CYCLIC AMP IN ISOLATED CORPORA LUTEA OF THE RAT: INFLUENCE OF GONADOTROPHINS AND PROSTAGLANDINS

1974 ◽  
Vol 77 (4) ◽  
pp. 737-752 ◽  
Author(s):  
Hans Herlitz ◽  
Lars Hamberger ◽  
Sten Rosberg ◽  
Kurt Ahrén
Keyword(s):  
Corpus Luteum ◽  
Incubation Medium ◽  
Time Course ◽  
First Generation ◽  
Prostaglandin E ◽  
Corpora Lutea ◽  
Bicarbonate Buffer ◽  
Camp Content ◽  
Camp Levels

ABSTRACT Corpora lutea were isolated from 34–39 day-old rats injected with 10 IU PMSG when 30 days old leading to ovulation early in the morning of day 33. Three day-old corpora lutea were incubated in Krebs bicarbonate buffer containing 5.5 mmol/l glucose. Addition of LH to the incubation medium increased accumulation of cyclic 3′, 5′-AMP (cAMP) in a dose-dependent way in the isolated tissue and caused a release of cAMP to the incubation medium. cAMP was determined by the protein binding technique of Gilman (1970). There was a continuous rise in the levels of cAMP in both tissue and medium with increasing incubation time in presence of LH. Neither prolactin nor FSH could influence the cAMP levels in the corpus luteum tissue in vitro. Prostaglandin E2 (PGE2) caused a stimulation with respect to cAMP formation. The time-course of the PGE2 effect was quite different from that of LH: PGE2 gave maximal tissue cAMP content within 5–15 min with decreasing levels thereafter, compared to the continuous rise with LH stimulation for at least 1 h. In experiments performed on corpora lutea of different ages (1–7 day-old), tissue levels of cAMP were determined after incubations with LH or PGE2. There was a dramatic increase of tissue cAMP in the very young corpus luteum and a marked decrease in sensitivity to LH with increasing age of the corpus luteum with a very small but still significant effect on the old corpora lutea. The stimulatory effect of PGE2 on cAMP levels on the other hand showed only a minimal change. A few experiments on the first generation of corpora lutea from mature cycling rats are also reported for comparison.

OOCYTE MATURATION AND GLYCOLYSIS IN ISOLATED PRE-OVULATORY FOLLICLES OF PMS-INJECTED IMMATURE RATS

1976 ◽  
Vol 82 (4) ◽  
pp. 809-830 ◽  
Author(s):  
Torbjörn Hillensjö
Keyword(s):  
Oocyte Maturation ◽  
Time Course ◽  
Polar Body ◽  
Prostaglandin E ◽  
Germinal Vesicle Breakdown ◽  
Bicarbonate Buffer ◽  
Lh Surge ◽  
Immature Rats ◽  
Lactate Formation

ABSTRACT Graafian follicles were isolated by microdissection from the ovaries of PMS-injected immature rats killed at specific times on the day before ovulation. They were incubated in Krebs bicarbonate buffer containing 5 mm glucose and 1 % bovine serum albumin. The oocytes were recovered after incubation and examined by Nomarski interference contrast microscopy. The amount of lactate and glucose in the incubation medium was analysed enzymatically. Oocytes recovered from follicles extirpated in the morning, i. e. before the endogenous LH surge, and incubated for 2–10 h were in the dictyate stage, while addition of LH or FSH to the medium resulted in oocyte maturation as revealed by germinal vesicle breakdown (GVB) and polar body formation. The time-course of GVB in vitro was similar to that seen in vivo after exogenous LH. Both gonadotrophins markedly enhanced lactate accumulation in the medium and, as studied for LH, glucose uptake by the follicles. The effects of LH but not those of FSH, on GVB and lactate formation were prevented by the presence of an antiserum against the β-subunit of LH. The gonadotrophic effects on GVB could be mimicked by the addition of prostaglandin E2 to the medium. When the follicles were extirpated in the late afternoon, i. e. after the LH surge, and incubated in hormone-free medium for 4 h the oocytes showed GVB in a progressively increasing proportion depending on the time of follicle extirpation. Lactate formation by this group of follicles was markedly enhanced compared to that of "morning" follicles, but it could be even more stimulated by in vitro exposure to LH. A preliminary series of experiments on the effect of phosphodiesterase inhibitors showed that theophylline and isobutylmethylxanthine at certain concentrations completely blocked the LH effect on GVB, whereas a newly developed compound, ICI 63.197, in itself induced GVB.

