Induce Oocyte Maturation Research Articles

The trigger of oocyte maturation with high dosage of hCG during IVF stimulation negatively affect oocyte / embryo quality in patients with peak serum estradiol level ≥ 4,000 pg/ml

10,000 IU hCG (10K) has been routinely used for the induction of oocyte maturation during IVF cycle, but 5,000 IU (5K) may be a safer dose in high responding patients. We compared embryo development, implantation rate, and ongoing pregnancy rate/live birth rate in high responding patients who received hCG 5K vs hCG 10K in donor and non-donor IVF cycles.

Molecular characterization and analysis of a putative 5-HT receptor involved in reproduction process of the pearl oyster Pinctada fucata

5-HT (5-hydroxytryptamine; serotonin) has been linked to a variety of biological roles including gonad maturation and sequential spawning in bivalve molluscs. To gain a better understanding of the effects of 5-HT on developmental regulation in the pearl oyster Pinctada fucata, the isolation, cloning, and expression of the 5-HT receptor was investigated in this study. A full-length cDNA (2541bp) encoding a putative 5-HT receptor (5-HTpf) of 471 amino acids was isolated from the ovary of the pearl oyster. It shared 71% and 51% homology, respectively, with the Crassostrea gigas 5-HT receptor and the Aplysia californica 5-HT1ap. The 5-HTpf sequence possessed the typical characteristics of seven transmembrane domains and a long third inner loop. Phylogenetic analysis also indicated that 5-HTpf was classified into the 5-HT1 subtype together with other invertebrate 5-HT1 receptors. Quantitative RT-PCR showed that 5-HTpf is widely expressed in all tissues tested, is involved in the gametogenesis cycle, embryonic and larval development stages, and expression is induced by E2 in ovarian tissues. These results suggest that 5-HTpf is involved in the reproductive process, specifically in the induction of oocyte maturation and spawning of P. fucata.

Dual trigger with gonadotropin-releasing hormone agonist and standard dose human chorionic gonadotropin to improve oocyte maturity rates

To evaluate the percentage (%) of mature oocytes retrieved in patients with a previous history of >25% immature oocytes retrieved who were triggered with gonadotropin-releasing hormone agonist (GnRH-a) and human chorionic gonadotropin (hCG) to induce oocyte maturation. Retrospective cohort study. A university-based tertiary fertility center. Patients with a history of >25% immature oocytes retrieved in a prior invitro fertilization cycle who were triggered with GnRH-a and hCG 5,000 IU or 10,000 IU in a subsequent cycle from January 2008 through February2012. Dual trigger of GnRH-a and hCG 5,000 or 10,000 IU. Percent of mature oocytes retrieved and fertilization rate. The proportion of mature oocytes retrieved was significantly higher with a dual trigger compared with the subject's previous cycle (75.0%, interquartile range 55.6%-80.0% vs. 38.5%, interquartile range 16.7%-55.6%). The odds of a mature oocyte retrieved for patients who received a dual trigger was 2.51 times higher after controlling for stimulation protocol, hCG dose, gonadotropin dose, and oocyte retrieval time interval (odds ratio 2.51; confidence interval 1.06-5.96). The implantation, clinical, and ongoing pregnancy rates for the dual trigger were 11.8%, 26.1%, and 17.4%, respectively. In patients with a low percentage of mature oocytes retrieved who are triggered with a combination of GnRH-a and hCG, the % of mature oocytes retrieved improved. invitro fertilization outcomes, however, remain poor, suggesting an underlying oocyte dysfunction.

Kisspeptin: a novel physiological trigger for oocyte maturation in in-vitro fertilisation treatment

