Immunocytochemical demonstration of oxytocin in bovine ovarian tissues

1985 ◽  
Vol 109 (4) ◽  
pp. 537-542 ◽  
Author(s):  
Th. A. M. Kruip ◽  
H. G. B. Vullings ◽  
D. Schams ◽  
J. Jonis ◽  
A. Klarenbeek
Keyword(s):  
Acetic Acid ◽  
Corpus Luteum ◽  
Granulosa Cells ◽  
Ovarian Tissue ◽  
Picric Acid ◽  
Corpora Lutea ◽  
Luteal Cells ◽  
Antral Follicles

Abstract. The presence of oxytocin in ovarian tissue was examined immunocytochemically. Bovine antral follicles and corpora lutea were fixed with glutaraldehyde, picric acid and acetic acid fixative and immuno-stained by the indirect peroxidase-antiperoxydase (PAP) technique. Immunoreactive oxytocin was demonstrated in the granulosa cells of small and large follicles, in the granulosalutein cells of the young corpus luteum and in the large luteal cells of the mature corpus luteum. The regressing corpus luteum was not stainable. It is discussed that these findings additionally support the view that oxytocin is actually synthesized in ovarian tissues.

Localization of inhibin/activin subunit mRNAs during the luteal phase in the primate ovary

1993 ◽  
Vol 10 (3) ◽  
pp. 245-257 ◽  
Author(s):  
H M Fraser ◽  
S F Lunn ◽  
G M Cowen ◽  
P T K Saunders
Keyword(s):  
Corpus Luteum ◽  
Granulosa Cells ◽  
Luteal Phase ◽  
Biologically Active ◽  
Corpora Lutea ◽  
Α Subunit ◽  
Antral Follicles ◽  
Subunit Mrna ◽  
High Level ◽  
Low Expression

ABSTRACT Localization of inhibin/activin subunit mRNAs within the macaque ovary from the immediate pre-ovulatory period of the menstrual cycle, when serum immunoreactive inhibin begins to rise, to day 9 of the luteal phase, when serum inhibin concentrations are maximal, was investigated using in-situ hybridization. Ovaries were studied on the day of the LH surge (day 0) and on days 2, 5, and 9 of the luteal phase by hybridizing frozen tissue sections with radio-labelled riboprobes specific to the inhibin/activin α-, βA-and βB-subunits. After autoradiographic exposure for 10 and 21 days, grain concentrations were quantified by image analysis. Moderate expression of α-, βA- and βB-subunit mRNA was present within the granulosa cells of the pre-ovulatory follicle (day 0). The granulosa-lutein cells of the corpora lutea expressed high levels of α-subunit at days 2, 5 and 9. mRNAs for βA and βB were detected at low but significant levels in all of the corpora lutea. All healthy antral follicles exhibited a high level of expression of βB-subunit mRNA in the granulosa cells. On day 2 after ovulation these follicles also expressed high α- and moderate βA-subunit mRNA. On day 9 the βB-inhibin mRNA in antral follicles was found in association with low expression of the other subunits. Small follicles in ovaries on day 2 expressed moderate α- and low levels of βB-subunit mRNA, while mRNA for βA was absent. α-subunit mRNA expression was present on day 5 while neither βA- nor βB-subunit mRNA was detected. On day 9 a proportion of small follicles expressed α- and βA-subunit mRNA. These results demonstrate that marked differences are present in the levels of expression of the three inhibin/activin subunit genes between follicles and the corpus luteum. The predominance of the βB-subunit mRNA within antral follicles would be consistent with the synthesis of activin. The predominance of the α-subunit combined with the low expression of the β-subunits in the corpus luteum suggests that both biologically active inhibin and free α-subunit are produced by the primate corpus luteum.

scholarly journals 11β-Hydroxysteroid dehydrogenase expression and activities in bovine granulosa cells and corpora lutea implicate corticosteroids in bovine ovarian physiology

