M. Viggiano

The involvement of oxytocin in ovulation and in the outputs of cyclo-oxygenase and 5-lipoxygenase products from isolated rat ovaries

The effects on ovulation of a specific anti-oxytocin rabbit serum (anti-OT) (50.0 microliters) given by intrabursal injection into the right ovaries of etherized adult female rats at proestrus, were explored by counting the number of ovulated ova present within the right oviducts. Left ovaries were not treated and served as control ovaries. Control rats were treated with male normal rabbit serum (NRS) (50.0 microliters) given by intrabursal injections into the right ovaries of animals at proestrus. Ovulation was induced by injection of human chorionic gonadotrophin (hCG). Anti-OT administered into the right ovarian bursae of proestrous rat ovaries evoked a significant 51% inhibition of ovulation in comparison with that observed in control non-injected left ovaries (p less than 0.01). Also, when the ovulation of right ovaries injected with anti-OT was compared with that of left ovaries injected with NRS, the number of ovulated ova in the right side was significantly smaller (30%) than on the contralateral side (p less than 0.02). However, in rats pre-treated with hCG the intrabursal injection of oxytocin (OT) (50.0 mU/ml) into right and left ovaries failed to alter the number of ovulated ova compared with that of rats receiving intrabursal injections of saline. The basal control and the OT-evoked synthesis and release of endogenous prostaglandin E2 (PGE2) and PGF2 alpha were explored in ovaries isolated from prepuberal rats injected with pregnant mares serum gonadotrophin (PMSG), two days prior to sacrifice. OT augmented the basal release of PGF2 alpha but did not influence that of PGE2. Moreover, the conversion of exogenous 14C-arachidonic acid (14C-AA) into different prostanoids and into 5-HETE, in the presence and in the absence of added OT (50.0 mU/ml), was studied in rat ovaries isolated in proestrus. The challenge with OT augmented the basal synthesis and release of PGF2 alpha and of 5-HETE from 14C-AA, but failed to influence the formation of products generated via the cyclo-oxygenase pathway, namely 6-keto-PGF1 alpha, PGE2 and thromboxane B2 (TXB2). Therefore, the present results suggest that ovarian OT may play a role in the ovulatory process, via generation of PGF2 alpha to enhance contractions of ovarian smooth muscle and of 5-HETE to promote follicular collagenolysis.

Influence of ova within rat oviducts on spontaneous motility and on prostaglandin production.

The present study was performed in order to explore the influence of ova present within rat oviducts on: a) tubal spontaneous motility and b) oviduct prostaglandin production. It was found that the isometric developed tension (IDT) of tubes isolated from proestrous rats (preovulatory oviducts) was significantly higher (P less than 0.01) than the IDT of tubes from rats at estrus and at metestrus (postovulatory oviducts). After flushing the oviducts with KRB solution (i.e., after removing existing ova) the IDT of the oviducts obtained from estrous rats increased significantly (P less than 0.01), whereas the IDT of tubes isolated from proestrous rats (i.e., preparations without ova) was not modified. On the other hand, isolated tubes containing their corresponding ova released into the suspending solution significantly more PGE1 than PGE2 or PGF2 alpha (P less than 0.005). It was particularly interesting to find that after flushing the oviducts, tissue production of PGE1, PGE2 and PGF2 alpha was similar. Finally, when dose response curves for PGE1 and for PGE2 on the spontaneous contractions of oviducts isolated from rats at proestrus, estrus and metestrus were constructed, both PGs evoked an inhibitory inotropic action. The ED50 for PGE1 in tubes from estrous rats was significantly smaller (P less than 0.01) than that for metestrous animals but significantly greater (P less than 0.01) than that observed in oviducts from proestrous rats. The ED50 for PGE2 did not change in the different tested periods of the sex cycle. Results reported herein suggest the possibility that the ova present within rat oviducts, may influence their own transport along the tubes by modifying the amount of prostaglandins produced by the oviducts or via their own prostaglandin synthesis.

Probable influence of ova and embryo prostaglandins in the differential ovum transport in pregnant and cycling rats

In this study we explored the possible underlying mechanism(s) of the differential transport of unfertilized and fertilized ova in cycling and pregnant rats. The number of ova recovered from rat oviducts and uterus was not significantly different in estrus, metestrus and diestrus but dropped sharply at proestrus. When estrus rats were injected with indomethacin (10(-6)), a well known inhibitor of cyclooxygenase, delivered into both ovarian bursae, and sacrificed next day at metestrus, the number of ova in the oviduct was significantly smaller (p less than 0.025) than in controls at metestrus. On the other hand, when diestrus rats were injected with PGE1 (10(-6)) delivered into both ovarian bursae, and sacrificed next day at proestrus, no ova were found in the oviducts, and only a few of them were in the uterus. When fertilized ova were recovered from oviducts and uteri at day 4 of pregnancy (corresponding to proestrus of cycling rats) an average of 4 embryos were still found in the oviducts, proving a differential ovum transport between cycling and pregnant rats. In order to establish if there exists any ova or embryo releasing factor responsible for this difference, the prostaglandins released to the incubation medium by ovum or 3-day embryo were measured. Unfertilized ova produced significantly more PGE1 (p less than 0.05) than PGE2 or PGF2 alpha. The same pattern of PG production was observed with incubated embryos, but in this case the amount of PGE1 released was significantly higher (p less than 0.01) that the PGE1 released by unfertilized ova.(ABSTRACT TRUNCATED AT 250 WORDS)