BackgroundAlthough in-vitro fertilisation (IVF) treatment allows infertile couples to conceive, it can result in a potentially life-threatening condition termed the ovarian hyperstimulation syndrome (OHSS). The major cause of OHSS is use of human chorionic gonadotropin (hCG) in current IVF protocols for initiating oocyte maturation. The development of a more physiological stimulus for oocyte maturation would avoid this dangerous side-effect and thus improve safety of IVF treatment. Kisspeptin is a recently identified hypothalamic hormone that acutely and potently increases endogenous secretion of luteinising hormone in a gonadotropin-releasing hormone (GnRH)-dependent manner. We aimed to investigate whether kisspeptin can induce oocyte maturation in IVF treatment. MethodsIn this single-centre prospective clinical trial at Hammersmith Hospital, London, women were recruited with the following inclusion criteria: age 18–35 years, body mass index less than 30 kg/m2, serum anti-Mullerian hormone 10–40 pmol/L, and no more than one previous IVF cycle. They underwent a recombinant follicle stimulating hormone plus GnRH antagonist IVF protocol with a single subcutaneous injection of kisspeptin. A control group was not recruited for ethical reasons. Primary outcome was production of mature oocytes (metaphase II oocytes) after egg collection. Participants and doctors giving IVF treatment were masked to the dose of kisspeptin administered. Women were independently randomised by the study statistician, initially to the lowest tier of kisspeptin doses (1·6 or 3·2 nmol/kg, n=2–3 per dose) and then as per protocol to a higher tier of doses (6·4 or 12·8 nmol/kg, n=21 per dose). Oocyte retrieval was done 36 h after kisspeptin injection. After intracytoplasmic sperm injection (ICSI) one or two embryos were transferred to the woman and pregnancy testing done 12 days later. Clinical pregnancy was confirmed on ultrasound scan at 6 weeks of gestation. Multiple means were compared by use of one-way ANOVA with post-hoc Bonferroni correction. Proportions were compared by χ2 test. All data were analysed on an intention-to-treat basis. Women gave written informed consent. The study received approval from the Hammersmith and Queen Charlottes Research Ethics Committee (application number 10/H0707/2) and the Medicines and Healthcare Products Regulatory Agency. The trial is registered with ClinicalTrials.gov, number NCT01667406. FindingsUp to Sept 1, 2013, 47 women completed the study protocol. All doses of kisspeptin resulted in a mean 9·0-fold (SD 7·5) increase in luteinising hormone release 12 h after injection. Oocyte maturation was observed at all doses of kisspeptin. 45 women (96%) had oocyte maturation (mean number of metaphase II oocytes 7·9, SD 4·1). Embryogenesis occurred in 43 women (91%) after treatment with kisspeptin (mean number of zygotes 5·7, SD 3·5). Complete pregnancy data are awaited, but up to Sept 1, 2013, 16 (36%) of 44 women had a positive pregnancy test at 12 days post embryo transfer and ten (23%) had clinical pregnancy confirmed on ultrasound examination at 6 weeks of gestation. Clinical follow-up during pregnancy is continuing, but already the first participant to have received kisspeptin to induce oocyte maturation gave birth to a healthy baby boy in May, 2013. No adverse events were noted at any time during the study. InterpretationThe results of this study suggest, for the first time, to our knowledge, that kisspeptin induces oocyte maturation in women undergoing IVF treatment. Kisspeptin might therefore offer a novel therapeutic option for fertility treatment. This small pilot study provides proof of concept that kisspeptin can stimulate oocyte maturation in women undergoing IVF treatment. Further work is now underway in a larger number of patients to determine the optimum protocol for kisspeptin administration to induce oocyte maturation in a population at high risk of OHSS. FundingUK Medical Research Council, National Institute for Health Research.

Reducing the Trigger Dose of Human Chorionic Gonadotrophin Does Not Affect Final Oocyte Maturation and Subsequently Pregnancy Outcome of Frozen-thawed Embryo Transfer