2007 ◽  
Vol 193 (2) ◽  
pp. 299-310 ◽  
Author(s):  
L M Thurston ◽  
D R E Abayasekara ◽  
A E Michael
Keyword(s):  
Molecular Mass ◽  
Granulosa Cells ◽  
Luteal Phase ◽  
Hydroxysteroid Dehydrogenase ◽  
Follicular Phase ◽  
Corpora Lutea ◽  
Luteal Cells ◽  
Antral Follicles ◽  
Follicle Diameter ◽  
Dependent Type

Cortisol–cortisone metabolism is catalysed by the bi-directional NADP(H)-dependent type 1 11β-hydroxysteroid dehydrogenase (11βHSD1) enzyme and the oxidative NAD+-dependent type 2 11βHSD (11βHSD2). This study related the expression of 11βHSD1 and 11βHSD2 enzymes (mRNA and protein) to net 11-ketosteroid reductase and 11β-dehydrogenase (11β-DH) activities in bovine follicular granulosa and luteal cells. Granulosa cells were isolated from follicles of < 4, 4–8, > 8 and > 12 mm in diameter in either the follicular or luteal phase of the ovarian cycle. Luteal cells were obtained from corpora lutea (CL) in the early non-pregnant luteal phase. Enzyme expression was assessed by reverse transcription-PCR and western blotting, while enzyme activities were measured over 1 h in cell homogenates using radiometric conversion assays with 100 nM [3H]cortisone or [3H]cortisol and pyridine dinucleotide cofactors. Irrespective of follicle diameter, the expression of 11βHSD2 and NAD+-dependent oxidation of cortisol predominated in granulosa cells harvested in the follicular phase. In contrast, in granulosa cells obtained from luteal phase follicles and in bovine luteal cells, expression of 11βHSD1 exceeded that of 11βHSD2 and the major enzyme activity was NADP+-dependent cortisol oxidation. Increasing follicular diameter was associated with progressive increases in expression and activities of 11βHSD2 and 11βHSD1 in follicular and luteal phase granulosa cells respectively. In follicular phase granulosa cells from antral follicles < 12 mm, 11βHSD1 migrated with a molecular mass of 34 kDa, whereas in the dominant follicle, CL and all luteal phase granulosa cells, a second protein band of 68 kDa was consistently detected. In all samples, 11βHSD2 had a molecular mass of 48 kDa, but in large antral follicles (> 8 mm), there was an additional immunoreactive band at 50 kDa. We conclude that 11βHSD2 is the predominant functional 11βHSD enzyme expressed in follicular phase granulosa cells from growing bovine antral follicles. In contrast, in bovine granulosa cells from dominant or luteal phase follicles, and in bovine luteal cells from early non-pregnant CL, 11βHSD1 is the major glucocorticoid-metabolising enzyme. The increasing levels of cortisol inactivation by the combined NADP+- and NAD+-dependent 11β-DH activities suggest a need to restrict cortisol access to corticosteroid receptors in the final stages of follicle development.

Induced luteal regression: differential effects on follicular and luteal inhibin/activin subunit mRNAs in the marmoset monkey

1995 ◽  
Vol 144 (2) ◽  
pp. 201-208 ◽  
Author(s):  
H M Fraser ◽  
S F Lunn ◽  
P F Whitelaw ◽  
S G Hillier
Keyword(s):  
Corpus Luteum ◽  
Granulosa Cells ◽  
Luteal Phase ◽  
Gnrh Antagonist ◽  
High Expression ◽  
Corpora Lutea ◽  
Ovulatory Cycle ◽  
Α Subunit ◽  
Luteal Regression ◽  
Antral Follicles

