GnRH-I类似物对离体大鼠胃壁细胞三磷酸肌醇含量的影响

目的：探讨促性腺激素释放激素（GnRH）-I类似物阿拉瑞林对离体大鼠胃壁细胞内三磷酸肌醇（IP3）含量的影响。方法：体外分离大鼠胃壁细胞并给予不同浓度的阿拉瑞林孵育,利用标记的[3H]-IP3采用放射免疫分析法检测了壁细胞内IP3的含量。结果：当加入终浓度分别为0．001、0．01、0．1、1μmol／L的阿拉瑞林时,大鼠胃壁细胞内IR的含量逐渐增加,呈剂量依赖性,当阿拉瑞林为1μmol／L时,IB的含量达到高峰;同时随着药物作用时间的延长,IR的含量也会增加,当阿拉瑞林作用5min时,IR的含量达到峰值,随后开始下降;磷酸二脂酶（PLC）抑制剂Compound48／80温育细胞后,再加入阿拉瑞林孵育,可使IB的含量轻度增加,但与PLC抑制剂组相比,无统计学意义（P〉0．05）;IP3受体阻滞剂heparin温育细胞后,再加入阿拉瑞林孵育,壁细胞内IB含量略有上升,与只加入Heparin组相比无统计学意义。结论：在体外GnRH—I类似物可使大鼠胃壁细胞内IR的含量明显增加,说明IR参与了GnRH—I类似物对大鼠胃壁细胞功能调控的信号转导过程。     

GnRH类似物对大鼠回肠组织胰高血糖素释放的影响

目的 :研究促性腺激素释放激素 (GnRH)类似物 (阿拉瑞林 )对大鼠回肠L细胞释放胰高血糖素的影响。方法 :应用放射免疫分析法对体内和体外大鼠回肠进行观察。结果 :大鼠回肠灌注GnRH类似物后 ,血中及肠液中胰高血糖素的含量较对照组明显升高 ;体外孵育大鼠回肠组织后 ,在一定浓度范围内 ,孵育液中胰高血糖素含量随GnRH类似物浓度升高而升高 ;当浓度高于一定范围时 ,则随浓度升高而降低。GnRH类似物浓度为 1 .0× 1 0 - 4mol/L时孵育液中胰高血糖素含量是升高的。结论 :GnRH可能对大鼠回肠L细胞分泌胰高血糖素呈现双向调节作用。但是GnRH类似物为 1 .0× 1 0 - 4mol/L ,不论是体内还是体外 ,都可能对肠道分泌胰高血糖素表现促进作用     

GnRH-Ⅰ类似物对离体大鼠胃壁细胞三磷酸肌醇含量的影响

目的:探讨促性腺激素释放激素(GnRH)-Ⅰ类似物阿拉瑞林对离体大鼠胃壁细胞内三磷酸肌醇(IP3)含量的影响.方法:体外分离大鼠胃壁细胞并给予不同浓度的阿拉瑞林孵育,利用标记的[3H]-IP3,采用放射免疫分析法检测了壁细胞内IP3的含量.结果:当加入终浓度分别为0.001、0.01、0.1、1 μmol/L的阿拉瑞林时,大鼠胃壁细胞内IP3的含量逐渐增加,呈剂量依赖性,当阿拉瑞林为1 μmol/L时,IP3的含量达到高峰;同时随着药物作用时间的延长,IP3的含鼍也会增加,当阿拉瑞林作用5 min时,IP3的含量达到峰值,随后开始下降;磷酸二脂酶(PLC)抑制剂Compound 48/80温育细胞后,再加入阿拉瑞林孵育,可使IP3的含量轻度增加,但与PLC抑制剂组相比,无统计学意义(P0.05);IP3受体阻滞剂heparin温育细胞后,再加入阿拉瑞林孵育,壁细胞内IP3含量略有上升,与只加入Heparln组相比无统计学意义.结论:在体外GnRH-Ⅰ类似物可使大鼠胃壁细胞内IP3的含量明显增加,说明IP3参与了GnRH-Ⅰ类似物对大鼠胃壁细胞功能调控的信号转导过程.     

GnRH类似物对培养的大鼠胃平滑肌细胞增殖的影响

目的　观察促性腺激素释放激素 (gonadotropin releasinghormone,GnRH)类似物阿拉瑞林 (alarelin)对培养的大鼠胃平滑肌细胞增殖的影响。　方法　应用离体培养的SD大鼠胃平滑肌细胞 (gastricsmoothmusclecell,GSMC) ,采用四唑盐 (MTT)比色法实验 ,3H 胸腺嘧啶核苷酸 (3H TdR)参入、免疫荧光化学检测增殖细胞核抗原 (pro liferatingcellnuclearantigen ,PCNA)表达的平均荧光值和流式细胞仪技术 ,观察阿拉瑞林对GSMC的增殖、DNA合成和细胞周期的影响。　结果　当加入终浓度为 10 - 9、10 - 7、10 - 5mol L的阿拉瑞林 2 4h后 ,与未用药的对照组相比 ,发现其MTT吸光度 (A值 )逐渐降低 (P <0 0 5 ) ;PCNA表达的平均荧光值逐渐减弱 (P <0 0 5 ) ;GSMC的3H TdR参入量依次减少 (P <0 0 5 ) ,且药物浓度越大 ,此三者下降越明显。在细胞周期中 ,与对照组相比 ,G1 期细胞所占比例明显增加 (P <0 0 5 ) ;S期细胞所占比例明显减少 ,且随药物浓度的增加而逐渐减少 (P <0 0 5 )。　结论　GnRH类似物可明显抑制大鼠GSMC的增殖作用 ,其作用途径可能是通过平滑肌细胞自身GnRH受体的直接介导而实现的     