Objective To compare the efficacy of human chorionic gonadotrophin (hCG) at reduced doses of 2 000 IU and 3 000 IU for moderate or high responders with the dose of 5 000 IU in term of inducing final oocyte maturation for IVF/ICSI and the subsequent pregnancy outcome in frozen-thawed embryo transfer (FET). Methods In the retrospective cohort study, 2 166 patients undergoing IVF/ICSI with moderate or high response were recruited and classified into three groups according to the trigger dose of hCG: 2 000 IU (group A, n=722), 3 000 IU (group B, n=722) and 5 000 IU (group C, n=722). The main outcome was the proportion of mature oocytes retrieved, fertilization rates, clinical pregnancy rates, cumulative pregnancy rates and incidence of ovarian hyperstimulation syndrome (OHSS). Results No evidence of statistically difference was found in the proportion of mature oocytes retrieved (89.92%, 91.40%, 90.20%, respectively) and fertilization rate (79.8%, 80.07%, 80.51%, respectively) among groups A, B and C. Serum E 2 level on the day of hCG injection, the number of mature oocytes retrieved and good-quality embryos in group A were significantly higher than those in group B and group C. Clinical pregnancy rates per transfer cycle (45.95%, 43.97% and 44.25%), ongoing pregnancy rates (43.17%, 40.91% and 42.53%), implantation rates (30.74%, 27.78% and 29.86%) and cumulative pregnancy rates per patient (58.31%, 53.6% and 54.85%) were comparable among groups A, B and C. The incidence of OHSS in groups A, B and C (0%, 0.14% and 0.28%) was low, with no significant difference. Conclusion A reduced hCG dose of 2 000 IU for moderate or high responders leads to similar efficacy compared with a dose of 5 000 IU in inducing oocyte maturation without adversely affecting the pregnancy outcome meanwhile eliminating the risk of OHSS.

Improvement of ovulation induction by additive injection of 17,20β-Dihydroxy-4-pregnen-3-one after human chorionic gonadotropin administration in a pelagic egg spawning marine teleost, nibe croakerNibea mitsukurii(Jordan & Snyder)

This study aimed to develop the consistent ovulation induction method in a pelagic egg spawning marine teleost, nibe croaker Nibea mitsukurii. Attempts to induce oocyte maturation and ovulation in nibe croaker using human chorionic gonadotropin (hCG; 0.5 IU g−1) resulted in the normal progression of oocyte maturation and hydration, but a failure to induce ovulation in many individuals. This ovulation disorder was similarly observed even when the dose of hCG was increased 10 times (5 IU g−1) or decreased to one tenth (0.05 IU g−1), indicating that it cannot be completely overcome solely by hCG administration. However, this ovulation disorder could be completely overcome by subsequent administration of 17,20β-dihydroxy-4-pregnen-3-one (DHP) at the appropriate dose (0.5 μg g−1) and time (20 h after hCG administration). An increase in the number of individuals that ovulated due to DHP administration led to an increase in individuals producing larvae, resulting in an approximately threefold increase in the estimated number of larvae produced compared with the group of fish administered hCG alone. Thus, this ovulation induction method using DHP administration after hCG was demonstrated to overcome the ovulation disorder in nibe croaker and could be applicable to commercially important species with similar ovulation problems.

Production of Fertile Offspring from Oocytes Grown In Vitro by Nuclear Transfer in Cattle1

Because of recent advancements in reproductive technology, oocytes have attained an increasingly enriched value as a unique cell population in the production of offspring. The growing oocytes in the ovary are an immediate potential source that serve this need; however, complete oocyte growth before use is crucial. Our research objective was to create in vitro-grown (IVG) oocytes that would have the ability to perform specialized activities, including nuclear reprogramming, as an alternative to in vivo-grown oocytes. Bovine oocyte-granulosa cell complexes with a mean oocyte diameter of approximately 100 μm were cultured on Millicell membrane inserts, with culture medium supplemented with 4% polyvinylpyrrolidone (molecular weight, 360,000), 20 ng/ml androstenedione, 2 mM hypoxanthine, and 5 ng/ml bone morphogenetic protein 7. Oocyte viability after the 14-day culture period was 95%, and there was a 71% increase in oocyte volume. Upon induction of oocyte maturation, 61% of the IVG oocytes extruded a polar body. Eighty-four percent of the reconstructed IVG oocytes that used cumulus cells as donor cells underwent cleavage, and half of them became blastocysts. DNA methylation analyses of the satellite I and II regions of the blastocysts revealed a similar highly methylated status in the cloned embryos derived from in vivo-grown and IVG oocytes. Finally, one of the nine embryos reconstructed from the IVG oocytes developed into a living calf following embryo transfer. Fertility of the offspring was confirmed. In conclusion, the potential of a proportion of the IVG oocytes was comparable to that of in vivo-grown oocytes.

