Melanoma risk after in vitro fertilization: A review of the literature.

BACKGROUND: The role of female sex hormones in the pathogenesis of malignant melanoma (MM) remains controversial. Although melanocytes appear to be hormonally responsive, the effect of estrogen on MM cells is less clear. Available clinical data does not consistently demonstrate that increased endogenous hormones from pregnancy or increased exogenous hormones from oral contraceptive pills and hormone replacement affect MM prevalence and outcome. OBJECTIVE: We sought to examine potential associations between in vitro fertilization (IVF) and melanoma. METHODS: A literature review was conducted. Primary outcomes were reported as associations between IVF and melanoma risk compared with the general population. Secondary outcomes included associations stratified by type of IVF regimen and subgroup, such as parous versus nulliparous patients. RESULTS: Eleven studies met our inclusion criteria. Five studies found no increased risk for MM among IVF users compared with the general population. Two studies found an increase in MM in clomiphene users, and 4 studies found an increase in MM among patients who were gravid or parous either before or after IVF. CONCLUSION: The reviewed studies do not reveal consistent patterns of association between IVF and MM among all infertile women. However, the data indicates a potential increased risk for MM in ever-parous patients treated with IVF. High-quality studies including a large number of MM cases that control for well-established MM risk factors are needed to adequately assess the relationship between IVF and MM, particularly among ever-parous women.

GnRH Neuron-Specific Ablation of Gαq/11 Results in Only Partial Inactivation of the Neuroendocrine-Reproductive Axis in Both Male and Female Mice: In Vivo Evidence for Kiss1r-Coupled Gαq/11-Independent GnRH Secretion.

The gonadotropin-releasing hormone (GnRH) is the master regulator of fertility and kisspeptin (KP) is a potent trigger of GnRH secretion from GnRH neurons. KP signals via KISS1R, a Gαq/11-coupled receptor, and mice bearing a global deletion of Kiss1r (Kiss1r(-/-)) or a GnRH neuron-specific deletion of Kiss1r (Kiss1r(d/d)) display hypogonadotropic hypogonadism and infertility. KISS1R also signals via ß-arrestin, and in mice lacking ß-arrestin-1 or -2, KP-triggered GnRH secretion is significantly diminished. Based on these findings, we hypothesized that ablation of Gαq/11 in GnRH neurons would diminish but not completely block KP-triggered GnRH secretion and that Gαq/11-independent GnRH secretion would be sufficient to maintain fertility. To test this, Gnaq (encodes Gαq) was selectively inactivated in the GnRH neurons of global Gna11 (encodes Gα11)-null mice by crossing Gnrh-Cre and Gnaq(fl/fl);Gna11(-/-) mice. Experimental Gnaq(fl/fl);Gna11(-/-);Gnrh-Cre (Gnaq(d/d)) and control Gnaq(fl/fl);Gna11(-/-) (Gnaq(fl/fl)) littermate mice were generated and subjected to reproductive profiling. This process revealed that testicular development and spermatogenesis, preputial separation, and anogenital distance in males and day of vaginal opening and of first estrus in females were significantly less affected in Gnaq(d/d) mice than in previously characterized Kiss1r(-/-) or Kiss1r(d/d) mice. Additionally, Gnaq(d/d) males were subfertile, and although Gnaq(d/d) females did not ovulate spontaneously, they responded efficiently to a single dose of gonadotropins. Finally, KP stimulation triggered a significant increase in gonadotropins and testosterone levels in Gnaq(d/d) mice. We therefore conclude that the milder reproductive phenotypes and maintained responsiveness to KP and gonadotropins reflect Gαq/11-independent GnRH secretion and activation of the neuroendocrine-reproductive axis in Gnaq(d/d) mice. SIGNIFICANCE STATEMENT: The gonadotropin-releasing hormone (GnRH) is the master regulator of fertility. Over the last decade, several studies have established that the KISS1 receptor, KISS1R, is a potent trigger of GnRH secretion and inactivation of KISS1R on the GnRH neuron results in infertility. While KISS1R is best understood as a Gαq/11-coupled receptor, we previously demonstrated that it could couple to and signal via non-Gαq/11-coupled pathways. The present study confirms these findings and, more importantly, while it establishes Gαq/11-coupled signaling as a major conduit of GnRH secretion, it also uncovers a significant role for non-Gαq/11-coupled signaling in potentiating reproductive development and function. This study further suggests that by augmenting signaling via these pathways, GnRH secretion can be enhanced to treat some forms of infertility.

