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1.
The nature distribution and associated GTP gamma S binding activity of phosphatidylinositol phospholipase C (PI-PLC) has been studied in non-pregnant and pregnant guinea pig uterine smooth muscle. Cytosolic fractions partially purified by Q-Sepharose and heparin-Agarose chromatography show two isoenzyme forms, one with an apparent molecular weight of 58 kD that crossreacts with PI-PLC alpha and a has Km for phosphatidylinositol of 292 +/- 72.6 microM, designated alpha, and a form that has an apparent molecular weight of 86 kD and a substrate Km of 54 +/- 20 microM designated delta. Approximately 80% of the total PI-PLC activity was recovered in the cytosolic fraction and this increased 8-10 fold for both isoenzymes from the non-pregnant to the late pregnant uterus and the proportion of the alpha isoenzyme increased from approximately 40% to 55% of the total. PI-PLC alpha but not delta activity had GTP gamma S binding activity associated with it after Q-Sepharose or heparin-Agarose chromatography. This associated activity accounted for 2% of the total GTP gamma S-binding activity in the non-pregnant uterus and 31% of that in the near-term uterus. On separation of the PI-PLCa-GTP gamma S-binding complex by gel filtration on Sephacryl S200 gave two peaks one of 118 kD accounting for two-thirds of all the binding and two-thirds of the enzyme activity and a 58 kD peak. The 118 kD peak could not be separated by treatment with 0.5% cholate, but in this form enzyme activity was protected from detergent inactivation found with the 58 kD form. In sodium dodecyl sulphate polyacrylamide-gel electrophoresis PI-PLC alpha was released from the 118 kD complex and showed an apparent molecular weight of 61.5 kD. All the activity in the residual membrane fraction could be released by washing with buffer followed by, 2 M KCl and then 2 M KCl plus 0.5% cholate. This released isoenzyme forms that appeared identical to those in the cytosolic fraction and with GTP gamma S-binding activity associated with PI-PLC alpha. It is concluded that in the near term guinea pig uterus there is a dramatic increase in the capacity for inositol polyphosphate production. Moreover the dramatic increase in GTP gamma S-binding activity associated with PI-PLC alpha implies large changes in the extent and possibly nature of the putative G-protein activation of this pathway.(ABSTRACT TRUNCATED AT 400 WORDS)  相似文献   

2.
The ability of GTP gamma S to activate release of inositol polyphosphates from isolated permeabilised guinea pig uterine smooth muscle cells and from partially purified PI-PLC alpha has been studied. Streptolysin O permeabilised and [3H]inositol prelabelled cells show a time dependent release of inositol polyphosphates, predominantly inositol 4-phosphate. Ca2+ stimulated IP release with a Ka of 161 +/- 1.1 nM and this was further enhanced in an additive manner by GTP gamma S between 1-100 microM; the Ka for Ca2+ in the presence of 0.1 mM GTP gamma S was 117 +/- 0.7 nM. GTP gamma S activation of IP production did not require Ca2+ in the medium. Permeabilisation of the uterine smooth muscle cells with Streptolysin O readily released PI-PLC activity into the medium. However, unlike studies with isolated membranes 63.4 +/- 6.4% of the enzyme activity remained associated with membranes and/or particulate fractions of the cell. Studies were undertaken with PI-PLC alpha, the predominant isoenzyme form, partially purified from uterine smooth muscle at different stages of pregnancy by Q-Sepharose and Heparin-Agarose chromatography. The enzyme co-purifies with firmly associated GTP-binding activity. Enzyme prepared from near-term uterus is activated by 0.1 mM GTP gamma S, up to 100% when Ca2+ is between 0.1-1 microM, while 10 microM AlF4- under those conditions caused complete inhibition of the enzymes. Responses for enzymes prepared from non-pregnant uteri were broadly similar. In contrast enzyme preparations from guinea pig uteri at 20-60 days of pregnancy show an inhibition of activity in response to 0.1 mM GTP gamma S addition.(ABSTRACT TRUNCATED AT 250 WORDS)  相似文献   