Trigger of oocyte maturation with low-dose human chorionic gonadotropin (hCG) reduces the risk of ovarian hyperstimulation syndrome (OHSS) and improves IVF outcome

10,000 IU hCG has been routinely used for the induction of oocyte maturation in the IVF treatment, but there is no solid scientific evidence to support whether 10,000 IU hCG is really needed to induce oocyte maturation. In this study, we compared the risk of OHSS development, oocyte maturation rate and IVF success rate in the high-risk patients after 5,000 IU hCG vs 10,000 IU hCG triggers in donor or non-donor IVF cycles.

Induction of an endogenous LH surge enhances progesterone rise and embryo yield during antagonist IVF cycles

Unlike GnRH agonist in vitro fertilization (IVF) cycles, antagonist cycles provide two options to stimulate final oocyte maturation (“trigger”): (1) recombinant hCG (recHCG) or (2) induction of an endogenous LH surge through administration of a GnRH agonist (GnRHa). The aim of this study was to compare progesterone rise, oocyte maturation, blastulation, and embryo yield in patients undergoing their first antagonist cycle who received recHCG versus GnRHa to induce oocyte maturation.

Aromatase inhibitor co-treatment with gonadotropin reduces the dose of gonadotropn and peak estradiol level without compromising IVF outcomes in high responder patients

WITHDRAWN E2 (pg/ml) on hCG day 3850 765 4272 1508 1.5 ng/ml) or low (P4 3000 pg/ml on the hCG day). Patients were stimulated with daily doses of 100-150 UI of recombinant FSH (Puregon , MSD) starting on day 2-3; GnRH antagonist was added when the leading follicle reached a diameter of 14 mm (Cetrotide, 0.25 mg/day, Merck-Serono) and a single bolus of 250 mg of rhCG (Ovitrelle, Merck-Serono) was administered to induce oocyte maturation. FERTILITY & STERILITY Chi-squared test and ANOVA were applied to detect statistical differences; p<0.05 was considered statistically significant. RESULTS: Regarding the influence of progesterone concentration on clinical outcomes in presence of high estradiol levels, we did not find significant differences in any of the clinical variables analyzed. However, we observed statistical differences in E2 and P4 levels on the day of hCG between the two groups of the study. CONCLUSION: Our data suggest that in high responder women with estradiol levels over 3000 pg/ml, progesterone levels do not affect IVF results. We confirmed the close link between E2 and P4 as we observed that high E2 levels are significant correlated with high P4 levels. These results lead us to conclude that, regardless the progesterone levels attached during the cycle in high responders, clinical outcomes are not affected. O-358 Wednesday, October 16, 2013 04:45 PM COMPARING THE EFFICACYAND SAFETY OF GnRH AGONIST (LEUPROLIDE) WITH HCG TRIGGER VERSUS GnRH ANTAGONIST (GANIRELIX)WITHTRIPTORELIN TRIGGER IN PATIENTS OFPCODUNDERGOING IVFTREATMENT. R. Sharma, F. Rehman, N. Nakshtramalini. IVF and Reproductive Medicine, Max Hospital, New Delhi, India. OBJECTIVE: To determine the ideal protocol for PCOD patients undergoing IVF treatment. DESIGN: Retrospective analysisJan 2010 –Jan 2013. MATERIALS AND METHODS: 70 PCOS patients (according to Rotterdam criteria), who underwent IVF/IVF-ICSI cycles between Jan 2010 – Jan 2013 were evaluated retrospectively for the type of protocol used, amount and duration of gonadotrophin use, cost, no.of oocytes, fertilization rate, cleavage rate,embryo quality, clinical pregnancy rate and complication rate. RESULTS: The profiles of the patients in both groupswere comparable. Cost was almost same in both groups though there was reduction in the total amount of gonadotrophins used in the antagonist cycles. There was Reduction in the treatment days with analogue in the antagonist group (average 4.5 in antagonist versus 21 days in the agonist group). Duration of stimulation (9.5 versus 11.2 days) and total gonadotrophin required (1425 versus 1680 IU) were lower in the antagonist compared with agonist protocol. Mean no. of oocytes (COC’s) were less,(16.26 in the antagonist group versus 19.45 in the agonist group) therefore mean number of fertilized oocytes and consequent no. of embryos per patient were less in the antagonist group though fertilization rate(57.17%versus 58.09%) and cleavage rate(88.53%versus89.38%) were similar. There was no evidence of a statistically significant difference in clinical pregnancy rate between the two groups (36.66%in antagonist group versus 37.5%in the agonist group). There was significantly lower incidence of OHSS in the antagonist group. In agonist group, 3 patients had severeOHSS; due towhich embryo transfer had to be postponed .In antagonist group no patient had severe OHSS. In antagonist group one patient had empty follicles at the time of Oocyte retrieval with no embryos to transfer subsequently. CONCLUSION: The use of GnRH antagonists may be considered more advantageous because of more patient comfort and decreased risk of OHSS with good clinical outcome in high responder patients. O-359 Wednesday, October 16, 2013 05:00 PM THE FIRST SUCCESSFUL LIVE BIRTH FOLLOWING PREIMPLANTATION GENETIC DIAGNOSIS FOR TYPE I CITRULLINEMIA USING POLYMERASE CHAIN REACTION. C.-H. Kim, K.-H. Lee, J.-H. Cho, J.-M. Kim, G.-H. Kim, H.-W. Yoo. Obstetrics

