Postovulatory Period Research Articles

Feed restriction and insulin treatment affect subsequent luteal function in the immediate postovulatory period in pigs: progesterone production in vitro and messenger ribonucleic acid expression for key steroidogenic enzymes.

Progesterone production and release in vitro, and mRNA expression for key steroidogenic enzymes, were studied in luteal tissue recovered in the immediate postovulatory period from cyclic gilts allocated to one of three treatments: moderate feed restriction during the first (RH) or second week of the estrous cycle, with (HR+I) or without (HR) concomitant injections of long-acting insulin. Time of feed restriction affected neither progesterone production or release, nor mRNA expression for several key steroidogenic enzymes. However, luteal tissue from RH but not from HR gilts responded to LH stimulation by increasing progesterone production and release (P: < 0.05). Insulin treatment increased progesterone production and release, restored luteal tissue responsiveness to LH, up-regulated steroidogenic enzyme mRNA expression, and down-regulated the tissue inhibitor of metalloproteinase-I mRNA expression in HR+I compared with HR gilts (P: < 0.05). In vitro progesterone production and gene expression were affected by time of tissue collection after ovulation in RH and HR gilts but not in HR+I gilts, and were correlated with temporal changes in oviductal and peripheral plasma progesterone concentrations. Inherent differences in luteal function therefore appear to mediate latent effects of nutrition and insulin treatment on circulating progesterone concentrations in the critical postovulatory period in gilts.

Distribution of NADPH-diaphorase and nitric oxide synthase (NOS) in different regions of porcine oviduct during the estrous cycle.

Nitric oxide synthase (NOS) is responsible for the biological production of nitric oxide (NO) in several organs, including those of the reproductive tract. We investigated potential changes in NADPH-diaphorase (NADPH-d) activity (marker for NOS activity) and the presence and distribution of NOS in the porcine oviduct. Tissues were obtained from gilts (n=16) on different days of the estrous cycle. One fallopian tube was used for histo- and immunohistochemistry and the other for Western blotting analysis. NADPH-d activity was much higher in the epithelium of the mucosa than in the myosalpinx. The highest activity of NADPH-d was always found in the epithelium of the isthmus. The intensity of the reaction (arbitrary units +/- SEM) in isthmus epithelium increased from the postovulatory period until early proestrus (96.2 +/- 11.2) and then gradually decreased. The lowest intensity of NADPH-d reaction in the epithelium of the isthmus was seen at estrus (58.4 +/- 7.7). The most intense NADPH-d activity in myosalpinx of all parts of the oviduct was observed at the postovulatory stage of the estrous cycle (isthmus 38.3 +/- 2.5; ampulla 35.6 +/- 4.2; infundibulum 24.7 +/- 0.8) and then decreased during the remaining stages of the estrous cycle (p< 0.001). The presence of endothelial NOS (eNOS) was detected in epithelial cells of mucosa and in endothelium of vascular tissues and myosalpinx during all studied days of the estrous cycle. The positive reaction for inducible NOS (iNOS) was restricted only to the endothelium of lymph vessels and some blood vessels. Because our Western blotting analysis revealed that porcine oviduct contains eNOS but not iNOS, we suggest that eNOS is the main isoform of NOS expressed in the porcine oviduct. We concluded that the different activity of NADPH-d in the various regions of the oviduct, accompanied by changes in its activity during the course of the estrous cycle, could indicate an important role of NO in regulation of tubal function.

Relationships between changes in plasma concentrations of leptin before and after parturition and the timing of first post-partum ovulation in high-producing Holstein dairy cows.

