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PubMed1990

Prostaglandin production by the largest preovulatory follicles in the domestic hen (Gallus domesticus).

An injection of 5 micrograms of gonadotropin-releasing hormone (GnRH) into hens 8 h prior to oviposition advanced the expected time of oviposition by approximately 1 h. The plasma concentration of progesterone increased approximately 1 h earlier in GnRH-injected hens in comparison to saline-injected hens. The plasma concentration of 13,14-dihydro-15-keto-prostaglandin F2 alpha (PGFM) increased significantly (p less than 0.05) at the time of oviposition in both the GnRH- and saline-injected hens. Significantly (p less than 0.05) greater concentrations of prostaglandin F2 alpha (PGF2 alpha) were assayed in media containing the largest preovulatory follicles collected at oviposition than in media containing the second and fifth largest preovulatory follicles collected at the same time. No prostaglandin was detected in media containing small, nonhierarchial follicles. The concentration of PGF2 alpha in media containing granulosa cells from the largest preovulatory follicle was significantly greater (p less than 0.05) than in media containing 4 times as many theca cells. Ovine luteinizing hormone (oLH) alone or in combination with arachidonic acid had no effect on PGF2 alpha output from granulosa cells collected 6 h before oviposition, whereas A23187 caused a small stimulation of PGF2 alpha output. However, treating cells first with oLH and then with A23187 stimulated a 15- to 20-fold increase in PGF2 alpha. None of these stimuli enhanced the already high output of PGF2 alpha when added to incubations of granulosa cells collected within 5 min after oviposition. These data suggest that the granulosa cells of the largest preovulatory follicle are the major intraovarian source of prostaglandin and that production of PGF2 alpha is associated with the preovulatory surges of gonadotropins and steroid hormones preceding oviposition.

Animals↗
PubMed2003

Oocyte maturation, follicle rupture and luteinization in human cryopreserved ovarian tissue following xenografting.

BACKGROUND: Previous studies have demonstrated development of antral follicles in cryopreserved human ovarian tissue after autografting and xenografting, thus indicating successful preservation of follicular function. The study aim was to assess whether these follicles could also undergo periovulatory changes in response to hCG. METHODS: Ovarian tissue from three patients were dehydrated in propanediol (PROH)/sucrose and cryopreserved using the slow cooling/rapid thaw procedure. Thawed tissue was placed under the kidney capsule in immunodeficient mice. Following growth (>20 weeks) in the presence of gonadotrophin, hCG was administered and ovarian tissue examined histologically. RESULTS: Thirty-two antral follicles (diameter range 0.6 to 5 mm) were examined. Histological evidence of a response to hCG was evident in all follicles. Disruption of the concentric layers of mural granulosa and theca cells was apparent in all antral cavities. In 17 (53%) follicles the exterior follicular wall had reduced to a few cells thick, and in eight (25%) the wall had ruptured. Mucified oocyte-cumulus cell complexes were present in 32 follicles, 17 of which had begun to detach from the pedicle. Resumption of meiosis had occurred in over half the oocytes (five metaphase II and seven metaphase I oocytes, eight germinal vesicle breakdown). Two corpora lutea were also detected. CONCLUSIONS: Follicles cryopreserved within human ovarian tissue using the PROH procedure, can develop to the antral stage and undergo periovulatory changes following xenografting and exposure to a luteinizing stimulus.

Adolescent↗
PubMed2002

Effect of co-culture with theca interna on nuclear maturation of horse oocytes with low meiotic competence, and subsequent fusion and activation rates after nuclear transfer.