In vitro induction of oocyte maturation in the Japanese sea cucumber Apostichopus japonicus by cubifrin and the developmental ability of the eggs

We describe the process of cubifrin-induced in vitro oocyte maturation in the Japanese sea cucumber Apostichopus japonicus, and we propose a procedure for practical hatchery application. Oocyte maturation occurred 40–50 min after stimulation of the ovaries by cubifrin. Concurrently, eggs were released from the ovarian tubules, initially by slow peristalsis, followed by active release after about 60 min. The first and second polar bodies were released from the eggs at 90–100 min and about 140 min, respectively. Cubifrin was more effective in inducing oocyte maturation at 1–100 nM than at 100 pM. Cubifrin at 1 pM–10 nM in rearing seawater had no effect on larval development. Delayed fertilization reduced the developmental ability of eggs. Broodstock could be maintained at 15–16 °C for up to about 50 days without affecting the developmental ability of eggs. Eggs matured in vitro and in vivo were similar in their developmental ability. In conclusion, the in vitro induction of oocyte maturation is effective in Japanese sea cucumber and represents a viable alternative to the induction of spawning by cubifrin injection.

In vitro induction of oocyte maturation and steroidogenesis by gonadotropins, insulin, calcitonin and growth factor in an estuarine flat head grey mullet, Mugil cephalus L.

In this article, an in vitro investigation was carried out to ascertain the roles of hormones and growth factor in the inductions of oocyte maturation and steroidogenesis of the postvitellogenic follicles in an Indian estuarine grey mullet, Mugil cephalus L. Oocyte maturation was evaluated by scoring the germinal vesicle breakdown (GVBD) percent of the postvitellogenic follicles. All the sex [17α,20β-dihydroxy-4-pregnane-3-one (DHP), estradiol 17β (E₂), progesterone (P), 17α-OH progesterone (17-OH-P) and testosterone] and other [bovine-insulin and salmon-calcitonin, human chorionic gonadotropin (hCG), luteinizing hormone (LH) or hCG+DHP] hormones and insulin-like growth factor-I (IGF-I) significantly increased GVBD% in 9 h culture. DHP had a maximum effect (75 %) compared to other effectors. Some effectors (hCG: 82.14 %, LH: 78.94 %, hCG plus DHP: 81.81 %, E₂: 80 % and IGF-I: 74.19 %) including DHP (79 %) further increased GVBD% in 15-h culture. All the hormones (except DHP) and IGF-I increased DHP, E₂ and testosterone productions by the postvitellogenic ovarian follicles in vitro. DHP and testosterone productions were increased with the increase of incubation time from 9 h through 15 h. E₂ production was not further increased beyond 12 h. DHP production was highest by hCG compared to other effectors. The hCG of all the test compounds was most effective in both the induction of GVBD% and steroid production. DHP is the most potent inducer of oocyte maturation in Indian estuarine flat head grey mullet. Involvement of estrogen in mullet oocyte maturation is indicated. hCG, like DHP, is equally potent and induces oocyte maturation via DHP production in vitro. hCG with DHP has synergistic action on oocyte maturation in mullet ovary. Interplay of several hormones (hCG, LH, and probably E₂ and testosterone) and IGF-I on oocyte maturation is suggested in the mullet.