Luteal phase support with progesterone in intrauterine insemination: a prospective randomized study.

OBJECTIVE: To determine the effect of vaginal progesterone as luteal support on pregnancy outcomes in infertile patients who undergo ovulation induction with gonadotropins and intrauterine insemination (IUI). DESIGN: Prospective randomized trial. SETTING: Tertiary referral center. PATIENT(S): About 398 patients with primary infertility were treated during 893 ovarian stimulation and IUI cycles from February 2010 to September 2012. METHODS: All patients underwent ovarian stimulation with gonadotropins combined with IUI. Patients in the supported group received vaginal micronized progesterone capsules 200 mg once daily from the day after insemination until next menstruation or continuing for up to 8 weeks of pregnancy. Women allocated in the control group did not receive luteal phase support. MAIN OUTCOME MEASURE(S): Livebirth rate, clinical pregnancy rate and early miscarriage rate per cycle. RESULT(S): Of the 893 cycles, a total of 111 clinical pregnancies occurred. There were no significant differences between supported with progesterone and unsupported cycle in terms of livebirth rate (10.2% versus 8.3%, respectively, with a p value = 0.874) and clinical pregnancy rate (13.8% compared with 11.0% in unsupported cycle with a p value = 0.248). An early miscarriage rate of 3.6% was observed in the supported cycles and 2.7% in the unsupported cycles, with no significant differences between the groups (p value = 0.874). CONCLUSION(S): In infertile patients treated with mildly ovarian stimulation with recombinant gonadotropins and IUI, luteal phase support with vaginal progesterone is not associated with higher livebirth rate or clinical pregnancy rate compared with patients who did not receive any luteal phase support.

Pre- and postnatal developmental exposure to the polychlorinated biphenyl mixture aroclor 1221 alters female rat pituitary gonadotropins and estrogen receptor alpha levels.

Polychlorinated-biphenyls (PCBs) are industrial compounds, which were widely used in manufacturing of electrical parts and transformers. Despite being banned in 1979 due to human health concerns, they persist in the environment. In humans and experimental model systems, PCBs elicit toxicity in part by acting as endocrine-disrupting chemicals (EDCs). Aroclor 1221 (A1221) is a weakly estrogenic PCB mixture known to alter reproductive function in rodents. EDCs can impact hormone signaling at any level of the hypothalamic-pituitary-gonadal (HPG) axis, and we investigated the effects of A1221 exposure during the prenatal and postnatal developmental periods on pituitary hormone and steroid receptor expression in female rats. Examining offspring at 3 ages, postnatal day 8 (P8), P32 and P60, we found that prenatal exposure to A1221 increased P8 neonate pituitary luteinizing hormone beta (Lhb) mRNA and LHß gonadotrope cell number while decreasing LH serum hormone concentration. No changes in pituitary hormone or hormone receptor gene expression were observed peri-puberty at P32. In reproductively mature rats at P60, we found pituitary follicle stimulating hormone beta (Fshb) mRNA levels increased by prenatal A1221 exposure with no corresponding alterations in FSH hormone or FSHß expressing cell number. Estrogen receptor alpha (ERα) mRNA and protein levels were also increased at P60, but only following postnatal A1221 dosing. Together, these data illustrate that exposure to the PCB A1221, during critical developmental windows, alters pituitary gonadotropin hormone subunits and ERα levels in offspring at different phases of maturation, potentially impacting reproductive function in concert with other components of the HPG axis.

