Endothelial cell-derived fibroblast growth factor-18 regulates ovarian function in sheep.

Increasing the efficiency of farm animal reproduction is necessary to reduce the environmental impact of food production systems. One approach is to increase the number of healthy eggs (oocytes) produced per female for fertilization, thus it is important to understand factors that decrease oocyte health. One paracrine factor that decreases ovarian follicle growth is fibroblast growth factor 18 (FGF18) secreted by cells in the theca layer of the ovarian follicle, however the factors that regulate FGF18 secretion are unknown. In this study we hypothesized that FGF18 secretion is controled by intrafollicular factors and is linked to fertility, which we tested by using cell culture and sheep genetic models in vivo. Separation of theca cell populations revealed that FGF18 messenger RNA (mRNA) is located mainly in thecal endothelial rather than endocrine cells, and immunohistochemistry localized FGF18 protein to microvessels in the theca layer in situ. Culture of ovine theca-derived endothelial cells was used to demonstrate stimulation of FGF18 mRNA and protein abundance by bone morphogenetic protein 4 (BMP4), a growth factor derived from theca endocrine cells. Taking advantage of a sheep genetic model, we demonstrate reduced ovarian and peripheral FGF18 concentrations in the hyperprolific Booroola ewe harboring the FecBB mutation in BMPR1B. These data suggest a novel control of fertility by follicular endothelial cells, in which theca endocrine cells secrete BMP4 that stimulates the secretion of FGF18 from thecal endothelial cells, which in turn diffuses into the granulosa cell layer and promotes apoptosis.

Prenatal programming by testosterone of follicular theca cell functions in ovary.

In mammalian ovaries, the theca layers of growing follicles are critical for maintaining their structural integrity and supporting androgen synthesis. Through combining the postnatal monitoring of ovaries by abdominal magnetic resonance imaging, endocrine profiling, hormonal analysis of the follicular fluid of growing follicles, and transcriptomic analysis of follicular theca cells, we provide evidence that the exposure of ovine fetuses to testosterone excess activates postnatal follicular growth and strongly affects the functions of follicular theca in adulthood. Prenatal exposure to testosterone impaired androgen synthesis in the small antral follicles of adults and affected the expression in their theca cells of a wide array of genes encoding extracellular matrix components, their membrane receptors, and signaling pathways. Most expression changes were uncorrelated with the concentrations of gonadotropins, steroids, and anti-Müllerian hormone in the recent hormonal environment of theca cells, suggesting that these changes rather result from the long-term developmental effects of testosterone on theca cell precursors in fetal ovaries. Disruptions of the extracellular matrix structure and signaling in the follicular theca and ovarian cortex can explain the acceleration of follicle growth through altering the stiffness of ovarian tissue. We propose that these mechanisms participate in the etiology of the polycystic ovarian syndrome, a major reproductive pathology in woman.

Anti-Müllerian hormone production in the ovary: a comparative study in bovine and porcine granulosa cells†.

In this study, we aimed to determine the origin of the difference, in terms of anti-Müllerian hormone production, existing between the bovine and porcine ovaries. We first confirmed by quantitative real-time-Polymerase-Chain Reaction, ELISA assay and immunohistochemistry that anti-Müllerian hormone mRNA and protein production are very low in porcine ovarian growing follicles compared to bovine ones. We then have transfected porcine and bovine granulosa cells with vectors containing the luciferase gene driven by the porcine or the bovine anti-Müllerian hormone promoter. These transfection experiments showed that the porcine anti-Müllerian hormone promoter is less active and less responsive to bone morphogenetic protein stimulations than the bovine promoter in both porcine and bovine cells. Moreover, bovine but not porcine granulosa cells were responsive to bone morphogenetic protein stimulation after transfection of a plasmidic construction including a strong response element to the bone morphogenetic proteins (12 repetitions of the GCCG sequence) upstream of the luciferase reporter gene. We also showed that SMAD6, an inhibitor of the SMAD1-5-8 pathway, is strongly expressed in porcine compared to the bovine granulosa cells. Overall, these results suggest that the low expression of anti-Müllerian hormone in porcine growing follicles is due to both a lack of activity/sensitivity of the porcine anti-Müllerian hormone promoter, and to the lack of responsiveness of porcine granulosa cells to bone morphogenetic protein signaling, potentially due to an overexpression of SMAD6 compared to bovine granulosa cells. We propose that the low levels of anti-Müllerian hormone in the pig would explain the poly-ovulatory phenotype in this species.