INSULIN ASPART CAN LEAD MEIOTIC RESUMPTION IN CATFISH, Clarias Grapienus OOCYTES

This study directly tested the hypothesis that the induction of oocyte maturation on catfish Clariasgrapienusby insulin aspartresulted in a transient decrease of oocyte cyclic AMP (cAMP) level due to the increase ofphosphodiesterase (PDE) activity. Under the influence of insulin the germinal vesicle (GV) of the oocyte migrated towards the animal pole (GVM), reached the micropyle, and then dissolved (GVBD). By using different concentrations of insulin aspart i.e., 0 U/ml, 0.01 U/ml, 0.1 U/ml, 1 U/ml and 10 U/ml, optimum amount required was found to be 1U/ml. The number of oocytes incubated for 24 hours with insulin aspartic treatment that undergo meiotic re-initiation can reach the percentage of meiotic resumption oocytes was 82%. The results suggest that catfish oocytes have responsive to sensitive adenylatecyclase and also an active phosphodiesterase system. Addition of the maturation-inducing steroid, a progesterone, stimulated (P&lt;0.01) GVBD of catfish oocytes and its combination with insulin aspart showed an additive effect. This information will be important in hormonal manipulation during induced breeding of Indonesian commercial catfish.

Involvement of PLA2, COX and LOX inRhinella arenarumoocyte maturation

In Rhinella arenarum, progesterone is the physiological nuclear maturation inducer that interacts with the oocyte surface and starts a cascade of events that leads to germinal vesicle breakdown (GVBD). Polyunsaturated fatty acids and their metabolites produced through cyclooxygenase (COX) and lipoxygenase (LOX) pathways play an important role in reproductive processes. In amphibians, to date, the role of arachidonic acid (AA) metabolites in progesterone (P4)-induced oocyte maturation has not been clarified. In this work we studied the participation of three enzymes involved in AA metabolism - phospholipase A2 (PLA2), COX and LOX in Rhinella arenarum oocyte maturation. PLA2 activation induced maturation in Rhinella arenarum oocytes in a dose-dependent manner. Oocytes when treated with 0.08 μM melittin showed the highest response (78 ± 6% GVBD). In follicles, PLA2 activation did not significantly induce maturation at the assayed doses (12 ± 3% GVBD). PLA2 inhibition with quinacrine prevented melittin-induced GVBD in a dose-dependent manner, however PLA2 inactivation did not affect P4-induced maturation. This finding suggests that PLA2 is not the only phospholipase involved in P4-induced maturation in this species. P4-induced oocyte maturation was inhibited by the COX inhibitors indomethacin and rofecoxib (65 ± 3% and 63 ± 3% GVBD, respectively), although COX activity was never blocked by their addition. Follicles showed a similar response following the addition of these inhibitors. Participation of LOX metabolites in maturation seems to be correlated with seasonal variation in ovarian response to P4. During the February to June period (low P4 response), LOX inhibition by nordihydroguaiaretic acid or lysine clonixinate increased maturation by up to 70%. In contrast, during the July to January period (high P4 response), LOX inhibition had no effect on hormone-induced maturation.

The Relationship between Body Mass Index and Quantitative Serum HCG Levels Following HCG Trigger in In Vitro Fertilization Cycles

During controlled ovarian hyperstimulation (COH) for in vitro fertilization (IVF), human chorionic gonadotropin (HCG) is administered to induce oocyte maturation approximately 34-36 hours before oocyte aspiration. One method to ensure the proper administration and absorption of HCG before aspiration is to measure serum HCG levels one day following HCG dosing. Two small studies have suggested that HCG may vary with body mass index (BMI), but the clinical significance has not been well described (1, 2).