We conducted this study to examine whether or not co-culture with theca cells improves the maturation rate of horse oocytes with compact cumuli and to evaluate the cytoplasmic competence of oocytes after maturation by assessing fusion, activation and cleavage rates after nuclear transfer. We collected oocytes by scraping follicles from slaughterhouse-derived ovaries and classified them as having an expanded or a compact cumulus. Expanded oocytes were matured in M199 supplemented with 10% FBS and 5 microU/ml FSH for 24 h: compact oocytes were cultured in the same medium, or they were co-cultured in the same medium with theca interna explants, for 24 or 42 h. Oocytes were held with or without 10 microg/ml cytochalasin B, before washing and micromanipulation. and they were fused with donor fibroblasts by electrical pulse. Fused oocytes were activated with Ca ionophore/cycloheximide, cultured for 5 days, and stained with Hoechst to assess nuclear development. We considered oocytes with an enlarged nucleus, or having cleavage with multiple nuclei, to be activated. There was no significant difference in overall maturation rate between compact oocytes cultured with theca and compact controls. When these two groups were combined, there was a significant increase in the proportion of oocytes in MII between 24 and 42 h (P < 0.05). Expanded oocytes had a significantly higher rate of maturation than did compact oocytes (64% versus 25-30%; P < 0.001). There were no significant differences in rates of successful enucleation, fusion, activation or cleavage between compact control and compact + theca oocytes, nor between compact and expanded oocytes; however, expanded oocytes treated with cytochalasin B had a significantly higher survival rate after enucleation than did untreated expanded oocytes (P < 0.05). Three embryos developed from recombined oocytes, with maximum cleavage to 10 cells. The results of this study indicate that co-culture with theca cells does not increase either nuclear or cytoplasmic maturation of compact oocytes. Cytochalasin B is helpful in increasing survival of horse oocytes during enucleation. In vitro matured equine oocytes have the potential to develop into embryos after nuclear transfer; this is the first full report of production of cloned embryos in this species.

Animals↗
PubMed2000

Messenger ribonucleic acids for MAC25 and connective tissue growth factor (CTGF) are inversely regulated during folliculogenesis and early luteogenesis.

Cell proliferation, terminal differentiation, and angiogenesis occur during cycles of follicular and luteal development. In other paradigms, mac25, a potent tumor inhibitor is strongly induced in senescent epithelial cells, whereas CTGF stimulates angiogenesis and wound healing. Using in situ hybridization and immunohistochemistry, we have examined the possibilities that mac25 is inhibited, whereas CTGF is induced during active periods of follicular development and luteogenesis. Ovaries were collected during the follicular and early luteal phases from prostaglandin F2alpha-treated mature pigs and from slaughterhouse sows. CTGF transcripts were induced during the late preantral stage in granulosa and theca cells concomitantly with the appearance of endothelial cells in the theca. CTGF mRNA expression increased in granulosa cells to a maximum (P < 0.01) in mid-antral follicles but was down regulated (P < 0.01) in preovulatory follicles. In contrast, granulosa cell mac25 mRNA expression was undetectable between the preantral and mid-antral stage but was strongly induced in terminally differentiated granulosa cells of preovulatory follicles. CTGF mRNA and peptide were also detected in the theca externa/interstitium and in vascular endothelial cells of ovarian blood vessels, whereas mac25 transcripts, which were also abundant in ovarian blood vessels increased in the theca interna with follicular development. Transcripts of cyclin D 1, a marker of cell proliferation, appeared during the early antral stage and were moderate in granulosa cells but abundant in capillary endothelial cells in the theca interna, underneath the basement membrane. Following ovulation, CTGF and cyclin D1 mRNAs were associated with the migration of endothelial cells into the CL. Subsequently, there was a marked up-regulation of CTGF mRNA expression in granulosa luteins concomitantly with an increase in endothelial cell proliferation within the CL. We hypothesize that CTGF may promote ovarian cell growth and blood vessel formation during follicular and luteal development whereas mac25, a tumor inhibitor, may promote terminal differentiation of granulosa cells in preovulatory follicles.

Animals↗
PubMed1987

[A study on cellular interaction between theca and granulosa cells in estradiol biosynthesis by the human ovary].

Cellular interaction in estradiol biosynthesis by two types of cells in the ovary has been suggested by previous studies. To verify this, steroidogenesis by theca and granulosa cells, separated or combined, was investigated using a two cell combined monolayer culture method. Granulosa and theca cells were harvested from preovulatory follicles of women with normal menstrual cycles. RIA measurements of steroidogenesis revealed the following: 1) Estradiol (E2) production by the combined culture after 24 h and 48 h was significantly higher than the total for each separated culture. 2) delta 4-androstenedione (delta 4) production by the combined culture after 24 h was significantly lower than total delta 4 production by each separated culture. 3) E2 production by the combined culture revealed higher values than the delta 4-added granulosa cell culture after 24 h. These data suggest that cellular interaction is of a synergistic character and it may be attributed not only to aromatase substrate (delta 4), but also to some intercellular E2 stimulating factor.