3.
The activity of NAD+-dependent PGDH was measured in the cytosolic fractions (100,000 x g) of uterine tissues obtained from transsexual, pregnant and non-pregnant women. The specific activity (mean +/- SD) of the enzyme at maximum velocity of the enzyme reaction in these three groups of women was 5.5 +/- 2.30, 0.53 +/- 0.27 and 0.54 +/- 0.25 mU/mg protein respectively using PGE2 as substrate, and with PGF2 alpha as substrate the respective values were 5.48 +/- 2.80, 0.49 +/- 0.41 and 0.51 +/- 0.30 mU/mg protein. These data suggest that, with either substrate, the uterine enzyme activity in the transsexuals was about 10-fold greater than in pregnant and non-pregnant women (p less than 0.001). However, the Km values of the enzyme for both PGE2 and PGF2 alpha were similar in all three groups, indicating the presence of same enzyme in the uterus of transsexual, pregnant and non-pregnant women. We speculate that PGDH activity was raised in the uterus of transsexual women because of the prolonged androgen therapy they received for the management of female-to-male transsexualism.  相似文献   

4.
Studies on the mechanism of action of antifertile PG in animal models   总被引:1,自引:0,他引:1  
Antifertile effects of PGF2 alpha, PGE2, PGE1, sulprostone and other PGs were evaluated in different pregnancy models in rats, guinea pigs and rhesus monkeys and the underlying mechanisms of action were investigated. Quantitative and qualitative species differences and pregnancy stage dependency were recorded. Basic regulatory differences of the pregnant uterus seem to exist in these species. In early pregnant rats, abortifacient effects were based on luteolytic effects, independent of the PG used. The myometrium was found to be refractory to the injected PG as long as serum progesterone levels were kept high. By contrast, in guinea pigs after the luteoplacental shift of progesterone secretion (tested after day 40 p.c.) and in rhesus monkeys even before this shift (tested day 20 p.c.) abortifacient effects were found to be exerted by direct stimulation of the myometrium. Uterine stimulation was possible in the presence of any level of serum progesterone. The induction of uterine PG synthesis was probably of importance supporting the expulsion. The role of obvious tissue damage within the conceptus remained uncertain. In contrast to rats there seems to be a pre-existing PG-sensitivity of the pregnant myometrium in guinea pigs and primates. In guinea pigs sensitivity slightly increased for E- but not for F-type PG toward term. Oxytocin sensitivity was found to increase by a factor of more than 100 between days 23-63 of pregnancy. Time dependent changes in uterine receptivity to PG and oxytocin may be considered as a regulatory principle which might permit parturition to occur in the presence of progesterone as an evolutionary adaptation to a placental progesterone secretion which cannot be abolished. It was concluded that in the presence of already established gradual uterine responsiveness to PG (and Oxytocin) during gestation efficient blocking mechanisms for uterine PG-formation must exist in order to explain uterine quiescence. Almost complete resistance of pregnancy to oestrogen which exists in humans, monkeys and guinea pigs was considered as to be pharmacological evidence of such a mechanism. The principles of endocrine control of the myometrium and its pharmacology seem similar in guinea pig and primate pregnancy. The guinea pig might therefore provide a relevant model to study potential drug effects on the regulatory balance of the pregnant uterus and also to achieve a better understanding of human uterine physiology.  相似文献   

5.
Radioimmunoassay of progesterone in systemic and placental blood of pregnant rabbits and guinea pigs. 1. The level of progesterone in pregnant rabbits and guinea pigs serum was measured directly (without extraction) using a RadioImmunoAssay (RIA). 2. Hormonal concentrations in systemic blood were shown to increase with gestational age, being at their highest half-way through pregnancy (16.03 +/- 2.63 ng/ml for rabbits; 319.01 +/- 42.10 ng/ml for guinea pigs) and decreasing at the end of the pregnancy. 3. Progesterone was not detectable in rabbit placental blood, whereas a high level of this hormone was found in guinea pig placental blood, which increased with gestational age. From the 28th to the 56th post-coital day, the level increased from 143.22 +/- 13.15 to 283.30 +/- 36.84 ng/ml. 4. The method used enables to measure correctly progesterone concentrations in rabbit and guinea pig serum without extraction.  相似文献   

