Effect of chronic alcohol consumption on rat uterine spontaneous motility, prostaglandin production and triglyceride metabolism

The spontaneous isometric developed tension (IDT), the synthesis and release of prostaglandins (PGs) into the incubating medium and the metabolism of triglycerides (TGs) in uterine strips isolated from controls and chronic ethanol fed rats, were studied. In order to observe how the uterus of rats fed alcohol reacts during a situation of metabolic emergency, the above mentioned studies were done in the presence or in the absence of glucose in the incubating medium. The decrement of IDT as time progressed was significantly greater in strips obtained from rats which had been drinking 20% ETOH than in controls. Nevertheless, the absolute magnitude of the initial IDT was similar in both groups. On the other hand, the decline of the frequency of contractions (FC) of uterine strips isolated from controls and from ETOH-exposed rats, after 60 min of spontaneous activity was similar. When the uterine strips isolated from ETOH-exposed and from control rats were suspended in glucose-free solution they exhibited the same decrement of IDT and FC after 60 min of activity. The basal release of PGE1 and PGE2 was similar in control tissues incubated in medium containing glucose, but the output of PGE2 was significantly smaller than that of PGE1 in uterine strips isolated from ETOH-exposed rats. The production of PGE1 and PGE2 by uteri suspended in glucose-free medium was similar in control preparations. On the contrary the release of both PGs differs in uterine strips from ETOH-exposed rats, i.e. the output of PGE2 was significantly smaller than in controls and the release of PGE1 increased around 4-fold in comparison with controls.(ABSTRACT TRUNCATED AT 250 WORDS)     

Histamine alters prostaglandin output from diestrous rat uteri. Involvement of H2-receptors and 9-keto-reductase

The effects of exogenous histamine (H) on prostaglandin (PG) generation and release in uteri isolated from diestrous rats and the influences of H2-receptors blockers (cimetidine and metiamide) on the output of uterine PGs, were explored. Moreover, the action of H on the uterine 9-keto-reductase, was also studied. Histamine (10(-4) M) failed to alter the basal output of PGE1 but reduced significantly the generation and release of PGE2 and augmented the output of PGF2 alpha. On the other hand, cimetidine (10(-5) M) enhanced the basal release of PGE2 but had no action on the outputs of PGs E1 or F2 alpha. The enhancing effect of H on the production and release of PGF2 alpha was abolished in the presence of cimetidine. Also, the antagonist reversed the influence of H on the output of PGE2. Metiamide, another H2-receptor antagonist, did not alter the basal control generation and release of uterine PGs, but antagonized the augmenting influence of H on PGF2 alpha uterine output, as much as cimetidine did, and prevented the depressive action of H on the release of PGE2 from uteri. Histamine (10(-4) M) significantly stimulated uterine formation of cyclic-adenosine monophosphate, an action which was antagonized by the presence of cimetidine (10(-5) M), a blocker of H2 receptors. Also, histamine (10(-5) M) and dibutyrylcyclic-adenosine monophosphate (DB-cAMP) at 10(-3) M, enhanced significantly the formation 3H-PGF2 alpha from 3H-PGE2. Results presented herein demonstrate that H is able to diminish the generation of PGE2 in uteri from rats at diestrus augmenting the synthesis of PGF2 alpha, apparently via the activation of H2-receptors, enhancing adenylate-cyclase. These effects appear to increase uterine 9-keto-reductase activity which transforms PGE2 into PGF2 alpha. Relationships between the foregoing results and those evoked by estradiol, are also discussed.     

Effect of chronic underfeeding on uterine glycogen of rats. Influence of indomethacin and nordihydroguaiaretic acid