During early lactation, dairy cattle are in negative energy balance and the delay to first post-partum ovulation depends on the time taken to recover from this situation. Lactating cows rely heavily on body fat to meet their requirements, leading to the suggestion that leptin, a hormone secreted mainly by adipocytes, is acting as a metabolic signal to sites that control the reproductive axis. The relationship between plasma leptin concentrations and the timing of the first ovulation post partum in 20 high-producing Holstein dairy cows, using a radioimmunoassay based on recombinant bovine leptin was studied. Plasma leptin concentrations declined after parturition, reached a nadir of 0.74 +/- 0.17 ng mL(-1) on 10.1 +/- 2.2 days after parturition (all values are mean +/- SEM). They then increased and became stable near the time of ovulation. Leptin concentrations averaged 1.81 +/- 0.31 ng mL(-1) in the 14 days prepartum, 1.32 +/- 0.21 ng mL(-1) in the post-partum preovulatory period and 1.61 +/- 0.24 ng mL(-1) in the post-ovulatory period. The differences between periods were significant (P<0.01). The interval from parturition to first ovulation averaged 25.9 +/- 2.0 days and was not correlated with the prepartum, preovulatory or post-ovulatory leptin values. However, the interval to first ovulation correlated significantly (r = 0.83 P < 0.0001) with the interval from parturition to the leptin nadir. These results show that plasma concentrations of leptin decrease in dairy cows in the early post-partum period and suggest that a delay in the recovery of leptin secretion increases the delay to the first ovulation.

Postovulatory follicle: A model for experimental studies of programmed cell death or apoptosis in teleosts

This is the first evidence of programmed cell death, or apoptosis, occurring in the postovulatory follicle (POF) of teleost fish. Females of Astyanax bimaculatus lacustris were submitted to induced ovulation through injecting pituitary extract. Ultrastructural analyses of POFs at time intervals varying from zero to four days postspawning showed several characteristic events of the apoptosis. Typical apoptotic figures, such as nucleus with chromatin condensation underlying the nuclear envelope in a crescent pattern and apoptotic bodies at different stages of formation and reabsorption, were observed in the follicular cells a few days after the onset of the postovulatory period. The results indicated that apoptosis is the major mechanism responsible for the elimination of the follicular cells in the POFs of A. bimaculatus lacustris during ovarian recovery postspawning. It is suggested that POFs might be used as an experimental model in dynamic studies involving cell death in teleosts.

Reproductive endocrinology of female elasmobranchs: Lessons from the little skate (Raja erinacea) and spiny dogfish (Squalus acanthias)

Conventional classification of reproductive modes in female elasmobranchs fails to account for the diversity in ovarian dynamics that operate during oviparous and viviparous cycles. Delineating this diversity is crucial for understanding the endocrine regulation of the manifold physiological mechanisms utilized to retain and protect eggs and developing embryos, to fuel embryogenesis, and to manage the intrauterine milieu. Oocyte development and follicular steroidogenesis overlap with egg retention and pregnancy in some species, whereas in others the follicular phase of the cycle is temporally separated from the gravid period. A luteal phase predominates the post-ovulatory period in viviparous species. In oviparous species, the luteal phase overlaps with the follicular cycle. This heterogeneity in ovulatory cycles suggests that the endocrine system evolved a transmutable system for regulating steroidogenesis and the control of the reproductive events. The reproductive biology and endocrinology of the oviparous little skate and lecithotrophic viviparous spiny dogfish are reviewed in order to derive a working hypothesis that explains the complex nature of endocrine patterns observed in species utilizing disparate reproductive modes. An understanding of the adaptations in ovarian dynamics to particular ovulatory cycles is key to developing theories about the evolution of reproductive strategies in female elasmobranchs. J. Exp. Zool. 284:557–574, 1999. © 1999 Wiley-Liss, Inc.

Distribution of extracellular matrix components in the developing ruminant corpus luteum: a wound repair hypothesis for luteinization.

