The observations that substance P (SP) is localized in the anterior pituitary gland (AP) and is regulated by the hormonal status of the animal, as well as the demonstration of SP binding sites in the AP, have led to the idea that SP may participate in the regulation of AP function. Numerous and sometimes contradictory reports of SP effects on AP hormone secretion, particularly on luteinizing hormone (LH), left the question of whether SP acts directly at the level of the AP to regulate LH secretion still unanswered. To investigate a possible physiological function of SP in the AP, the effects of exogenous SP on LH secretion from AP cells from adult and prepubertal male and female rats in short term culture were studied. It was found that SP (100nM-1μM) significantly stimulates LH release in cultured AP cells and that this effect varies as a function of age and sex. SP has no significant effect on LH release from AP cells of male and female prepubertal rats. After day 30 a sharp increase in the response to SP occurs in both sexes. This level of responsiveness continues through adulthood in AP cells from the female rat. In contrast, AP cells from male rats failed to respond during adulthood (over 50 days of age) but were highly responsive during the peripubertal period (30-35 days). The possibility that the responsiveness to SP is influenced by the endocrine status of the animal was investigated by exposing AP cells from responding animals to androgens in vivo and in vitro. It was found that AP cells from female rats treated with androgen were less responsive to 100nM SP but did respond at higher doses of SP. SP effects on AP function were further analyzed in experiments using radioligand binding assays to assess possible changes in SP receptor number or affinity as related to age and sex. In AP membranes from female rats, maximum binding is 8-fold higher (Bmax=4.2 pmo1/mg membrane protein) than in AP membranes from male rats (Bmax=560fmo1/ mg membrane protein). These studies suggest a role for SP as a secondary regulator of LH secretion with possible physiological significance for reproductive function.

No description available.

Introduction: Current health literature suggests that there has been a decline in the age of pubertal onset and that pubertal onset/duration of puberty may, besides weight status, be influenced by socioeconomic context. Objective: The goal of this study was to determine whether pubertal onset/ duration and puberty-triggering hormones luteinizing hormone (LH) and follicle-stimulating hormone (FSH) vary according to socioeconomic status (SES). Moreover, we aimed to propose cutoff values of serum LH and FSH for predicting gonadarche in boys. Methods: 2,657 apparently healthy children and adolescents between 5.5 and 18 years from the area of Leipzig were recruited from the LIFE Child study. Age at pubertal onset/end of puberty was given in 738/573 children, respectively. Anthropometric parameters of puberty, blood measurements of LH and FSH, and questionnaires assessing SES were evaluated. Results: Lower SES was associated with earlier thelarche and longer duration of puberty in overweight/obese girls, whereas age of menarche was not affected. In boys with low SES, a trend versus earlier puberty onset can be seen. Lower SES was significantly associated with boys’ age at mutation. No significant differences in boys’ and girls’ serum levels of LH and FSH during puberty according to SES were observed. Serum LH levels of 0.56 IU/L and serum FSH levels of 1.74 IU/L showed the best prediction of gonadarche in boys. Conclusion: Puberty onset/duration and boys’ age at mutation is affected by SES. The proposed cutoff levels for serum LH and FSH could provide a serological tool to determine gonadarche in boys

No description available.

Ce travail a consisté à analyser, par des approches toxicocinétiques (TK) et mécanistiques, les effets oestrogéniques du Bisphenol A (BPA) sur un biomarqueur précoce et sensible : la sécrétion de l'hormone lutéinisante (LH) chez la brebis prépubère ovariectomisée. La plus faible concentration plasmatique en BPA induisant une inhibition de LH s'est avérée proche des concentrations maximales décrites chez l'Homme. Cette inhibition de LH pourrait impliquer une inhibition des systèmes neuronaux à kisspeptine. L'approche TK comparative d'espèces a montré que la clairance du BPA est toujours élevée, proche du débit sanguin hépatique. Pour une exposition à la dose journalière admissible, cette approche permet de prédire chez l'Homme des concentrations en BPA très inférieures à celles associées à une inhibition de LH dans notre modèle. / The goal of this thesis was to analyse through toxicokinetic (TK) and mechanistic approaches the estrogeno-mimetic effects of bisphenol A (BPA) on a precocious and sensitive biomarker: LH secretion in ovariectomized female lambs. The lowest plasma BPA concentrations associated to an inhibition of LH secretion appeared to be close to the highest one reported in human. LH suppression could be mediated by an inhibition of hypothalamic kisspeptin systems. The multispecies TK approach showed that BPA clearance is always high and equivalent to the liver blood flow. For an exposure scheme corresponding to the tolerable daily intake, this approach allows to predict human BPA concentration much lower than the one associated to LH inhibition in our highly sensitive lamb model.