Glycogen values in uterine strips isolated from normal-fed estrous or diestrous rats, or from rats fed a restricted diet (50% of normal food intake for 25 days) were measured. Determinations were made immediately after killing (0 time or post-isolation) as well as after incubation in glucose-free medium (60 min time or post-incubation). The post-incubation levels of glycogen in the uteri from normal-fed animals diminished significantly in comparison to post-isolation values, and this decrement was not modified by the addition of indomethacin, nordihydroguaiaretic acid or exogenous prostaglandins E1, E2 or F2 alpha. In rats fed a restricted-diet, the initial glycogen values (0 time) were significantly lower than in normal-fed controls, but did not decline further after incubation in glucose-free medium (60 min time). The addition of indomethacin, acetylsalicylic acid or of nordihydroguaiaretic acid led to a significant fall in the glycogen levels, and exogenous PGE1, PGE2 or PGF2 alpha failed to alter the effects of the inhibitors. The values of PGE and PGF prostaglandins release to the medium by the uterus from restricted-diet rats did not differ from those obtained in the experiments with normal-fed animals. Administration of 17-beta estradiol to restricted-diet rats led to suppression of the effects of this diet on the glycogen concentration. The above results indicate that in rats subjected to a prolonged period of dietary restriction, the uterine glycogen becomes responsive to the effects of cyclooxygenase and lipoxygenase inhibitors, suggesting the operation of some regulatory mechanism during critical periods of nutrition.     

The biosynthesis of the 3-series prostaglandins in rat uterus after alpha-linolenic acid feeding: Mass spectroscopy of prostaglandins E and F produced by rat uteri in tissue culture

The abnormal uterine activity associated with dietary n-3 fatty acids may result from competitive inhibition of PG2 production. Uterine synthesis of 2- and 3-series prostaglandins F(PGF) and E(PGE) was studied using mass spectrophotometry in rats fed diets containing predominantly n-3 fatty acid, n-6 fatty acid, or control pelleted diet. Mass spectra of PGF (Me, TMS and Me, TBDMS derivatives) synthesised by uteri of n-3 fed rats were characterised by 8 ions containing the n-3 double bond, and m.i.d. of the 651/653 ions of PGF-Me, TBDMS indicated PGF3 alpha synthesis (44 +/- 8% and 13 +/- 2% of PGF release by uteri incubated + or -5 micrograms/ul calcium ionophore A23187 respectively). In uteri from the control diet group incubated with ionophore, PGF3 alpha ions were detected and PGF 3 alpha represented 9.5 +/- 1.0% of PGF alpha release. Similarly, analysis of PGE from uteri of n-3 fed rats indicated that PGE3 (16 +/- 6% of PGE) was released in the presence of ionophore A23187. Synthesis of 3-series PG by rat uteri was detected after only 3 weeks of n-3 diet. The capacity to synthesise 3-series PG increased at intracellular calcium concentrations which mimicked cell calcium during decidual autolysis at parturition. These experiments suggest that uterine synthesis of 3-series PG is regulated by the specifity of enzymes incorporating fatty acids, rather than by the cyclooxygenase enzyme.     

Oxytocin enhances the basal release of uterine prostaglandin F2 alpha, but not that of PGE1, or of PGE2, and changes the metabolism of exogenous arachidonate, favouring the formation of prostaglandin F2 alpha and 5-HETE. Relationships with its uterotonic action and modulation by estradiol

We attempted to explore possible mechanism(s) subserving the influence of oxytocin on uterine motility by studying the action of the hormone on: 1) the contractile activity of isolated rat uteri in the presence or absence of indomethacin; 2) the synthesis and release of prostaglandins (PGs) into the solution incubating the uterine tissue as well as the metabolism of labelled arachidonic acid; 3) the uptake of 45Ca2+ by uterine strips. The experiments were bone with uterine preparations isolated from spayed rats treated or not with 17-beta-estradiol. The values of isometric developed tension (IDT) and of frequency of contractions (FC) induced by oxytocin in uterine strips isolated from spayed and spayed-estrogenized rats, were not modified by indomethacin at 10(-6) M. On the other hand, uterine strips from untreated spayed rats, release into the incubating medium approximately equal amounts of PGE1, PGE2 and PGF2 alpha. The in vitro presence of oxytocin (50 mU/ml) increased significantly (p 0.05) the output of PGF 2 alpha without changing the release of PGE1 or PGE2. Uteri from spayed rats injected prior to sacrifice with 17-beta-estradiol released significantly less PGE1 and PGE2 (p less than 0.005) than preparations from non-injected animals, whereas the output of PGF2 alpha in the suspending solution remained unchanged. Following estrogenization the addition of oxytocin to preparations obtained from spayed-estrogenized rats also increased the output of uterine PGF2 alpha (p less than 0.001) without changing that of PGs E1 or E2.(ABSTRACT TRUNCATED AT 250 WORDS)     

