Mutations in the luteinizing hormone/chorionic gonadotropin receptors (LH/CGRs), representatives of the G protein-coupled receptor family, have been rapidly identified over the last 20 years. This review aims to compare and analyze the data reported the activating and inactivating mutations of the LH/CGRs between human, rat, equine and fish, specifically (Japanese eel Anguilla japonica). Insights obtained through detailed study of these naturally-occurring mutations provide a further update of structure-function relationship of these receptors. Specifically, we present a variety of data on eel LH/CGR. These results provide important information about LH/CGR function in fish and the regulation of mutations of the highly conserved amino acids in glycoprotein hormone receptors.

Equine chorionic gonadotropin (eCG) is a heavily glycosylated glycoprotein composed of non-covalently linked α- and β-subunits. To study the function and signal transduction of tethered recombinant-eCG (rec-eCG), a single chain eCG molecule was constructed, and the rec-eCG protein was prepared. In this study, we constructed 5 mutants (Δ1, Δ2, Δ3, Δ4, and Δ5) of rec-eCG using data about known glycoprotein hormones to analyze the role of specific follicle stimulating homone (FSH)-like activity. Three amino acids of certain specific sites were replaced with alanine. The expression vectors were transfected into CHO cells and subjected to G418 selection for 2～3 weeks. The media were collected and the quantity of secreted tethered rec-eCGs was quantified by ELISA. The LH- and FSH-like activities were assayed in terms of cAMP production by rat LH/CG and rat FSH receptors. Then, the metabolic clearance rate analyzed by the injection of rec-eCG (5 IU) into the tail vein was analyzed. The mutant eCGs (Δ1, Δ4, and Δ5) were transcripted, but not translated into proteins. Rec-eCG Δ2 was secreted in much lower amounts than the wild type. Only the rec-eCG Δ3 (β-subunit: Gln94-Ile95-Lys96→Ala94-Ala95-Ala96) was efficiently secreted. Although activity is low, its LH-like activity was similar to that of tethered eCGβα. However, the FSH-like activity of rec-eCGβαΔ3 was completely flat. The result of the analysis of the metabolic clearance rate shoed the persistence of the mutant in the blood until 4 hours after the injection. After then, it almost disappeared at 8 hours. Taken together, these data suggest that 94～96 amino acid sequences in eCG β-subunit appear to be of utmost importance for signal transduction of the FSH receptor.

The objective of this study was to examine the effect of eCG and various concentrations (20, 40, and 80 ) of porcine FSH on nuclear maturation and intracellular glutathione (GSH) level of oocytes, and embryonic development after parthenogenetic activation (PA) and somatic cell nuclear transfer (SCNT) in pigs. Immature pig oocytes were matured in TCM-199 supplemented with porcine follicular fluid, cysteine, pyruvate, EGF, insulin, and hormones (10 IU/ml hCG and 10 IU/ml eCG or FSH) for the first 22 h and then further cultured in hormone-tree medium for an additional 22 h. Nuclear maturation of oocytes () was not influencem foreCG and various concentrations FSH. Embryonic development to the cleavage stage () and mean number of cells in blastocyst ( cells) after PA were not altered but blastocyst formation e-treignificaddlor(p<0.05) improvem forthe supplementation eith 80 FSHr(64%) compared to 47%, io8%, iand 47% in oocytes that were treated with eCG, 20,i and 40 FSH,i numectivelo. In SCNT, fusion () of cell-cytoplast couplets and siosequent embryo cleavage () were not influencem fordifferent gonadotropins but blastocyst formation tended to increase forthe supplementation eith 80 FSHr(25% vs. ). Our nuults demonstrated that oocyte maturation and embryonic development after PA and SCNT e-frinfluencem fortype of gcem fortype of gits concentration. In this study, supplementation of maturation medium eith 80 FSHrimproved preimplantation development of PA and SCNT pig embryos, probably by increasing intracellular GSH concentration of matured oocytes.

Ovarian cancer is a significant cause of cancer-related death in women, but the main biological causes remain open questions. Hormonal factors have been considered to be an important determinant causing ovarian cancer. Recent studies have shown that gonadotropin-releasing hormone (GnRH)-I and its analogs have clinically therapeutic value in the treatment of ovarian cancer. In addition, numerous studies have shown that the potential of GnRH-II in normal reproductive system or reproductive disorder. GnRH-I receptors have been detected in approximately 80% of ovarian cancer biopsy specimens as well as normal ovarian epithelial cells and immortalized ovarian surface epithelium cells. GnRH-II receptors have also been found to be more widely expressed than GnRH-I receptors in mammals, suggesting that GnRH receptors may have additional functions in reproductive system including ovarian cancer. The signal transduction pathway following the binding of GnRH to GnRH receptor has been extensively studied. The activation of protein kinase A/C (PKA/PKC) pathway is involved in the GnRH-I induced anti-proliferative effect in ovarian cancer cells. In addition, GnRH-I induced mitogen-activated protein kinase (MAPK) activation plays a role in anti-proliferative effect and apoptosis in ovarian cancer cells and the activation of transcriptional factors related to cellular responses. However, the role of GnRH-I and II receptors, there are discrepancies between previous reports. In this review, the role of GnRH in ovarian cancer and the mechanisms to induce anti-proliferation were evaluated.

