Regulation by nitric oxide of prostaglandin E synthesis and spontaneous motility in rat uterine tissue.

We explored the role of endogenous nitric oxide (NO) in the spontaneous motility of uterine tissue from pseudopregnant (psp) rats and the correlation between this action and the uterotonic prostaglandin (PG) E production. We worked in the early psp (on day 5 of psp), and in late psp (on day 8 and day 9). Treatment with N(G)-monomethyl-L-arginine L-NMMA (300 microM), a competitive nitric oxide synthase (NOS) inhibitor, did not modify isometric developed tension (IDT) and frequency of contractions (FC) on day 5 of psp; on day 8, tissue pretreated with L-NMMA showed an increase in the IDT and FC compared with controls, while on day 9 of psp, both IDT and FC showed a lower stability after treatment with the inhibitor. These data suggest that NO modulates uterine motility on day 8 (decreasing it) and on day 9 of psp (enhancing it). We also evaluated the total NOS activity and that of its isoforms at the three times mentioned, demonstrating that total NOS activity was higher on day 5 of psp and decreased with psp development. On day 5 of psp, calcium-dependent and calcium-independent NOS each forms around 50% of total NOS activity. On day 8 of psp, the calcium-dependent was the predominant NOS form, while on day 9 of psp, the uterine tissue showed a higher calcium-independent form of the enzyme. In view of the fact that we found an inhibitor effect of the endogenous NO in uterine contractility on day 8 of psp and an inverse action on day 9 of psp (enhancing uterine contractility), we suggest that the NOS calcium-dependent form could be responsible for uterine contractility in psp rats. Finally, we evaluated the relationship between endogenous NO and PGE production. We observed that on days 5 and 8 of psp, the L-NMMA (300 microM) treatment did not affect PGE production, but on day 9 of psp, the preincubation with the NOS inhibitor diminished PGE synthesis, suggesting that at this time endogenous NO can upregulate uterine PGE production. These results confirm that NO can modulate uterine motility by means of PGE production. In summary, we suggest that in uterine tissue from psp rats, the NO system can alternatively decrease or increase uterine contractions, this last effect by enhancing uterine PGE synthesis.

Nitric oxide mediates platelet-activating factor stimulatory action on uterine prostaglandin production.

Accumulated evidence suggests that platelet-activating factor (PAF) may have a role in implantation by stimulating prostaglandin (PG) production. Since we had demonstrated that nitric oxide (NO) can increase uterine PGs, the aim of this study was to explore whether or not NO could mediate rat uterus responses to PAF on day 5 of gestation, when implantation takes place. Uterine motility was enhanced by PAF as compared to controls. This action was abolished by either the arginine analogue, N-monomethyl L-arginine (L-NMMA) or the cyclooxygenase inhibitor, indomethacin. On the other hand, NOS activity was detected in uterine strips and could be stimulated by PAF. The cyclooxygenase product PGE2 was also significantly stimulated by PAF. Inhibition of endogenous NO formation abolished the PAF effect on PG synthesis. Our results suggest that NO is an important intermediate in the interaction between PAF and PGs.

Role of nitric oxide on oxytocin-evoked contractions and prostaglandin synthesis in isolated pregnant rat uterus.

Uterine contractions elicited by oxytocin (OT), possibly linked with uterus prostaglandin (PG) release, are involved in the final pathway of labor. It is known that nitric oxide (NO) may contribute to the maintenance of uterine contractile quiescence during gestation. Therefore in this study the effect of the inhibition of NO synthase (NOS), with N-monomethyl L-arginine (L-NMMA), on the ability of OT to stimulate uterine contractions and PG synthesis was investigated in isolated rat uterus at days 13 and 21 of pregnancy. L-NMMA did not modify the frequency and the force of contractions elicited by OT at day 13. On day 21 the frequency of contractions evoked by OT were better sustained in the presence of L-NMMA. PGs were not affected by OT on day 13. OT stimulated PGF2alpha on day 21 when NOS had been inhibited with L-NMMA, but not in the absence of L-NMMA. NOS activity was stimulated by OT at day 21 of gestation. In summary these findings indicate that near term NO can regulate OT PGF2alpha induced contractions and PG synthesis in isolated pregnant rat uterus.