6.
Summary Neuropeptide Y (NPY) is a recently discovered neuropeptide with vasoconstrictor effects when given in vivo. It occurs in many sympathetic neurons, where it appears to coexist with noradrenaline (NA). It is wellknown that profound changes in the levels of uterine NA occur in many species during pregnancy. Therefore we have investigated the distribution of catecholamine neurons and NPY by immunohistochemistry in the pregnant and nonpregnant guinea pig uterus. In the virgin uterus NPY-like immunoreactivity was present in nerve fibres and terminals in the smooth muscle layers of the uterine horns and around blood vessels. The distribution of NPY fibres was very similar to that of noradrenergic nerves visualized with antibodies against the catecholamine synthesizing enzyme tyrosine hydroxylase (TH). In the pregnant uterus, NPY- and TH-like immunoreactivity disappeared almost completely. In the cervix, a slight decrease of immunoreactivity was observed, whereas in the ovaries no changes were noted between the pregnant and nonpregnant condition. The results indicate that NPY and catecholamines coexists in the adrenergic neurons of the guinea pig uterus, cervix and ovary and that they vary together in the myometrium during pregnancy. We suggest that NPY may be of functional importance for the pregnant uterus.  相似文献   

7.
Neuropeptide Y (NPY) is a recently discovered neuropeptide with vasoconstrictor effects when given in vivo. It occurs in many sympathetic neurons, where it appears to coexist with noradrenaline (NA). It is wellknown that profound changes in the levels of uterine NA occur in many species during pregnancy. Therefore we have investigated the distribution of catecholamine neurons and NPY by immunohistochemistry in the pregnant and nonpregnant guinea pig uterus. In the virgin uterus NPY-like immunoreactivity was present in nerve fibres and terminals in the smooth muscle layers of the uterine horns and around blood vessels. The distribution of NPY fibres was very similar to that of noradrenergic nerves visualized with antibodies against the catecholamine synthesizing enzyme tyrosine hydroxylase (TH). In the pregnant uterus, NPY- and TH-like immunoreactivity disappeared almost completely. In the cervix, a slight decrease of immunoreactivity was observed, whereas in the ovaries no changes were noted between the pregnant and nonpregnant condition. The results indicate that NPY and catecholamines coexists in the adrenergic neurons of the guinea pig uterus, cervix and ovary and that they vary together in the myometrium during pregnancy. We suggest that NPY may be of functional importance for the pregnant uterus.  相似文献   

8.
F A Moustafa 《Acta anatomica》1988,132(4):310-316
The present study was undertaken to investigate the structural changes in both cholinesterase (ChE)-positive nerve fibers and adrenergic nerves with formaldehyde-induced fluorescence in pregnant and postpartum uteri of both the albino rat and guinea pig. Particular attention was directed to the relationship between these changes and the local factors associated with the growing fetus. ChE reaction was absent in the control and pregnant uterus of the guinea pig. In the albino rat, there were signs of degeneration in pregnancy. These were evidenced by vacuolation of large nerve trunks and the presence of focal segments with very faint reaction along the course of the nerve bundles. Myometrial segments from fetus-containing horns showed some fragmented nerve fibers, but at the same time some other normal ones. Most of the fine nerve bundles gave a weak reaction. Three weeks after delivery, multiple ChE fibers were found in the uterus of the albino rat. The normal appearance was, however, not regained and some nerve fibers were still fragmented. Noradrenergic (NA) nerve fibers were disintegrated and markedly reduced in number in the myometrium of the pregnant uterus of both the guinea pig and albino rat, particularly in the uterine horns that were distended by fetuses. The number of NA fibers was not significantly reduced in the tubal ends of the albino rat uterus. Three weeks after delivery, normal NA fibers were seen in the myometrium of both the albino rat and guinea pig uterus. Nerves with reduced fluorescence reaction were observed less frequently.  相似文献   

9.
The fine structure of myometrium from pigs collected at well-defined stages during pregnancy was investigated by transmission electron microscopy. The morphology of the pregnant myometrium resembled that in non-pregnant pigs. Thick myofilaments were conspicuous during early pregnancy, inapparent during mid-pregnancy and visible again by days 80-84 and towards parturition. Gap junctions were extremely rare throughout pregnancy. Only occasionally were nerve axons observed.  相似文献   