[Stimulation in vitro of oocytes of acipenserids with progesterone and homologous gonadotropic hormone of hypophysis].

We showed that the percentage of oocytes of acipenserids ovulating in vitro in Ringer solution modified for sturgeons (RMS) considerably depends on the concentration of sodium bicarbonate and the concentration of progesterone. Under optimal conditions (0.5 g/L of sodium bicarbonate and 30 ng/mL of progesterone), it can be higher than 80. Oocytes that matured and ovulated under such conditions are capable of normal development. In the best case, approximately 70% of developing embryos (of the number of ovulated oocytes) reach the stage of hatching (dead-line of observation). This method of producing offspring based on the insemination of oocytes that have matured and ovulated in vitro can be used in work with single females of rare and disappearing species of acipenserids.

Seasonal Related Multifactorial Control of Pituitary Gonadotropin and Growth Hormone in Female Goldfish: Influences of Neuropeptides and Thyroid Hormone.

Female reproduction is under multifactorial control of brain-pituitary-peripheral origin. The present study provides information on seasonal changes in circulating LH and GH concentrations, as well as transcript levels for a number of genes involved in the regulation of reproduction and growth in female goldfish. We also provide information on the effects of treatments with GnRH and/or GnIH, and their interaction with T3, at three stages of gonadal recrudescence. Maximum basal concentration of LH was observed at late recrudescence (Spring) while no seasonal changes in basal serum GH levels was detected. Serum LH and GH levels were stimulated by GnRH as expected, depending on the season. GnIH stimulated basal GH concentrations in gonadally regressed fish. GnIH inhibitory action on GnRH-induced LH response was observed in late, but not in mid recrudescence. T3 actions on basal and GnRH- or GnIH-induced GH secretion were generally inhibitory, depending on season. Administration of T3 attenuated GnRH-induced LH responses in mid and late stages of gonadal recrudescence, and the presence of GnIH abolished inhibitory actions of T3 in fish at mid recrudescence. Our results also demonstrated seasonal patterns in basal and GnRH- and/or GnIH-induced transcript levels for ERα, ERßI, FSHR, aromatase, TRαI, TRß, IGF-I, and Vtg in the liver and ovary. However, there were no clear correlations between changes in transcript levels and circulating levels of LH and GH. The results support the hypothesis that GnRH, GnIH, and T3 are contributing factors in complex reciprocal control of reproduction and growth in goldfish.

Ovarian steroid cells. I. Differentiation of the lutein cell from the granulosa follicle cell during the preovulatory stage and under the influence of exogenous gonadotrophins.

The granulosa follicle cell of the Graafian follicle of the rabbit ovary differentiates into a lutein cell involved in steroid synthesis. Cytological events which occur within the granulosa cell of the normally stimulated follicle prior to ovulation have been duplicated by the intrafollicular injection of exogenous gonadotrophin. The luteinization of the granulosa cells involves the accumulation of 250- to 300-A, electron-opaque, spherical granules, dispersed within the cytoplasmic matrix, which have been identified as glycogen with the PAS-staining procedure. Further development of the granulosa cell following ovulation involves an increase in cell size, a decrease in the number of RNP particles, and an accumulation of an abundant system of intracellular membranes (agranular endoplasmic reticulum). Glycogen granules first appear in the granulosa cells as the separate, monoparticulate form. After follicle rupture and the formation of agranular endoplasmic reticulum, glycogen particles are present in a rosette arrangement within membrane-bounded vacuoles. The rosette arrangement of glycogen particles is also found dispersed within the cytoplasmic matrix of the lutein cell during the later stages of the cell life-span. Injection of luteinizing hormone or human chorionic gonadotrophin into a mature follicle also produces a marked accumulation of monoparticulate glycogen in the majority of granulosa cells, within 30 min. Cytoplasmic extensions which contain the glycogen masses are noticeably free of RNP particles.