DIFFERENCES IN ACTION OF LH AND FSH ON THE FORMATION OF CYCLIC AMP IN THE PREPUBERTAL RAT OVARY

1976 ◽  
Vol 81 (1) ◽  
pp. 150-164 ◽  
Author(s):  
Gunnar Selstam ◽  
Sten Rosberg ◽  
Jan Liljekvist ◽  
Lena Grönquist ◽  
Torsten Perklev ◽  
...  
Keyword(s):  
Cyclic Amp ◽  
Incubation Medium ◽  
Time Course ◽  
Tissue Levels ◽  
Rat Ovary ◽  
Camp System ◽  
High Concentrations ◽  
Camp Levels

ABSTRACT The actions of LH (NIH-LH-B8) and FSH (NIH-FSH-S9) on the cyclic AMP (cAMP) system in ovaries of 23–24 day old rats have been analyzed. An intravenous injection of LH increased ovarian cAMP levels in vivo after only 20 seconds. Maximal cAMP levels were seen after 15 min. Addition of LH or FSH in vitro to the isolated ovaries produced dose dependent increases of cAMP in the tissue as well as in the incubation medium. Low concentrations of LH caused a release of cAMP into the incubation medium without any detectable change in the tissue levels. The levels of cAMP in the incubation media for all concentrations of FSH were lower than the tissue levels, whereas for LH the opposite was found. In time-course experiments where the concentrations of LH (10 μg/ml) and FSH (100 μg/ml) were chosen to give similar tissue levels of cAMP, the release of the cyclic nucleotide into the incubation medium was approximately 2–3 times greater for LH than for FSH at the time periods studied (5–240 min). When LH and FSH were tested together in high concentrations, their effects were additive. When the ovaries were first incubated with FSH for 120 min followed by an incubation with LH, the stimulatory effect of LH was considerably reduced. When the order of the incubations was reversed, however, LH did not change the response to FSH. The results show that both LH and FSH have intrinsic effects on the cAMP system in the prepubertal rat ovary, but that the effects of the two gonadotrophins are not identical.

RELEASE OF PROTEIN WHICH BINDS PROGESTERONE FROM THE BOVINE CORPUS LUTEUM

1982 ◽  
Vol 92 (1) ◽  
pp. 51-61 ◽  
Author(s):  
D. L. WILLCOX ◽  
M. R. ALISON
Keyword(s):  
Corpus Luteum ◽  
Incubation Medium ◽  
Binding Protein ◽  
Equilibrium Dialysis ◽  
Corpora Lutea ◽  
Gas Liquid Chromatography ◽  
Cytosol Fraction ◽  
Major Band ◽  
Bovine Corpus Luteum

The nature of the proteins released together with progesterone from the bovine corpus luteum was investigated. Slices of bovine corpus luteum obtained at the mid-luteal stage of the oestrous cycle were incubated in vitro in the presence of bovine LH. Radioactive monitoring of compounds separated by gas–liquid chromatography showed that added [14C]acetate was converted to sterols and progesterone within the 2-h incubation period. Electrophoresis of the proteins recovered from the incubation medium showed the presence of a major band which migrated at the same rate as a progesterone-binding protein isolated from pooled corpora lutea. Densitometry revealed that the proteins in the incubation medium were enriched threefold in binding protein compared to those in an homogenate of corpus luteum. Equilibrium dialysis of the medium after addition of [3H]progesterone revealed the presence of a progesterone-binding activity (association constant = 106l/mol) analogous to that contained in the cytosol fraction from bovine corpus luteum. The finding that progesterone-binding protein is released concomitantly with the hormone has implications for the granule hypothesis of steroid secretion. The existence of an intracellular steroid-binding protein complex to maintain progesterone in high intragranular concentration would be predicted from this hypothesis.

Blended Effects of Herbal Components of Tokishakuyakusan on Somatomedin C/Insulin-like Growth Factor 1 Level in Rat Corpus Luteum

1991 ◽  
Vol 19 (01) ◽  
pp. 61-64 ◽  
Author(s):  
Satoshi Usuki
Keyword(s):  
Growth Factor ◽  
Corpus Luteum ◽  
Corpora Lutea ◽  
Insulin Like Growth Factor ◽  
Somatomedin C ◽  
Factor I ◽  
Peony Root ◽  
Growth Factor I ◽  
Herbal Components

The effect of herbal components of Tokishakuyakusan on somatomedin C/insulin-like growth factor I (IGF-1) level in medium from rat corpora lutea incubated in vitro was examined. Hoelen + peony root + Japanese angelica root, hoelen + peony root, hoelen + Japanese angelica root or peony root + Japanese angelica root decreased the IGF-1 level. The data suggest that constituent herbal components of Tokishakuyakusan regulate the IGF-1 level by rat corpora lutea.