Efficacy of a single blood anti-Müllerian hormone (AMH) concentration measurement for the selection of Japanese Black heifer embryo donors in herd breeding programs.

We evaluated the relationship between plasma anti-Müllerian hormone (AMH) concentrations in Japanese Black (JB) heifers at 7-10 months of age and the number of embryos recovered after superovulation treatment in selected ovum pick-up donors, concomitantly with changes in their AMH concentrations before and after parturition. Plasma AMH concentrations in heifers were positively correlated with the total number of follicles (r = 0.647, P < 0.01) and embryos (r = 0.681, P < 0.01) recovered from the animals postpartum, when selected as donor cows, but did not correlate with the total number of transferable embryos. No difference was observed between the plasma AMH concentration at the heifer period and the postpartum period. Additionally, serum AMH concentrations of heifers weakly correlated with the number of follicles and embryos recovered by virgin flush after superovulation treatment at 13-15 months of age. Therefore, a single blood AMH concentration measurement may accelerate intensive JB cattle breeding.

Efficacy of a single measurement of plasma anti-Müllerian hormone concentration for ovum pick-up donor selection of Japanese Black heifers in herd breeding programs.

In this study, we evaluated the efficiency of a single measurement of plasma anti-Müllerian hormone (AMH) concentration in heifers in determining the number of oocytes recoverable by ovum pick-up (OPU), and compared AMH concentrations among sister heifers from the same parents. For this, blood samples from 50 embryo-transfer-derived female Japanese Black (JB) heifers (mean: 8.7 age in months) were collected and plasma AMH concentration was measured. At 13-15 months of age, both the number of follicles (2-9 mm) and the number of collected oocytes after OPU were counted and compared. Results indicated that the heifers with the highest AMH concentration had the highest number of follicles in their ovaries and gave the highest number of collected oocytes with OPU, thereby indicating that a single measurement of plasma AMH concentration is informative for the selection of OPU-donor heifers in herd breeding programs. The practice of performing a single AMH measurement may accelerate the intensive breeding of JB herds.

Differentially expressed genes and gene networks involved in pig ovarian follicular atresia.

Ovarian folliculogenesis corresponds to the development of follicles leading to either ovulation or degeneration, this latter process being called atresia. Even if atresia involves apoptosis, its mechanism is not well understood. The objective of this study was to analyze global gene expression in pig granulosa cells of ovarian follicles during atresia. The transcriptome analysis was performed on a 9,216 cDNA microarray to identify gene networks and candidate genes involved in pig ovarian follicular atresia. We found 1,684 significantly regulated genes to be differentially regulated between small healthy follicles and small atretic follicles. Among them, 287 genes had a fold-change higher than two between the two follicle groups. Eleven genes (DKK3, GADD45A, CAMTA2, CCDC80, DAPK2, ECSIT, MSMB, NUPR1, RUNX2, SAMD4A, and ZNF628) having a fold-change higher than five between groups could likely serve as markers of follicular atresia. Moreover, automatic confrontation of deregulated genes with literature data highlighted 93 genes as regulatory candidates of pig granulosa cell atresia. Among these genes known to be inhibitors of apoptosis, stimulators of apoptosis, or tumor suppressors INHBB, HNF4, CLU, different interleukins (IL5, IL24), TNF-associated receptor (TNFR1), and cytochrome-c oxidase (COX) were suggested as playing an important role in porcine atresia. The present study also enlists key upstream regulators in follicle atresia based on our results and on a literature review. The novel gene candidates and gene networks identified in the current study lead to a better understanding of the molecular regulation of ovarian follicular atresia.

BMP15 regulates the inhibin/activin system independently of ovulation rate control in sheep.