Tissue levels of prostaglandins and what do they mean? Studies on the guinea-pig uterus

The ratios of the concentrations of PGF2 alpha, PGE2 and 6-keto-PGF1 alpha in guinea-pig uterine horns, which were removed and placed in ethanol in 1.5 to 2 min, were 0.3:1.0:0.6 on day 7 and 13.8:1.0:0.8 on day 15 of the oestrous cycle. Adding indomethacin (10 micrograms/ml) to the ethanol had no significant effect on the tissue levels observed. These ratios were similar to the ratios of the outputs of PGF2 alpha, PGE2 and 6-keto-PGF1 alpha from the guinea-pig uterus (0.6:1.0:0.9 on day 7 and 7.6:1.0:1.5 on day 15), but were different (particularly on day 7, but only for 6-keto-PGF1 alpha on day 15) to the ratios of the amounts of the three PGs synthesized by homogenates of the guinea-pig uterus (7.2:1.0:2.4 on day 7 and 11.7:1.0:3.3 on day 15). Consequently, the measurement of tissue levels of PGs in the guinea-pig uterus reflects PG synthesis by intact tissue and changes in this synthesis, rather than PG synthesis by homogenates (broken cell preparations). Therefore, it appears meaningful to measure levels of PGs in the guinea-pig uterus since they reflect uterine PG output. Separation of the endometrium from the myometrium, which involved handling and mild trauma, stimulated uterine PG levels, but the ratio of the levels of PGF2 alpha, PGE2 and 6-keto-PGF1 alpha in the endometrium was still similar to that found in the non-separated uterus.     

Electric field stimulation alters the outputs of prostaglandins from isolated rat urinary bladder preparations. Influences of papaverine and tetradotoxin

The effects of electric field stimulation (EFS) on the outputs of prostaglandins (PGs) E1, E2 and F2 alpha from isolated contracting rat urinary bladders, were explored. Also, the influences of papaverine (5.10(-6) M) and of tetradotoxin (TTX = 5.10(-7) M) on PGs released by spontaneously contracting or electrically driven preparations, were tested. The basal control outputs of the three PGs in spontaneously contracting urinary bladders, had a comparable magnitude. On the contrary, in electrically stimulated preparations the output of PGE2 rose significantly; that of PGF2 alpha presented a significant reduction and the release of PGE1 was similar to that in controls. Papaverine failed to modify the profile of basal control PG release in non-stimulated bladders, abolished contractile responses to EFS and blocked the augmented output of PGE2 elicited by EFS. The presence of TTX in the suspending solution had no action on the basal control release of PGs from non-stimulated spontaneously contracting preparations, depressed between 80-90% the inotropic responses triggered by EFS and completely antagonized the enhanced output of PGE2 evoked by EFS. Results are discussed in terms of the relative participations of the evoked inotropism of the detrusor muscle or by the stimulation of nerve endings, accounting for the greater release of tissue PGE2 after EFS.     

Uterine and placental prostaglandins and their modulation of oxytocin sensitivity and contractility in the parturient uterus

The parturient uterus develops a markedly enhanced sensitivity to the uterotonic action of oxytocin (OT). The mechanism leading to this enhanced OT sensitivity is not known. Our previous work suggested that prostaglandins (PGs) may be involved. To define the relationship between OT sensitivity and uterine PG production, we measured uterine sensitivity to OT by a quantitative dose-response procedure in rats on Days 19, 20, 21 and 22 of pregnancy and monitored uterine and placental tissue concentrations of PGF2 alpha and PGE2. In addition, we determined the effects of inhibition of endogenous PG synthesis on OT sensitivity and uterine contractility. We found that both OT sensitivity and spontaneous contractility are positively related to uterine PGF2 alpha production. An abrupt increase in OT sensitivity was observed on Days 21 and 22 of pregnancy. The increase in OT sensitivity was coincidental with the marked increase in PGF2 alpha production in the uterus on Days 21 and 22 of pregnancy. Suppression of in vivo PG synthesis caused a reduction in both spontaneous uterine contractility and OT-induced contractions. Uterine PGE2 concentrations and release were 3-5 times lower than PGF2 alpha. There were no significant fluctuations of uterine PGE2 concentration measured on these last 4 days of gestation. Placental PG levels were also found not to be related to uterine contractility. Placental PGE2 levels were higher than PGF2 alpha and may play a regulatory role in placental perfusion. However, placental PGs did not vary with gestational age.     