Water temperature manipulation can induce oocyte maturation and ovulation in the common Japanese conger, Conger myriaster

To develop seed production techniques for aquaculture of the common Japanese conger, Conger myriaster, we attempted to induce ovarian maturation and ovulation without exogenous hormone treatment and examined the changes in steroid hormones to understand the endocrine condition of the fish during their maturation process. Juveniles caught in November 2001 (20 to 30cm in total length) were reared for 1.5years in seawater at temperatures ranging from 10 to 20°C before the experiments. Then, 141 randomly selected female eels (58 to 82cm in total length) were maintained at a water temperature of 6°C from April 2003 to March 2004. The initial GSI values (5.0 to 12.0) and oocyte diameters (300 to 400μm) recorded in April 2003 had increased to maximum values of 30.0 and 650μm, respectively, at the end of experiment, in March 2004. In late December 2003, 12 fish with oocytes measuring over 600μm in diameter were selected and divided into an elevated temperature group from 6 to 10°C and a control group of 6°C for 43days. The mean oocyte diameter in the elevated temperature group increased significantly, and one fish (estimated GSI, 47.0) ovulated eggs with an average diameter of 995±46μm on the 41st day at a water temperature of 10°C, in contrast, the control group showed no significant changes in oocyte diameter, and no fish ovulated. Plasma T levels, examined via ELISA, increased with oocyte maturation, with the highest level detected at the migratory nucleus stage, and declined after ovulation. In contrast, plasma E2 was maintained at high levels during the vitellogenic stage, exhibiting the highest value after ovulation. The present study demonstrates that ovarian maturation and ovulation in the common Japanese conger could be achieved through water temperature manipulation.

Neuropeptides trigger oocyte maturation and subsequent spawning in the hydrozoan jellyfish Cytaeis uchidae

Oocyte maturation and subsequent spawning in hydrozoan jellyfish are generally triggered by light-dark cycles. To examine if the initiation of the maturation process after light stimulus is mediated by neurotransmitters, neuropeptides isolated originally from Hydra magnipapillata were applied to sexually mature female medusae of the hydrozoan jellyfish Cytaeis uchidae. Among the Hydra neuropeptides tested, Hym-53 (NPYPGLW-NH2 ), as well as a nonphysiological peptide, CGLWamide (CGLW-NH2 ), were most effective in inducing oocyte maturation and spawning. Hym-355 (FPQSFLPRG-NH2 ) also triggered these events, but the stimulatory effect was weaker. Since Hym-53-OH (NPYPGLW) and Hym-355-OH (FPQSFLPRG) had no effect, amidation at the C-terminus may be critical for the stimulatory activities of the peptides. Exposure to Hym-53 for 2 min was sufficient to trigger of oocyte maturation, and the spawned eggs were able to be fertilized and to develop normally. Transmission electron microscopy confirmed that bundles of axon-like structures that contain dense-core synaptic vesicles and microtubules are present in the ovarian ectodermal epithelium overlying the oocytes. In addition, immunohistological analyses revealed that some of the neurons in the ectodermal epithelium are GLWamide- and PRGamide-positive. These results suggest that a neuropeptide signal transduction pathway is involved in mediating the induction of oocyte maturation and spawning in this jellyfish.

Spatial Distribution and Receptor Specificity of Zebrafish Kit System - Evidence for a Kit-Mediated Bi-Directional Communication System in the Preovulatory Ovarian Follicle

Consisting of Kit ligand and receptor Kit, the Kit system is involved in regulating many ovarian functions such as follicle activation, granulosa cell proliferation, and oocyte growth and maturation. In mammals, Kit ligand is derived from the granulosa cells and Kit receptor is expressed in the oocyte and theca cells. In the zebrafish, the Kit system contains two ligands (Kitlga and Kitlgb) and two receptors (Kita and Kitb). Interestingly, Kitlga and Kitb are localized in the somatic follicle cells, but Kitlgb and Kita are expressed in the oocyte. Using recombinant zebrafish Kitlga and Kitlgb, we demonstrated that Kitlga preferentially activated Kita whereas Kitlgb specifically activated Kitb by Western analysis for receptor phosphorylation. In support of this, Kitlgb triggered a stronger and longer MAPK phosphorylation in follicle cells than Kitlga, whereas Kitlga but not Kitlgb activated MAPK in the denuded oocytes, in agreement with the distribution of Kita and Kitb in the follicle and their specificity for Kitlga and Kitlgb. Further analysis of the interaction between Kit ligands and receptors by homology modeling showed that Kitlga-Kita and Kitlgb-Kitb both have more stable electrostatic interaction than Kitlgb-Kita or Kitlga-Kitb. A functional study of Kit involvement in final oocyte maturation showed that Kitlga and Kitlgb both suppressed the spontaneous maturation significantly; in contrast, Kitlgb but not Kitlga significantly promoted 17α, 20β-dihydroxy-4-pregnen-3-one (DHP) -induced oocyte maturation. Our results provided strong evidence for a Kit-mediated bi-directional communication system in the zebrafish ovarian follicle, which could be part of the complex interplay between the oocyte and the follicle cells in the development of follicles.

