Attila Török

Measurement of Noradrenaline, Dopamine and Serotonin Contents in Follicular Fluid of Human Graafian Follicles after Superovulation Treatment

We measured the noradrenaline (NA), serotonin (5-HT) and dopamine (DA) contents of 47 normally maturated and 16 cystically degenerated follicular fluid samples obtained from patients involved in the in vitro fertilization and gamete transfer program. The patients were given human menopausal gonadotropin (HMG), as a superovulation treatment, and 7,500 IU human chorionic gonadotropin (HCG) to induce ovulation 34-36 h prior to the follicular puncture done by laparoscope. The NA content of the normally developed follicles was 11.4 + 8.4 micrograms/100 ml on average. For cystically degenerated follicles, the following data were obtained: 1.1 + 0.7 micrograms/100 ml (p less than 0.001). 5-HT and DA contents in the preovulatory follicles are 14.3 +/- 8.9 and 19.3 +/- 8.2 micrograms/100 ml, respectively; at the same time, 5-HT and DA contents in the cystically degenerated follicles were 12.2 +/- 6.2 and 12.7 +/- 6.8 micrograms/100 ml, respectively. They suggest that the higher amount of NA in the follicular fluid might play an important role in the mechanism of ovulation, the regulation of postovulatory tubal motility and the release of progesterone from granulosa cells.

Modulatory Effect of Acetylcholine on Gonadotropin-Stimulated Human Granulosa Cell Steroid Secretion

The aim of this study was to explore the direct action of acetylcholine on gonadotropin-stimulated progesterone (P) and estradiol (E2) secretion of human granulosa cells (GCs) cultured in serum-free medium. Human GCs were isolated from preovulatory follicular fluid aspirated from 22 women undergoing in vitro fertilization at the University Women’s Hospital of Tübingen. The production of progesterone and E2 was measured in the presence and absence of acetylcholine, carbachol, atropine, luteinizing hormone (LH) or follicle-stimulating hormone (FSH) using radioimmunoassay. Statistical analysis of the data was performed by ANOVA and Newman-Keuls test. Administration of acetylcholine or carbachol (10–5M) resulted in a significant increase in P and E2 secretion. This response was specifically blocked by the muscarinic receptor antagonist atropine. Similarly, carbachol resulted in a significant increase in P and E2 output, though the response to it was somewhat reduced when compared to that evoked by acetylcholine. Acetylcholine did not show any additive effect on LH-stimulated P secretion, while it augmented the stimulatory effect of FSH on P release. In contrast, carbachol markedly diminished the stimulatory effect of LH on P secretion, while it caused no change in FSH-induced P output. When administered together, acetylcholine did not modify the stimulatory effect of FSH on E2 secretion, however, it markedly elevated LH-induced E2 output. Similar to this, carbachol significantly increased LH-induced E2 release, however it decreased FSH-stimulated E2 secretion. We suggest that acetylcholine has a direct modulatory effect on gonadotropin-stimulated steroid production of GCs, an effect that is mediated via muscarinic receptors. This effect may have a physiological role in the regulation of GC function during the menstrual cycle.

Influence of melatonin on basal and gonadotropin-stimulated progesterone and estradiol secretion of cultured human granulosa cells and in the superfused granulosa cell system

The aim of this study was to explore the direct action of melatonin (Me) on basal and gonadotropin-stimulated progesterone (PG) and estradiol (E2) secretion of human granulosa cells (GCs) cultured in serum-free medium and in a superfused GC system. Human GCs were isolated from preovulatory follicular fluid aspirated from 34 women undergoing in vitro fertilization at the University Women’s Hospital of Tübingen. PG and E2 production was measured in the presence and absence of Me, propranolol, LH or FSH using radioimmunoassay. Statistical analysis of the data was performed by ANOVA and Newman-Keuls test. Me stimulated E2 secretion in a dose-dependent manner. Propranolol did not cause any change in E2 secretion, and when given with Me, it only partially blocked but could not entirely prevent E2 output. There was no statistically significant effect of Me on PG production when Me was administered at concentrations between 10–4 and 10–8 M. However, at 10–3 M Me significantly suppressed PG output of granulosa cells. LH and FSH significantly stimulated the secretion of both steroid hormones. Me significantly reduced LH- and FSH-induced E2 secretion, as well as LH-stimulated PG output, while it caused only a slight, yet significant decrease in PG secretion. In the superfused GC system, FSH and LH resulted in a significant stimulatory effect on PG release. Me did not modify the stimulatory effect of FSH on PG, while it caused some delay in LH-stimulated PG release. Propranolol and Me had no stimulatory effect on PG release. On the basis of our results we suggest that Me has a direct modulatory effect on basal E2 and gonadotropin-stimulated E2 and PG secretion of human GCs. The observed effect may play a physiological role in the regulation of GC function during the menstrual cycle.