Polymorphisms in the gene encoding bone morphogenetic protein 15 (BMP15) have been associated with multiple ovulations in sheep. As BMP15 regulates inhibin expression in rodents, we assumed that the ovarian inhibin/activin system could mediate part of the effect of BMP15 mutations in the regulation of ovulation rate in sheep. To answer this question, we have studied the effects of two natural loss-of-function mutations of BMP15 on the expression of components of this system. The FecXR and the FecXGr mutations, when present respectively in Rasa Aragonesa ewes at the heterozygous state and in Grivette ewes at the homozygous state, were associated with a twofold increase in ovulation rate. There were only small differences between mutant and wild-type ewes for mRNA expression of INHA, INHBA, ACVR1B, ACVR2A, FST or TGFBR3 in granulosa cells and inhibin A or activin A concentrations in follicular fluid. Moreover, the effects of mutations differed between breeds. In cultures of granulosa cells from wild-type ewes, BMP15, acting alone or in synergy with GDF9, stimulated INHA, INHBA and FST expression, but inhibited the expression of TGFBR3 Activin A did not affect INHBA expression, but inhibited the expression of ACVR2A also. The complexity of the inhibin/activin system, including positive and antagonistic elements, and the differential regulation of these elements by BMP15 and activin can explain that the effects of BMP15 mutations differ when present in different genetic backgrounds. In conclusion, the ovarian inhibin/activin system is unlikely to participate in the increase of ovulation rate associated with BMP15 mutations in sheep.

Molecular evidence that follicle development is accelerated in vitro compared to in vivo.

In this study, we systematically compared the morphological, functional and molecular characteristics of granulosa cells and oocytes obtained by a three-dimensional in vitro model of ovine ovarian follicular growth with those of follicles recovered in vivo Preantral follicles of 200 µm diameter were recovered and cultured up to 950 µm over a 20-day period. Compared with in vivo follicles, the in vitro culture conditions maintained follicle survival, with no difference in the rate of atresia. However, the in vitro conditions induced a slight decrease in oocyte growth rate, delayed antrum formation and increased granulosa cell proliferation rate, accompanied by an increase and decrease in CCND2 and CDKN1A mRNA expression respectively. These changes were associated with advanced granulosa cell differentiation in early antral follicles larger than 400 µm diameter, regardless of the presence or absence of FSH, as indicated by an increase in estradiol secretion, together with decreased AMH secretion and expression, as well as increased expression of GJA1, CYP19A1, ESR1, ESR2, FSHR, INHA, INHBA, INHBB and FST There was a decrease in the expression of oocyte-specific molecular markers GJA4, KIT, ZP3, WEE2 and BMP15 in vitro compared to that in vivo Moreover, a higher percentage of the oocytes recovered from cultured follicles 550 to 950 µm in diameter was able to reach the metaphase II meiosis stage. Overall, this in vitro model of ovarian follicle development is characterized by accelerated follicular maturation, associated with improved developmental competence of the oocyte, compared to follicles recovered in vivo.

Multi-scale modelling of ovarian follicular development: From follicular morphogenesis to selection for ovulation.

In this review, we present multi-scale mathematical models of ovarian follicular development that are based on the embedding of physiological mechanisms into the cell scale. During basal follicular development, follicular growth operates through an increase in the oocyte size concomitant with the proliferation of its surrounding granulosa cells. We have developed a spatio-temporal model of follicular morphogenesis explaining how the interactions between the oocyte and granulosa cells need to be properly balanced to shape the follicle. During terminal follicular development, the ovulatory follicle is selected amongst a cohort of simultaneously growing follicles. To address this process of follicle selection, we have developed a model giving a continuous and deterministic description of follicle development, adapted to high numbers of cells and based on the dynamical and hormonally regulated repartition of granulosa cells into different cell states, namely proliferation, differentiation and apoptosis. This model takes into account the hormonal feedback loop involving the growing ovarian follicles and the pituitary gland, and enables the exploration of mechanisms regulating the number of ovulations at each ovarian cycle. Both models are useful for addressing ovarian physio-pathological situations. Moreover, they can be proposed as generic modelling environments to study various developmental processes and cell interaction mechanisms.

Inhibitors of c-Jun phosphorylation impede ovine primordial follicle activation.