Gonadotropin-induced anomalies of the zona pellucida of the baboon ovum.

Vesiclulation and other anomalies were observed in the zona pellucida about the ovum of female baboons (Papio anubis) that had received treatment with Pergonal and human chorionic gonadotropin; and in somiie instances the zona pellucida was absent.

Application of RNAi technology to the inhibition of zebrafish GtHalpha, FSHbeta, and LHbeta expression and to functional analyses.

Zebrafish (Danio rerio) were used as a model fish, and the technique of RNA interference (RNAi) was employed to knockdown three subunits of the gonadotropin alpha (GtHalpha, common alpha), follicle-stimulating hormone beta (FSHbeta), and luteinizing hormone beta (LHbeta) genes. Three short-hairpin RNA (shRNA) expression vectors and three mismatched shRNA expression vectors as controls for each subunit gene were constructed, and the depression efficiency was tested in vivo by microinjection; the RNA or protein expression levels of the GtH genes were monitored by RT-PCR, Southern blotting, and green fluorescent protein (GFP) analyses. Expression of GtH mRNA was obviously and more efficiently depressed by GtHalpha RNAi expression compared with the other two subunits. A GtHalpha morpholino analysis showed that the GtHalpha morpholino led to suppression of embryonic development and the production of embryonic mutants as a result of an injection of GtHalpha -shRNA. Taken together, these results show that GtHalpha-shRNA, which more efficiently targets RNAi, may have an essential role in the further development of sterility technology of transgenic fish for biosafety purposes.

[Endocrinology of in vitro maturation of oocytes].

In-vitro maturation (IVM) of oocytes is an alternative to conventional in-vitro fertilization (IVF) treatment. Significant progress has been made to improve pregnancy and implantation rates from in-vitro matured oocytes. However oocyte maturation in vitro is profoundly affected by culture conditions. Most IVM protocols supplement gonadotropins--folicle stimulating hormone (FSH) and luteinizing hormone (LH) into a culture medium for oocyte maturation. The addition of these hormones, as well as some growth factors is based on their physiological role in oocyte maturation in vivo. However, it is possible, they not to play the same role in promoting oocyte maturation in vitro. The aim of this survey is to examine the hormones and growth factors, which participate in oocyte maturation in vivo and to summarize briefly the available information about the effect they have on the in vitro maturation. The researches on effects of these endocrine factors on oocyte maturation and subsequent fertilization, as well as on early embryonal development proceed.

Novel evidence that pituitary sex hormones regulate migration, adhesion, and proliferation of embryonic stem cells and teratocarcinoma cells.

The pituitary sex hormones (SexHs): follicle­stimulating hormone (FSH), luteinizing hormone (LH), and prolactin (PRL) regulate several functions crucial for reproduction, including oogenesis, spermatogenesis, and lactation. An important source of prolactin-like hormones, known as lactogens, is the placenta, and lactogens bind to the PRL receptor (PRLR) with high affinity and thereby mimic the actions of PRL. Recently, it has been demonstrated that pituitary SexHs were involved in metastatic lung cancer, certain sarcomas, and leukemia. In the present study we aimed to investigate whether FSH, LH, and PRL were able to stimulate stem cells involved in early development. To address this issue we employed a murine embryonic stem cell line (ES-D3) as well as two teratocarcinoma cell lines, P19 (murine) and NTera2 (human). We determined that all these cells expressed SexH receptors at the mRNA and protein levels and that stimulation of these receptors induced phosphorylation of p42/44 MAPK, p38 MAPK, and AKT. Moreover, ES-D3, P19, and NTera2 cells responded with increased migration and adhesion to physiological concentrations of pituitary SexHs. In view of these findings we proposed that maternal-derived pituitary SexHs regulate the biology of stem cells involved in early development.

The effect of withholding gonadotropin (a coasting period) on the outcome of in vitro fertilization cycles.