Effect of acute surgical vagotomy on the mucosal content of 6-keto-PGF1 alpha, PGE2 and glutathione after intragastric 96% ethanol treatment in rats.

It has been observed earlier that gastric cytoprotection produced by PGI2, beta-carotene, small doses of atropine or cimetidine has failed in surgically vagotomized rats. This phenomenon may be in connection with endogenous prostaglandins (PGs) and glutathione (GSH) level of the gastric mucosa. The aims of the study were to evaluate the effect of vagus nerve on the gastric mucosal 6-keto-PGF1 alpha, PGE2 and glutathione after intragastric 96% ethanol (ETOH) treatment. The observations were carried out on CFY rats. The gastric mucosal damage was produced by intragastric administration of 1 ml 96% ETOH. Acute bilateral surgical vagotomy (ASV) was carried out 30 min prior to ETOH application. The animals were sacrificed 1, 5, 15 or 60 min after ETOH installation. The number and the severity of gastric mucosal lesions were noted and 6-keto-PGF1 alpha, PGE2 an GSH contents of gastric mucosa were measured. It has been found that: 1. the number and the severity of gastric mucosal lesions were increased after ASV compared to those with intact vagal nerve, 2. 96% ETOH treatment increased both the gastric mucosal PGs and GSH levels, 3. 6-keto-PGF1 alpha peaked at 5 min PGE2 and GSH peaked at 15 min after ETOH treatment, 4. ASV decreased the gastric mucosal PGs content and delayed the peaks of PGE2 and GSH. It has been concluded that the decreased content of PGs and the delayed GSH increase may play a pathological role in the failure of gastric cytoprotection of rats after ASV.     

Effects of experimental diabetes on spontaneous contractions, on the output of prostaglandins and on the metabolism of labelled arachidonic acid, in uteri isolated from ovariectomized rats. Influences of estradiol

Spontaneous changes in isometric developed tension (IDT) as a function of time after isolation (contractile constancy) in uteri from control-castrated and castrated chronic streptozotocin-diabetic rats, were explored. The effects of injecting 17-beta estradiol (Eo) were also studied. No differences in the minor changes of contractile constancy, between control and diabetic preparations, during a period of 60 min, were detected, whereas uteri from non-diabetic Eo injected animals (0.5 + 1.0 ug, prior to sacrifice), exhibited a profound reduction of IDT, significantly greater than in tissues obtained from Eo injected-diabetic rats. Moreover, basal generation and outputs into the suspending solution of prostaglandins (PGs) E1, E2 and F2 alpha, were explored in the same groups, at 60 min following tissue isolation. The basal outputs of these three PGs were similar in castrated control rats, but preparations from castrated-diabetics released significantly more PGE1. The administration of Eo to castrated-diabetics, failed to alter the releases of the three PGs explored. In addition, the metabolism of labelled arachidonic acid (AA) into different prostanoids (6-keto-PGF1, PGF2, PGE2 and thromboxane B2-TXB2), was also investigated. The non-diabetic spayed rat uterus converted AA into these four prostanoids, the transformation into 6-keto-PGF1 alpha (as an index of PGI2 formation) being the most prominent. In preparations from diabetic rats the formation) being the most prominent. In preparations from diabetic rats the formation of 6-keto-PGF1 alpha, PGF2 alpha and PGE2, was significantly smaller than in controls, whereas a greater % of TXB2 formation (as an index of TXA2), was detected. On the other hand uterine preparations from non-diabetic spayed rats injected with Eo formed less 6-keto-PGF1 alpha and PGE2 and similar amounts of PGF2 alpha or of TXB2 from AA, than Eo injected controls, whereas uteri from castrated diabetic animals injected with Eo, formed a similar % of 6-keto-PGF1 alpha, PGF2 alpha and PGE2 from AA, than tissue preparations from non-estrogenized controls. However, the enhanced transformation of the labelled fatty acid precursor (AA) into TXB2 in the diabetic group, was significantly reduced by the steroid. The role of the augmented generation and release of PGE1 in uteri from diabetic rats is discussed in terms of precedents indicating the relevance of PGs type E supporting rat uterine motility. In addition the influence of Eo is attractive, because its reducing effect on TX production, in diabetes, a disease known to be accompanied by enhanced synthesis of vasoconstrictor and platelet aggregation TXA2, and by frequent obstructive circulat     

