Metabolism of arachidonic acid in vitro by ovine conceptuses recovered during early pregnancy

Metabolism of [³H] arachidonic acid ([³H] AA) and synthesis of prostaglandins were examined with ovine conceptuses and endometrial slices collected on various days after mating. Tissues were incubated for 24 hr with or without 5 μCi of [³H] AA and with 200 μg radioinert AA. In experiment 1, results of chromatography indicated that conceptuses collected on days 14 and 16 after mating metabolized [³H] AA to PGE₂, PGF_2α, PGFM, 6-keto-PGF_1α, and to unidentified compounds in three chromatographic regions. One of these regions (region 1) contained triglycerides. Endometrial slices metabolized only small amounts of the [³H] AA to prostaglandins. In experiment 2, results of radioimmunoassays indicated that day 14 conceptuses released somewhat similar amounts (ng/mg tissue) of PGF_2α (32.1 ± 17.9), PGFM (8.4 ± 6.2), PGE₂ (12.3 ± 7.5) and 6-keto-PGF_1α (41.4± 4.8), whereas day 16 conceptuses released more (P<.05) PGF_2α (9.0 ± 4.1) and 6-keto-PGF_1α (15.9 ± 2.7) than PGE₂ (0.9 ± 0.2) or PGFM (0.5 ± 0.08). Day 14 and 16 endometrial slices released (ng/mg tissue) more (P<.05) PGFM (3.0 ± 0.2) and 6-keto-PGF_1α (4.0 ± 0.4) than PGF_2α (0.5 ± 0.08) or PGE₂ (0.05 ± 0.02). In experiment 3, conceptuses were recovered on days 16, 20 and 24 of pregnancy and incubated with [³H] AA to determine the effects of indomethacin on [³H] AA metabolism. In general, indomethacin (Id; 4 × 10⁻⁴ M) reduced (P<.05) the percentage of total dpm recovered as prostaglandins, but Id increased the release of chromatographic region I. Experiment 4 was conducted with day 16, 20 and 24 conceptuses to evaluate the time course of metabolism of [³H] AA, and the appearance of region I and of prostaglandins. In general, the percentage of total dpm in region I increased as the percentage of dpm as [³H] AA decreased. The percentage of dpm as prostaglandins increased as the percentage of dpm in region I decreased. Prostaglandins, probably essential for embroynal survival and development, were synthesized in vitro by ovine conceptuses.     

Prostaglandin E2 opunteracts the effects of PGF2α in indomethacin treated cycling gilts

Twenty crossbred gilts with at least 2 consecutive estrous cycles of 18 to 21 days in length were used to study the effects of prostaglandins E₂ and F₂α (PGE₂ and PGF₂α) on luteal function in indomethacin (INDO) treated cycling gilts. Intrauterine and jugular vein catheters were surgically palced before day 7 of the treatment estrous cycle and gilts were randomly assigned to 1 of 5 treatment groups (4/groups). With exception of the controls (Group I) all gilts received 3.3 mg/kg INDO every 8 h, Groups III, IV and V received 2.5 mg PGF₂; 2.5 mg PGF₂α + 400 μg PGE₂ every 4 hr, or 400μg PGE₂ every 4 h, respectively. All treatments were initiated on day 7 and continued until estrus or day 23. Jugular blood for progesterone analysis was collected twice daily from day 7 to 30. Estradiol-17β (E₂-17β) concentrations were dtermined in samples collected twice daily, from 2 d before until 2 d following the day of estrus onset. When compared to pretreatment values, estrous cycle length was unaffected (P>0.05) in Group I, prolonged (P<0.05) in Groups II, IV and V; and shortened (P<0.05) in Group III. The decline in plasma progesterone concentration that normally occurs around day 15 was unaffected (P>.05) in Group I; delayed (P<0.05) in Groups II, IV and V; and occurred early (P<0.05) in Group III. Mean E₂-17β remained high (31.2 ± 4.9 to 49.3 ± 3.1 pg/ml) in Groups III and IV, while the mean concentrations in Groups III and V varied considerably (17.0 ± 2.0 to 52.2 ± 3.5 pg/ml). The results of this study have shown that PGE₂ will counteract the effects of PGF₂α in INDO treated cycling gilts. The inclusion of PGF₂α appeared to either stimulate E₂-17β secretion or maintain it at a higher level than other treatments.     