OBJECTIVE: The purpose of this study was to evaluate the effects of coasting (withdrawing gonadotropin administration) on the cycle outcomes, including total number and quality of oocytes and embryos and pregnancy rate in patients undergoing in vitro fertilization. STUDY DESIGN: Sixty-seven patients undergoing coasting were evaluated in a retrospective study at the Royan Institute between May 2002 and June 2003. All patients underwent standard long protocol with gonadotropin-releasing hormone (GnRH) analogue and human menopausal gonadotropin (HMG) and then in vitro fertilization and embryo transfer (IVF-ET). Coasting was considered when the estradiol level was more than 3000 pg/ml and the number of follicles >10 in each ovary. HCG was administered whenever estradiol reached < or =3000 pg/ml and then the (IVF-ET) program was carried out. According to coasting periods, patients were divided into two groups: coasting period < or =3 days and coasting period >3 days. Statistical comparisons were performed using Student's t test and Fisher's exact test. RESULTS: There were no significant differences between the two groups with regard to mean age, body mass index (BMI), number of polycystic ovary syndrome (PCOS) patients, number of HMG ampoules and stimulation duration. The total number of retrieved oocytes decreased significantly in patients with more than 3 days of coasting (P=0.04). The number of high quality oocytes also decreased in this group; however, this did not reach significant levels. There were no significant differences between the two groups with regard to fertilization and pregnancy rate. None of the patients developed severe ovarian hyperstimulation syndrome (OHSS). CONCLUSION: This study reveals that a long coasting period (>3 days) has a negative effect on the number of oocytes, although the fertilization and pregnancy rates are not affected. Prospective randomized studies with larger sample sizes are needed to compare coasting with other procedures.

Meat and Livestock Association Plenary Lecture 2005. Oocyte signalling molecules and their effects on reproduction in ruminants.

Sheep (Ovis aries) are a highly diverse species, with more than 900 different breeds that vary significantly in their physiological characteristics, including ovulation rate and fecundity. From examination of inherited patterns of ovulation rate, several breeds have been identified with point mutations in two growth factor genes that are expressed in oocytes. Currently, five different point mutations have been identified in the BMP15 (GDF9b) gene and one in GDF9. Animals heterozygous for the GDF9 and/or the BMP15 mutations have higher ovulation rates than their wild-type counterparts. In contrast, those homozygous for any of the aforementioned BMP15 or GDF9 mutations are sterile owing to arrested follicular development. In bovine and ovine ovaries, GDF9 was expressed exclusively in oocytes throughout follicular growth from the primordial stage of development, whereas in sheep BMP15 was expressed exclusively in oocytes from the primary stage: no data for the ontogeny of BMP15 expression are currently available for cattle. In vitro, ovine growth differentiation factor 9 (oGDF9) has no effect on (3)H-thymidine incorporation by either bovine or ovine granulosa cells, whereas ovine bone morphogenetic protein 15 (oBMP15) has modest (1.2- to 1.6-fold; P < 0.05) stimulatory effects. Ovine GDF9 or oBMP15 alone inhibited progesterone production by bovine granulosa cells, whereas in ovine cells only oGDF9 was inhibitory. The effects of oGDF9 and oBMP15 together were often cooperative and not always the same as those observed for each factor alone. Active immunisation of ewes with BMP15 and/or GDF9 peptides affected ovarian follicular development and ovulation rate. Depending on the GDF9 and/or BMP15 vaccine formulation, ovulation rate was either increased or suppressed. A primary and single booster immunisation of ewes with a BMP15 peptide in a water-based adjuvant has led to 19-40% increases in lambs born per ewe lambing. Collectively, the evidence suggests that oocyte signalling molecules have profound effects on reproduction in mammals, including rodents, humans and ruminants. Moreover, in vivo manipulation of these oocyte signalling molecules provides new opportunities for the management of the fertility of ruminants.