The aim of this study was to investigate corpus luteum development by visualization of extracellular matrix proteins in the tissue at sequential stages of the luteal phase. Corpora lutea were collected from oestrus-synchronized sheep and from bovine material from an abattoir. The distributions of collagen types I and IV, fibronectin and von Willebrand factor were determined using immunohistology and semi-quantitative image analysis. During the post-ovulatory period, a fibronectin- and von Willebrand factor-rich matrix occurred centrally, adjacent to the inner parenchymal surface, whereas during early luteal development a clear border of fibronectin separated the inner parenchyma from the lumen. The inner parenchyma had abundant fibronectin initially, but the amount decreased as the rate of organ growth decreased. Over the same period, the amount of collagen type I first increased and then decreased. Collagen type I and fibronectin were less abundant in other regions of the parenchyma, and the general pattern was of slightly increasing amounts of collagen type I and decreasing amounts of fibronectin as luteal development proceeded. In contrast to earlier studies, only a small percentage of large luteal cells was found to have an associated layer of collagen type IV (presumed basal lamina). It is concluded that luteal growth and maturation require organized sequences of tissue remodelling. The central meshwork of fibronectin and von Willebrand factor and sequential deposition of collagen type I and fibronectin are strongly reminiscent of events in granulation tissue. This indicates that luteinization may be best understood as a wound repair-like process that succeeds the inflammation-like events of ovulation.

Gastric emptying in hyperemesis gravidarum and non-dyspeptic pregnancy.

Emesis and hyperemesis are significant problems associated with early pregnancy. However, gastric emptying of solids has never been studied during early pregnancy in humans. To investigate gastric emptying of solids in patients recovering from hyperemesis gravidarum and in non-dyspeptic pregnant women and to compare these results with a group of healthy non-pregnant women. Fourteen patients with hyperemesis gravidarum, 10 non-dyspeptic pregnant women and 36 non-pregnant women in the first half of the menstrual cycle underwent a gastric emptying study. Seven non-pregnant women repeated the test in the post-ovulatory period. Gastric emptying of solids was not significantly delayed in non-dyspeptic pregnant women compared with non-pregnant women. The emptying rate tended to be impaired in the post-ovulatory period of the menstrual cycle. Solid emptying was significantly accelerated in patients recovering from hyperemesis gravidarum, correlating well with thyroid function in the latter group. Pregnancy in humans is not associated with decreased solid gastric emptying. In subjects recovering from hyperemesis gravidarum, solid emptying is increased, correlating well with thyroid function abnormalities. Nausea and vomiting in hyperemesis are therefore probably not due to upper gastrointestinal disorders.

Ultrastructural changes in the oviductal mucosa throughout the sexual cycle inBufo arenarum

In Bufo arenarum the oviduct exhibits conspicuous changes throughout the sexual cycle. In the present study, we analyzed the optical and ultrastructural characteristics of the oviductal pars convoluta mucosa, the portion responsible for jelly secretion, during both the preovulatory and postovulatory periods. Secretory epithelial cells, ciliated cells, basal cells, and glandular cells are described. Secretory epithelial cells are characterized by the presence of secretory granules, the size, shape and electron density of which vary markedly. Their contents are mainly released by exocytosis into the oviductal lumen. Moreover, in the preovulatory period, apocrine, and holocrine secretion processes frequently occur. During the postovulatory period, these cells exhibit a marked diminution of secretory granules. Ciliated cells show a typical ultrastructural organization. Basal cells are distinguished in the lower part of the epithelium by their heterochromatic nuclei and electron-lucent cytoplasm. These cells, to the best of our knowledge, are reported for the first time in Amphibia. Glandular cells exhibit oval, round, or polyhedric granules, most of them with one or more cores. Our results indicate that the contents of epithelial and glandular secretory cells are partially secreted during the preovulatory period. Additional secretion occurs during the transit of the oocytes. J. Morphol. 239:61–73, 1999. © 1999 Wiley-Liss, Inc.

Endocrinology of the ovarian cycle and pregnancy in the Asian (Elephas maximus) and African (Loxodonta africana) elephant.