Abstract not available

In sheep, large ovarian antral follicles grow in waves with a periodicity of every 4 to 5 days; each wave is initiated by a peak in serum concentrations of follicle stimulating hormone (FSH). In the present thesis, follicular data and hormone estimations acquired from daily ultrasonography and blood samples, respectively, were used to study mechanisms regulating the number of follicular waves per estrous cycle. Using additional approaches such as implants releasing estradiol-17â and or progesterone, immunization against gonadotropin releasing hormone (GnRH), and injections of GnRH, the role of pulsed luteinizing hormone (LH) secretion and FSH peaks in follicular wave emergence and growth and the dependency of FSH peaks on pulsed GnRH secretion, were studied in sheep. The viability of aged follicles was also addressed.<p> The results of the present studies showed that ewes with three or four waves per cycle had cycles of the same length. The inter-wave interval was longer for the first and the last or ovulatory wave of the cycle in three compared to four wave cycles. The length of the lifespan and regression phase of the largest follicle of a wave declined across the cycle as FSH peak concentration and amplitude decreased. The maximum follicular diameter of the largest follicle growing in the first wave and the last or ovulatory wave of the cycle was greater compared to other waves of the cycle. Treatment of anestrous ewes with estradiol releasing implants alone completely abolished pulsed LH secretion and suppressed follicular wave development; however, FSH secretion was only minimally affected and the pool of small follicles was not affected. When pulsed secretion of LH was restored by frequent injections of GnRH, follicular waves were re-established. Treatment of anestrous ewes with implants releasing estradiol and progesterone, decreased FSH peak amplitude and abolished LH pulses and follicular waves; the size of the pool of small follicles increased. Immunization against GnRH in anestrous ewes abolished pulsatile LH secretion and suppressed follicular wave emergence; however, FSH peaks continued to occur for several weeks. In cyclic ewes, creating an LH pulse frequency typical of the follicular phase, during the luteal phase of the cycle by giving GnRH, increased maximum diameter of the largest follicle in a wave and serum concentrations of estradiol and progesterone. The enhanced growth of follicles in a wave blocked the next expected FSH peak and its associated follicular wave. Decreasing LH pulse frequencies lower than the minimal frequency seen in the luteal phase, by implants releasing progesterone, did not affect the growth of follicular waves.<p> It was previously demonstrated that treatment of non-prolific WWF ewes with Prostaglandin F2á (PGF2á) and medroxy progesterone acetate (MPA) increased the ovulation rate by adding ovulations from the penultimate wave in addition to the final wave of the cycle; however, fertility was not improved. In the last study of my thesis, we collected follicles, with an extended lifespan, from the penultimate wave of the cycle in ewes given the PGF2á and MPA treatment. We compared their quality with follicles from the final wave of the cycle by looking at the expression of markers of follicular development. The results showed that theca cells of follicles from the final wave had significantly higher mRNA expression for vascular endothelial growth factor (VEGF) compared to follicles from the penultimate wave. Granulosa cells of follicles from the final wave had significantly higher mRNA expression for connexion 43 (Cx43) compared to follicles from the penultimate wave. Protein expression for Cx43, proliferating cell nuclear antigen (PCNA) and Factor VIII was greater in follicles from the final compared to the penultimate wave.<p> We concluded from the present studies that: 1) the mechanism that makes a three wave or four wave cycle is unclear; 2) some level of pulsatile LH secretion is required for an FSH peak to trigger emergence of follicular waves in anestrous ewes; 3) progesterone enhances the inhibitory effects of estradiol on FSH secretion in anestrous ewes, suppressing specifically FSH peak amplitude; 4) an endogenous rhythm may exist that drives the peaks in FSH secretion independent of secretory products from the follicles growing in a wave and pulsed GnRH secretion; 5) follicular waves in ewes, when exposed to an LH pulse frequency similar to the follicular phase, during the luteal phase of the cycle, when serum progesterone concentrations are high, can grow and function like ovulatory follicles growing in the follicular phase of the cycle; 6) expression of some markers of vascularization/ angiogenesis, gap-junctional communication and cell proliferation, appeared to be decreased in follicles from the penultimate compared to the final wave of an estrous cycle, when the lifespan of follicles from the penultimate wave was extended such that they were present in the ovary with follicles from the final wave of the cycle.