Relationship between the Monoamine, Progesterone and Estradiol Content in Follicular Fluid of Preovulatory Graafian Follicles after Superovulation Treatment

We measured the progesterone, estradiol (E2), serotonin (5-HT), noradrenaline (NA) and dopamine (DA) contents of follicular fluid (FF) samples obtained from 35 patients undergoing in vitro fertilization and embryo transfer. Progesterone and E2 were determined by radioimmunoassay, and monoamines were measured by a spectrofluorimetric method. Significantly higher progesterone and NA levels were found (p < 0.01) in FF from cycles in which the oocyte cleaved and resulted in pregnancy compared to FF containing uncleaved oocytes. NA and DA contents were significantly higher in FF from cycles resulting in pregnancy than in FF containing cleaved oocytes not resulting in pregnancy (p < 0.01). There were significantly lower NA and 5-HT levels in FF containing uncleaved oocytes compared to the latter group (p < 0.05). Significant positive correlations were found between progesterone and 5-HT (r = 0.67) and between NA and DA (r = 0.93) in the pregnant group; between progesterone and E2 (r = 0.84) in FF containing uncleaved oocytes; between progesterone and E2 (r = 0.71) and between NA and DA (r = 0.62) in FF from cycles in which the oocyte cleaved but did not result in pregnancy. The results suggest that the follicular hormonal changes associated with oocyte maturation may be locally modulated by monoamines.

Scavenger Capacity of Follicular Fluid, Decidua and Culture Medium with Regard to Assisted Reproduction: An in vitro Study Using Electron Paramagnetic Resonance Spectroscopy

The natural scavenger capacity of follicular fluid of women pre-treated for in vitro fertilization (IVF) embryo transfer, Ham’s F10 nutrient mixture used for oocyte culture and endometrium samples were studied in a hydroxyl free radical generating system, using electron paramagnetic resonance (EPR) spectroscopy. The EPR signals appearing after the addition of samples to the reaction mixture show a diversely decreasing production of phenyl-tertier-butylnitrone spin adducts. Presumably, the follicular fluid and endometrium samples contain active factors which function as radical scavengers. The findings suggest that appropriate augmentative antioxidative therapy might be advised for IVF patients.

Organic Hydroperoxide-Induced Chemiluminescence of Follicular Fluid and Blood Serum Samples Obtained from Women Pretreated for in vitro Fertilization

The organic hydroperoxide-induced chemiluminescence of follicular fluid obtained from in vitro fertilized patients and its differently separated fractions were evaluated. Peroxidative stress causes a different photo-emission in the samples which alludes to some factors playing a role in the maintenance of the pro-oxidant/antioxidant balance. Interactions between the protein compounds of the samples and the organic hydroperoxide associate with formation of excited species contributing to the distinctive light emission processes. The technique offers a special re-interpretation of the scavenger state relating to the components of follicular fluid.

Significant differences in capillary electrophoretic patterns of follicular fluids and sera from women pre-treated for in vitro fertilization

Human ovarian follicular fluids and sera obtained from women pre-treated for in vitro fertilization (IVF) were investigated by capillary zone electrophoresis. Comparison of the matching physiological liquids showed substantial differences in the electrophoretic patterns. Significant decrease in the alpha(1)- and gamma-fractions of follicular fluids of every woman were observed, whereas other fractions of the samples did not show such alterations. Since follicular fluid is a product of both, secretion by granulosa cells and diffusion from the theca capillaries, we can assume that the forced production of follicular fluid upon hormone stimulation (with gonadotropin releasing hormone (GnRH), follicle stimulating hormone (FSH) and corionic gonadotroph hormone (hCG)) may play role in the uneven presence of the proteins.

Screening in Pregnancy with Unconjugated Serum Estriol Compared with Urinary Estriol

The diagnostic value of estriol determinations in high-risk pregnancies is frequently discussed in the literature. Estriol levels of urine and serum samples of 48 asymptomatic pregnant women were analyzed using direct radioimmunoassay. A correlation was found (r = 0.5997) between urine and serum samples, nevertheless the correlation was stronger (r = 0.8278) if 4 cases with extreme deviation were omitted. In these cases the serum estriol levels were high and the urine estriol levels were low. The outcome of the pregnancies was without any complication. Our observations suggest that determination of unconjugated serum estriol levels is more reliable in the monitoring of high-risk pregnancies, since they do not depend on renal and hepatic functions.