Adult↗
PubMed2001

Potential local regulatory functions of inhibins, activins and follistatin in the ovary.

The changing pattern of granulosa cell expression of inhibin/activin subunits and follistatin during follicle development and their differential regulation by extrinsic and intraovarian factors supports evidence from functional studies, mostly in vitro, that these proteins have important roles in folliculogenesis, oocyte maturation and corpus luteum function. Gonadal inhibins function as negative feedback hormones to regulate the synthesis and secretion of pituitary FSH, a key determinant of follicle development, but there is little supportive evidence for a peripheral endocrine role for ovary-derived activins or follistatin in this regard. However, activins and follistatin are expressed in numerous other tissues, including anterior pituitary, and they are firmly implicated as local intrapituitary regulators of FSH secretion. Intraovarian actions of granulosa cell-derived activins include the promotion of granulosa cell proliferation and upregulation of FSH receptors, P450arom, oestrogen synthesis, granulosa cell LH receptors and enhancement of oocyte maturation. Through its activin-binding role, follistatin can reverse each of these activin-induced responses. In addition to their endocrine feedback role, granulosa-derived inhibins can sensitize theca cells to LH, thereby enhancing the production of androgens, an essential requirement for follicular oestrogen synthesis. Activins can oppose this effect and suppress thecal androgen production. Granulosa cells overproduce inhibin a subunit precursor relative to betaA/betaB subunit precursors and evidence indicates that different parts of the inhibin a subunit precursor have intrinsic biological activities distinct from inhibin alphabetaA/B dimer, and serve as additional local modulators of follicle and corpus luteum function.

Activins↗
PubMed2000

Response of porcine theca and granulosa cells to GH during short-term in vitro culture.

In Experiment 1, the influence of exogenous GH on steroid secretion by granulosa and theca interna cells recovered from small (1-3 mm), medium (4-6 mm) and large (8-12 mm) follicles was tested. In the second experiment, theca cells (Tc) and granulosa cells (Gc) obtained from large follicles were cultured separately or in two types, Tc/Gc co-culture, where both types of cells were mixed in one well or Gc and Tc were separated by cell culture membrane inserts. In the third experiment, the influence of GH on the morphology of Gc and Tc cells and activity of Delta(5),3beta-hydroxysteroid dehydrogenase (3beta-HSD) was studied. Cells were grown in the control medium (M199+5% of calf serum) or supplemented with 100 ng/ml GH. Testosterone (10(-7) M) was added as the aromatase substrate to granulosa cells cultures. The media were assayed after 48 h of culture for progesterone and oestradiol by RIA. GH added to the culture media had no effect on oestradiol and progesterone secretion by granulosa cells isolated from small and medium follicles while it stimulated both oestradiol and progesterone secretion by Gc isolated from large preovulatory follicles. A stimulatory effect on oestradiol secretion by Tc isolated from all size follicles was observed. GH did not stimulate progesterone secretion by Tc isolated from small follicles but stimulated progesterone secretion by Tc isolated from medium and large preovulatory follicles. Both co-culture systems exhibited synergistic effect on oestradiol secretion. The stimulatory effect on progesterone secretion under the influence of GH was observed in Gc cultured alone and Tc cultured alone. In contrast, the secretion of progesterone was attenuated in both co-culture systems and the addition of GH further augmented this attenuation. A statistically significant increase in oestradiol secretion was observed in all culture conditions. The addition of GH to the culture medium stimulated the activity of 3beta-HSD compared with the control culture from both types of cells. In conclusion, the present studies indicate that there are direct and follicular development stage dependent actions of GH on steroidogenesis of porcine follicular cells.

3-Hydroxysteroid Dehydrogenases↗
PubMed1982

Occurrence of preantral intra-ovarian oocyte release in the rat and the Syrian hamster (Mesocricetus auratus).