Treatment success in IVF-ICSI is mainly limited by female age, but differences in ovarian reserve (OR; the remaining pool of oocytes and their quality) between individuals modify treatment prerequisites among women of similar age. OR may be assessed by OR tests (ORTs). The main aims of this work were to study menstrual cycle length (MCL), basal levels of circulating gonadotrophins, antral follicle count (AFC) and serum Anti-Müllerian hormone (AMH) levels and their associations with and prognostic capacities regarding IVF-ICSI outcome in large cohorts of unselected women. Age-adjusted MCL was positively and linearly associated with pregnancy rates (PRs), live-birth rates (LBRs) and ovarian response to controlled ovarian hyperstimulation. An MCL of &gt;34 days almost doubled the LBR compared with an MCL of &lt;26 days. The grouped variable ‘combined FSH and LH levels’ was superior to both individual gonadotrophin levels and the LH:FSH ratio. The highest mean PR was seen in connection with a combination of FSH &lt;6.7 U/l with LH &gt;4.9 U/l; PRs were lowest when FSH-LH levels were opposite to this (high-low) and intermediate when FSH-LH levels were low-low or high-high. Associations with LBR and ovarian response were similar as those for PR. AFCs and serum AMH levels were positively and log-linearly associated with PR, LBR and ovarian response. Success rates levelled out above AFC 30 or AMH 5 ng/ml. Treatment outcome was superior among women with polycystic ovaries. Among the studied ORTs, logAFC and logAMH concentration correlated most strongly. After multivariate testing, entering all studied ORTs, AMH and female age remained independently associated with LBR. AMH + AFC + age predicted both poor and excessive ovarian responses with high accuracy. Adjusting for age and oocyte yield, all ORTs remained significant for LBR, implying that ORTs also capture information on oocyte quality. In conclusion, measures of OR are strongly associated with PR, LBR and ovarian response in a log-linear fashion, and partly reflect oocyte quality. The OR spectrum is continuous, from small ‘oligofollicular’ ovaries (the low extreme) to polycystic ovaries (the high extreme). Among the studied ORTs, AMH together with age provide the most powerful basal estimate for IVF/ICSI outcome.

In sheep, large ovarian antral follicles grow in waves with a periodicity of every 4 to 5 days; each wave is initiated by a peak in serum concentrations of follicle stimulating hormone (FSH). In the present thesis, follicular data and hormone estimations acquired from daily ultrasonography and blood samples, respectively, were used to study mechanisms regulating the number of follicular waves per estrous cycle. Using additional approaches such as implants releasing estradiol-17â and or progesterone, immunization against gonadotropin releasing hormone (GnRH), and injections of GnRH, the role of pulsed luteinizing hormone (LH) secretion and FSH peaks in follicular wave emergence and growth and the dependency of FSH peaks on pulsed GnRH secretion, were studied in sheep. The viability of aged follicles was also addressed.<p> The results of the present studies showed that ewes with three or four waves per cycle had cycles of the same length. The inter-wave interval was longer for the first and the last or ovulatory wave of the cycle in three compared to four wave cycles. The length of the lifespan and regression phase of the largest follicle of a wave declined across the cycle as FSH peak concentration and amplitude decreased. The maximum follicular diameter of the largest follicle growing in the first wave and the last or ovulatory wave of the cycle was greater compared to other waves of the cycle. Treatment of anestrous ewes with estradiol releasing implants alone completely abolished pulsed LH secretion and suppressed follicular wave development; however, FSH secretion was only minimally affected and the pool of small follicles was not affected. When pulsed secretion of LH was restored by frequent injections of GnRH, follicular waves were re-established. Treatment of anestrous ewes with implants releasing estradiol and progesterone, decreased FSH peak amplitude and abolished LH pulses and follicular waves; the size of the pool of small follicles increased. Immunization against GnRH in anestrous ewes abolished pulsatile LH secretion and suppressed follicular wave emergence; however, FSH peaks continued to occur for several weeks. In cyclic ewes, creating an LH pulse frequency typical of the follicular phase, during the luteal phase of the cycle by giving GnRH, increased maximum diameter of the largest follicle in a wave and serum concentrations of estradiol and progesterone. The enhanced growth of follicles in a wave blocked the next expected FSH peak and its associated follicular wave. Decreasing LH pulse frequencies lower than the minimal frequency seen in the luteal phase, by implants releasing progesterone, did not affect the growth of follicular waves.<p> It was previously demonstrated that treatment of non-prolific WWF ewes with Prostaglandin F2á (PGF2á) and medroxy progesterone acetate (MPA) increased the ovulation rate by adding ovulations from the penultimate wave in addition to the final wave of the cycle; however, fertility was not improved. In the last study of my thesis, we collected follicles, with an extended lifespan, from the penultimate wave of the cycle in ewes given the PGF2á and MPA treatment. We compared their quality with follicles from the final wave of the cycle by looking at the expression of markers of follicular development. The results showed that theca cells of follicles from the final wave had significantly higher mRNA expression for vascular endothelial growth factor (VEGF) compared to follicles from the penultimate wave. Granulosa cells of follicles from the final wave had significantly higher mRNA expression for connexion 43 (Cx43) compared to follicles from the penultimate wave. Protein expression for Cx43, proliferating cell nuclear antigen (PCNA) and Factor VIII was greater in follicles from the final compared to the penultimate wave.<p> We concluded from the present studies that: 1) the mechanism that makes a three wave or four wave cycle is unclear; 2) some level of pulsatile LH secretion is required for an FSH peak to trigger emergence of follicular waves in anestrous ewes; 3) progesterone enhances the inhibitory effects of estradiol on FSH secretion in anestrous ewes, suppressing specifically FSH peak amplitude; 4) an endogenous rhythm may exist that drives the peaks in FSH secretion independent of secretory products from the follicles growing in a wave and pulsed GnRH secretion; 5) follicular waves in ewes, when exposed to an LH pulse frequency similar to the follicular phase, during the luteal phase of the cycle, when serum progesterone concentrations are high, can grow and function like ovulatory follicles growing in the follicular phase of the cycle; 6) expression of some markers of vascularization/ angiogenesis, gap-junctional communication and cell proliferation, appeared to be decreased in follicles from the penultimate compared to the final wave of an estrous cycle, when the lifespan of follicles from the penultimate wave was extended such that they were present in the ovary with follicles from the final wave of the cycle.