Chloroquine stimulates absorption and inhibits secretion of ileal water and electrolytes

1982 ◽  
Vol 243 (2) ◽  
pp. G117-G126
Author(s):  
R. Fogel ◽  
G. W. Sharp ◽  
M. Donowitz
Keyword(s):  
Cholera Toxin ◽  
Calcium Content ◽  
Intracellular Camp ◽  
Rabbit Ileum ◽  
Camp Content ◽  
Serosal Surface ◽  
Ileal Absorption ◽  
Camp Levels

The effects of chloroquine diphosphate, a drug with "'membrane-stabilizing" properties, were studied on basal ileal absorption and on ileal secretion induced by increased intracellular cAMP levels and calcium (serotonin). The studies were performed on rat (in vivo) and rabbit ileum (in vitro). Intraluminal chloroquine (10(-4) M) reversed cholera toxin- and theophylline-induced secretion in rat ileum but did not alter the cholera toxin- and theophylline-induced increases in cAMP content. Addition of chloroquine (10(-4) M) to the mucosal surface of rabbit ileum did not alter basal active electrolyte transport or the serotonin-induced decreased Na and Cl absorption but inhibited the theophylline-induced C1 secretion. Addition of chloroquine (10(-4)) M) to the serosal surface stimulated net Na and Cl absorption. This effect may involve intracellular calcium. Chloroquine increased the rabbit ileal calcium content and decreased 45Ca2+ influx from the serosal surface. Both the mucosal and serosal effects of chloroquine described led to a net increase in absorptive function of the intestine and should prove useful in developing treatment of diarrheal diseases.

Reversible effects of phenothiazines on frog gastric stimulation

1984 ◽  
Vol 247 (4) ◽  
pp. G366-G376
Author(s):  
N. Raphael ◽  
E. B. Ekblad ◽  
T. E. Machen
Keyword(s):  
Adenylate Cyclase ◽  
Time Course ◽  
Phosphodiesterase Inhibitor ◽  
Secretory Activity ◽  
Gastric Stimulation ◽  
Camp Content ◽  
Oxyntic Cell ◽  
Camp Generation ◽  
Stimulus Secretion Coupling

The calmodulin inhibitors trifluoperazine (TFP), chlorpromazine (CPZ), and promethazine (PZ) were tested for effects on stimulus-secretion coupling in in vitro bullfrog gastric mucosa. When added to histamine-stimulated tissues, the drugs caused H+ secretion to decrease and transepithelial resistance to increase over a 2-h time course. The potency sequence was TFP (IC50 = 40 microM) greater than CPZ (IC50 = 72 microM) congruent to PZ (IC50 = 72 microM). Anesthetics and other phenothiazines with weak anticalmodulin activity had no effect on secretory parameters. In the presence of histamine, further addition of isobutylmethylxanthine (IBMX, a phosphodiesterase inhibitor) plus dibutyryl cAMP (DBcAMP), IBMX alone, or forskolin (a specific activator of adenylate cyclase) to phenothiazine-inhibited tissues caused full resumption of secretory activity. If TFP (50 microM) was added before stimulation with histamine, the normal increases in tissue cAMP content (which occurs primarily in oxyntic cells), oxyntic cell apical membrane elaboration (morphometric analysis of electron micrographs), and H+ secretion were all blocked. Subsequent addition of IBMX or IBMX plus DBcAMP completely reversed the TFP effect. These results indicate that the histamine-sensitive adenylate cyclase may be the site of TFP inhibition and Ca2+-calmodulin regulation; since these drugs inhibited stimulation by DBcAMP plus IBMX, they may also be exerting additional effects distal to cAMP generation.

scholarly journals Temporal gene expression in bovine corpora lutea after treatment with PGF2alpha based on serial biopsies in vivo

Reproduction ◽  
2001 ◽  
pp. 905-913 ◽  
Author(s):  
SJ Tsai ◽  
K Kot ◽  
OJ Ginther ◽  
MC Wiltbank
Keyword(s):  
Gene Expression ◽  
Corpus Luteum ◽  
Time Course ◽  
Regulatory Protein ◽  
Corpora Lutea ◽  
Multiple Time ◽  
Lh Receptor ◽  
Time Points ◽  
Multiple Biopsies ◽  
Fp Receptor