In reviewing the literature, this paper assesses the current level of understanding of the hormonal control of the ovarian cycle and pregnancy in the elephant. Data are compiled from two genera, Elephas maximus and Loxodonta africana, recognizing differences where known to exist. Measurements of various steroid and peptide hormones, but most notably immunoreactive progesterone (iP), indicate an overall cycle length of 15-16 weeks, comprising an 8-11-week luteal phase and a shorter inter-luteal (follicular) period of 4-6 weeks. Oestrogen related phenomena suggest the occurrence of two (possibly more) waves of follicular development with a 3-week periodicity, although oestrogen levels are low and fluctuate without clear pattern. The inter-luteal period is characterized by two discrete LH peaks also 3 weeks apart, but only the second is associated with an iP rise. Ovulation rate is variable and additional luteal structures lacking ovulation points are probably formed each cycle. The post-ovulatory period is associated with elevated progestogen levels, mainly 5alpha-reduced compounds, while progesterone itself is a minor secretory product. A revised model for the elephant ovarian cycle is presented. Detailed information on the endocrinology of pregnancy is lacking. Elevated progestins beyond the normal luteal phase indicate that conception extends the functional lifespan of the CL, with maximum steroidogenic activity between 3-15 months. The nature of the luteotrophic support is not known and placental gonadotrophins have not been conclusively demonstrated. Progestins fluctuate at or slightly above luteal phase values throughout pregnancy. There is a marked increase in prolactin levels after 16-24 weeks and a relationship with oestrogen secretion may exist since circulating and urinary levels show a progressive increase from 6-8 months. A placental contribution to progestin secretion is likely, although direct evidence is lacking. Considerable enlargement of foetal gonads during the second half of pregnancy in African elephants suggests a possible steroidogenic function, but none has been defined. The endocrine events leading to parturition are unknown. In the Asian elephant, a fall in iP precedes parturition; oestrogen levels decline and cortisol increases at the time of birth. The paper concludes with a brief selection of priorities for future research.

Changes in sperm ultrastructure and localisation in the porcine oviduct around ovulation.

Changes in the morphology and localisation of spermatozoa in the utero-tubal junction (UTJ) and three equal segments of the isthmus (lower, middle and upper isthmus) within +/-8 h of ovulation were studied with scanning and transmission electron microscopy. Spermatozoa were located at specific sites within the UTJ-isthmic environment. Two sperm subpopulations were evident, one with epithelial contact and one without such contact. Most of the sperm population with epithelial contact maintained intact plasma membrane during the pre-ovulatory period and showed acrosome reacted-like membrane changes during the post-ovulatory period. In contrast, plasma membranes in most of the spermatozoa with no epithelial contact were already broken before ovulation. Ovulation was also accompanied by a relocation of spermatozoa from the mucosal crypts/interfolds towards the more central part of the mucosal surface.

Motility of spermatozoa in hydrosalpingeal and follicular fluid of pigs.

Hydrosalpinges were created to collect adequate volumes of fluid during pre-, peri- and postovulatory intervals from the ampulla, ampullary-isthmic junction and the isthmic-utero-tubal junction of the oviducts from Large White gilts that had exhibited at least two natural oestrous cycles. The accumulated fluids, follicular fluid and Butschwiler's medium were compared for their effects on various parameters of boar sperm motility using the CellSoft, computer-assisted, digital image analysis system. Sperm velocity (micron s-1 +/- SEM) was significantly higher (P < 0.05) in follicular fluid (84 +/- 3; n = 5) than in fluids from the ampulla during peri- and early postovulatory intervals, and from the isthmic-utero-tubal junction during pre- and early postovulatory intervals. It was also higher (P < 0.05) than in the fluid from the ampullary-isthmic junction during pre- and early postovulatory intervals; however, sperm velocity in follicular fluid was not significantly different from that in the periovulatory fluid from the ampullary-isthmic junction. The mean lateral head displacement (ALHmean) of spermatozoa was significantly greater in follicular fluid (3.9 +/- 0.3 microns; n = 5) than in fluid from the ampulla during peri- and early postovulatory intervals and from the isthmic-utero-tubal junction during pre- and early postovulatory intervals, and was also higher (P < 0.05) than in fluid from the ampullary-isthmic junction during the preovulatory period, but was not different from the peri- and postovulatory ampullary-isthmic junction fluids. The proportion of spermatozoa exhibiting circular motion was significantly higher (P < 0.05) in the periovulatory fluid from the ampullary-isthmic junction (24 +/- 3%) compared with fluids obtained during preovulatory and early postovulatory periods. Follicular fluid had no effect on the proportion of spermatozoa exhibiting circular motion. The average radius of sperm movement in circular trajectories was higher in follicular fluid than in the periovulatory fluids from the ampulla and ampullary-isthmic junction (P < 0.05). In hydrosalpingeal fluids collected 2-5 days after ovulation, the average radius of movement was greater in the ampulla fluid and ampullary-isthmic junction fluid than in fluid from the isthmic-utero-tubal junction (P < 0.05). These results demonstrate that follicular fluid and oviductal fluids have considerable influences on boar sperm motility. Furthermore, the immediate effect of periovulatory ampullary-isthmic junction fluid in increasing the percentage of spermatozoa swimming in circles (hyperactivated) is relevant, since it is at this time and within this region that fertilization occurs.