Effect of exogenous gonadotrophins on ovarian morphology and oocyte maturation in the southern hairy nosed wombat Lasiohinus latifrons during the breeding season.

The effect of the exogenous administration of porcine follicle-stimulating hormone (pFSH) and pregnant mare serum gonadotrophin (PMSG) on ovarian follicular development and oocyte maturation in the southern hairy nosed wombat Lasiorhinus latifrons was investigated. Three experimental groups were administered pFSH at various doses and for different treatment lengths, followed by 25 mg porcine luteinising hormone (pLH) 12 h after the last dose of pFSH. Another group was given PMSG followed 72 h later by 25 mg pLH. Animals were killed 24 h after pLH. The left ovary was fixed for histology and the morphology of the antral follicles was determined, whereas follicular oocytes in the right ovary were aspirated, fixed, stained with 42,62-diamidino-2-phenylindole, and viewed for nuclear maturation. There was no significant difference in the mean number of ovarian follicles >1 mm, or in the size class of follicles assessed between control and experimental groups. However, a trend was observed suggesting a possible increase in follicles >3.0 mm in experimental groups compared with control animals. In all females administered exogenous porcine gonadotrophins, but not controls, some of the mural granulosa cells of large tertiary antral follicles had markedly enlarged nuclei (approximately 14 microm in diameter). All oocytes from the control group remained at the germinal vesicle stage, whereas approximately 40% of oocytes retrieved from the pFSH groups and 82.4% retrieved from the PMSG-primed animals had undergone germinal vesicle break down, with a small number reaching meiosis II. The present study shows that exogenous administration of either pFSH or PMSG to hairy nosed wombats can induce follicular growth and oocyte maturation. Such findings could be useful in the development of reproductive technology in this species.

Aromatase cytochrome P450 in rat ovarian granulosa cells before and after luteinization: adenosine 3',5'-monophosphate-dependent and independent regulation. Cloning and sequencing of rat aromatase cDNA and 5' genomic DNA.

The complete nucleotide sequence for rat ovarian aromatase cytochrome P450 (P450arom) has been derived from four cDNA clones isolated from three rat granulosa/luteal cell lambda gt11 cDNA expression libraries. The composite P450arom cDNA extends 1597 basepairs, encodes a protein of 508 amino acids (calculated mol wt = 58,263), and hybridizes to three mRNA transcripts (3.3, 2.6, and 1.9 kilobases in size) in rat ovarian tissues. A 5' genomic fragment was isolated from a rat genomic library and shown to contain exon I and 538 basepairs of 5' flanking sequences, including putative promoter elements. Further, we document that P450arom mRNA and estradiol (E) biosynthesis are regulated by cAMP-dependent mechanisms in granulosa cells of preovulatory (PO) follicles, but are maintained by cAMP-independent mechanisms after LH/hCG-induced luteinization. The transition of the PO granulosa cell to the luteal cell (PO + hCG) phenotype requires 5 h of exposure to hCG in vivo. Once the luteal cell phenotype is programed, P450arom mRNA and E biosynthesis are maintained in the luteinized cells for up to 10 days in a constitutive manner in the absence of hormones or agents that increase intracellular cAMP. Furthermore, when PO + hCG (7 h) follicles were isolated and incubated for 1-3 h with reversible inhibitors of transcription (actinomycin-D) or translation (cycloheximide) before harvesting the granulosa cells, neither morphological nor functional luteinization of granulosa cells in culture was impaired. Thus, rapid cellular and molecular events occur in granulosa cells within 5-7 h after an ovulatory LH/hCG surge that alter the hormonal regulation of the aromatase gene.

Developmental competence of immature pig oocytes under the influence of EGF, IGF-I, follicular fluid and gonadotropins during IVM-IVF processes.