10.
Two peaks of phosphoinositide-specific phospholipase C (PI-PLC) activity were resolved when guinea pig uterus cytosolic proteins were chromatographed on a DEAE-Sepharose column. The first peak of enzyme activity eluting from the DEAE-Sepharose column (PI-PLC I) was further purified to homogeneity, whereas the second peak of enzyme activity was enriched 300-fold. PI-PLC I migrated as a 62-kDa protein on sodium dodecyl sulfate-polyacrylamide gels. Antibodies prepared against PI-PLC I failed to react with PI-PLC II. PI-PLC I hydrolyzed all three phosphoinositides, exhibiting a greater Vmax for phosphatidylinositol 4,5-bisphosphate greater than phosphatidylinositol 4-phosphate greater than phosphatidylinositol. Hydrolysis of phosphatidylinositol was calcium-dependent, whereas significant hydrolysis of phosphatidylinositol 4-phosphate and phosphatidylinositol 4,5-bisphosphate occurred in the presence of 2.5 mM EGTA. At physiological concentrations of calcium, phosphatidylinositol 4-phosphate and phosphatidylinositol 4,5-bisphosphate were the preferred substrates. Antibodies specific for PI-PLC I reacted with a 62-kDa protein in both the cytosol and membrane fractions from guinea pig uterus. Quantitation of the immunoblots revealed that 25% of the 62-kDa protein was membrane-associated, whereas only 5% of the total enzyme activity was membrane-associated. Approximately 20% of the membrane-bound phospholipase C activity and immunoreactive material were loosely bound, whereas the remainder required detergent extraction for complete solubilization. The 62-kDa protein associated with the membrane fractions did not bind lectin affinity columns, suggesting that it was not glycosylated. PI-PLC I was identified as a phosphoprotein in [32P]orthophosphate-labeled rat basophilic leukemia (RBL-1) cells by two-dimensional gel electrophoresis followed by immunoblotting. In untreated cells, 32P-labeled PI-PLC I was found in the cytosolic fraction. Treatment of RBL-1 cells with those phorbol esters which are known to activate the Ca2+/phospholipid-dependent enzyme protein kinase C, resulted in a time-dependent increase in the phosphorylation of both membrane-bound and cytosolic PI-PLC I. Thus, in RBL-1 cells, protein kinase C may play an important role in the regulation of phospholipase C through protein phosphorylation.  相似文献   

11.
The specific binding of [3H]oxytoxin to uterine membrane preparations derived from different species at late pregnancy was examined. The highest receptor density (bmax value) was found in membranes derived from the myometria of guinea pigs between day 60 post-conception (bmax = 3.6 +/- 0.1 pmol/mg) and day 65 (bmax = 4.4 +/- 0.1 pmol/mg). The similarity of Kd values for oxytocin binding (Kd = 2.6 +/- 0.2 nM) and for vasopressin binding (Kd = 2.1 +/- 0.4 nM) to the same membranes derived from a guinea pig myometrium indicate a homogeneous population of high-affinity binding sites which do not discriminate between these two hormones. Competitive binding experiments with specific oxytocin agonists containing either sarcosine or N-methylalanine in the place of Pro7 demonstrated that these myometrial receptors have the pharmacological properties of oxytocin receptors. The analogue of 1-deamino-[8-lysine]vasopressin containing a photoreactive azidophenylamidino group at the sidechain of Lys8 retained roughly the same receptor affinity as oxytocin. In photoaffinity labelling experiments with the tritium-labelled analogue a membrane protein from guinea pig myometrium with an apparent relative molecular mass Mr of 78,000 +/- 5000 (n = 13) was preferentially labelled. The labelling of this protein was completely suppressed by a 100-fold molar excess of either oxytocin, or [Sar7]oxytocin or [Thr4, Sar7]oxytocin, but not by other peptide hormones. These results provide evidence that the labelled 78,000-Mr protein is a myometrial oxytocin-receptor protein.  相似文献   