This study examined pregnancy and fetal loss rates according to different estrus synchronization protocols and injection of gonadotropin releasing hormone (GnRH) after transfer of Korean Native Cattle embryos to Holstein recipients. In Experiment 1, recipients received no treatment (Control, n = 119); two injections of prostaglandin ( ) 11 days apart (PGF group, n = 120); GnRH (day 0)- (day 7)-GnRH (day 9) (Ovsynch group, n = 120); and CIDR (day 0)- and CIDR removal (day 7)-GnRH (day 9) (CIDR group, n = 110). In Experiment 2, the control group was received no treatment of GnRH. The treatment groups were received GnRH at embryo transfer (ET) (day 0), 7 days later, 14 days later, ET and 7 days later, 7 and 14 days later, or ET, 7 and 14 days later. Recipients were assigned to treatment randomly and received two in vitro produced blastocysts. Pregnancy was diagnosed at day 60 by palpation per rectum. Fetal loss to term was determined by palpation every 90 days thereafter. In Experiment 1, the pregnancy rate in the CIDR group (59.1%) were higher than in the Control group (42.0%) (p<0.01); fetal loss rates were similar for all groups (12.0 to 18.5%). In Experiment 2, the pregnancy rate in Day 0+7+14 group was higher (60.2%) than the control (40.2%) (p<0.01) and resulted in a lower fetal loss (p<0.05) than the control (4.6 vs. 11.4%). There were no significant difference between other treatment and the control (p>0.05). These results show that pregnancy rates of bovine embryos can be enhanced by CIDR insertion or GnRH treatment. Additionally, fetal loss may be reduced with GnRH treatment after ET.

본 연구에서는 GnRH가 과배란 처치된 래트의 초기 난포기와 후기 난포기에서 난소기능에 어떠한 영향을 미치는지를 이해하기 위해서, 30IU PMSG와 10IU hCG로 전처치된 미성숙 래트에 있어서 배란반응, 배란 난자의 형태학적 이상 유무 및 핵 성숙도, 난소 중량, 난소의 조직학적인 변화 및 혈중 스테로이드 호르몬 (17-estradiol, progesterone 및 testosterone) 농도에 대하여 GnRH agonist의 효과를 검사하였다.

The objective of this study was to enhance the pregnancy rate of repeat-breeder Hanwoo with gonadotropin-releasing hormone(Gn-RH) at the time, dose and site of administration.The results obtained were summaried as fallows:1.Ovulation time and pregnancy rate following GnRH administration time was 46.0, 27.4, 42.0 and 43.2hr and 33.3, 57.1, 37.5 and 40.0% at non-treatment, estus, 1st A' and 2nd Al treatment, respectively.2. Ovulation in repeat-breeder was induced 100% within 24hr with GnRH administration at the time of estrus.3. Ovulation time and pregnancy rate following GnRH adminstration dose and site was 25.2, 32.6, 17.6 and 27.6hr, and 28.6, 42.9, 75.0 and 66.7% at 50g+IU, 50g+IM, 100g+IU and 100g+IM treatments, respectively. It is concluded that GnRH administration for repeat-breeder was enhanced the pregnancy rate when treated with 100g intrauterine at the time of estrus.

미성숙 래트의 외경정맥에 카테타를 장착하고, 다음날 (28일령) 대조군에는 4IU, 다배란 처치 군에는 20IU의 PMSG를 피하주사하였다. 각 실험동물은 혈중의 LH농도 변화를 측정하기 위하여 PMSG 투여 직전 (0시간), 투여후 12시간, 그 이후 6시간 간격으로 혈액을 채취하고 72시간에 희생시켰다. 그 결과 다배란 용량의 PMSG 투여는 먼저 배란반응 및 난소중량을 대조군에 비하여 각각 4,7배 및 2.1배나 현저하게 (P<0.05) 증가시켰다