Production rates of prostaglandin F, 6-keto-PGF1α and thromboxane B2 by perifused human endometrium

Human endometrium obtained from fresh hysterectomy specimens was perifused for 7 hr in 95% O₂/5% CO₂ at 37°C. The phase of the menstrual cycle was determined by histological examination. The concentrations of PGF, 6-keto-PGF_1α and TxB₂ in 20 min fractions of the perifusion medium were measured by radioimmunoassay and production rates were calculated in terms of dry weight of tissue. Biphasic patterns of production were observed; high initial values fell to about 20% at 2 hr and then increased to relatively stable values at about 4 hr which were maintained for the next 2 hr. During this latter period, production rates in endometria taken at different phases of the cycle differed markedly from each other; the production rates of PGF in secretory and early proliferative endometria were low (15.8 ± 2.6, mean ± SEM and 67.2 ± 8.3 ng/min/g respectively) whereas they were high in late proliferative and premenstrual endometria (188.0 ± 16.7 and 196.4 ± 16.9 ng/min/g respectively). The patterns of production of 6-keto-PGF_1α and TxB₂ were similar to those of PGF but the absolute values were much lower (<10%). We conclude that the observed rates of production of prostaglandins by perifused human endometrium are consistent with synthesis being stimulated either by estrogen or withdrawal of hormonal support and being inhibited by progesterone.     

Seminal prostaglandins in men with subnormal sperm motility and therapeutic treatment

The contents of prostaglandins in seminal plasma from a total of 73 men were evaluated. The subjects were grouped as follows: normospermic men, patients with impaired motility, patients with small untreated varicocelle and patients with impaired motility and Kallikrein therapy. Sperm density, morphology and motility were examined. High performance reversed phase liquid chromatography (HPLC) in combination with specific radioimmunoassays were used for the determination of PGE₂, PGI₂ and PGF_2α. There was a significant difference (p < 0, 025; F-test) between the PGI₂ concentrations in patients with impaired motility (5,6 ± 1,4 pg/mg protein) and normal men (8,8 ± 3,7 pg/mg protein). PGE₂ and PGF_2α were significantly different in patients with varicocele (p < 0,025, F-test). Wide ranges of prostaglandins occured in the Kallikrein-group with no significant differences. We conclude that: a) PGI₁ is an additional prostaglandin compound in seminal plasma. b)its measurement may not be useful as diagnostic parameter in subfertile men and c) Kallikrein has no influence on the prostaglandin content in seminal plasma and other seminal parameters as motility, motility index and sperm counts.     

Prostaglandin synthesis by the cochlea

The exogenous and endogenous syntheses of prostaglandins (PG's) by the cochlea of adult mongolian gerbils were studied . After incubation of the whole membraneous cochlea with [³H]-arachidonic acid (AA), syntheses of PGF_2α, 6-keto PGF_1α, PGE₂, thromboxane (TX) B₂ and PGD₂ were evidenced in this order. The synthesis of radioactive PG's was almost completely inhibited by incubation with 10⁻⁵ M indomethacin. No significant amounts of those PG's were detected by radioimmunoassay (RIA) in the cochlea obtained from animals killed by microwave irradiation at 5.0 kw for 0.8 sec. However, when the homogenate of the whole membraneous cochlea obtained from animals without microwave irradiation was incubated at 37°C for 0–15 min, PGD₂, PGE₂, PGF₂α and 6-keto PGF₁α were found to be formed from endogenous AA in the cochlea by RIA. PG's were formed already at 0 time to considerable level (PGD₂, PGF₂α and 6-keto PGF₁α, 90–120 pg/cochlea; PGE₂, 370 pg/cochlea), reached to the maximum level (PGD₂, PGF₂α and 6-keto PGF₁α, 170–200 pg/cochlea; PGE₂, 500 pg/cochlea) at a 5-min incubation, and then gradually decreased. On the other hand, the amount of TXB₂ was lower than the detection limit by RIA (<50 pg/cochlea) even after the incubation. The cochlea was dissected into three parts: organ of Corti + modiolus (OC + M), lateral wall (LW), and cochlear nerve (CN), and then PG's formed by these tissues were determined after a 5-min incubation of the homogenates. In the CN and OC + M, PGE₂ was the major PG (100 and 160 pg/tissue, respectively), and the amounts of PGD₂, PGF₂α and 6-keto PGF₁α were about of those of PGE₂. In the LW, the amounts of PGD₂, PGE₂, PGF₂α and 6-keto PGF₁α were about the same level (70–100 pg/LW).     