12.
A comparative study of the enzymes delta5-3beta-HSD, cytochrome oxidase and peroxidase has been made in the ovaries and uterus of mammals (mouse, guinea pig, cat and dog) during various reproductive phases. The granulosa cells of developing follicles, hypertrophied interstitial cells of thecal origin and the luteal cells show intense delta5-3beta-HSD and cytochrome oxidase activity. Peroxidases are found to be present in the corpus luteum and the epithelial cords of thecal origin. delta5-3beta-HSD and cytochrome oxidase activity is localized to the endometrium and myometrium of mature and pregnant uterus of mouse and guinea pig, while peroxidase is seen only in the decidua and endometrial glands of pregnant animals. The significance of these enzymes is discussed in relation to the cellular basis of luteinization and steroid hormone synthesis.  相似文献   

13.
The innervation of the myometrium in virgin, pregnant and post-partum ewes was examined using the SPG histofluorescence method and a high-performance liquid chromatographic catecholamine assay. In non-pregnant ewes, fluorescent axons were visible in all regions of the uterus. At 50 days of pregnancy the innervation was unchanged, but at 100 days axons were scarce over the whole uterus with the exception of the tubal extremities of the uterine horns. Noradrenaline concentrations were also significantly lower in late pregnancy, and significant variations occurred in different regions of the uterus, with the greatest concentrations present in the tubal extremities. At 13-16 weeks post partum, the density of innervation was variable, although the noradrenaline concentration was only slightly less than in virgin animals. Dopamine was also present in substantial quantities, but the mean concentrations remained unchanged during pregnancy.  相似文献   

14.
Lipid content and steroid sulfatase activities were determined in liver and uterus microsomes of non-pregnant guinea pigs. The results were compared with values obtained in pregnant and cortisol-treated animals. Steroid sulfatase activities were always higher in pregnant animals, and we supposed that the increase in circulating cortisol in pregnant guinea pigs before parturition has an influence on the membrane-bound sulfatase activities. Sulfatase activities were identical in cortisol-treated and untreated non-pregnant females, although cortisol induced changes in microsomal lipid composition. These results lead us to three conclusions: in intact female guinea pigs, cortisol induces variations in the lipid content of uterus and liver microsomes, especially in the cholesteryl sulfate to phospholipid ratios; the variations of the lipid composition in pregnant animals do not appear to be cortisol-dependent; membrane-bound steroid sulfatase activities are not directly influenced by the lipid composition of microsomes.  相似文献   

15.
Aquaporin-1 Increases in the Rat Myometrium During Early Pregnancy   总被引:3,自引:0,他引:3  
Immunofluorescence and immunogold techniques were used to determine the presence and distribution of aquaporin-1 (AQP1) within the rat uterus. Uterine tissue from non-pregnant (proestrus) as well as pregnant (days 1, 3, 6 and 7) rats were used. It was found that this water channel was present in the myometrium of the pregnant rat uterus with the intensity of AQP1 immunoreactivity increasing from day 1 to day 6 of pregnancy. In particular, an increase was also observed in mesometrial as compared to antimesometrial myometrium. Immunolocalization at the electron microscope level indicated that AQP1 was localized to the plasma membrane of smooth muscle cells found within the inner circular layer. It is suggested that AQP1 plays a role in stromal oedema, uterine closure and orientation of the blastocyst.  相似文献   

16.
Progesterone withdrawal is essential for parturition, but the mechanism of this pivotal hormonal change is unclear in women and other mammals that give birth without a pre-labor drop in maternal progesterone levels. One possibility suggested by uterine tissue analyses and cell culture models is that progesterone receptor levels change at term decreasing the progesterone responsiveness of the myometrium, which causes progesterone withdrawal at the functional level and results in estrogen dominance enhancing uterine contractility. In this investigation we have explored whether receptor mediated functional progesterone withdrawal occurs during late pregnancy and labor in vivo. We have also determined whether prostaglandins that induce labor cause functional progesterone withdrawal by altering myometrial progesterone receptor expression. Pregnant guinea pigs were used, since this animal loses progesterone responsiveness at term and gives birth in the presence of high maternal progesterone level similarly to primates. We found that progesterone receptor mRNA and protein A and B expression decreased in the guinea pig uterus during the last third of gestation and in labor. Prostaglandin administration reduced while prostaglandin synthesis inhibitor treatment increased progesterone receptor A protein abundance. Estrogen receptor-1 protein levels remained unchanged during late gestation, in labor and after prostaglandin or prostaglandin synthesis inhibitor administration. Steroid receptor levels were higher in the non-pregnant than in the pregnant uterine horns. We conclude that the decreasing expression of both progesterone receptors A and B is a physiological mechanism of functional progesterone withdrawal in the guinea pig during late pregnancy and in labor. Further, prostaglandins administered exogenously or produced endogenously stimulate labor in part by suppressing uterine progesterone receptor A expression, which may cause functional progesterone withdrawal, promote estrogen dominance and foster myometrial contractions.  相似文献   