STUDY HYPOTHESIS: Is the c-Jun-N-terminal kinase (JNK) pathway implicated in primordial follicle activation? STUDY FINDING: Culture of ovine ovarian cortex in the presence of two different c-Jun phosphorylation inhibitors impeded pre-antral follicle activation. WHAT IS KNOWN ALREADY: Despite its importance for fertility preservation therapies, the mechanisms of primordial follicle activation are poorly understood. Amongst different signalling pathways potentially involved, the JNK pathway has been previously shown to be essential for cell cycle progression and pre-antral follicle development in mice. STUDY DESIGN, SAMPLES/MATERIALS, METHODS: Ovine ovarian cortex pieces were cultured with varying concentrations of SP600125, JNK inhibitor VIII or anti-Mullerian hormone (AMH) in the presence of FSH for 9 days. Follicular morphometry and immunohistochemistry for proliferating cell nuclear antigen (PCNA), apoptosis and follicle activation (Foxo3a) were assessed. MAIN RESULTS AND THE ROLE OF CHANCE: Inhibition of primordial follicle activation occurred in the presence of SP600125, JNK inhibitor VIII and AMH when compared with controls (all P < 0.05) after 2 days of culture. However, only in the highest concentrations used was the inhibition of activation associated with induction of follicular apoptosis (P < 0.05). In growing follicles, PCNA antigen expression was reduced when the JNK inhibitors or AMH were used (P < 0.05 versus control), indicating reduced proliferation of the somatic compartment. LIMITATIONS, REASONS FOR CAUTION: Although we evaluated the effects of inhibition of c-Jun phosphorylation on primordial follicle development, we did not determine the cellular targets and mechanism of action of the inhibitors. WIDER IMPLICATIONS OF THE FINDINGS: These results are the first to implicate the JNK pathway in primordial follicle activation and could have significant consequences for the successful development of fertility preservation strategies and our understanding of primordial follicle activation. LARGE SCALE DATA: n/a. STUDY FUNDING AND COMPETING INTERESTS: Dr Michael J. Bertoldo and the laboratories involved in the present study were supported by a grant from 'Région Centre' (CRYOVAIRE, Grant number #320000268). There are no conflicts of interest to declare.

Postnatal pituitary and follicular activation: a revisited hypothesis in a sheep model.

The importance of postnatal pituitary activation as regards female reproductive development is not yet understood. By taking advantage of the experimental model developed in a previous study, i.e. ewe lambs expressing markedly different ovarian phenotypes at 50 days of age, we designed this study to determine whether differences found in ovarian status during the early prepubertal period are due to different patterns of postnatal pituitary activation, and to assess whether these differences have long lasting effects on subsequent reproductive performance. Results showed that ewe lambs with high antral follicle count (AFC) at 50 days of age had significantly lower plasma FSH concentrations and higher anti-Mullerian hormone (AMH) concentrations during the first 9 weeks of age compared with low AFC ewe lambs (P<0.0001). With a longitudinal experiment we showed that a high AFC in the early prepubertal period is associated with consistently higher AMH concentrations and numbers of antral follicles up to the postpubertal period, and with higher pregnancy rates in the first breeding season. In addition, the effect of age in decreasing AMH concentrations was more marked in the low AFC group. Results of the present study demonstrate that ewe lambs undergo different patterns of postnatal pituitary activation. A high AFC at 50 days of age indicates an advanced phase of ovarian maturation, which was accompanied by constantly higher AMH concentrations up to the postpubertal period, a greater ovarian response to FSH stimulation and by higher pregnancy rates at first mating, as compared with the low AFC group.

The highly prolific phenotype of Lacaune sheep is associated with an ectopic expression of the B4GALNT2 gene within the ovary.