Effects of endometrium on metabolismof arachidonic acid by bovine blastocysts

Metabolism of radiolabeled arachidonic acid (^*AA) by blastocysts and endometrial slices recovered from five gilts 16 days after detection of estrus was studies . Blastocysts from each gilt were divided into four 216 ± 18 mg, and each portion was placed into a separate petri dish containing 15 ml modified minimum essential medium (MEM)_. The incubates from each gilt received either 25, 50, 100 or 200 μg radioinert arachidonic acid (AA). Endometrium was dissected from each uterin horn, sliced and duplicate 509 ± 3 mg portions from each gilt were placed into petri dishes containing 15 ml MEM and 200 μm AA. All incubates received 5 νCi of ^*AA (either [¹⁴C]-arichidonic acid or [³H]-arichidonic acid). The incubates were rocked at 37°C for 24 h in an atmosphere of 50% n₂:45% O₂:5% CO₂. After incubation, tissues and MEM were separated by centrifugation. Metabolism of ^*AA was assessed in extracts of MEM and tissue homogenates by separating ^*AA and its metabolites on columns of Sephades LH-20. Blastocysts produced compounds that migrated with [³H]-13,14-dihydro-15-keto-PGF_2^α (^*PGFM), [³H]-PGE₂ (^*PGE₂) and [³H]-PGF_2^α (^*PGF_2^α). The greatest (P<.05) proportion (35.7 ± 1.8%) of the radioactivity in blastocyst MEM was recovered as PGE₂. In blastocyst homogenates, most (66.2 ± 3.3%; P<0.05) of the radioactivity was in a nonporal peak assumed to be arachidonate esters. The concentration of AA ni MEM did not alter metabolism of ^*AA by blastocysts. Endometrial slices produced ^*PGFM and ^*PGE₂ but only in small amounts, and they were capable of producing nonpolar, probably esterified, forms of ^*AA. It was concluded that porcine blastocysts produced and metabolized prostaglandins and that they make a contribution to the uterine milieu during early pregnancy.     

Effect of estradiol-17β on peripheral plasma concentration of 15-keto,14-dihydro PGF2α and luteolysis in cyclic cattle

Friesian heifers (n = 10) were assigned randomly to receive an intravenous injection of estradiol-17^β (E₂; 3 mg) or saline: ethanol vehicle solution (6 ml; 1:1) on day 13 of the estrous cycle. Blood was collected collected from the jugular vein by venipuncture into heparinized vacutainer tubes at 30 minute intervals for 2 hours (h) preinjection, 10.5 h postinjection and then at 3 h intervals until estrus. Repeated hormone measurements of 15-keto-13,14-dihydro-PGF_2α (PGFM) and progesterone (P₄) were evaluated by split-plot analysis of variance. Mean concentration of PGFM for the 12.5 h acute sampling phase was 164.1 ± .14 pg/ml. A treatment by time interaction was detected (P < .01). After treatment with E₂, PGFM concentrations began to increase at approximately 3.5 h, reached a mean peak of 330.4 ± 44.5 pg/ml (n = 5) at 5.5 ± .3 h, and returned to basal concentration by 9.0 ± .6 h. Vehicle treatment did not alter concentrations of PGFM. Injections of E₂ on day 13 of the estrous cycle caused luteolysis (P₄ concentration < 1 ng/ml) to occur earlier following injection (96.9 ± 10.6 h < 153.6 ±17.7 h; P, 0.05) than did the vehicle control treatment. During the chronic sampling phase of 3 h intervals, 39 of 606 samples (6.4%) were classified as PGFM spikes (323.0 ± 50.0 pg/ml); 21 (53%) of the spikes occurred at a mean interval of 18.9 ± 3.86 h before the time of completed luteolysis. Exogenous E₂ induced an acute increase in PGFM that may be indicative of uterine PGF_2α production. Peaks of PGFM in plasma were temporally associated with luteolysis on a within cow basis.     