Prostaglandin translocation from the lumen of the rabbit uterus in vitro in relation to day of pregnancy or pseudopregnancy

Transport of 3H-labeled prostaglandins (PGs) E2 and F2 alpha from the uterine lumen across the uterine wall has been studied in rabbit uteri in vitro in incubations lasting up to 180 min, in relation to sexual state of the rabbit, incubation temperature, intraluminal PG concentration, addition of metabolic inhibitors and time of incubation. PG accumulation by the tissue increased rapidly up to 30 min and then remained relatively constant. By 30 min, radioactivity was found in the external incubation medium, and this increased linearly with time. The translocation of PGF2 alpha was significantly greater in pseudopregnant than in pregnant animals on Day 6, whereas that of PGE2 was significantly higher in pregnant than in pseudopregnant animals on Day 6.8. In pregnant animals, both PGF2 alpha and PGE2 were translocated to the exterior more rapidly on Day 6.8 than on Days 5 or 6. Transport of PGs was reduced by low temperature, unaffected by metabolic inhibitors and only that of PGE2 increased with increased (5 microM) intraluminal concentrations. During incubation, the tissue remained viable as judged by T/M ratios (dpm tissue/dpm medium) for 204 thallium. Transport of [14C] sucrose was much slower than that of [14C] urea, which was greater than the fastest rates exhibited by the PGs. In general, amounts of radioactivity found in antimesometrial, mesometrial and lateral portions of the uterine wall, or in implantation and interimplantation areas did not differ, but more was found in the endometrium than the myometrium. PGF2 alpha was translocated unmetabolized to the external medium, while only two-thirds of the PGE2 was translocated unchanged, and one-third converted to PGF2 alpha. It is concluded that the rabbit uterus shows some selectivity in handling PGs in relation to stage of pregnancy.     

Progesterone reduces erectile dysfunction in sleep-deprived spontaneously hypertensive rats

Background

The rate-limiting step in prostaglandin (PG) biosynthesis is catalyzed by phospholipase A2 (PLA2) enzymes which hydrolyze arachidonic acid from membrane phospholipids. Despite their importance in uterine PG production, little is known concerning the specific PLA2 enzymes that regulate arachidonic acid liberation in the uterine endometrium. The objectives of this study were to evaluate the expression and activities of calcium-independent Group VI and Group IVC PLA2 (PLA2G6 and PLA2G4C) and calcium-dependent Group IVA PLA2 (PLA2G4A) enzymes in the regulation of bovine uterine endometrial epithelial cell PG production.

Methods

Bovine endometrial epithelial cells in culture were treated with oxytocin, interferon-tau and the PLA2G6 inhibitor bromoenol lactone, alone and in combination. Concentrations of PGF2alpha and PGE2 released into the medium were analyzed. Western blot analysis was performed on cellular protein to determine the effects of treatments on expression of PLA2G4A, PLA2G6 and PLA2G4C. Group-specific PLA2 activity assays were performed on cell lysates following treatment with oxytocin, interferon-tau or vehicle (control), alone and in combination. To further evaluate the role of specific PLA2 enzymes in uterine cell PG biosynthesis, cells were transfected with cDNAs encoding human PLA2G6 and PLA24C, treated as described above and PG assays performed.

Results

Constitutive cell production of PGF2alpha was about two-fold higher than PGE2. Oxytocin stimulated production of both PGs but the increase of PGF2alpha was significantly greater. Interferon-tau diminished oxytocin stimulation of both PGs. The PLA2G6 inhibitor, bromoenol lactone, abolished oxytocin-stimulated production of PGF2alpha. Treatments had little effect on PLA2G4A protein expression. In contrast, oxytocin enhanced expression of PLA2G6 and this effect was diminished in the presence of interferon-tau. Expression of PLA2G4C was barely detectable in control and oxytocin treated cells but it was enhanced in cells treated with interferon-tau. Oxytocin stimulated PLA2 activity in assays designed to evaluate PLA2G6 activity and interferon-tau inhibited this response. In assays designed to measure PLA2G4C activity, only interferon-tau was stimulatory. Cells overexpressing PLA2G6 produced similar quantities of the two PGs and these values were significantly higher than PG production by non-transfected cells. Oxytocin stimulated production of both PGs and this response was inhibited by interferon-tau. Bromoenol lactone inhibited oxtocin stimulation of PGF2alpha production but stimulated PGE2 production, both in the absence and presence of oxytocin. Cells over-expressing PLA2G4C produced more PGE2 than PGF2alpha and interferon-tau stimulated PGE2 production.