Effect of exogenous phospholipase A2 and triacylglycerol lipase on the synthesis and release of monoenoic and bisenoic prostaglandins from isolated rat uterus.

The possible existence of a selective and independent mechanism subserving the formation of prostaglandin E1 (PGE1) and of prostaglandin E2 (PGE2) has been reported in previous studies from our group. In the present experiments we have demonstrated that neutral lipid lipases play an important role yielding dihomo-gamma-linolenic acid for the formation of PGE1. Indeed, exogenous triglyceride lipase added to the incubation bathing solution at a concentration of 150 U/ml increased several fold the production of PGE1 by isolated uterine strips obtained from spayed rats. Nevertheless the presence of the enzyme did not modify significantly the synthesis and release of bisenoic PGs (PGE2 and PGF2 alpha). When triarachidonin was added, as an artificial substrate into the incubating medium in order to detect the presence of endogenous triacylglycerol lipase, we observed a significant increment in the generation of PGE2 (p less than 0.005) and of PGF2 alpha (p less than 0.001) without evident changes in the basal release of PGE1. On the other hand, the addition of phospholipase A2 (PLA2) at 0.2 U/ml, increased significantly the production of PGE2 (p less than 0.001) but failed to alter the concentration of PGE1 in the incubating solution. Surprisingly, PLA2 did not enhance the synthesis of PGF2 alpha in the present experiments, a situation for which we do not have a clear explanation. Exogenous bradykinin (10(-6) M), a well known stimulant of PLA2 activity in several tissues, also increased significantly (p less than 0.001) the production of PGE2 without altering that of PGE1.(ABSTRACT TRUNCATED AT 250 WORDS)

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)

Morphine diminishes the constancy of spontaneous uterine contractions, antagonizes the positive inotropic effects of prostaglandin E2, but not of prostaglandin F2 alpha and inhibits prostaglandin E and F outputs from the uterus of ovariectomized rats.

The effects of morphine on the constancy of spontaneous contractions (isometric developed tension = IDT and contractile frequency = CF), in uterine strips isolated from ovariectomized rats and the influence of naloxone, were explored. The inotropic responses to added prostaglandins (PGs) E2 and F2 alpha and the influences of morphine and of morphine in the presence of naloxone on PG actions, were also determined. Moreover, the synthesis and outputs of PGs E and F from uteri and the effects of morphine alone and of morphine plus naloxone, were studied. Morphine (10(-6) M) significantly depressed uterine constancy of IDT during the first hours following delivery, but its action on CF did not differ from controls. Naloxone, neither at 10(-8) M nor at 10(-6) M, altered the negative inotropic influence of morphine on IDT. Exogenous PGs E2 and F2 alpha, stimulated uterine inotropism in a concentration-dependent fashion. Morphine altered dose-response curves for exogenous PGE2, evoking a parallel surmountable shift to the right, but did not affect the inotropic action of added PGF2 alpha. This antagonistic effect of the opioid was not altered by preincubation with naloxone. Basal synthesis and outputs of PGs E and F in uteri from ovariectomized rats were significantly depressed by morphine (10(-6) M) but not altered by incubating tissues with morphine in presence of naloxone. Results are discussed in terms of a presumptive dual action of morphine on uterine motility, i.e., antagonizing PGE2 receptors and inhibiting the synthesis of some PGs by the uterus. These influences of morphine do not appear to be subserved by the activation of mu opioid receptors.(ABSTRACT TRUNCATED AT 250 WORDS)

Norepinephrine alters PGE2/PGE1 output ratio in isolated uterus from ovariectomized rats.