Prolific sheep have proven to be a valuable model to identify genes and mutations implicated in female fertility. In the Lacaune sheep breed, large variation in litter size is genetically determined by the segregation of a fecundity major gene influencing ovulation rate, named FecL and its prolific allele FecL(L) . Our previous work localized FecL on sheep chromosome 11 within a locus of 1.1 Mb encompassing 20 genes. With the aim to identify the FecL gene, we developed a high throughput sequencing strategy of long-range PCR fragments spanning the locus of FecL(L) carrier and non-carrier ewes. Resulting informative markers defined a new 194.6 kb minimal interval. The reduced FecL locus contained only two genes, insulin-like growth factor 2 mRNA binding protein 1 (IGF2BP1) and beta-1,4-N-acetyl-galactosaminyl transferase 2 (B4GALNT2), and we identified two SNP in complete linkage disequilibrium with FecL(L) . B4GALNT2 appeared as the best positional and expressional candidate for FecL, since it showed an ectopic expression in the ovarian follicles of FecL(L) /FecL(L) ewes at mRNA and protein levels. In FecL(L) carrier ewes only, B4GALNT2 transferase activity was localized in granulosa cells and specifically glycosylated proteins were detected in granulosa cell extracts and follicular fluids. The identification of these glycoproteins by mass spectrometry revealed at least 10 proteins, including inhibin alpha and betaA subunits, as potential targets of B4GALNT2 activity. Specific ovarian protein glycosylation by B4GALNT2 is proposed as a new mechanism of ovulation rate regulation in sheep, and could contribute to open new fields of investigation to understand female infertility pathogenesis.

The ovarian reserve of primordial follicles and the dynamic reserve of antral growing follicles: what is the link?

The growing follicles develop from a reserve of primordial follicles constituted early in life. From this pre-established reserve, a second ovarian reserve is formed, which consists of gonadotropin-responsive small antral growing follicles and is a dynamic reserve for ovulation. Its size, evaluated by direct antral follicular count or endocrine markers, determines the success of assisted reproductive technologies in humans and embryo production biotechnologies in animals. Strong evidence indicates that these two reserves are functionally related. The size of both reserves appears to be highly variable between individuals of similar age, but the equilibrium size of the dynamic reserve in adults seems to be specific to each individual. The dynamics of both follicular reserves appears to result from the fine tuning of regulations involving two main pathways, the phosphatase and tensin homolog (PTEN)/phosphatidylinositol-3 kinase (PI3K)/3-phosphoinositide-dependent protein kinase-1 (PDPK1)/v-akt murine thymoma viral oncogene homolog 1 (AKT1) and the bone morphogenetic protein (BMP)/anti-Müllerian hormone (AMH)/SMAD signaling pathways. Mutations in genes encoding the ligands, receptors, or signaling effectors of these pathways can accelerate or modulate the exhaustion rate of the ovarian reserves, causing premature ovarian insufficiency (POI) or increase in reproductive longevity, respectively. With female aging, the decline in primordial follicle numbers parallels the decrease in the size of the dynamic reserve of small antral follicles and the deterioration of oocyte quality. Recent progress in our knowledge of signaling pathways and their environmental and hormonal control during adult and fetal life opens new perspectives to improve the management of the ovarian reserves.

Progesterone improves the maturation of male-induced preovulatory follicles in anoestrous ewes.

The first ovulation induced by male effect in sheep during seasonal anoestrus usually results in the development of a short cycle that can be avoided by progesterone priming before ram introduction. In elucidating the involvement of the hypothalamic-pituitary-gonadal axis in the occurrence of short cycles, the effects of progesterone and the time of anoestrus on the development of male-induced preovulatory follicles were investigated in anoestrous ewes using morphological, endocrine and molecular approaches. Ewes were primed with progesterone for 2 (CIDR2) or 12 days (CIDR12) and untreated ewes used as controls during early (April) and late (June) anoestrus. The duration of follicular growth and the lifespan of the male-induced preovulatory follicles were prolonged by ∼1.6 days in CIDR12 ewes compared with the controls. These changes were accompanied by a delay in the preovulatory LH and FSH surges and ovulation. Intra-follicular oestradiol concentration and mRNA levels of LHCGR and STAR in the granulosa and theca cells of the preovulatory follicles were higher in CIDR12 ewes than the control ewes. The expression of mRNA levels of CYP11A1 and CYP17A1 also increased in theca cells of CIDR12 ewes. CIDR2 ewes gave intermediate results. Moreover, ewes ovulated earlier in June than in April, without changes in the duration of follicular growth, but these effects were unrelated to the lifespan of corpus luteum. Our results give the first evidence supporting the positive effect of progesterone priming on the completion of growth and maturation of preovulatory follicles induced by male effect in seasonal anoestrous ewes, thereby preventing short cycles.

Anti-Müllerian hormone plasma concentration in prepubertal ewe lambs as a predictor of their fertility at a young age.