Epidermal growth factor (EGF) and insulin like growth factor-I (IGF-I) were evaluated for their effects on in vitro maturation and fertilization in presence or absence of gonadotropin and porcine follicular fluid. Four groups were made with the addition of growth factors: none (control), EGF, IGF-I or EGF+IGF-I. Each group underwent four predefined treatments with gonadotropin (FSH and LH), follicular fluid, a combination of both, or none (as control). Porcine cumulus-oocyte complexes (COCs) were matured in media containing the above-mentioned treatments for 42-44 h prior to fertilization with fresh sperm capacitated for 2.5 h. At the end of the fertilization period, the presumable embryos were fixed, stained and examined as whole-mounts to ascertain their nuclear status. The addition of EGF alone or in combination with IGF-I, significantly increased the proportion of monospermic oocytes forming 2 normal pronuclei. Also, supplementation with both growth factors together enhanced the percentages of pronucleus formation and total penetration. In addition, treatments with EGF+IGF-I significantly decreased (P<0.01) the incidence of degeneration in fertilized oocytes. However, no significant differences in the proportions of COCs undergoing polyspermy were observed among all treatments. These results suggest a stimulatory effect of tested growth factors in maturation and fertilization of pig oocytes. Furthermore, gonadotropins and follicular fluid can be replaced by the addition of EGF and IGF-I to the maturation media with positive effects on fertilization rate.

Ovarian cyclic adenosine monophosphate-dependent protein kinase activity: ontogeny and effect of gonadotropins.

The ontogeny of ovarian cyclic AMP-binding and protein kinase activities during the postnatal development of the rat, as well as the effect of LH and FSH administration on ovarian cyclic AMP-binding and protein kinase activities in 5-day-old and in hypophysectomized rats was examined. Ovaries of 4 to 8-day-old rats possessed little or no measureable cyclic AMP-binding and protein kinase activities. Subsequent postnatal development occurred in three distinct phases. During the first phase, ovarian cyclic AMP-binding and protein kinase activities increased progressively from age 8 days to age 23 days, when adult levels were observed. Protein kinase activity declined markedly during the second postnatal developmental phase from days 24 to 26, lost its cyclic AMP-dependency, and became refractory to stimulation by cyclic AMP. Studies employing a heat-stable protein kinase inhibitor protein isolated from rabbit skeletal muscle suggest that ovarian protein kinase activity during the refractory period was largely of the cyclic AMP-independent variety. During the third postnatal phase, comprising days 30 to 40, ovarian cyclic AMP-binding and protein kinase activities increased to levels seen in sexually mature rats. Protein kinase cyclic AMP-dependency which was lost during the refractory second postnatal period was fully restored during the third phase. Administration of FSH or LH led to a marked increase of ovarian cyclic AMP-binding and protein kinase activities in 5-day-old rats. Hypophysectomy of 20-day-old rats caused a significant reduction of the cyclic AMP-binding and protein kinase activities in a 27,000 X g supernatant fraction, as well as in the mitochondrial, microsomal, and 105,000 X g supernatant fraction. The decreased cyclic AMP-binding and protein kinase activities of these fractions could be partially restored by FSH or LH treatment of the hypophysectomized rats. The results indicate that ovarian cyclic AMP-binding and protein kinase activities, as well as the ability of ovarian protein kinase to respond to cyclic AMP are gradually acquired after the first postnatal week. The postnatal development of ovarian protein kinase and cyclic AMP-binding activities presumably involves the participation of FSH and LH, although the precise mechanism of LH and FSH action remains to be established.

Dynamic pituitary and ovarian changes occurring during the anestrus to estrus transition in the golden hamster.

