGF_2α. It is concluded that for the production of PGF_2α both estrogen and progesterone are necessary and that the pretreatment with progesterone followed by estrogen results in the maximum production of PGF_2α.     

Prostaglandin F2α receptors in bovine corpus luteum cell membranes. Effect of enzymes and protein reagents

Various enzymes and proteins reagents inhibited [³H]prostaglandin F₂α binding to bovine corpus luteum cell membranes. Studies were undertaken (a) to explore further on the dose response relationships with the above agents, (b) to investigate the mechanism of inhibition of binding with respect to receptor affinities and number and (c) to assess whether decreased binding reflected changes in receptors and/or other membrane components.Preincubation of membranes with phoshpolipase A, trypsin, pronase, lipase, tetranitromethane, dinitrofluorobenzene, acetic anhydride and $\text{N-}\text{ethylmaleimide}$ resulted in moderate to drastic inhibitions of [³H]prostaglandin F₂α binding. The dose-dependent inhibition of binding by enzymes, but not by protein reagents (except for $\text{N-}\text{ethylmaleimide}$), exhibited a biphasic pattern: at lower concentrations, the loss of binding was low and relatively plateaued, but at higher concentrations, the losses were dramatic. The drastic reduction in binding by trypsin was due to destruction rather than solubilization of receptors from membranes. Phospholipase A was intrinsically more effective than phospholipases C and Ca²⁺ was not required for its inhibition of [³H]prostaglandin F₂α binding. Protein reagents inhibition of binding was differently influenced by added Ca²⁺ i.e., loss of binding increased with some ($\text{N-}\text{ethylmaleimide}$), decreased with others (tetranitromethane, dinitrofluorobenzene and azobenzene sulfenylbromide). These results are interpreted to indicate that Ca²⁺ induced conformational changes in membranes which may result in exposure of new groups and burying of already exposed modifiable groups.Treatment of membranes wiht trypsin and $\text{N-}\text{ethylmaleimide}$ selectively abolished high affinity prostaglandin F₂α receptors. The low affinity receptors were present but their numbers as well as their affinity were decreased. Lipase, phospholipase A, acetic anhydride, dinitrofluorobenzen and tetranitromethane appear to decrease binding by totally abolishing all prostaglandin F₂α receptors or by severely reducing their affinities.The occupancy of receptors by prostaglandin F₂α afforded considerable protection against trypsin, phospholipase A, lipase and dinitrofluorobenzene. These data indicated that the inhibition of binding by the above agents, at least in part, can be attributable to changes in receptor sites alone.