Conclusion

Results from these studies indicate that oxytocin stimulation of uterine PGF2alpha production is mediated, at least in part, by up-regulation of PLA2G6 expression and activity. In addition to its known inhibitory effect on oxytocin receptor expression, interferon-tau represses oxytocin-stimulated PLA2G6 expression and activity and this contributes to diminished PGF2alpha production. Furthermore, endometrial cell PGE2 biosynthesis was associated with PLA2G4C expression and activity and interferon-tau was stimulatory to this process.     

Biosynthesis and catabolism of prostaglandin F2alpha (PGF2alpha) are controlled by progesterone in the rat uterus during pregnancy

Myometrial quiescence is a key factor in all species to accomplish a successful gestation. PGs play a crucial role in mediating parturition events, and their synthesis and metabolism are regulated by cyclooxygenases (COXs) and NAD(+)-dependent 15-hydroxy-PG dehydrogenase (PGDH), respectively. Progesterone (P(4)) is the hormone responsible for maintaining uterine smooth muscle quiescence during pregnancy. In this work, we have studied the effect of P(4) on the activity of COXs and PGDH, the uterine enzymes involved in the biosynthesis and metabolism of prostanoids in the rat. We found that during pregnancy PGF(2alpha) production and also protein levels of COX-1 and COX-2 were decreased. The exogenous administration of P(4) significantly inhibited the uterine production of PGF(2alpha) and also the protein level of COX-2. PGF(2alpha), metabolism was assessed by PGDH activity, which resulted high during pregnancy and increased as a result of P(4) administration. These results indicate that PGs levels were negatively modulated by P(4), which could be exerting its effect by increasing PGs metabolism through stimulation on PGDH activity and an inhibition on COX and that is a major mechanism for maintain uterine quiescence in pregnancy.     

Influence of sex hormones on prostaglandin dehydrogenase activity in the rat uterus

The present experiments report the effects of estradiol or of progesterone on the activity of 15-prostaglandin-dehydrogenase (PGDH) in the uterus of spayed rats. When the substrate was PGF2 alpha the treatment with progesterone (4 mg X day-1, two days) or with estradiol-17-beta (0.5 ug + 1 ug) did not show any effect on the activity of the enzyme. On the contrary, uteri from ovariectomized rats injected with a higher dose of estradiol-17-beta (0.5 ug + 50 ug) exhibited a significant increment. When the substrate was PGE2, progesterone failed again to modify the enzyme activity, whereas estradiol, both at a low and at a high doses, enhanced significantly the uterine PGDH activity. The possibility of two different PGDHs for each PG and the role of estradiol in enhancing PGE2 catabolism into 15-keto-PGE2 as a mechanism subserving the effect of estrogens on the output of this PG in the rat uterus, are discussed.     

The release of PGF2 alpha and PGE2 from separated cells of human endometrium and decidua

The capacity of separated glandular and stromal cells from endometrium and first trimester decidua to release prostaglandins (PGs) was studied over 48 hours in culture. Glandular preparations released more PGs than stromal preparations in all tissues. Stromal release of PGs did not alter throughout the cycle or in early pregnancy but the capacity of glandular preparations to release PGs varied considerably. Proliferative glands released most PGF2 alpha and PGE2 followed by secretory glands and decidua. Histamine (10(-5)) stimulated PG release from endometrial and decidual glands but the response of proliferative glands was greatest. Actinomycin D stimulated release of PGF2 alpha and PGE2 from glandular cells of secretory endometrium and decidua. These results suggest that in vitro release of PGs is suppressed after ovulation and is in part due to inhibition of PG release by a protein or proteins synthesized in the glandular fraction of secretory endometrium or decidua.     