BACKGROUND: In mammals, the ovarian follicular reserve is highly variable between individuals and impacts strongly on ovarian function and fertility. Nowadays, the best endocrine marker of this reserve in human, mouse and cattle is the anti-Müllerian hormone (AMH). The objectives of this work were to determine whether AMH could be detected in the plasma of prepubertal ewe lambs and to assess its relationship with their fertility at a young age. RESULTS: Plasma was taken from 76 Rasa Aragonesa ewe lambs at 3.6 months of age for AMH determination. Simultaneously, 600 IU equine chorionic gonadotropin (eCG) was administered and the number of ovulations recorded 6 days later. AMH was detected in 93% of the lambs, and the concentrations were about 3-4-fold higher in ovulating than in non-ovulating lambs (P < 0.004). Ewes aged around 10 months were mated, giving an overall fertility of 29%, and those failing to conceive were mated again 4 months later. Fertility at first mating was significantly correlated with plasma AMH concentration at 3.6 months (Spearman's ρ = 0.34; P < 0.01). To use plasma AMH concentration as a screening test, a value of 97 pg/mL was determined as the optimum cutoff value to predict fertility at first mating (sensitivity = 68.2%; specificity = 72.2%). Fertility at first mating was 34.8 percentage points higher in ewe lambs with an AMH ≥ 97 pg/mL than in those with lower AMH concentrations (50% vs. 15%; P < 0.001). CONCLUSIONS: Plasma AMH concentration might be a reliable marker of the ovarian status of prepubertal ewe lambs, reflecting their ability to respond to eCG stimulation. A single AMH measurement performed on ewe lambs early in age could be useful to select for replacement ewes with a higher predicted fertility at first mating.

Determination of anti-Müllerian hormone concentrations in blood as a tool to select Holstein donor cows for embryo production: from the laboratory to the farm.

High between-animal variability in the number of embryos produced by multiple ovulation and embryo transfer (MOET) and ovum pick-up and in vitro production (OPU-IVP) methods remains a major limit to the development of embryo biotechnologies in cattle. The measurement of anti-Müllerian hormone (AMH) endocrine concentrations in cows can help to predict their follicular and ovulatory responses to gonadotrophin treatment. The present study aimed to provide practical information for a simple prognostic method based on AMH measurement in Holstein cows. Accurate AMH concentrations could be measured with ELISA in blood or plasma. In cows undergoing repeated OPU protocols over 1 year, the AMH concentrations measured in plasma samples collected before each gonadotrophin treatment were found to be highly repeatable and were tightly correlated with follicular responses. From data obtained at both an experimental station and farm settings, it was possible to propose AMH cut-off values to identify low-responding cows. Gonadotrophin-stimulated cows producing fewer than 15 large follicles at oestrus and fewer than 10 embryos in MOET protocols could be discarded efficiently with plasma AMH concentrations below 87 and 74 pg mL(-1), respectively. In conclusion, we propose a prognostic method based on a single AMH measurement to improve the results of embryo biotechnologies.

Regulation of anti-Müllerian hormone production in domestic animals.

In mammals, anti-Müllerian hormone (AMH) expression is detected in the granulosa cells of all growing follicles and is highest in healthy small antral follicles, which contribute most significantly to AMH endocrine levels. AMH is a reliable endocrine marker of this population of gonadotrophin-responsive follicles in ruminants and, over the longer term, plasma AMH concentrations are characteristic of individual animals. In the cow, plasma AMH concentrations follow specific dynamic profiles throughout the prepubertal period, the oestrous cycle and the change from gestation to the post partum period, with the alterations most likely reflecting numerical changes in the population of high AMH-producing follicles. In granulosa cells, bone morphogenetic proteins (BMP) enhance AMH gene expression and AMH synthesis, with these effects antagonised by FSH. BMP could both support follicular growth and contribute significantly to the induction and/or maintenance of AMH expression in small growing follicles. AMH expression decreases sharply in large follicles when they become oestrogenic, suggesting a role for FSH and/or oestradiol in these changes, but the underlying mechanisms remain hypothetical. A better understanding of the factors and mechanisms regulating AMH production is needed to propose new strategies for managing the reserve of primordial and small growing follicles, as well as for improving embryo production.