Serial sections of 62 rat ovaries and 66 ovaries from Syrian hamsters (Mesocricetus auratus) have been examined by light microscopy. The relative and absolute percentage of preantral intra-ovarian oocyte release (PIOR) was recorded. PIOR was incomplete when the rupture of the basal membrane was not followed by the release of the oocyte from its preantral follicle into the ovarian stroma; conversely, a PIOR was regarded to be complete if the oocyte was released from the preantral follicle into the adjacent stroma. PIOR occurred in 23% of rat ovaries and in 32% of hamster ovaries; meiosis was not resumed. In rat ovaries the frequency of complete PIOR was 71%; the oocyte disappeared quickly after its release and herniated granulosa cells could invade capillaries. By contrast, incomplete PIOR was prevalent in hamster ovaries (67%); oocytes survived in the interstitial cortical tissue after the disappearance of their granulosa and theca cells ("free oocytes"). It is concluded that PIOR reflects the onset of follicular atresia and that different local factors are probably responsible for species differences.

Animals↗
PubMed1994

Oxytocin gene expression and oxytocin immunoactivity in the ovary of the common marmoset monkey (Callithrix jacchus).

Oxytocin was identified in ovaries recovered on Day 5 (+/- 1) of the luteal phase from three female marmoset monkeys (Callithrix jacchus). With use of a reverse transcription-polymerase chain reaction assay, expression of mRNA for oxytocin and oxytocin receptor was detected in both luteal tissue and in the ovarian remnant. Evidence for ovarian synthesis of oxytocin was provided by immunohistochemistry, which showed positive staining for oxytocin and neurophysin in the cytoplasm of the luteal cells. Some luteal cells had a more intensely stained perinuclear region than others for oxytocin immunoreactivity, whereas the staining for neurophysin was evenly distributed. Granulosa and theca cells of antral follicles also showed positive staining for oxytocin immunoreactivity; no reactivity was found in fibroblast or endothelial cells. Oxytocin immunoreactivity was also detected in the luteal tissue of all animals by immunoassay, with values ranging from 2.8 to 12.1 ng/g wet weight. The oxytocin concentration for the ovarian remnant was either very low (0.55-0.75 ng/g wet weight) or nondetectable (< 0.5 ng/g wet weight). Local production of oxytocin within the ovary was suggested by the measurement of higher oxytocin concentrations in the blood from ovaries containing corpora lutea compared with peripheral blood. Collectively, these results provide evidence for ovarian biosynthesis of oxytocin and suggest the possibility of a paracrine role in the regulation of primate ovarian function.

Animals↗
PubMed2006

Cell junctional proteins in the human corpus luteum: changes during the normal cycle and after HCG treatment.

BACKGROUND: Regulation of tissue remodelling and ovarian permeability by intercellular adhesion complexes may be involved in normal and pathological ovarian function. Therefore, the occurrence, distribution and hormonal control of the adherens junction protein vascular endothelial cadherin (VE-cadherin) and the tight junction proteins occludin and claudin in the human corpus luteum (CL) were investigated. METHODS: CLs from patients undergoing hysterectomy for benign reasons were enucleated during early, mid- and late stages of the functional luteal phase and after HCG rescue in vivo. Immunostaining for occludin, claudins 1 and 5 and VE-cadherin was carried out on fixed tissue. Endothelial cells, granulosa lutein cells and theca lutein cells were identified by reference to serial sections immunostained for CD34, 17alpha-hydroxylase and 3beta-hydroxy-steroid-dehydrogenase. Quantitative analyses were performed using image analyses. RESULTS: Occludin was localized to the plasma membrane of granulosa lutein cells and endothelial cells but was absent in theca lutein cells. Claudin 1 was exclusively localized to the plasma membrane of steroidogenic cells. Claudin 5 and VE-cadherin were only present in endothelial cells. After HCG administration in vivo, adherens and tight junction proteins were significantly down-regulated (P < 0.05). CONCLUSIONS: The decrease of junctional proteins after HCG treatment suggests a hormonal control of tight and adherens junctions in the CL associated with tissue remodelling and an increase in luteal permeability during early pregnancy.

Antigens, CD↗
PubMed2003

Targeting of big stanniocalcin and its receptor to lipid storage droplets of ovarian steroidogenic cells.