Gamma-linolenic acid improves the constancy of contractions in uteri from sprayed rats and is metabolized to prostaglandin E1 but not to bisenoic prostanoids

The effects of gamma-linolenic acid (GLA) on the time-dependent constancy of spontaneous contractions (isometric developed tension = IDT and frequency of contractions = FC) in uterine strips isolated from spayed rats, were explored. Moreover, the influence of the unsaturated fatty acid on the basal generation and release of tissue prostaglandins (PGs) as well as the conversion of labelled GLA into prostanoids by the uterine tissue and the effects of p-bromo-phenacyl-bromide (BPB), were also studied. GLA (10(-7)M), attenuated significantly the spontaneous decrement of contractile constancy exhibited by control preparations during a period of 180 min of activity in isolation, whereas BPB (10(5) M) resulted in an augmented and faster decrement of inotropic constancy. Spontaneous changes in the constancy of uterine motility as time progressed involved similarly both IDT and FC. After 180 min of activity in isolation a basal generation and release of PGs E and F of the series 1 and 2, were detected. The challenge with 10(-7) M GLA (delivered immediately after isolation) enhanced significantly the output of PGE1 but did not influence the generation and release of PGE2 or PGF2 alpha. BPB (10(-5) M) had no significant effect on the basal output of PGE1, PGE2 or PGF2 but completely prevented the enhancing action of GLA on the synthesis and release of PGE1. Labelled GLA was mainly converted to PGE1 by rat uterine segments and negligible counts in the 2-series of prostanoids, were observed. In presence of BPB (10(-5) M) the conversion of 1-14C-GLA, to PGE1 was almost completely abolished. The foregoing evidences suggest that exogenous GLA is metabolized by the spayed rat uterus via an elongase, forming di-homogamma-linolenic acid (DHLA), which in turn is substrate for cyclo-oxygenase peroxidase reactions yielding finally PGE1. No evidence of a delta 5-desaturase activity, converting DHLA into arachidonate and further derivatives, was detected. Coincidently, exogenous GLA was able to support a better contractile constancy as a function of time than that evidenced in untreated uterine strips isolated from castrated rats.     

Bovine placental prostaglandin synthesis: principal cell synthesis as modulated by the binucleate cell

Bovine placentomes were collected during late gestation, prepartum, and immediately postpartum. Postpartum tissues were collected prior to fetal placental release. A procedure for separating fetal placental principal cells from fetal binucleate cells (BNC) was developed. Dispersed fetal placental cells (mixed types), principal cells, and BNC were each examined for their ability to produce prostaglandins (PGs) from arachidonic acid (AA) and to metabolize prostaglandin E2 (PGE2) and prostaglandin F2 alpha (PGF2 alpha) in vitro. Dispersed fetal placental cells obtained prepartum produced predominantly PGs of the E series (PGEs) from AA (p less than 0.05). PGE synthesis predominated (p less than .05) in cells from postpartum tissue if the fetal placental membranes were subsequently retained, whereas synthesis of PGs of the F series (PGFs) predominated (p less than 0.05) if the fetal membranes were subsequently released. Principal cells were the primary source of fetal placental PG synthesis from AA (p less than 0.05). BNC exhibited a lesser ability to synthesize PGs from AA (p less than 0.05), but were able to convert PGF2 alpha to PGE2. Dispersed fetal placental cells exhibited greater ability to convert PGF2 alpha to PGE2 (p less than 0.05) than did the separated cells. These data suggest the function of a two-cell system within the fetal villi such that the BNC modulate the output of principal cell PG synthesis and/or metabolism.     

Patterns of prostaglandin synthesis and degradation in isolated superficial and proliferative colonic epithelial cells compared to residual colon