There is growing evidence to indicate that PGF(2alpha)-induced luteolysis involves altered gene expression in the corpus luteum. Concentrations of mRNA encoding nine different gene products were quantified at three time points from corpora lutea in situ. Serial luteal biopsies (2.1-5.5 mg per biopsy) were collected using an ultrasound-guided transvaginal method and mRNA concentrations were quantified with standard curve quantitative competitive RT-PCR. In the first experiment, three luteal biopsies were collected from three heifers and analysed in multiple assays to evaluate the repeatability of the methods. Concentrations of mRNA for glyceraldehyde 3-phosphate dehydrogenase (GAPDH), PGF(2alpha) receptor (FP receptor) and LH receptor were found to be highly repeatable between assays, between multiple biopsies and between animals (coefficients of variation 1.3-17.3%). In the second experiment, heifers on days 9-11 after ovulation were assigned randomly to receive saline only (n = 6), saline with biopsies taken at t = 0, 0.5 and 4.0 h after injection (n = 6), PGF(2alpha) only (n = 6) or PGF(2alpha) with biopsies taken at t = 0, 0.5 and 4.0 h after treatment (n = 7). Biopsy alone did not change corpus luteum diameter, serum progesterone concentrations or days to next ovulation within the saline- or PGF(2alpha)-treated groups. Concentrations of mRNA for steroidogenic acute regulatory protein, FP receptor, 3beta-hydroxysteroid dehydrogenase, cytosolic phospholipase A(2) and LH receptor were decreased at 4.0 h after PGF(2alpha) injection. In contrast, PGF(2alpha) increased mRNA concentrations for prostaglandin G/H synthase-2, monocyte chemoattractant protein-1 and c-fos but the time course differed for induction of these mRNAs. Concentrations of mRNA for GAPDH did not change after PGF(2alpha) treatment. In conclusion, the techniques allowed analysis of multiple, specific mRNAs in an individual corpus luteum at multiple time points without altering subsequent luteal function. Use of these techniques confirmed that luteolysis involves both up- and downregulation of specific mRNA by PGF(2alpha).

Effect of gonadotrophin-releasing hormone (GnRH) and GnRH agonists upon accumulation of progesterone, cAMP and prostaglandin in isolated preovulatory rat follicles

1982 ◽  
Vol 101 (4) ◽  
pp. 603-610 ◽  
Author(s):  
Torbjörn Hillensjö ◽  
William J. LeMaire ◽  
Martin R. Clark ◽  
Kurt Ahrén
Keyword(s):  
Phosphodiesterase Inhibitor ◽  
Fold Increase ◽  
Control Group ◽  
Prostaglandin E ◽  
Gnrh Agonists ◽  
Bicarbonate Buffer ◽  
Preovulatory Follicles ◽  
Camp Levels ◽  
Dose Dependent ◽  
Stimulation Of

Abstract. To study the acute and direct effects of GnRH agonists preovulatory follicles were isolated from PMSG-treated immature rats and incubated for 15–360 min in modified Kreb's bicarbonate buffer. The levels of cAMP, prostaglandin E, and progesterone were analysed in the tissue and/or incubation media. GnRH and two GnRH agonists produced a dose-dependent stimulation of progesterone production with maximal levels 5–6-fold higher than the control group. As compared to LH the magnitude of this effect was small and was detected only after 240–360 min of incubation. GnRH also stimulated prostaglandin E accumulation and this effect was as pronounced as for LH. There were no detectable changes in cAMP levels for any concentration of GnRH when the incubation time varied between 15 and 120 min whether or not a phosphodiesterase inhibitor was present, but after 240 min of incubation a 2-fold incease in cAMP was found. Consistent with previous results, LH caused a pronouced (40–50-fold) increase in follicular cAMP which was already detectable after 15 min of incubation. Indomethacin abolished the rise in prostaglandin E induced either by GnRH or LH but did not affect the response in terms of cAMP or progesterone, and did not affect the stimulation of meiotic maturation of the follicle-enclosed oocytes caused by the hormones. It is concluded that GnRH can exert acute and LH-like stimulatory effects on the preovulatory rat follicle but that the mechanism of GnRH action is different from that of LH.