Abstract During the luteal phase of the primate ovulatory cycle the predominant inhibin/activin subunit mRNAs produced by the corpus luteum and antral follicles are those for the α- and βB-subunits respectively. The control of expression of these mRNAs and the resultant nature of the endocrine and paracrine signals which they may potentially generate has yet to be elucidated. Inhibin/activin subunit mRNAs may have a role in both the paracrine regulation of follicular and luteal function and modulation of FSH secretion. The aim of this study was to investigate the expression of inhibin/activin subunit mRNAs following luteal regression induced by either withdrawal of LH support (GnRH antagonist treatment), or by a direct inhibitory action (prostaglandin administration). Marmoset monkeys with regular ovulatory cycles were treated on day 8 and 9 of the luteal phase with either GnRH antagonist, prostaglandin or vehicle (n=3 per group). Ovaries were studied 48 h after onset of treatment (on day 10 of the luteal phase) by hybridizing frozen tissue sections with radiolabelled riboprobes specific to the inhibin/activin α-, βA- and βB-subunit mRNAs. After autoradiographic exposure, grain concentrations were quantified by image analysis. In corpora lutea from control marmosets there was high expression of α-mRNA with only marginal expression of βB-mRNA. Corpora lutea in animals treated with GnRH antagonist or prostaglandin had markedly reduced expression of α-mRNA while βB-mRNA was unchanged. In controls, all healthy antral follicles exhibited a high level of expression of βB-mRNA in the granulosa cells and low expression of α-mRNA in theca cells. This was unaffected by either treatment. βA-mRNA was found at a low level in granulosa cells but was not evident at a significant level in the corpora lutea of any of the groups. These results demonstrate (1) the marmoset corpus luteum is a source of high expression of α-subunit mRNA, (2) this α-mRNA is dependent upon LH support, (3) the process of luteal regression takes place without alteration of βB-mRNA. Antral follicle α- and βB-mRNAs are independent of the process of luteal regression or gonadotrophic withdrawal during the period of the luteal-follicular phase transition. Journal of Endocrinology (1995) 144, 201–208

Enzymic Correlates of Development, Secretory Function and Regression of Follicles and Corpora Lutea in the Bovine Ovary. PART II: Formation, Development and Involution of Corpora Lutea

1968 ◽  
Vol 59 (2_Suppl) ◽  
pp. S35-S51 ◽  
Author(s):  
B. L. Lobel ◽  
E. Levy
Keyword(s):  
Corpus Luteum ◽  
Vascular System ◽  
Hydrolytic Enzymes ◽  
Sharp Decrease ◽  
Cellular Growth ◽  
Corpora Lutea ◽  
Cell Nuclei ◽  
Steroid Synthesis ◽  
Luteal Cells ◽  
Vascular Walls

ABSTRACT Activities of various hydrolases and dehydrogenases were studied during the formation, development and involution of cyclic corpora lutea and in the corpora lutea of early pregnancy. At 24 hours postovulation the luteal cells, whether of granulosal or thecal origin, contained demonstrable levels of Δ5-3β-hydroxysteroid dehydrogenase and the NADP and NADPH2 diaphorases. During the period of proliferation and cellular growth, enzymic activities in the luteal cells were moderate at first, and then increased. In the mature corpus luteum, activities of the dehydrogenases occurred in all luteal cells but were most intense in the large polymorphic luteal cells. Activities of hydrolytic enzymes, low in the immediate postovulatory period, increased with the development of the vascular system. Enzymic characteristics of corpora lutea of gestation were similar to those of cyclic corpora, except for phosphorylase activity which was observed in luteal cells in gestational corpora, but confined to the vascular walls in cyclic corpora. No increase in activities of 17β- and 20β-hydroxysteroid dehydrogenases (above those seen in pre-ovulatory follicles) were observed after incubation of sections of either mature cyclic or gestational corpora. Involution of cyclic corpora lutea began with degenerative changes in the blood vessels: pyknosis of the endothelial cell nuclei and a sudden decline in activities of hydrolytic enzymes in the vascular walls. Subsequently, the luteal cells showed a sharp decrease in activities of the dehydrogenases as well as other signs of regressive change. The cytochemical findings are discussed in relation to biochemical observations on steroid synthesis by the bovine corpus luteum.