Anestrous hamsters exhibit daily afternoon gonadotropin and progesterone surges, but little estrogen secretion. In the first experiment, short day anestrous females were transferred to long days to detect hormonal changes associated with recovery of cyclicity. Morning and afternoon blood samples were taken at increasing durations in long days. Females autopsied at their first vaginal estrus after transfer to long days differed from long day estrous controls only in their lower uterine weights. Some females at all durations exhibited signs of recovery, though they had not yet shown estrus. They did not display afternoon gonadotropin surges and had low circulating progesterone but high estradiol levels, stimulated uteri, and enhanced follicular development. Results of the second and third experiments provided evidence that the daily gonadotropin surges are not the cause per se of anestrus, and that changes in estrogen secretion are essential for the transition to and from anestrus. Phenobarbital blockade of the daily surges in anestrous females did not result in increased follicular growth and estrogen secretion. Furthermore, daily afternoon injections of gonadotropins it appears that the daily surges in anestrous hamsters simply reflect low estradiol levels. It is still not known what signal promotes rapid follicular maturation during the recovery from anestrus. Once initiated, however, this recovery appears to occur within a few days, with a rapid cascade of events. First, follicular development and estrogen secretion resume, and the daily LH surges cease. Then, progesterone levels decline, and an ovulatory surge of gonadotropins is triggered.

Insulin-like growth factor binding protein -2 and -4 messenger ribonucleic acid expression in bovine ovarian follicles: effect of gonadotropins and developmental status.

This work is concerned with the role of insulin-like growth factor binding protein (IGFBP)-2 and -4 in the regulation of IGF bioactivity in bovine follicles during the development of dominance. We measured the expression of IGFBP-2 and -4 messenger RNA (mRNA) in small (1-4 mm) gonadotropin-sensitive follicles and medium (4-8 mm) and large (>8 mm) gonadotropin-dependent follicles using in situ hybridization. In healthy nonatretic bovine follicles, IGFBP-2 and -4 mRNA expression was confined to granulosa and theca tissue, respectively. Moreover, during the development of follicular atresia, there were distinct changes in the temporal and spatial expression of these genes. IGFBP-2 immunoactivity was localized in granulosa tissue and the basement membrane of healthy preantral follicles, whereas IGFBP-4 immunoactivity was localized in both theca and granulosa tissue. Of particular interest was the lack of IGFBP-2 mRNA expression in large (>8 mm) gonadotropin-dependent follicles, an observation that was confirmed by the lack of immunoreactive IGFBP-2 in these follicles. The regulation of IGFBP-2 and -4 mRNA expression in granulosa and theca cells was analyzed using a serum-free cell culture system. FSH inhibited the expression of IGFBP-2 mRNA in granulosa cells, whereas LH stimulated IGFBP-4 mRNA expression in theca cells. Our results provide evidence for the existence of different roles for IGFBP-2 and -4 in the developing follicle.

Serum follicle-stimulating hormone, luteinizing hormone, and prolactin during the induction of ovulation with exogenous gonadotropin.

To examine the gonadotropic milieu presiding over recruitment and selection of a dominant follicle during gonadotropin induction of ovulation, four patients were studied over nine cycles of human pituitary gonadotropin (hPG) therapy. These hypogonadotropic subjects received a routine schedule of hPG injections monitored by daily urinary estrogen and pregnanediol determinations. Serum FSH, LH, and PRL profiles were measured in daily morning blood samples throughout each menstrual cycle. hPG therapy produced markedly abnormal gonadotropin patterns. Mean serum FSH levels were above the upper limit of the normal serum FSH range and no early or midfollicular FSH peaks occurred. The FSH-LH ratio was abnormally high for 8 days before ovulation. Progressive and marked elevations of serum PRL developed during hPG treatment. A bimodal luteal phase serum PRL profile appeared with peak values of 40.7 +/- 5.6 ng/ml (mean +/- SE) 1 day and 42.0 +/- 3.0 ng/ml 9 days after the LH peak. We conclude that: 1) Current gonadotropin treatment regimens to induce ovulation produce radioimmunoassayable serum FSH, LH, and PRL profiles which are qualitatively and quantitatively abnormal, and 2) Excessive FSH levels and the elevated FSH-LH ratio orchestrate aberrant folliculogenesis and result in the clinical problems of multiple ovulation and hyperstimulation.