Role of interleukin-1β in the regulation of porcine corpora lutea during the late luteal phase of the cycle and during pregnancy

2012 ◽  
Vol 60 (3) ◽  
pp. 395-407 ◽  
Author(s):  
Agata Zmijewska ◽  
Anita Franczak ◽  
Genowefa Kotwica
Keyword(s):  
Secretory Activity ◽  
Oestrous Cycle ◽  
Interleukin 1Β ◽  
Prostaglandin E ◽  
Corpora Lutea ◽  
Prostaglandin Synthase ◽  
Late Luteal Phase ◽  
Prostaglandin F ◽  
Hormonal Milieu

Interleukin-1β (IL-1β) may regulate ovarian physiology. In this study, the influence of IL-1β on secretory activity within the corpora lutea (CL) of cyclic and gravid pigs was determinedin vitroduring different stages of the CL lifespan, e.g. on Days 10–11, 12–13 and 15–16 of the oestrous cycle and pregnancy. IL-1β (10 ng/ml) increased prostaglandin E2(PGE2) secretion from CL of the cyclic and gravid pigs during studied days of the oestrous cycle and pregnancy. Increase (P < 0.05) of prostaglandin F2α(PGF2α) in IL-1β-treated CL was demonstrated only on Days 10–11 of the oestrous cycle. More potent stimulatory effect of IL-1β on PGE2than PGF2αsecretion resulted in the enhancement of the PGE2:PGF2αratio in cyclic and early pregnant CL. IL-1β increased (P < 0.05) progesterone (P4) secretion only in gravid CL and had no effect on oestradiol-17β (E2) release. Expression of cyclooxygenase-2 (COX-2) mRNA was stimulated (P < 0.05) in IL-1β-treated cyclic and gravid CL. Expression of prostaglandin synthase mRNAs in response to IL-1β did not increase. In conclusion, IL-1β modulates PGE2, PGF2αand P4secretion from porcine CL, depending on luteal stage and the surrounding hormonal milieu. The cytokine may act locally in porcine CL for luteotrophic support throughout the PGE2-mediated synthesis and secretion.

PROPERTIES OF HUMAN GONADOTROPHINS ELUTED FROM HUMAN CORPUS LUTEUM AND MOUSE LUTEOMA LH-HCG RECEPTORS

1977 ◽  
Vol 84 (1) ◽  
pp. 142-154 ◽  
Author(s):  
F. E. Cole ◽  
P. C. Arquembourg ◽  
B. F. Rice
Keyword(s):  
Corpus Luteum ◽  
Electrophoretic Mobility ◽  
Present Report ◽  
Corpora Lutea ◽  
Fresh Tissue ◽  
Mouse Tissues ◽  
Tissue Receptors ◽  
Human Corpus Luteum

ABSTRACT Studies were performed to try to determine if gonadotrophins are altered during their interaction with tissue receptors. Immunologic, electrophoretic and binding properties of lactoperoxidase labelled [125I]HLH and [125I]HCG were examined before and after elution from mouse luteoma and human corpora lutea receptor preparations. The anti-HCG used in these studies at a 1:10 000 dilution precipitated 92% of a freshly iodinated [125I]HCG preparation. Receptor eluted [125I]HCG, derived from the same batch of labelled ligand, was virtually quantitatively precipitated by the same dilution of anti-HCG. [125I]HCG eluted from the human corpus luteum was electrophoretically more homogenous when compared to its heterogenous parent labelled preparation and migrated to a position similar to that of native HCG. In Ouchterlony double diffusion experiments against anti-HCG antiserum, corpus luteum eluted [125I]HCG and [125I]HLH showed immunologic identity with each other as well as with native HCG and HLH. Receptor eluted [125I]HCG from the mouse luteoma, following in vivo administration via tail vein injection or after incubation in vitro with labelled hormones, was immunologically indistinguishable from native HCG. The electrophoretic mobility of HCG was retarded when HCG was added to extracts of mouse luteoma, liver and kidney. Eluates of mouse luteoma, applied to Bio-Gel columns previously equilibrated with [125I]HCG showed the ability to concentrate [125I]HCG in the high molecular weight column fractions. Similar results were obtained with columns equilibrated with [125I]TSH and [125I]HGH. [125I]HCG eluted from the mouse luteoma was able to bind to fresh luteoma homogenate but, in contrast to an earlier report with [125I]HCG eluted from rat testis, no enhancement of binding of the eluted [125I]HCG was observed with fresh tissue. These results could be explained by the extraction of non-dialyzable intracellular component during the [125I]HCG elution procedure from the luteoma homogenate which combines with HCG to lower its binding and alter its electrophoretic mobility. This component could be extracted from other mouse tissues and combines with other labelled peptide hormones. Data in the present report support in part the hypothesis that gonadotrophins eluted from mouse luteoma and human corpus luteum are not altered by their interaction with tissue receptors.

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