Sow (Sus scrofa) follicular fluid: Prostaglandin content and effect on the motility of isolated oviducts

The present study provides information regarding the effects of the sow follicular fluid (FF) on the motility of isolated segments of swine and rabbit oviducts. In addition, the concentration of prostaglandins (PGs) F_{2 α}, E₂ and E₁ in the follicular fluid of sow ovaries isolated at different stages of the sex cycle as well as the generation of the same PGs by walls of ovarian follicles in early and late proestrus, in estrus, in metestrus and in diestrus, were explored. The stimulatory contractile effect of proestrous FF in isolated segments of sow fimbria was antagonized by polyphloretin phosphate (PPP), a PG receptor blocker and by indomethacin, an inhibitor of PG synthesis. The positive inotropism evoked by the FF was mimiked by bradykinin and the influences of both interventions were similarly antagonized by PPP. It appears plausible that the inotropic effect of the preovulatory FF on the sow fimbria could be not only by PGs already present in the fluid, but also by the stimulation of the synthesis of tubal PGs by follicular fluid bradykinin. The FF also stimulated the ampullary tubal segments isolated from proestrous sows whereas the same volume of FF depressed significantly the isometric developed tension of rabbit ampulla. The total concentration of the three PGs in the FF from late proestrous follicles was significantly greater than that of the same PGs in the other two stages of the sex cycle (early proestrus and diestrus), whereas the concentration of each PG (PGE₂, PGF_{2 α} or PGE₁), did not differ within any of the stages of the cycle. Furthermore, the total amount of the three PGs produced by the walls of follicles from late proestrous ovaries was also significantly greater than that generated by ovarian follicles from early proestrus, estrus, metestrus and diestrus.     

Effects of inhibitors of thromboxane synthesis on reactions of the cat bronchus in situ to prostaglandins

Cats were anesthetized by chloralose, relaxed by suxamethonium, and ventilated. The influence of 3 inhibitors of thromboxane synthesis, N.0096 (±α-benzyl-α-p-chlorobenzyl-4-hydroxyacetophenon phenylhydrogen-phosphonate), imidazole and dipyridamole were studied on bronchial reactions to prostaglandins E₂ and F₂α and arachidonic acid. During a bronchoconstriction maintained by infusion of 5-HT, N.0096 and imidazole dose-dependently potentiated the intensity and duration of the bronchodilator effect of PGE₂. This effect was maximal at a total dose of 17 mg/kg N.0096 or 25–50 mg/kg imidazole, but decreased after higher doses which are supposed to inhibit cyclo-oxygenase also. Bronchoconstrictor reactions to single injections of arachidonic acid were inhibited by N.0096 in the same dose range. Reactions to PGF_2α were unaltered by both drugs. Dipyridamole was devoid of a comparable activity.     

In vitro prostaglandin synthesis by human glomeruli and papillae

We have investigated in vitro prostaglandin synthesis by human isolated glomeruli and papillary homogenates and compared the results with those obtained in parallel studies using rat material. Prostaglandins were measured by two methods, namely radiometric high performance liquid chromatography after incubation with ¹⁴C arachidonic acid and radioimmunoassay. The relative abundance of various prostaglandins synthesized by glomeruli was different in man (6 keto PGF_1α > TXB₂ > PGF_2α > PGE₂) and in the rat (PGE₂ TXB₂ > 6 keto PGF_1α). Unidentified peaks eluting between 6 keto PGF_1α and TXB₂ were observed only in rat glomeruli. These peaks were suppressed by indomethacin. Direct radioimmunoassay of prostaglandins in the incubation medium of human glomeruli confirmed the predominance of 6 keto PGF_1α synthesis and showed its stimulation by arachidonic acid, its progressive decrease with time and its linear relationship with glomerular protein at low concentrations. On the contrary, the profile of prostaglandin synthesis by the papilla was similar in man and in the rat, PGE₂ and PGF_2α being the major products in both species. However, related to one mg of protein, papillary synthesis of these two prostaglandins was greater in the rat. These results show that PGI₂ is the major prostaglandin synthesized in human glomeruli and suggest a role for this prostaglandin in glomerular physiology in man.     