Localization of oxytocin and neurophysin in baboon (Papio anubis) corpus luteum by immunocytochemistry

1986 ◽  
Vol 113 (4) ◽  
pp. 570-575 ◽  
Author(s):  
Firyal S. Khan-Dawood
Keyword(s):  
Corpus Luteum ◽  
Rabbit Serum ◽  
Corpora Lutea ◽  
Positive Staining ◽  
Normal Rabbit Serum ◽  
Fallopian Tubes ◽  
Papio Anubis ◽  
Luteal Cells ◽  
Luteal Tissue ◽  
Immunocytochemical Reaction

Abstract. Immunoreactive oxytocin is detectable in the corpora lutea of women and cynomolgus monkeys by radioimmunoassay. To localize the presence of oxytocin and neurophysin I in ovarian tissues of subhuman primates, three corpora lutea and ovarian stromal tissues and two Fallopian tubes obtained during the menstrual cycle of the baboon and decidua from two pregnant baboons were examined using highly specific antisera against either oxytocin or neurophysin I and preoxidase-antiperoxidase light microscopy immunohistochemistry. Oxytocin-like as well as neurophysin I-like immunoreactivities were found in some cells of all the corpora lutea only, but could not be demonstrated in ovarian stromal tissues, Fallopian tubes and decidua. Specificity of the immunocytochemical reaction was further confirmed by immunoabsorption of the antiserum with excess oxytocin or neurophysin, after which the immunoreactivities for both oxytocin and neurophysin in the luteal tissue were negative. Similar controls using normal rabbit serum gave no positive staining for either oxytocin or neurophysin. Counterstaining of the positive immunoreactivities for oxytocin and neurophysin I with Mayer's haematoxylin and eosin demonstrated clearly that the oxytocin and neurophysin I appeared as granular material mainly within the cytoplasm of the luteal cells. The localization of immunoreactive oxytocin and neurophysin I in the corpus luteum of the baboon demonstrates directly the presence of these two neurohypophysial peptides within primate luteal cells and suggests their local production.

scholarly journals Immunohistological Localization and Expression of α-Actin in the Baboon (Papio anubis) Corpus Luteum

1997 ◽  
Vol 45 (1) ◽  
pp. 71-77 ◽  
Author(s):  
Firyal S. Khan-Dawood ◽  
Jun Yang ◽  
M. Yusoff Dawood
Keyword(s):  
Corpus Luteum ◽  
Luteal Phase ◽  
Adherens Junctions ◽  
Tunica Albuginea ◽  
Corpora Lutea ◽  
Papio Anubis ◽  
Lh Surge ◽  
Luteal Cells ◽  
Steroidogenic Cells ◽  
E Cadherin

We have recently shown the presence of E-cadherin and of α- and γ-catenins in human and baboon corpora lutea. These are components of adherens junctions between cells. The cytoplasmic catenins link the cell membrane-associated cadherins to the actin-based cytoskeleton. This interaction is necessary for the functional activity of the E-cad-herins. Our aim therefore was to determine the presence of α-actin in the baboon corpus luteum, to further establish whether the necessary components for E-cadherin activity are present in this tissue. An antibody specific for the smooth muscle isoform of actin, α-actin, was used for these studies. The results using immunohistochemistry show that (a) α-actin is present in steroidogenic cells of the active corpus luteum, theca externa of the corpus luteum, cells of the vasculature, and the tunica albuginea surrounding the ovary. The intensity of immunoreactivity for α-actin varied, with the cells of the vasculature reacting more intensely than the luteal cells. A difference in intensity of immunoreactivity was also observed among the luteal cells, with the inner granulosa cells showing stronger immunoreactivity than the peripheral theca lutein cells. There was no detectable immunoreactivity in the steroidogenic cells of the atretic corpus luteum. However, in both the active and atretic corpora lutea, α-actin-positive vascular cells were dispersed within the tissue. (b) Total α-actin (luteal and non-luteal), as determined by Western blot analyses, does not change during the luteal phase and subsequent corpus luteum demise (atretic corpora lutea). (c) hCG stimulated the expression of α-actin and progesterone secretion by the early luteal phase (LH surge + 1–5 days) and midluteal phase (LH surge + 6–10 days) cells in culture, but only progesterone in the late luteal phase (LH surge + 11–15 days). The data show that α-actin is present in luteal cells and that its expression is regulated by hCG, thus suggesting that E-cadherin may form functional adherens junctions in the corpus luteum.