Stanniocalcin (STC) is a large polypeptide hormone that is widely distributed in tissues such as kidney, adrenal, and ovary. In most tissues, STC exists as a 50-kDa homodimer (STC50). The ovaries produce a higher molecular weight variant (big STC) in androgen-producing theca cell and interstitial cell compartments. Luteal cells, which do not express the STC gene, nonetheless contain high levels of STC protein, suggesting they are targeted by and sequester big STC through a receptor-mediated process. Recently, an STC.alkaline phosphatase fusion protein was used to characterize mitochondrial targeting and sequestration of STC50 and its receptor in liver and kidney. The main objective of the present study was to characterize big STC and its receptor in mammalian ovary and determine whether the ovarian STC variant was similarly targeted to luteal cell mitochondria. By in situ ligand binding, we identified large numbers of STC receptors on corpus luteal cells. However, a more detailed analysis of sub-cellular fractions revealed that both STC and its receptor were not preferentially targeted to mitochondria but instead to cholesterol/lipid storage droplets, which was more indicative of a role in steroidogenesis. Functional studies revealed that additions of big STC had concentration-dependent inhibitory effects on both basal and stimulated progesterone output by primary cultured luteal cells. Furthermore, STC receptor levels were up-regulated in luteal cells in response to protein kinase A activation. Taken together, these findings indicate that theca cell-derived big STC is targeted to the cholesterol/lipid storage droplets of luteal cells to regulate steroidogenesis. This constitutes the first reported description of polypeptide hormone and receptor targeting to cholesterol/lipid droplets and the first biological role for the big STC variant.

Animals↗
PubMed2000

Two methods of inducing low plasma progesterone concentrations have different effects on dominant follicles in cows.

The effects of two methods of inducing low progesterone concentrations on the shape of the plasma progesterone curve and on follicular characteristics in lactating cows were studied. A low ascending progesterone curve was elicited by three PGF2alpha injections on d 3 to 4 of the estrous cycle; a low constant curve by induction of corpus luteum regression on d 6 and insertion of two progesterone-containing intravaginal devices from d 6 to 15 of the cycle. Plasma progesterone concentration was highest in the untreated control group, intermediate in low ascending group, and lowest in the low constant group. On d 15, both control and low ascending groups had one large healthy and one large atretic follicle, suggesting a turnover of follicular waves; in the low constant group, the presence of only one very large healthy follicle indicated follicular persistence. Estradiol concentration in the follicular fluid and its production by granulosa cells were highest in the low constant, intermediate in the low ascending, and lowest in the control group. Androstenedione concentration in the follicular fluid and its production by theca cells were higher in the low constant than in the low ascending and control groups. The results indicate that the low ascending progesterone curve affected follicular development and steroidogenesis differently from the low constant curve. We suggest that the low ascending curve mimics the effects of naturally occurring low plasma progesterone concentrations better, and it might, therefore, be used as a model for studying the effects of low plasma progesterone on fertility.

Androstenedione↗
PubMed2001

Inhibin A is a follicle stimulating hormone-responsive marker of granulosa cell differentiation, which has both autocrine and paracrine actions in sheep.

The aim of these studies was to examine the origin, control and local actions of inhibin A in monovular species, using the sheep as a model. Experiment 1 examined the pattern of mRNA expression for the inhibin subunits in relation to follicular size and pattern of expression to other differentiative markers in granulosa (P450 aromatase) and thecal cells (P450 17alpha-hydroxylase). Experiment 2 examined the pattern of inhibin A production, in relation to oestradiol, by granulosa cells induced to differentiate in vitro with follicle-stimulating hormone (FSH). Experiment 3 examined possible paracrine and autocrine actions of inhibin A by determining the effect of addition of human recombinant inhibin A and/or antiserum to inhibin on gonadotrophin-stimulated cellular differentiation. The results of Experiment 1 showed that expression of mRNA encoding inhibin subnuits alpha, beta(A) and beta(B) is greater (P<0.05) in large oestrogenic follicles than in small follicles but that only expression of the inhibin beta(A) subunit differs (P<0.05) between large oestrogenic and non-oestrogenic follicles. Expression of 17alpha-hydroxylase, but not of the luteinising hormone (LH) receptor, in thecal cells was related to both the size and the oestrogenicity of antral follicles, in a manner similar to that of the inhibin subunits. Experiment 2 demonstrated that the production of inhibin A by sheep granulosa cells is FSH-responsive after prolonged exposure (P<0.001) and precedes the production of oestradiol by around 48 h in the differentiative cascade induced in granulosa cells by FSH. The results of Experiment 3 showed that inhibin A can augment gonadotrophin-stimulated steroid production by both granulosa and theca cells (P<0.01), and that the addition of antiserum to inhibin can inhibit FSH-stimulated oestradiol production by granulosa cells from both small and large follicles (P<0.001). We conclude that inhibin A is an FSH-responsive marker of granulosa cell differentiation which has both autocrine and paracrine actions in sheep.