Regulation of anti-Müllerian hormone production in the cow: a multiscale study at endocrine, ovarian, follicular, and granulosa cell levels.

Anti-Müllerian hormone (AMH) is an endocrine marker that can help predict superovulatory responses to treatments administered to cows for embryo production. However, the optimal time of the estrous cycle at which a blood test should be performed for a highly reliable prognosis has not yet been established. Moreover, little is known about the regulation of AMH production. To answer these questions, a study was designed to investigate the regulation of AMH production in cows selected for their high or low ovulatory responses to superovulation. At the granulosa cell level, AMH production was inhibited by follicle-stimulating hormone but enhanced by bone morphogenetic proteins. At the follicular level, the expression of AMH within the follicle was dependent on the stage of follicular development. At the ovarian level, the size of the pool of small antral growing follicles determined ovarian AMH production. At the endocrine level, AMH followed a specific dynamic profile during the estrous cycle, which occurred independently of the follicular waves of terminal follicular development. Cows selected for their high or low responses to superovulation did not differ in the regulation of AMH production, but cows with higher responses had higher plasma AMH concentrations throughout the cycle. The optimal period of the estrous cycle at which to measure AMH concentrations with the aim of selecting the best cows for embryo production was found to be at estrus and after Day 12 of the cycle. Based on this multiscale study, we propose a model that integrates the different regulatory levels of AMH production.

Anti-Mullerian hormone as a predictive endocrine marker for embryo production in the goat.

Recently, we demonstrated the relationship between anti-Müllerian hormone (AMH) circulating concentrations, ovarian follicles, and embryo production in cattle. However, they have not yet been established in a species with a seasonal breeding activity. Thus, goats were subjected to repeated in vivo embryo production during the breeding season, at the end of the breeding season, and at the end of the anestrus season. Embryo production after FSH treatment was highly repeatable for each goat. Plasma AMH concentrations, measured before the first FSH treatment, were highly correlated with the number of collected, transferable, and freezable embryos, resulting from the three sessions of embryo production. Plasma AMH concentrations transiently decreased after each exogenous FSH treatment, but they showed little change with season, and no relationship was observed between AMH and endogenous FSH concentrations during seasonal transitions. Follicles of 1-5 mm in diameter were the main target of the FSH treatment and were major contributors to circulating AMH concentrations. Granulosa cell AMH expression decreased as the follicle approached terminal development, while the expression of maturation markers (CYP19A1 and FSHR) increased. In conclusion, circulating AMH concentrations can be predictive of the capacity of a donor goat to produce high or low numbers of high-quality embryos. This prediction could be accurately made from a single blood measurement of AMH during either breeding or anestrus seasons. Variability in the number of gonadotropin-responsive follicles of 1-5 mm in diameter between individuals resulted in the differences in circulating AMH concentrations measured between individuals.

The Crazy Ovary.

From fetal life until senescence, the ovary is an extremely active tissue undergoing continuous structural and functional changes. These ever-changing events are best summarized by a quotation attributed to Plato when describing motion in space and time-'nothing ever is but is always becoming…'. With respect to the ovary, these changes include, at the beginning, the processes of follicular formation and thereafter those of follicular growth and atresia, steroidogenesis, oocyte maturation, and decisions relating to the number of mature oocytes that are ovulated for fertilization and the role of the corpus luteum. The aims of this review are to offer some examples of these complex and hitherto unknown processes. The ones herein have been elucidated from studies undertaken in vitro or from normal in vivo events, natural genetic mutations or after experimental inactivation of gene function. Specifically, this review offers insights concerning the initiation of follicular growth, pathologies relating to poly-ovular follicles, the consequences of premature loss of germ cells or oocytes loss, the roles of AMH (anti-Müllerian hormone) and BMP (bone morphogenetic protein) genes in regulating follicular growth and ovulation rate together with species differences in maintaining luteal function during pregnancy. Collectively, the evidence suggests that the oocyte is a key organizer of normal ovarian function. It has been shown to influence the phenotype of the adjacent somatic cells, the growth and maturation of the follicle, and to determine the ovulation rate. When germ cells or oocytes are lost prematurely, the ovary becomes disorganized and a wide range of pathologies may arise.