Effect of indomethacin on the adrenal renin response to nephrectomy in the rat

The role of prostaglandins in the control of adrenal renin in vivo was evaluated in nephrectomized rats. Nephrectomy increased adrenal renin from 13.2 ± 1.37 ng angiotensin I/mg protein/hr to 166.5 ± 17.3 ng angiotensin I/mg protein/hr. Indomethacin treatment significantly suppressed the adrenal renin response to nephrectomy. (47.8 ± 5.22 ng angiotensin I/mg protein/hr). Adrenal aldosterone was also suppressed by indomethacin. Adrenal prostaglandin E₂ increased after nephrectomy and decreased after indomethacin.Plasma corticosterone and serum potassium did not change after indomethacin. These data indicate that inhibition of prostaglandin synthesis by indomethacin partially blocks the adrenal renin response to nephrectomy, suggesting that prostaglandins may play a role in the adrenal response to nephrectomy.     

Indomethacin inhibits the uptake of sodium by ovine trophoblastic tissue in vitro

Blastocysts from several species synthesize prostaglandins in vitro, but the exact functions of the prostaglandins are unknown. The purpose of this study was to determine if indomethacin, an inhibitor of prostaglandin synthesis, would inhibit the uptake of ²²sodium ([²²Na]) by ovine trophoblastic tissue. To determine the concentration of indomethacin that would inhibit the synthesis of PGF_2α and 13,14-dihydro-15-keto-PGF_2α (PGFM) by blastocysts, blastocysts were collected from ewes 16 days after mating, sliced into pieces approximately 2 mm in length and incubated for 48 h at 37°C in 2 ml of medium containing either 0, 0.2, 0.4, 0.8 or 1.6 mM of indomethacin. Concentrations of indomethacin mM reduced (P<.01) trophoblastic release (ng/μg DNA) of PGF_2α from

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, reduced PGFM from

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, and inhibited formation of trophoblastic vesicles. In a second experiment, blastocysts were recovered from ewes 16 days after mating and pieces of trophoblast were incubated with [²²Na] and either 0 or 0.4 mM of indomethacin. Indomethacin reduced the uptake of [²²Na], which is an indirect measure of the transport of water across epithelia, from 3680 ± 1118 to 934 ± 248 cpm/μg DNA (P<.03) and prevented formation of trophoblastic vesicles. Prostaglandins produced by ovine blastocysts might be involved in controlling uptake of water, which is essential for expansion of blastocysts.     

Simple direct radioimmunoassay of the F prostaglandins

A simplified and accurate method of determining the F prostaglandins in 0.1 ml of serum without previous extraction is described. The procedure involves addition of anti-prostaglandin F_2α to serum followed by tritiated prostaglandin, equilibration for 4 hours, removal of unbound prostaglandin with dextran-coated charcoal and subsequent liquid scintillation counting of the supernatant. The mean ± S.D. concentration of prostaglandin F_2α in the serum of 15 healthy men was 90 ± 33 pg/ml and in 20 women 108 ± 43 pg/ml.     

Influence of the estrous cycle and prostaglandins on utero-ovarian vein blood flow in the rat

The blood flow rate in the utero-ovarian vein (UOV) has been measured in adult female rats during the different phases of the estrous cycle. It was observed that the blood flow rate in the UOV is high at proestrus and at estrus and low during diestrus days 1 and 2. The intravenous injection of 10 μg PGF_2α or PGE₂ diminishes the blood flow rate in the UOV. The efficacy of the two PGs in reducing blood flow is different in the various phases of the estrous cycle, being maximal during the day of estrus.     

Influence of the estrous cycle and prostaglandins on utero-ovarian vein blood flow in the rat

The blood flow rate in the utero-ovarian vein (UOV) has been measured in adult female rats during the different phases of the estrous cycle. It was observed that the blood flow rate in the UOV is high at proestrus and at estrus and low during diestrus days 1 and 2. The intravenous injection of 10 μg PGF_2α or PGE₂ diminishes the blood flow rate in the UOV. The efficacy of the two PGs in reducing blood flow is different in the various phases of the estrous cycle, being maximal during the day of estrus.     