Androstenedione↗
PubMed2003

Expression of LRH-1 and SF-1 in the mouse ovary: localization in different cell types correlates with differing function.

Steroid biosynthesis in ovary is enhanced by the orphan nuclear receptor, steroidogenic factor-1 (SF-1); however, we reported that liver receptor homolog-1 (LRH-1), a closely related receptor to SF-1, is also expressed in mouse ovary. To further investigate the role of LRH-1 in mouse ovary, we used in situ hybridization to identify the cell types that express LRH-1 versus SF-1, and carried out functional studies to determine the role of LRH-1 in the regulation of the human (h) ovary-specific CYP19 promoter. LRH-1 expression was found to be abundant and highly restricted to cells involved in estrogen biosynthesis-granulosa cells during the estrous cycle, and in corpora lutea (CL) of pregnancy. In contrast, SF-1 was expressed most highly in C(19)-steroid-producing theca cells and interstitium, and at low levels in granulosa and luteal cells. Transfection studies using granulosa cells demonstrated that LRH-1 is a potent regulator of both basal and forskolin-induced transcription of the ovary-specific hCYP19 promoter. This activity was dependent upon two nuclear receptor half-sites within the proximal hCYP19 promoter. Based on these findings, we propose that LRH-1 plays an important role as a competence factor in regulating aromatase, and thus estrogen biosynthesis, in ovary.

Animals↗
PubMed1998

Follicular dominance in cattle is associated with divergent patterns of ovarian gene expression for insulin-like growth factor (IGF)-I, IGF-II, and IGF binding protein-2 in dominant and subordinate follicles.

A decrease in insulin-like growth factor (IGF) binding protein (BP) amount occurs within the follicular fluid of dominant ovarian follicles. At the same time, concentrations of follicular fluid IGF-I do not change. The mRNA for IGF-I, IGF-II, IGFBP-2, and IGFBP-3, in dominant and subordinate follicles were measured to determine if changes in IGF or IGFBP gene expression are associated with follicular dominance. Heifers were ovariectomized during a follicular wave, either during early-dominance (emerging dominant follicle, 9 mm diameter) or mid-dominance (established dominant follicle, 14-16 mm diameter). Follicles were classified as either dominant (DF), subordinate (SF), or not-recruited (NRF; small antral follicles). mRNA was localized by in situ hybridization and measured by image analyses. The IGF-I mRNA (granulosa cells) was greatest in DF and increased in DF, SF, and NRF from early- to mid-dominance. Likewise, IGF-II mRNA (theca cells) was greatest in DF compared with SF or NRF. The IGFBP-2 mRNA (granulosa cells), however, was nearly undetectable in DF, whereas adjacent SF expressed abundant IGFBP-2 mRNA. The NRF were not uniform in their IGFBP-2 expression because only 5 of 13 NRF had IGFBP-2 mRNA. The IGFBP-3 mRNA (granulosa cells) was found only in two NRF, suggesting that local synthesis is not a predominant source of follicular fluid IGFBP-3. These data show that changes in gene expression for IGFBP-2 are opposite to those for IGF-I or IGF-II. Increased IGF-I and IGF-II mRNA and decreased IGFBP-2 mRNA within the DF may be one mechanism leading to follicular dominance. The opposite pattern of IGFBP-2 gene expression in SF and some NRF may lead to follicular atresia.

Animals↗
PubMed1990

Catecholamines inhibit steroidogenesis by cultured porcine thecal cells.