Characterization of luteinizing hormone and luteinizing hormone receptor and their indispensable role in the ovulatory process of the medaka.

The molecular properties and roles of luteinizing hormone (Lh) and its receptor (Lhcgrbb) have not been studied for the medaka (Oryzias latipes), which is an excellent animal model for ovulation studies. Here, we characterized the medaka Lh/Lhcgrbb system, with attention to its involvement in the ovulatory process of this teleost fish. In the medaka ovary, follicle-stimulating hormone receptor mRNA was expressed in small and medium-sized follicles, while lhcgrbb mRNA was expressed in the follicle layers of all growing follicles. Experiments using HEK 293T cells expressing medaka Lhcgrbb in vitro revealed that gonadotropin from pregnant mare’s serum and medaka recombinant Lh (rLh) bound to the fish Lhcgrbb. The fish gonadotropin subunits Gtha, Fshb, and Lhb were essentially expressed at fairly constant levels in the pituitary of the fish during a 24-h spawning cycle. Using medaka rLh, we developed a follicle culture system that allowed us to follow the whole process of oocyte maturation and ovulation in vitro. This follicle culture method enabled us to determine that the Lh surge for the preovulatory follicle occurred in vivo between 19 and 15 h before ovulation. The present study also showed that oocyte maturation and ovulation were delayed several hours in vitro compared with in vivo. Treatment of large follicles with medaka rLh in vitro significantly increased the expression of Mmp15, which was previously demonstrated to be crucial for ovulation in the fish. These findings demonstrate that Lh/Lhcgrbb is critically involved in the induction of oocyte maturation and ovulation.

Predicting successful induction of oocyte maturation after gonadotropin-releasing hormone agonist (GnRHa) trigger

Are there factors predicting the number of total and mature oocytes retrieved after controlled ovarian hyperstimulation (COH) utilizing a gonadotropin-releasing hormone (GnRH) antagonist protocol and a GnRH agonist (GnRHa) to induce oocyte maturation? Peak estradiol (E₂) level, post-trigger LH and progesterone and the magnitude of LH rise are independent predictors of the total number of oocytes and mature oocytes retrieved. Despite multiple follicular development in high responders, oocyte retrieval after a GnRHa trigger in a small subset of patients fails to obtain a substantial number of total oocytes or mature oocytes. A retrospective chart review of all autologous and oocyte donation cycles utilizing a GnRHa antagonist protocol where GnRHa was used for the induction of oocyte maturation between 1 April 2003 and 31 December 2011. A total of 508 autologous and donor IVF/ICSI cycles utilizing a GnRH antagonist protocol for COH and GnRHa for the induction of oocyte maturation at a university-based tertiary fertility center. Peak E₂ on the day of trigger (r = 0.19, P < 0.001), post-trigger LH (r = 0.12, P = 0.009) and progesterone (r = 0.47, P < 001) and LH rise (r = 0.18, P < 0.001) all positively correlated with the number of total and mature oocytes retrieved. The true incidence of empty follicle syndrome was 1.4% (7/508). There was no post-trigger LH or progesterone cut-off value for the prediction of oocyte yield. However, all cases of empty follicle syndrome occurred in patients with post-trigger LH <15 IU/l and P ≤ 3.5 ng/ml. The findings of this study may also be due to chance since it was a retrospective study and not prospectively designed. This is a retrospective chart review and therefore subject to bias. Serum hormone measurements were performed between 8 and 12 h after GnRHa trigger rather than a standardized time period following trigger administration. Therefore, peak levels of LH may have been missed due to the short ascending limb of LH rise lasting approximately 4 h after GnRHa trigger. The results of this study can be generalized to high responders utilizing a GnRH antagonist protocol for COH and a GnRHa for the induction of oocyte maturation. The use of alternative stimulation regimens or medications will limit the ability to generalize the results of this study to other populations. This study was not funded, and there are no conflicts of interest. n/a.