Patterns of secretion of GnRH, LH and FSH during the postovulatory period in mares: mechanisms prolonging the LH surge.

To study the mechanisms responsible for the unusually slow decline of the ovulatory LH surge in mares, secretion patterns of GnRH, LH and FSH were monitored in pituitary venous blood collected every 2 or 5 min for 10.5-18.0 h from five mares on the third (n = 4) or fifth day after ovulation (first sampling period). To determine the effectiveness of progesterone negative feedback, mares were then given a luteolytic dose of a prostaglandin analogue (PGF2 alpha) and pituitary venous sampling (every 2 or 5 min for 16 h) recommenced 20-22 h later (second sampling period). During the declining arm of the LH surge, large peaks (detected by the Cluster algorithm) of concurrent LH and FSH secretion occurred infrequently, with four peaks being detected in a combined sampling period of 75 h. Outside the peaks, LH or FSH secretion continued (as assessed by a pituitary to jugular-venous concentration ratio > or = 1.25) during 46% +/- 13 or 40% +/- 10, respectively, of the sampling period. GnRH immunoactivity was detected during each spontaneous gonadotrophin peak, but at other times was generally at assay sensitivity. After PGF2 alpha, plasma progesterone fell (ng ml-1, mean +/- SEM; first sampling period: 8.6 +/- 0.8; second; 2.0 +/- 0.3; P = 0.001) and the frequency of LH (P < 0.05) and FSH (P < 0.02) peaks rose, with 28 peaks detected for each hormone in a total of 80 h sampling. Peaks in LH were smaller during the second period, with decreases observed in maximum (P = 0.027) and mean (P = 0.025) secretion rates. Maximum GnRH secretion rate during peaks also declined (P = 0.010); however, the decrement (-30 +/- 6%) was less than that in maximum LH secretion rate (-82 +/- 5%; P = 0.040), suggesting that other factors contribute to the reduced LH peak amplitude. In summary, gonadotrophin peak frequency during the downswing of the surge in mares is slow, as in the midluteal phase, and the slow rate of decline in peripheral gonadotrophin concentrations is due, at least in part, to continued secretion between pulses. Moreover, progesterone negative feedback is highly effective in early dioestrus, in that lessening it without complete removal markedly accelerates gonadotrophin pulse frequency.

Distribution, number and membrane integrity of spermatozoa in the pig oviduct in relation to spontaneous ovulation.

The pattern of distribution, number and membrane integrity of spermatozoa in the utero-tubal junction (UTJ) and isthmus during three oestrous stages were related to spontaneous ovulation in flushed and fixed oviducts of multiparous sows. Three unrelated boars were each used once to mate or artificially inseminate (neat ejaculate) six out of 18 sows, 18 h prior to expected ovulation. The sows were slaughtered 6-8 h before, during or 6-8 h after ovulation. The ad-uterine oviductal region (UTJ and isthmus) was divided into UTJ, lower isthmus, middle isthmus and upper isthmus segments. A higher fraction of middle and upper isthmus segments contained spermatozoa during the peri- and post-ovulatory periods than during the pre-ovulatory period. The distribution, numbers and membrane integrity of spermatozoa in the UTJ-isthmus region were influenced by the ovulation event. Numbers and distribution of spermatozoa varied depending on the boar used. The flushing technique allowed a better assessment of the distribution, number and membrane integrity of tubal spermatozoa than in situ observation with a scanning electron microscope (SEM).