Is there a role for dopamine in the regulation of motility of sow oviducts

Dose-response curves for norepinephrine (NE), dopamine (DA) and 2-bromo-alpha-ergocryptine (BEC) in the isthmic and ampullary regions isolated from proestrous sow oviducts, were drawn. NE, DA and BEC depressed the contractile activity of ampullary segments in dose-dependent fashion. In the isthmic region, NE induced a dose-related stimulation whereas DA and BEC evoked relaxation. Propranolol (10(-6)M) did not modify significantly the effect of DA on the ampulla, whereas spiperone, chlorpromazine and haloperidol, shifted to the right DA dose-response curves. Haloperidol and spiperone were able to partially antagonize responses to DA of the isthmus. The catecholamine contents of the ampullary (a) and isthmic (i) regions of sow oviducts during proestrus (P), estrus (E) and metestrus (M) proved to be quite different and differently affected by the variations of the sex cycle: NE, DA and epinephrine (EPI) in (i) higher than in (a); NE in (i) and (a) during M higher than during E; NE in (i) lower during E than during P. The probable presence of specific dopamine receptors in sow oviducts as well as their eventual role in modulating tubal motility are discussed in this paper.

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)

Prostaglandins and the contractile effect of angiotensin i and ii in the uterus isolated from ouariectomized rats. influence*of indo methacin and 17-β estradiol

The purpose of the present study was to explore whether, the stimulating contractile action of angiotensin II (AII) or of angiotensin I (AI), in uterine strips isolated from rats, either spayed or spayed and injected with 17-beta estradiol, was somehow linked to tissue generated prostaglandins (PGs). Indomethacin (10(-6)M), shifted to the left the dose-response curve for AII in preparations from ovariectomized animals; augmenting both, the efficacy and the potency of the agonist. Dose-response curves for AII were also explored in the presence of subthreshold concentrations of several exogenous PGs. PGE2 (10(-9) M) completely abolished the influence of indomethacin on the curve for AII; PGD2 (10(-9) M) did not evoke effects and PGF2 alpha (10(-9) M) only altered the threshold dose, the rest of the curve remaining similar to that obtained with indomethacin alone. The cumulative dose-response curve for AI, in strips isolated from ovariectomized rats, was comparable to that for AII and, the presence of indomethacin, only reduced the concentration for the threshold action of AI, in comparison to controls. Captopril (100 ng/ml), shifted to the right the dose-response curve for AI and indomethacin failed to alter the effects of AI in the presence of captopril. In strips isolated from ovariectomized rats, injected with 17-beta estradiol, the low concentration points of the dose-response curve for AII were shifted to the left of those for the agonist in control preparations from spayed rats not treated with estradiol. Furthermore, indomethacin failed to alter the responses to AII in strips from spayed animals injected with 17-beta estradiol. The foregoing results document that an inhibitor of tissue PGs synthesis, namely indomethacin, is able to modify the dose-response curve for AII, rather than that for AI, in the uterus from ovariectomized rats, but not in preparations from spayed animals treated with estradiol. This, as well as the effect of a subthreshold dose of PGs, suggest that a minimum of PGE2 is required for the contractile influence of AII and not for that of AI. Moreover, the absence of estrogen dominance appears, as well, as a crucial factor for the action of AII.

Prostaglandin F2α fails to alter the number of beta adrenoceptive specific binding sites in metestrous rat uteri but diminishes noradrenaline tissue uptake

3H-dihydroalprenolol (DHA) -- receptor binding was studied in membrane preparations from metestrous uterine tissue, both in presence and absence of exogenous prostaglandin (PG) F2 alpha at 10(-9) M. In addition, the uptake of 3H-noradrenaline (NA) by uterine segments from estrous and metestrous rats and the influences of PGF2 alpha (10(-9) M), cocaine (10(-5) M) corticosterone (5.10(-5) M), normetanephrine (10(-6) M) or acetylsalicylic acid (ASA: 10(-4) M), were explored. The Scatchard analysis of experimental data with 3H-DHA with or without added PGF2 alpha indicates the existence of a single class of high affinity receptors and no differences were found, in presence of PGF2 alpha, regarding the control dissociation constant or the control maximal sites of specific binding. On the other hand, the uptake of 3H-NA by uterine segments at metestrus was significantly greater than at estrus. In metestrous uteri PGF2 alpha (10(-9) M) reduced significantly NA uptake. ASA enhanced NA uptake by uteri from estrous rats, an influence prevented by PGF2 mu. In uterine segments isolated at estrus, cocaine, corticosterone and normetanephrine failed to alter 3H-NA uptake, whereas in preparations isolated at metestrus, corticosterone and normetanephrine reduced the uptake, but cocaine did not evoke any influence. Results are discussed in terms of previous findings documenting an amplification of the negative inotropic influence of NA mediated by the activation of beta-adrenoceptors, both in estrous or in metestrous preparations incubated with PGF2 alpha. Such previous findings cannot be explained by changes in the number of NA receptors or by a greater affinity of tissue receptors for the agonist, but rather by differences in NA uptake controlling its effective concentration at the biophase, near receptor sites. Interrelationships along sex hormones (estradiol), prostaglandins (PGF2 alpha) and catecholamines (NA) in uteri, are also discussed.

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.