Prostaglandin (PG) analysis in urine of humans and rats by different radioimmunoassays: Effect on PG-excretion by PG-synthetase inhibitors, laparotomy and furosemide

Thw radioimmunological (RIA) determination of prostaglandin (PG) E₂ and of PGF_2α in urine humans and rats is described in detail. After extraction and chromatography PGE₂ was determined by using a PGE specific antibody or by using either PGB or PGF_2α specific antibodies after the respective conversion procedures. The three different RIA procedures were compared to each other. PGF_2α was determined by a specific antibody to PGF_2α. Basal excretion of PGE₂ and of PGF_2α in healthy women on free diet was 9.3 ng/hour ± 0.96 and 18.3 ng/hour ± 2.5 respectively. Furosemide increased the excretion of PGE₂ and of PGF_2α in humans significantly, while PG-excretion rates decreased on indomethacin. In rat urine PGE₂ and PGE_2α increased markedly from 46.2 pg/min ± 9.3 and 27 ± 3.4 to 253.8 ± 43.3 and 108 ± 12.6 pg/min (per one kidney) in the anesthetized-laparotomized animal. This increase was abolished after giving two different PG synthetase inhibitors.     

Effects of superovulation, arachidonic acid or endometrium on release of prostaglandins and synthesis of proteins by bovine trophoblast

This study was conducted in vitro to examine factors that may regulate prostaglandin release by bovine trophoblast and endometrial slices. Trophoblastic tissues and endometrial slices were recovered from superovulating and normally-ovulating cattle on day 16 or 20 of pregnancy and incubated for 24 h. Release of PGF_2^α and 13,14-dihydro-15-keto-PGF_2^α (PGMF), and incorporation of [¹⁴C]-leucine into proteins were quantified and expressed per μg DNA, which gives a measure of cellular activity. Activity of trophoblastic tissue for synthesizing protein was decreased (P<.05) and for releasing PGMF was increased (P<.05) on day 20 compared to day 16 of pregnancy. Neither supercovulation nor day of pregnancy altered trophoblastic activity for releasing PGF_2^α. Supercovulation increased (P<.05) endometrial release of PGF_2^α. Endometrial release of PGF_2^α was less (P<.05) on day 20 than on day 16 of pregnancy. When arachidonic acid (0, 100, 200 or 400 μg) was added at the start of incubation, trophoblastic release of PGF_2^α changed (P<.05) quadratically with dose of arachidonic acid. When arachidonic acid was added 8 h after the start of incubation, triphoblastic release of PGF_2^α increased linearly (P<.01) with dose of arachidonic acid. Adding arachidonic acid to incubation medium did not affect trophoblastic or endometrial protein synthesis. Endometrial slices suppressed (P<.05) trophoblastic protein synthesis and release of PGF_2^α. Apparently, endometrium can modulate trophoblastic release of prostaglandins and synthesis of proteins in vitro, and trophoblastic tissue from supercovulated cattle 16 or 20 days pregnant can be used to study trophoblastic synthesis of prostaglandins and proteins.     

Renal sodium handling and stimulation of medullary Na---K---ATPase during blockade of prostaglandin synthesis