STEROID METABOLIC PATHWAYS IN THE OVARY OF THE CHINCHILLA (CHINCHILLA LANIGER)

1972 ◽  
Vol 52 (1) ◽  
pp. 37-50 ◽  
Author(s):  
W. H. TAM
Keyword(s):  
Corpus Luteum ◽  
Metabolic Pathways ◽  
Ovarian Tissue ◽  
Controlling Factor ◽  
Steroid Metabolism ◽  
Interstitial Tissue ◽  
Corpora Lutea ◽  
Luteal Tissue ◽  
Kinetic Investigations ◽  
High Yields

SUMMARY The ovarian tissue components of the pregnant chinchilla were incubated with equimolar amounts of [7α-3H]pregnenolone and [4-14C]progesterone. The greater contribution by [7α-3H]pregnenolone than by [4-14C]progesterone towards the formation of 17α-hydroxyprogesterone and androstenedione, and the relatively high yields of 17α-hydroxypregnenolone and dehydroepiandrosterone showed that both the 4-ene and 5-ene pathways of steroid metabolism were used in the interstitial tissue. No significant amount of 17α-hydroxylation was observed in the primary and accessory corpora lutea. The results of kinetic investigations using [7α-3H]pregnenolone as substrate also demonstrated a precursor—product relationship between dehydroepiandrosterone and androstenedione in the interstitial tissue, but this was not apparent in the luteal tissue. The results indicated that the interstitial tissue was capable of synthesizing progesterone and oestrogens as major products, and that the lack of 17α-hydroxylation in the luteal tissue was a controlling factor ensuring the synthesis of progesterone as its principal hormonal product. A small amount of [4-14C]dehydroepiandrosterone was always isolated with a much larger amount of the tritiated compound. This implied the conversion of 14C-labelled 4-en-3-oxosteroids into 5-ene-3β-hydroxysteroids which has generally been regarded as impossible. The isolation of this product, which may be an artifact, and the possibility that progesterone and oestrogens may be synthesized by different cells (granulosa and theca lutein cells) in the corpus luteum, or that there may be a third pathway for oestrogen synthesis, as suggested by the results of the kinetic experiments, are discussed.

scholarly journals Immunolocalization of steroidogenic enzymes in the corpus luteum and placenta of the Japanese black bear, Ursus thibetanus japonicus, during pregnancy

Reproduction ◽  
2001 ◽  
pp. 587-594 ◽  
Author(s):  
T Tsubota ◽  
S Taki ◽  
K Nakayama ◽  
JI Mason ◽  
S Kominami ◽  
...  
Keyword(s):  
Cytochrome P450 ◽  
Corpus Luteum ◽  
Fetal Development ◽  
Black Bear ◽  
Corpora Lutea ◽  
Luteal Cells ◽  
Ursus Thibetanus ◽  
Delayed Implantation ◽  
Post Implantation ◽  
Implantation Period