The ovaries of many species contain catecholamines and beta-adrenergic receptors. The present studies were done to determine if catecholamines play a role in the regulation of androgen production by porcine theca cells. Basal and luteinizing hormone (LH)-stimulated androstenedione production was significantly inhibited by noradrenaline and isoproterenol. The inhibitory effects were dose-dependent and were enhanced when the cultures contained the carboxy-O-methyl transferase inhibitor, U-0521. The inhibitory effect of isoproterenol was reversed by the beta-adrenergic antagonist, metoprolol. Isoproterenol caused a generalized inhibition of LH-stimulated steroidogenesis, decreasing the accumulation of pregnenolone, progesterone, androstenedione and estradiol in the culture medium. These studies suggest that catecholamines may be important regulators of thecal androgen production.

Androstenedione↗
PubMed1992

Demonstration of oxytocin release by bovine luteal cells utilizing the reverse hemolytic plaque assay.

Corpora lutea (CL) of a number of species produce oxytocin (OXT). In the present experiments we studied basal, prostaglandin (PG) F2 alpha-stimulated and ascorbate-stimulated OXT release from individual bovine luteal cells utilizing the reverse hemolytic plaque assay (RHPA). Using a mixture of C- and N-terminus-specific antisera against OXT, we were able to demonstrate OXT plaque formation by individual luteal cells. CL consist of two steroidogenic cell types: large luteal cells (LLC), believed to derive from granulosa cells and to produce and secrete OXT, and small luteal cells (SLC), thought to derive from theca cells. To distinguish between these two cell types, we designated cells greater than 20 microns as LLC and those less than 20 microns as SLC. On the basis of this morphological parameter, OXT release from both LLC and SLC was demonstrable. After an incubation period of 15 h, 7% of both cell types formed OXT plaques. PGF 2 alpha and ascorbate increased the size of plaques surrounding both LLC and SLC to more than 200% and 240%, respectively (basal plaque size = 100%). The number of plaque-forming cells increased only slightly in the presence of either PGF 2 alpha or ascorbate in comparison to basal conditions. We suggest that the RHPA can be used to demonstrate peptide release from luteal cells. It is concluded that LLC may be subdivided into functional subclasses because less than 10% of bovine luteal cells release OXT. Known OXT secretagogues increased the amount of OXT released. It appears that not only LLC but also SLC secrete this peptide.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1993

Effects of progesterone and anti-progesterone RU486 on ovarian 3 beta-hydroxysteroid dehydrogenase activity during ovulation in the gonadotrophin-primed immature rat.

The effects of progesterone and RU486, a synthetic anti-progesterone, on ovarian 3 beta-hydroxysteroid dehydrogenase (3 beta-HSD) activity, a key enzyme of progesterone production, were studied during ovulation in immature 22-day-old rats primed with pregnant mares' serum gonadotrophin (PMSG) and human chorionic gonadotrophin (hCG). Ovarian 3 beta-HSD activities had increased significantly 4 h after hCG injection. These increases were inhibited at 4 and 6 h after hCG when 20 mg RU486 kg-1 was administered 2 h before hCG. However, RU486 had no influence on the activity of 3 beta-HSD when administered at the same time as hCG injection. A histochemical study revealed that 3 beta-HSD activities in the granulosa cell layer, but not in the theca cell layer, were inhibited when RU486 was given 2 h before hCG. Serum progesterone concentrations, but not oestradiol concentrations, were significantly suppressed by RU486 treatment 4 and 6 h after hCG. The effect of progesterone on ovarian 3 beta-HSD activity was tested by administering graded doses of progesterone exogenously to rats 2 h before hCG injection. Ovarian 3 beta-HSD activity was increased in a dose-dependent manner, and more than 20 mg progesterone kg-1 significantly stimulated the activity. Although 10 mg progesterone kg-1 did not stimulate ovarian 3 beta-HSD activities, the RU486-inhibited activities were recovered by the concomitant administration of 10 mg progesterone kg-1 with RU486. These results indicate that ovarian 3 beta-HSD activity depends on progesterone concentrations, and suggest an autocrine regulation of progesterone production during ovulation in immature rat ovaries stimulated with PMSG and hCG.

3-Hydroxysteroid Dehydrogenases↗