Advanced maternal age has been shown to influence follicular and luteal dynamics in bovine ovary resulting in reduced fertility. The overall objective of the four studies presented in this thesis is to identify the maternal age-associated transcriptional changes in granulosa cells of the dominant follicles during follicle development. In the first study, mRNA expression levels of housekeeping genes were measured by real–time quantitative PCR (RT-qPCR) in granulosa cells of dominant follicles and FSH-stimulated follicles to select and validate suitable reference genes for relative gene expression analyses during maternal and follicular aging. Stability of six reference genes (GAPDH, ACTB, EIF2B2, UBE2D2, SF3A1 and RNF20) was analyzed using GeNorm, DeltaCT and NormFinder programs and comprehensive ranking order was determined based on these programs. Geometric mean of multiple genes (UBE2D2, EIF2B2, GAPDH and SF3A1) was more appropriate reference control than individual genes for the comparison of relative gene expression among dominant and FSH-stimulated follicles during maternal and/or follicular aging studies. In the second study, maternal age-associated changes in the transcriptome of granulosa cells recovered at the time of selection of the dominant follicle from aged (n=3) and young cows (n=3) were determined by EmbryoGENE bovine oligo-microarrays (EMBV3, Agilent Technology). The mRNA expression of five transcripts (CYP19A1, PCNA, GJA1, TPM2, and VNN1) was confirmed in a different set of granulosa cell samples by RT-qPCR to validate microarray data. A total of 169 genes/isoforms were differentially expressed (≥ 2-fold-change; P ≤ 0.05) in aged cows vs. young cows. These transcripts revealed inefficient 1) control of gonadotropins, and gonadotropin-induced changes in the cytoskeleton and extracellular matrix, 2) lipid metabolism and steroidogenesis 3) cell proliferation, cell cycle control and intercellular communication, and 4) higher oxidative stress responses in aged cows vs. young cows. In the third study, changes in the transcriptome of granulosa cells of the preovulatory follicle 24 h after LH treatment from aged (n= 3) and young (n=3) were determined. A total of 1340 genes were expressed differentially (≥ 2-fold change; P ≤ 0.05) in aged cows vs. young cows. The mRNA expression of five transcripts (RGS2, PTGS2, TNFAIP6, VNN1, NR5A2 and GADD45B) was confirmed in a different set of granulosa cell samples to validate microarray data. These transcripts were related to delayed 1) response to LH treatment 2) cellular differentiation and luteinization and 3) progesterone synthesis. Intra-follicle levels of progesterone were lower (P < 0.05) in aged cows compared to young and mid-aged cows. The fourth study compared the aged-associated changes in the transcriptome of granulosa cells during follicle development from the time of dominant follicle selection to preovulatory stage (24 h after LH). In comparison to young cows, aged cows expressed fewer differentially expressed genes/isoforms (1206 vs. 2260, respectively) at ≥ 2-fold-change (P ≤ 0.05) in the granulosa cells of the preovulatory (24 h after LH treatment) vs. the dominant follicle at selection. These transcripts in aged cows were related to late and inefficient 1) organization of cytoskeleton and cytoplasm, 2) differentiation, 3) lipid and cholesterol metabolism, 4) proliferation and 5) higher response to oxidative stress and free radical scavenging in the preovulatory follicles vs. the dominant follicle at selection. In conclusion, maternal age-alters the gene expression of granulosa cells of the dominant follicles during follicle development and results in a compromised follicular environment.