Prostaglandin (PG) synthesis and degradation were examined in different regions (epithelial versus non-epithelial structures) of the rat distal colon by both HPLC analysis of [14C] arachidonate (AA) metabolites and by specific radioimmunoassays. Intact isolated colonic epithelial cells synthesized mainly PGF2 alpha and TXA2, as monitored from the formation of its stable degradation product TXB2 (PGF2 alpha greater than TXB2 greater than 6-keto-PGF1 alpha, the stable degradation product of PGI2 = PGD2 = PGE2 = 13,14-dihydro-15-keto-PGF2 alpha). The profile of PG products of isolated surface epithelial cells was identical to that of proliferative epithelial cells. However, generation of PGs by surface epithelium was 2 to 3-fold higher than by proliferative cells both basally and in the presence of a maximal stimulating concentration (0.1 mM) of AA. The latter implied a greater synthetic capacity of surface epithelium, rather than differences due to endogenous AA availability. The major sites of PG synthesis in colon clearly resided in submucosal structures; the residual colon devoid of epithelial cells accounted for at least 99% of the total PGs produced by intact distal colon. The profile of AA metabolites formed by submucosal structures also differed markedly from that of the epithelium. The dominant submucosal product was PGE2. PGE2 and its degradation product 13,14-dihydro-15-keto-PGE2 accounted for 63% of the PG products formed by submucosal structures (PGE2 much greater than PGD2 greater than 13,14-dihydro-15-keto-PGE2 greater than PGF2 alpha = TXB2 = 6-keto-PGF1 alpha greater than 13,14-dihydro-15-keto-PGF2 alpha). By contrast, epithelial cells, and particularly surface epithelium, contributed disproportionately to the PG degradative capacity of colon, as assessed from the metabolism of either PGE2 or PGF2 alpha. When expressed as a percentage, epithelial cells accounted for 71% of total colonic PGE2 degradative capacity but only 23% of total colonic protein. Approximately 15% of [3H] PGE2 added to the serosal side of everted colonic loops crossed to the mucosal side intact. Thus, at least a portion of the PGE2 formed in the submucosa reaches, and thereby can potentially influence functions of the epithelium.     

Prostaglandin E receptor enhancement of catecholamine release may be mediated by phosphoinositide metabolism in bovine adrenal chromaffin cells

In the presence of ouabain, prostaglandin (PG) E2 stimulated a gradual secretion of catecholamines from cultured bovine adrenal chromaffin cells. PGE2 or ouabain alone evoked a marginal secretory response. The synergism of ouabain was also observed with muscarine. PGE2, like muscarine, induced a concentration-dependent formation of inositol phosphates: rapid rises in inositol trisphosphate and inositol bisphosphate followed by a slower accumulation of inositol monophosphate. This effect on phosphoinositide metabolism was accompanied by an increase in cytosolic free Ca2+. The potency of PGs (PGE2 greater than PGF2 alpha greater than PGD2) to stimulate catecholamine release was well correlated with that to affect phosphoinositide metabolism and that to increase the level of intracellular Ca2+. PGE2 did not stimulate cAMP generation significantly in bovine chromaffin cells. The effect of PGE2 on catecholamine release was mimicked by 12-O-tetradecanoylphorbol 13-acetate and A23187, but not by the cAMP analogue dibutyryl cAMP nor by forskolin. These results indicate that PGE2 may enhance catecholamine release from chromaffin cells by activating protein kinase C in concert with the increment of intracellular Ca2+.     

Effect of dietary restriction on triglyceride levels in the uterus isolated from pregnant rats. Influences of prostaglandins and indomethacin

Triglyceride (TGs) concentrations in uterine strips isolated from 14 or from 21 days-pregnant rats, either normal-fed or following a restricted-diet rats (50% food intake for 14 days), were measured. Determinations were made immediately after killing (0 min time or post-isolation) as well as after a period of incubation in glucose-free medium (60 min time or post-incubation). The post-isolation levels of TGs (0 min) in the uterus from normal-fed animals at 14 or at 21 days of pregnancy, were significantly higher in implantation sites than in the interembryonic segments. These values of TGs (0 time) did not change, in comparison to post-incubation concentrations (60 min), either without additions or in the presence of indomethacin (5 X 10(-6) M) or of prostaglandins (PGs) E1, E2 or F2 alpha (10(-7) M). At 0 time, uterine TGs of rats subjected to dietary restriction, increased as pregnancy progressed, more than in normal-fed controls. The post-incubation (60 min) pattern was different depending on the days of pregnancy; i.e. at 14 days, incubation in Krebs-Ringer Bicarbonate-medium (KRB) led to a significant fall unaffected by the addition of propranolol (10(-6) M). However, in the presence of indomethacin, TGs values had a level similar to the initial one (0 time). Furthermore, exogenous PGE1 or PGE2 failed to alter the effect of indomethacin, as PGF2 alpha did.(ABSTRACT TRUNCATED AT 250 WORDS)