Central precocious puberty (CPP) is less common in boys than girls; very little data is reported on effect of gonadotropin-releasing hormone analog (GnRHa) treatment in boys with CPP. The aim of the study was to evaluate growth changes in boys with CPP and early puberty (EP) treated with GnRHa therapy for 1 year. In 60 (39 CPP and 21 EP) boys with confirmed diagnosis of CPP and EP, auxological [height, height standard deviation score (Ht SDS), bone age (BA), Predicted adult height (PAH)] and endocrinological parameters were obtained at baseline, at 6 months, and at 1 year after GnRHa treatment in boys with CPP and EP. During the treatment a decline in Ht SDS and growth velocity, luteinizing hormone (LH), follicle stimulating hormone (FSH) and testosterone levels were observed (p＜0.01); and a deceleration in the maturation of bones after 1 year GnRHa treatment was observed (p＜0.01). PAH SDS was increased during treatment with GnRHa (p＜0.01). There was significant difference in PAH SDS between organic CPP and non-organic CPP (p＜0.05). The present data indicate that GnRHa therapy significantly improves growth prognosis in boys with CPP and EP.

GnRH (gonadotropin-releasing hormone) is a supreme hormone regulating reproductive activity in most animals. The sequences of amino acid and nucleic acid of GnRH reported up to now are examined from the evolutionary framework of Chordata. All identified GnRH are classified into GnRH1, GnRH2, or GnRH3. In all three forms of GnRH both N-terminal and C-terminal are conserved, which allows for effective binding to their receptors. The three amino acids in the middle of GnRH1 sequence have altered diversely from the primitive Chordata, which is indicative of the adaptation process to the ambient environment. GnRH2 and GnRH3 sequences are well conserved. There are more diverse modifications in the nucleic acids than in amino acid sequence of GnRH1. These variations can result from meiosis, mutation, or epigenetics and indicate that GnRH is the product of natural selection.

Previous studies showed that recombinant equine chorionic gonadotropin (rec-eCGβ/α) exhibits both folliclestimulating hormone (FSH) and luteinizing hormone (LH)-like activities in rat LHR- and FSHR-expressing cells. In this study, we analyzed signal transduction by eelFSHR and eelLHR upon stimulation with rec-eCGβ/α and native eCG. The cyclic adenosine monophosphate (cAMP) stimulation in CHO-K1 cells expressing eelLHR was determined upon exposure to different doses (0–1,450 ng/mL) of rec-eCGβ/α and native eCG. The EC50 values of rec-eCGβ/α and native eCG were 172.4 and 786.6 ng/mL, respectively. The activity of rec-eCGβ/α was higher than that of native eCG. However, signal transduction in the CHO PathHunter Parental cells expressing eelFSHR was not enhanced by stimulation with both agonist rec-eCGβ/α and native eCG. We concluded that rec-eCGβ/α and native eCG were completely active in cells expressing eelLHR, similar to the activity in the mammalian cells expressing LHRs. However, rec-eCGβ/α and native eCG did not invoke any signaling response in the cells expressing eelFSHR. These results suggest that eCG has a potent activity in cells expressing eelLHR. Thus, we also suggest that rec-eCGβ/α can induce eel maturation by administering gonadotropic reagents (LH), such as salmon pituitary extract.

Gonadotropin-releasing hormone (GnRH) is a key neuropeptide regulating reproduction in humans and other vertebrates. Here, we evaluated the reproductive control system mediated by GnRH in the Pacific abalone Haliotis discus hannai. We cloned a prepro-GnRH cDNA (Hdh-GnRH) from the pleural-pedal ganglion (PPG) in H. discus hannai, and analyzed its spatiotemporal gene expression pattern. The open reading frame of Hdh-GnRH encodes a protein of 101 amino acids, consisting of a signal peptide, a GnRH dodecapeptide, a cleavage site, and a GnRH-associated peptide. This structure and sequence are highly similar to GnRH-like peptides reported for mollusks and other invertebrates. Quantitative polymerase chain reaction demonstrated that Hdh-GnRH mRNA was more strongly expressed in the ganglions (PPG and cerebral ganglion [CG]) than in other tissues (gonads, gills, intestine, hemocytes, muscle, and mantle) in both sexes. In females, the expression levels of Hdh-GnRH mRNA in the PPG and branchial ganglion (BG) were significantly higher at the ripe and partial spent stages than at the early and late active stages. In males, Hdh-GnRH mRNA levels in the BG showed a significant increase in the partial spent stage. Unexpectedly, Hdh-GnRH levels in the CG were not significantly different among the examined stages in both sexes. These results suggest that Hdh-GnRH mRNA expression profiles in the BG and possibly the PPG are tightly correlated with abalone reproductive activities.