The Japanese black bear, Ursus thibetanus japonicus, is a seasonal breeder and shows delayed implantation for several months during pregnancy. The objective of this study was to clarify the steroidogenic capability of the corpus luteum and placenta during pregnancy, including both delayed implantation and fetal development, by immunolocalization of steroidogenic enzymes in these organs of the Japanese black bear. Ovaries and placentae from 15 wild Japanese black bears, which had been killed legally by hunters and were thought to be pregnant, were used in an immunocytochemical study to localize the cholesterol side chain cleavage cytochrome P450 (P450scc), 3beta-hydroxysteroid dehydrogenase (3betaHSD), 17alpha-hydroxylase cytochrome P450 (P450c17) and aromatase cytochrome P450 (P450arom) by the avidin-biotin-peroxidase complex method using polyclonal antisera raised in mammals against P450scc, 3betaHSD, P450c17 and P450arom. P450scc and 3betaHSD were localized in all luteal cells throughout pregnancy. P450c17 was present in a few luteal cells, especially in the outer area of the corpus luteum throughout pregnancy, but the number of positively immunostained cells decreased during the post-implantation period. Cells positively immunostained for P450c17 were significantly smaller than negatively immunostained cells (P < 0.01). P450arom was present sporadically in a few luteal cells throughout pregnancy, but the number of positively immunostained cells decreased during the post-implantation period. The size of cells positively immunostained for P450arom was not significantly different from that of negatively immunostained cells. The whole placenta was negatively immunostained for P450scc, 3betaHSD and P450c17, but P450arom was present in the syncytiotrophoblasts and endothelial cells of maternal blood vessels. These results indicate that, in the Japanese black bear, corpora lutea are a source of progesterone which may play an important role in the maintenance of delayed implantation and fetal development during pregnancy. Corpora lutea have a minimum capability to synthesize androgen in small luteal cells and oestrogen in normal-sized luteal cells during pregnancy, and placentae have the ability to synthesize oestrogen during late pregnancy.

β-Carotene and protein complex found in microsomal membranes of bovine corpora lutea

1996 ◽  
Vol 1996 ◽  
pp. 69-69
Author(s):  
AJ Holt ◽  
RG Rodway ◽  
JBC Findlay ◽  
HS Sands ◽  
DN Batchelder
Keyword(s):  
Corpus Luteum ◽  
High Performance ◽  
Gel Filtration ◽  
Biological Action ◽  
Corpora Lutea ◽  
Luteal Cells ◽  
Bovine Corpus Luteum ◽  
Microsomal Membranes ◽  
Ruminant Animals ◽  
Β Carotene

The role of β-carotene in the fertility of ruminant animals has long been acknowledged (Friesecke, 1978). Initially this was thought to be due to its action as a vitamin A precursor but recently β-carotene itself has been considered responsible for improving reproductive performance (Hurley & Doane, 1989). The mechanism by which β-carotcne acts is unclear, but as its concentration in the corpus luteum was typically found to be 70μg per gram of tissue, its biological action is probably exerted in this area.β-Carotene has been studied in the bovine corpus luteum using Raman spectroscopy, high performance liquid chromatography (HPLC) and gel filtration chromatography. The structure of β-carotene produces a characteristic Raman spectrum and by utilising an imaging technique, photographs of isolated luteal cells were obtained indicating the regions of β-carotene within them. Differential centrifugation was used to obtain pure subcellular fractions of luteal cells.

Oxytocin may play a role in the control of the human corpus luteum

1982 ◽  
Vol 95 (1) ◽  
pp. 65-70 ◽  
Author(s):  
G. J. S. Tan ◽  
R. Tweedale ◽  
J. S. G. Biggs
Keyword(s):  
Menstrual Cycle ◽  
Corpus Luteum ◽  
Inhibitory Action ◽  
Chorionic Gonadotrophin ◽  
Human Chorionic Gonadotrophin ◽  
Corpora Lutea ◽  
Luteal Cells ◽  
Progesterone Production ◽  
Human Corpus Luteum

The effects of oxytocin on dispersed luteal cells from human corpora lutea of the menstrual cycle were studied. Oxytocin at a concentration of 4 mi.u./ml produced a slight increase in basal progesterone production. However, higher oxytocin concentrations (400 and 800 mi.u./ml) markedly inhibited both basal and human chorionic gonadotrophin-induced progesterone production. These data provide evidence for an effect of oxytocin on the human corpus luteum. In view of the inhibitory action of oxytocin, increased secretion of this hormone may be important in the demise of the corpus luteum at the end of the menstrual cycle.

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