The effect of excess rumen degradable protein during the pre, peri or post-ovulatory period on conception rate and conceptus interferon secretion in naturally cycling ewes

Low conception rates in high producing dairy cows have been linked to the feeding of excessive quantities of rumen degradable protein (Elrod &amp; Butler, 1993). Similarly, a recent study in non-lactating ewes, superovulated during seasonal anoestrous, has shown that high rumen degradable protein intakes throughout the preovulatory and early pregnancy period have an adverse effect on embryo survival (Bishonga et al., 1994). The present study aimed to identify the window of oocyte/embryo development which is sensitive to variations in the degradability of maternal protein intake in naturally cycling ewes expressing their normal ovulation rate. Nutritional treatments were applied at three crucial developmental time points spanning (1) preovulatory oocyte maturation (2) fertilisation and early cell division in the oviduct and (3) entry of the embryo into the uterus, hatching and the transition from a spherical blastocyst to an elongating conceptus.

The effect of excess rumen degradable protein during the pre, peri or post-ovulatory period on conception rate and conceptus interferon secretion in naturally cycling ewes

Low conception rates in high producing dairy cows have been linked to the feeding of excessive quantities of rumen degradable protein (Elrod &amp; Butler, 1993). Similarly, a recent study in non-lactating ewes, superovulated during seasonal anoestrous, has shown that high rumen degradable protein intakes throughout the preovulatory and early pregnancy period have an adverse effect on embryo survival (Bishonga et al., 1994). The present study aimed to identify the window of oocyte/embryo development which is sensitive to variations in the degradability of maternal protein intake in naturally cycling ewes expressing their normal ovulation rate. Nutritional treatments were applied at three crucial developmental time points spanning (1) preovulatory oocyte maturation (2) fertilisation and early cell division in the oviduct and (3) entry of the embryo into the uterus, hatching and the transition from a spherical blastocyst to an elongating conceptus.

Heterogeneous distribution of fibronectin, tenascin-C, and laminin immunoreactive material in the cumulus-corona cells surrounding mature human oocytes from IVF-ET protocols--evidence that they are composed of different subpopulations: an immunohistochemical study using scanning confocal laser and fluorescence microscopy.

Monoclonal antibodies and immunofluorescence microscopy, including laser confocal microscopy, were used in this study to point out the production of fibronectin, tenascin-c, and laminin in the cumulus-corona (CC) cells surrounding mature human oocytes from IVF-ET protocols in view of their presumptive importance in the coordination of the processes leading to fertilization and early embryo cleavage, including the final maturation of the ovum, the sperm-egg interaction, and the "complex biochemical dialogue" between the gamete and the oviduct through the tubal luminal environment. One hundred fifty mature oocyte-CC complexes were obtained from IVF-ET protocols and fixed in 4.0% buffered paraformaldehyde. Specimens were incubated with a panel of primary monoclonal antibodies (mabs) recognizing different epitopes of fibronectin, tenascin-c, and laminin and then with fluorescein isothiocyanate-conjugated goat anti-mouse IgG. Observations were made by a scanning confocal microscope (Sarastro 2000) and a photomicroscope (Polyvar, Reichert-Jung) equipped with epifluorescence optics. The immunohistochemical data demonstrated that human CC cells are capable of producing fibronectin and tenascin-c but that their production is not homogeneous in the CC population. In fact, fibronectin immunoreactivity was shown mostly by inner CC cells (mainly corona cells), whereas tenascin was produced by some cells scattered in the entire cumulus mass. Moreover, fibronectin and tenascin-c immunoreactive material was observed in the intracytoplasmic areas, at the plasma membrane level as well as in the extracellular matrix. On the contrary, laminin immunofluorescent material was found around plasma membranes of almost all CC cells, but a clear intracytoplasmic reaction was never observed. This leads us to assume that laminin in the extracellular matrix remains entrapped once produced by granulosa follicular cells and that in the postovulatory period no active secretion occurs in CC cells. Even though the functional role of these extracellular matrix proteins remains still unclear, it is reasonable to suggest that they are necessary in various steps of the reproductive process, i.e., from the pick-up of the oocyte, its transport through the oviduct, and fertilization, up until the early cleavage of the embryo. Finally, functional differences between "corona radiata" and "cumulus" cells during the oocyte denudation may be accounted for particular distribution of these adhesive proteins.