GnRH는 국부적으로 난소에서 합성되며, 난소내 과립 및 황체세포에 직접적으로 작용하여 난소의 기능을 조절하는 것으로 알려져 있으며, 특히, GnRH는 난소내 과립-황체화 세포의 세포자연사를 유도하는 것으로 보고하고 있다. 그러나 GnRH에 의한 세포자연사가 FSH에 의해 회복될 수 있는지는 명확히 밝혀져 있지 않다. 따라서 본 실험에서 난자 채취시 획득한 사람 과립-황체화 세포를 배양한 후 5, 50, 100 ng/ GnRH와 1 IU/ FSH를 처리

연어과 어류인 열목어 재조합 생식선자극호르몬(r-mtFSH 또는 r-mtLH) 투여에 따른 암컷 뱀장어(Anguilla japonica)의 성성숙 유도 효과를 조사하였다. 양식산 암컷 뱀장어를 해수에 적응시킨 후 매주 복강에 재조합 호르몬을 농도별(0.1, 1, 10 ㎍/㎖/fish)로 10주 반복 주사하였다. 매주 증체중을 측정하였고, 생식소중량지수 [GSI;(생식소중량/체중량×100]와 혈중 성호르몬 농도 변화를 조사하였다. 그 결과, 모든 실험군에서 GSI는 서서히 감소하는 경향을 나타내었다. 또한, 대조구에서 혈중 testosterone (T)과 estradiol-17β(E2)는 유의적인 증가를 보이지 않았지만, 재조합 호르몬을 투여한 실험군에서 투여 2주와 4주 후에 T와 E2 농도가 증가하였다. 또한, mt-rFSH(1, 10 ㎍/㎖/fish) 또는 mt-rLH(0.1, 1, 10 ㎍/㎖/fish)을 투여한 실험군에서는 난경이 대조구에 비해 유의하게 증가하였다. 이러한 결과는 열목어 재조합 생식선자극호르몬이 암컷 뱀장어의 초기 난성숙 발달을 유도함을 시사한다.

최근 난포에서 GnRH와 그 수용체의 발현이 확인되면서 GnRH가 국소적으로 난소의 기능을 조절하고,특 히 과립세포의 세포자연사(apoptosis)를 유도하는 것으로 보고되고 있다. 그러나 황체에서 GnRH와 그 수용체의 발현과 기능에 대해서는 잘 알려져 있지 않다. 따라서 본 연구는 임신한 흰쥐의 황체세포에서 GnRH와 그 수용체가 발현되는지를 확인하고, 또한 GnRH가 황체세포의 세포자연사를 직접적으로 유발시킬 수 있는지를 알아보고자 시행하였다. 임

GnRH는 10개의 아미노산으로 구성된 호르몬으로서 생식기능을 조절, 관장하는 중요한 역할을 담당하는 것으로 알려져 있다. 특히 임신 중에는 태반에서 hCG의 분비를 조절하는 중요한 역할을 한다. 최근 사람의 두 번째 GnRH 유전자가 발견되었으며 그 10개의 아미노산 서열은 닭에서 두 번째로 발견된 GnRH (chicken GnRH-II)와 동일한 것으로 확인되었다. 이제까지 사람에서의 두 번째 GnRH (GnRH-II)의 발현은 중뇌와 신장에서 보고

The hypothalamic peptide GnRH plays a central role in the regulation of the mammalian reproductive axis. Recent studies suggested that GnRH stimulates or inhibits the ovarian steroidogenesis and gametogenesis directly. Our previous report indicated that GnRH gene is expressed in adult rat ovary as well as in hypothalamus and that the expressed GnRH may induce the follicular atresia and apoptosis of ovarian granulosa cells in rat. Therfore, we studied whether GnRH gene is expressed in the mouse fetal ovary, when the germ cells are degenerating by apoptosis during gonad diffeerentiation. Mouse fetal gonads were obtained on the 12, 15,18 and 20th day of gestation from the mother mice superovulated (10 IU PMSG and 10 IU hCG) and mated. The morphological changes of fetal ovaries were examined histochemically by hematoxylin-eosin staining. The fetal sex was confirmed by PCR methods for sexing. RT-PCR methods were used to examine the expression of GnRH gene and the sex steroid hormones were determined by conventional radioimmunoassays. The levels of estradiol (E) and progesterone (P) were increaseduntil 18th day of gestation and then E was decreased just before parturition. The morphological changes of fetal gonadal tissue sections showed the ovarian development and coincided with the result of PCR analysis for sexing using ovary- or testis- specific oligonucleotide primers. Immunoreactive GnRH in placenta was decreased gradually until the end of gestation but fetal brain and ovarian GnRH were increased. The level of GnRH gene expression was increased during fetal ovarian development from 12 till 18th day and decreased suddenly on 20th day just before birth. From these results, it is suggested that ovarian GnRh may play a regulatory role on the germ cell differentiation of fetal ovary.