The rat uterus generates and releases prostaglandins (PGs) of the series 2 as well as PGs of the series 1. The main purposes of the present study are to compare the effects of norepinephrine (NE) on the production and outputs of PGE1, PGE2 and PGF2 alpha by the uterus isolated from ovariectomized rats, treated or not with 17-beta-estradiol and to explore also, whether the effects of NE on PG synthesis are mediated through alpha, beta or both types of adrenoreceptors. Segments of control uterine horns obtained from ovariectomized rats generated and released into the incubating solution, equal amounts of PGE1, PGE2 and PGF2 alpha and propranolol (10(-6) M) or phentolamine (10(-6) M) failed to alter this basal production of PGs. Norepinephrine (3 X 10(-6) M) significantly depressed the outputs of PGE1 and PGF2 alpha but enhanced, also significantly, the release of PGE2. In the presence of the beta-adrenoreceptor blocker, propranolol, the reduction induced by NE on the output of PGE1 was not altered, but the stimulatory influence of NE on the release of PGE2 as well as the depression on the output of PGF2 alpha, were abolished. On the other hand the diminution evoked by NE on the release of PGF1 and PGF2 alpha as well as the increment induced on PGE2 output, were inhibited by the presence of phentolamine in the incubating solution. Uterine horns from ovariectomized rats treated with 17-beta-estradiol released into the incubating solution significantly more PGF2 alpha than PGE1 or PGE2. NE, either alone of in the presence of alpha 0 or beta-adrenoceptor blockers, did not modify this pattern of PG production. A possible mechanism(s) of action for NE on PG synthesis is discussed.

Biological profile of a bioactive aortic substance (BAS), released by vessel rings from indomethacin-treated rats.

The biological activity of a stable unknown material(s), generated by aortic rings (bioactive aortic substance = BAS) isolated from rats injected with a high dose of indomethacin, was explored on contractions of several smooth muscle preparations from normal rats and its effects compared with those elicited by prostacyclin (PGI2) or by 6-keto-prostaglandin F1 alpha (6-k-PGF1 alpha). The BAS evoked, as did PGI2 or 6-k-PGF1 alpha, positive inotropism in strips from rat stomach, ileum and urinary bladder, but failed to influence uterine contractions as did prostacyclin or its non-enzymatic metabolite. When tested in rat aortic strips both, PGI2 and the BAS produced relaxation, whereas 6-k-PGF1 alpha was not active. Moreover, lipid substances present in the incubates of aortic rings, were extracted and explored for effects on contractions of rat aortic strips and on arachidonate-evoked human platelet aggregation. These extracts were devoid of influence on both parameters. On the contrary, dried aqueous residues, after the lipid extraction of the supernatants of aortic ring incubates, exhibited human platelet antiaggregatory capacity as well as the ability to evoke positive and negative inotropism similar to those triggered by the BAS in different smooth muscle preparations. Experiments with BAS were also performed employing smooth muscle strips exposed to indomethacin, atropine, propranolol, phentolamine and cyproheptadine. The presence of these antagonists of several neuromodulators and of indomethacin failed to alter de BAS-induced inotropic capacity observed in controls. The findings suggest that the effects attributable to the BAS are not subserved by prostacyclin or other prostanoids, nor by acetylcholine, norepinephrine, histamine or 6-OH-tryptamine.

On the inotropic effects of leukotrienes in the isolated urinary bladder of guinea pigs and rats.

Leukotrienes (LTs) B4, C4 and D4 were tested on the motility of rings isolated from the urinary bladder of guinea pigs and rats. LTB4 did not evoke inotropic influences in any of the preparations, whereas LTC4 and LTD4 augmented the magnitudes of tonic and phasic contractions in the guinea pig but not in the rat detrusor muscle, LTC4 evoking a greater enhancement than LTD4. On molar bases, acetylcholine induced smaller positive inotropic effects. In the presence of 10(-4) acetylsalicylic acid (ASA), cumulative dose-response curves of phasic and tonic contractions for LTC4 were shifted to the right of controls, whereas curves of the phasic motility for LTD4 remained unaltered. However, ASA shifted to the left the dose-response curve of tonic contractions for LTD4 and in addition evoked an augmentation of the absolute developed tension. The initial (postequilibrium) contractile levels of basal phasic contractions did not differ between controls and preparations incubated with ASA (10(-4) M), nordihydroguaiaretic acid (10(-7) M) or FPL-55712 (3 X 10(-6) M). The findings suggest that some musculo-active prostanoid(s) could be modulating, in opposite directions, the smooth muscle contractile reactivity of the guinea pig urinary bladder when challenged in vitro with LTC4 and LTD4. Our data reported hereing also suggest that the role, if any, of endogenous prostanoids and leukotrienes for the normal basal contractile functioning of the guinea pig urinary bladder, remains obscure.