The effect of suppression of prostaglandin synthesis on renal sodium handling and microsomal Na---K ATPase was studied in control and indomethacin treated intact rats maintained on a normal sodium diet (series A) and chronically salt loaded (series B). Indomethacin administration resulted in a decreased GFR and a significantly depressed urinary excretion and an increased fractional reabsorption of sodium in animals fed the normal sodium diet or chronically salt loaded. In rats maintained on a nomral Na diet, the activity of the renal medullary Na---K ATPase after indomethacin was 206.3±6.4 ug Pi./mg protein, i.e. significantly higher as compared with the enzyme activity in the medullary renal fraction from control animals in which it averaged 148±7.79 ug Pi/mg protein (p<0.001). While after chronic salt load a similar increment in the activity of renal medullary Na---K ATPase was observed, no additional stimulation was elicited by subsequent indomethacin administration. The addition of exogenous PGE₂, mM to microsomal fractions obtained from kidneys of normal rats, was associated with a moderate suppression of the medullary Na---K---ATPase activity, from a basal level of 170±16 to 151.3±13 umol Pi/mg protein/hr (p<0.005. In isolated segments of medullary thick ascending limb of Henle's loop (MTAL) addition of PGE₂ to the incubation medium resulted in a significant inhibition of Na---K--- ATPase from 37.2±2 to 21.25 ± 1.17 × 10⁻¹¹ mol/mm/min (p<0.0001.These findings suggest that the increased renal Na reabsorption after inhibition of PG synthesis might be related, at least partly, to stimulation of medullary Na---K ATPase. In parallel, the reported natriuretic effect of prostaglandins might imply a direct inhibitory effect of these mediators on renal Na---K ATPase.     

Modulation of human myometrial PGE2 receptor by GTP characterization of receptor subtype

We studied PGE₂ specific binding sites in human myometrial microsomes prepared from uterine specimens obtained by hysterectomy (women between 38 and 55 years of age). Competition experiments showed that the potency order for various prostaglandins (PGs) was : PGE₂ ≥ PGE₁ PGF_2α > Iloprost ≥ Carbacyclin ZK 110841 (PGD₂ analogue). These relative affinities indicated that the receptor was of the EP type.In kinetic experiments GTP, GppNHp and GTPγS increased the rate of PGE₂ binding (steady state was reached more rapidly in the presence of nucleotides) but maximal specific binding was not significantly different. Complete dissociation could not be obtained, even in the presence of GTP. Only 50% of maximal binding was readily dissociable. The dissociation rate was 4.56.10⁻⁴ sec⁻¹ (half time of about 660 sec) and in the presence of GTP analogues it was slightly increased (k−1 = 7.16 10⁻⁴ sec⁻¹ half time 420 sec.). Scatchard analysis of saturation curves showed an increase in ligand receptor affinity in the presence of GTP or nucleotide analogues: the Kd shifted from 9.66 ± 2.8.10⁻⁹ M to 4.96 ± 1.25.10⁻⁹M, but the number of binding sites did not change significantly (310 ± 37 to 350 ± 17 fmol/mgP). The effect of GTP was observed at a concentration of 5.10⁻⁴M. GppNHp and GTPγS were effective at 1.10⁻⁵M. Pretreatment of myometrial membranes with pertussis or cholera toxins had no effect on PGE₂ binding to membrane sites. Our conclusion is that GTP induced conversion of a population of low affinity sites into a population of higher affinity sites. This effect of guanine nucleotides was described in adipocytes and kidney medulla.Competition studies with PGE₂ analogues (sulprostone, 17-phenyl-ω-trinor PGE₂, M&B 28,767, misoprostol, butaprost) showed that this receptor mediates a contractile response and is probably an EP₃ subtype.     

Luteolysis, estrus and ovulation, and blood prostaglandin F after intramuscular administration of 15, 30 or 60 mg prostaglandin F2α

During diestrus in three consecutive estrous cycles, each of six heifers was given (im) 30 mg, 15 mg (twice at 6-hr intervals) and 60 mg prostaglandin F_2α (PGF_2α) tham salt. Neither the decline in blood progesterone, the increase in blood estradiol, the duration or the peak of the LH surge, the interval to onset of estrus, nor the interval to ovulation was affected significantly by dose of PGF_2α. Thus, relative to that after 30 mg PGF_2α im, two injections of 15 mg at 6-hr intervals or 60 mg PGF_2α did not hasten luteolysis. Thirty mg was an ample im dose of PGF_2α to cause luteolysis. Regardless of im dose of PGF_2α, blood PGF peaked at about 6.0 ng/ml within 10 minutes and returned to basal values (<1.0 ng/ml) within 90 minutes. In another trial, after a single iv injection of 5 mg PGF_2α, blood PGF peaked (25 ng/ml) within 5 minutes and returned to basal values within 15 minutes. During a 30-minute infusion (0.5 mg/minute) of PGF_2α, blood PGF plateaued at 29.5 ng/ml with a metabolic clearance rate of 17.0 liters per minute.