17.
Endometrium of GnRH treated group resembled with pregnant group and endometrial thickness in these groups significantly increased in comparison with non-pregnant group. In GnRH treated animals, most of histomorphological changes in epithelial cells, glands and stroma of uterus was similar to pregnant group. The results revealed that mammalian form of GnRH exerted endometrial change in guinea pig almost similar to those occur in normal pregnant animals and its administration prior to implantation may improve pregnancy rate following embryo transfer.  相似文献   

18.
To assess the degree of circulatory fullness and to evaluate the influence of peripheral and cardiac factors in the regulation of cardiac output during pregnancy, the following studies were conducted using pentobarbital-anesthetized, open-chest nonpregnant and late term pregnant guinea pigs. Mean circulatory filling pressure was taken as the equilibrium pressure when the pulmonary artery was constricted. Total vascular compliance was assessed by +/- 5-mL changes in blood volume performed while this constriction was maintained. A separate group of guinea pigs was prepared with a pulmonary artery electromagnetic flow probe and right atrial catheter. Rapid infusion of saline was used to increase right atrial pressure while the cardiac output was determined. Pregnancy was characterized by the following changes relative to nonpregnant controls: 51Cr-labelled RBC blood volume increased from 55 +/- 3 to 67 +/- 3 mL/kg; mean circulatory filling pressure increased from 7.1 +/- 0.2 to 8.0 +/- 0.5 mmHg (1 mmHg = 133.322 Pa); right atrial pressure decreased from 3.4 +/- 0.2 to 2.1 +/- 0.3 mmHg; and cardiac output increased from 71.8 +/- 3.9 to 96.8 +/- 3.3 mL.min-1.kg-1. Total vascular compliance was not changed (2.1 +/- 0.1 mL.kg-1.mmHg-1) and most of the expanded blood volume was accommodated as unstressed volume. The cardiac function curve was shifted upwards in pregnant animals. The resistance to venous return, as determined from the slope of the venous return curves, was not changed. These data suggest that the circulation of the pregnant guinea pig is slightly overfilled.(ABSTRACT TRUNCATED AT 250 WORDS)  相似文献   

19.
The influence of pregnancy on the density and pattern of the peptidergic innervation of the guinea pig uterine artery was studied. Whole mount stretch preparations of the uterine artery from estrus and late pregnant guinea pigs were processed for the immunohistochemical demonstration of neuropeptide Y (NPY)-, vasoactive intestinal polypeptide (VIP)-, calcitonin gene-related peptide (CGRP)- and substance P (SP)- immunoreactive nerve fibres. In late pregnancy the density of NPY- and CGRP- containing nerve fibres was remarkably decreased, while that of VIP- and SP- immunoreactive nerves showed a moderate reduction. The meaning and the possible physiological relevance of the decreased density of peptide-immunoreactive nerves in the uterine artery in late pregnancy are discussed.  相似文献   

20.
The ultrastructure of the porcine myometrium collected at well-defined stages during parturition was investigated by transmission electron microscopy. The morphology of the parturient myometrium resembled in general that in pregnant and non-pregnant pigs. The diameter of the smooth muscle cells was, however, about twice that of non-pregnant myometrium. Thick myofilaments were numerous. The number of caveolae seemed to be higher in parturient compared with non-pregnant and pregnant cells. Gap junctions occurred richly and were large, while the intrinsic innervation was very scanty. To conclude, the endocrinological changes in the pig taking place just prior to parturition, are translated into morphological changes by stimulating the formation of uterine gap junctions. This provides low-resistance pathways between the muscle cells and activates the myometrium for the delivery process.  相似文献   

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