The effects of post-ovulatory administration of onapristone on the developmnent of a secretory endometrium.

The purpose of this study was to assess the ability of the anti-progestin onapristone administered in the immediate post-ovulatory period to disrupt endometrial differentiation as a potential method of fertility control. In all, 10 healthy female volunteers were given 400 mg onapristone 2 days after the mid-cycle luteinizing hormone (LH) surge in urine (LH+2). An endometrial biopsy was taken 4 or 6 days after the LH surge (i.e. LH+4 or LH+6) in a control cycle and on the corresponding day of the treatment cycle. Biopsies were assessed for histological dating and immunolocalization of oestrogen receptors, progesterone receptors and 15-hydroxyprostaglandin dehydrogenase (PGDH). On day LH+12, blood was taken for the measurement of insulin-like growth factor binding protein-1 (IGFBP-1) and placental protein 14 (PP14). Hormonal measurements in blood and urine were used to monitor the effects on the menstrual cycle. In addition, the concentration of cortisol in plasma was measured to determine if this dose of onapristone exerted significant anti-glucocorticoid activity. Treatment with onapristone retarded the development of secretory changes within the endometrium without affecting the length of the luteal phase. Intense nuclear immunostaining of oestrogen and progesterone receptors was evident in glands and stroma after treatment, suggesting that the progesterone-dependent down-regulation of steroid receptors was inhibited by the anti-progestin. Onapristone also affected the production of luteal phase endometrial proteins, as judged by the pronounced reduction in immunostaining of PGDH within the glands and the significant reduction in plasma concentrations of PP14. However, plasma concentrations of IGFBP-1 did not differ between cycles. Onapristone did not appear to exert significant anti-glucocorticoid activity because concentrations of cortisol were unaffected. These findings suggest that onapristone could potentially be used as a method of postovulatory fertility control.

Insulin-like growth factor-I (IGF-I) and IGF-binding proteins in bovine sera and pituitaries at different stages of the estrous cycle.

The objective of this study was to determine whether concentrations of hypophyseal and serum insulin-like growth factor-I (IGF-I) and binding activities of serum and hypophyseal IGF-binding proteins (IGFBPs) differ with stage of the estrous cycle in mature beef cows. Cows were assigned to the following stages of the estrous cycle based on serum concentrations of progesterone (P4) and ovarian structures at death: days 1-5 (day 0 = estrus; n = 18), days 6-10 (n = 24), days 11-16 (n = 39), and days 17-21 (n = 7). Serum samples collected at death and anterior pituitary homogenates were analyzed for IGF-I, LH, and FSH by RIA. Serum and pituitary IGFBPs were evaluated by ligand and immunoblot analyses. Serum samples contained IGFBP activity at 44 and 40 kilodaltons (kDa; IGFBP-3), 34 kDa (IGFBP-2), 30 kDa (IGFBP-5), 28 kDa, and 24 kDa. The intensity of binding by the different sized proteins in serum remained constant throughout the estrous cycle. IGFBPs detected in anterior pituitaries included a 36-/40-kDa doublet (IGFBP-3), a 32-kDa protein (IGFBP-2), and a 29-kDa IGFBP (IGFBP-5). The intensity of [125I]IGF-I binding to IGFBPs was greater (P < 0.05) during days 11-16 than days 1-5 or days 6-10 of the estrous cycle and was intermediate (P > 0.05) during days 17-21. Serum concentrations of LH were low (< 0.4 ng/ml) during days 1-16 of the estrous cycle, but increased (P < 0.05) approximately 3- to 4-fold during days 17-21. LH concentrations in the anterior pituitary increased (P < 0.05) from the postovulatory period (548 +/- 52 micrograms/g; days 1-5) to the late luteal phase (791 +/- 39 micrograms/g; days 11-16) and were intermediate (P > 0.05) during the preovulatory phase (707 +/- 85 micrograms/g; days 17-21). Concentrations of IGF-I and FSH in the anterior pituitary and serum did not differ (P > 0.05) by stage of the estrous cycle. A positive correlation among the different IGFBPs in the pituitary (P < 0.01) and between each pituitary IGFBP and serum P4 (P < 0.05) existed. The significance (P < 0.0002) of the correlation between pituitary IGFBP-3 and P4 was greater (P < 0.02) than that between the other pituitary IGFBPs and P4. In summary, IGFBP activity in the anterior pituitary, but not that in serum, changed with stage of the estrous cycle in association with serum concentrations of P4. The absence of similar changes in serum or anterior pituitary concentrations of IGF-I supports the hypothesis that IGFBPs may regulate the actions of IGF-I on gonadotropin release from the anterior pituitary gland during the estrous cycle of beef cattle.

Immunocytochemical Localization of Aromatase in The Ovary of Superovulated Cattle, Pigs and Sheep

The localization of aromatase, an enzyme converting androgens to estrogen, in the ovaries of superovulated cattle, pigs and sheep was studied immunocytochemically in the preovulatory and postovulatory period using anti-human placental aromatase cytochrome P-450 antiserum. Immunostaining for aromatase was detected in the granulosa cells of preovulatory follicles of all species studied. Theca interna cells were stained in preovulatory follicles in the pig but not in cattle and sheep. Interstitial gland cells, cumulus cells and oocytes were unstained in all species. In cattle and pig the corpora lutea were unstained whereas they displayed staining in the sheep. Preantral and small antral follicles were unstained during both the preovulatory and postovulatory period in all species. It is concluded that granulosa cells of preovulatory follicles are the main residence for aromatase activity in superovulated cattle, pig and sheep, whereas the activity of theca interna and corpora lutea is species specific.

Parameters of Fibrinolysis in Peritoneal Fluid and Plasma in Different Stages of the Menstrual Cycle

The purpose of this study was to investigate differences in fibrinolytic activity in peritoneal fluid and plasma of women in the first and second part of the menstrual cycle. Given the classic concept of decreased fibrinolytic activity as a cause of adhesion formation, and if such differences are found, the stage of women's menstrual cycle should be taken into consideration when scheduling a laparotomy. We measured fibrinolytic parameters in peritoneal fluid and plasma in eight women in the pre-ovulatory period and in eleven women in the post-ovulatory period of the menstrual cycle. There were no differences in t-PA-Ag, t-PA-Act, u-PA-Ag and scu-PA concentrations in peritoneal fluid between the pre- and post-ovulatory group. Nevertheless, PAI-1-Ag in peritoneal fluid was three-fold higher in the post-ovulatory phase (p < 0.02). In peritoneal fluid the concentrations of both TDP and FbDP were three-fold higher at the same phase (p < or = 0.05). Plasma u-PA-Ag and scu-PA concentrations were significantly lower (30%, p < 0.05) in the post-ovulatory phase and also lower than plasma u-PA-Ag and scu-PA (measured with the same assay) in a group of 50 healthy individuals. No differences in t-PA and PAI concentration were found. In conclusion, the intraperitoneal fibrinolytic capacity might be impaired in the second part of the menstrual cycle, regarding the elevated levels of PAI-1-Ag, leading to an increased risk for post-ovulatory adhesion formation. The low plasma u-PA-Ag and scu-PA levels post-ovulatory may have clinical relevance.