脑室注射L-谷氨酸对成年大鼠下丘脑GnRH含量的影响

目的 :观察脑室注射L -谷氨酸 (L -Glu)对成年雄性Wistar大鼠下丘脑促性腺激素释放激素(GnRH)含量的影响。方法 :摘取下丘脑组织 ,匀浆化 ,用RIA法检测匀浆上清液中GnRH的含量。结果 :脑室分别注射 0 0 1176、0 1176、1 176 0 μg/ 2 0 μl-1 -1L -Glu后 40分钟 ,下丘脑GnRH含量依次为 1 5 9± 0 41、0 88± 0 34、0 70± 0 42ng/ 10mg湿重 ,均显著低于盐水对照组 (P <0 .0 1) ;脑室注射 0 1176 μg/ 2 0 μl-1·-1L -Glu后 2 0、40、12 0分钟 ,下丘脑GnRH含量依次为 0 99± 0 37、0 88± 0 34、1 2 6± 0 39ng/ 10mg湿重 ,亦均显著低于盐水对照组 (P <0 .0 1)。脑室注射L -Glu对下丘脑GnRH含量的降低作用呈现剂量与时间依从关系。而脑室注射3H -Glu 2 μCi/ 2 0 μl-1·-1后 40分钟发现 ,大脑、小脑、垂体、下丘脑内侧基底部 (MBH)和视前区 (POA) 5个不同部位脑组织中以MBH对3H -Glu的摄取量最大 (10 6 9 82± 490 33cpm/ 10mg湿重 )。结论 :L -Glu可能参与了大鼠下丘脑GnRH神经元功能活动的调节。     

促性腺激素释放激素类似物对大鼠胃肠道5-羟色胺分泌的影响

目的 研究GnRH对胃肠道内5-HT分泌的影响。方法胃腔直接注射GnRH类似物阿拉瑞林(A1arelin GnRH—A)以模拟外分泌产生的GnRH，并以免疫组织化学、高压液相色谱电化学检测(HPLC-ECD)的方法对阿拉瑞林刺激后胃及十二指肠内5-HT免疫反应细胞、血清中5-HT含量进行检测。结果 胃腔内注射GnRH—A后，大鼠胃及十二指肠内5-HT免疫反应阳性细胞密度显著增多，但血清中5-HT含量显著减少。结论 外分泌的OnRH对于5-HT的释放起明显抑制作用，但对5-HT的合成可能不产生影响。     

乙酰唑胺对水孔蛋白-1表达的抑制与细胞内pH及Ca2+的关系

为明确乙酰唑胺对水孔蛋白-1(aquaporin-1,AQP1)基因表达的抑制是否与其对细胞内pH和Ca2+浓度的影响有关,采用激光共聚焦扫描显微镜的技术观察不同培养条件下(培养液中含或不含NaHCO3),10-5 mol*L-1乙酰唑胺对原代培养的大鼠肾脏近曲小管上皮细胞内pH和Ca2+浓度的影响,同时用免疫荧光法观察给药1d、3d、7d后AQP1表达的变化.结果显示在含NaHCO3组,乙酰唑胺处理7d后,使细胞内的H+探针的荧光强度由91.81±5.44降至39.58±3.10, Ca2+探针荧光强度由18.23±1.02降至11.5±1.03,AQP1蛋白荧光强度由44.4±1.86减少到25.91±1.97;在不含NaHCO3组,细胞内pH和Ca2+浓度均无明显变化;但AQP1蛋白的荧光强度由43.15±2.97明显减少到23.85±1.92.结果表明乙酰唑胺对细胞内pH和Ca2+浓度的调节依赖于细胞外液HCO3-的存在,而它对AQP1表达的抑制与细胞内pH和Ca2+浓度的改变无关.     

培养的大鼠胃平滑肌细胞GnRH受体的免疫组织化学及原位杂交研究

目的　观察培养的大鼠胃平滑肌细胞中是否能表达GnRH受体 ,为进一步研究GnRH对胃平滑肌细胞的功能提供形态学依据。　方法　采用免疫组织化学SABC和原位杂交方法。　结果　培养的大鼠胃平滑肌细胞呈GnRH受体免疫反应阳性 ,阳性物质分布于细胞浆和细胞膜上 ,细胞核呈阴性反应。培养的胃平滑肌细胞同样含有GnRH受体mRNA杂交信号 ,信号物质亦分布于细胞浆内 ,细胞核呈阴性反应。　结论　胃平滑肌细胞能自身表达GnRH受体 ,它可能也是GnRH的靶细胞     

耳蜗外毛细胞两种胞内钙库的初步探讨

为探讨毛细胞胞内钙库的种类 ,本文观察了在无钠、无钙和含镧液体中 ,在不受胞外 Ca2 +内流和质膜上 Ca2 +转运机制影响的条件下 ,和在三磷酸肌醇敏感钙库的工具药 thapsigargin和 ryanodine敏感钙库的工具药 caffeine作用下 ,毛细胞胞内游离钙([Ca2 + ] i)的变化过程。分离的豚鼠耳蜗外毛细胞经钙敏荧光染料 5μmol/L fluo-3染色后 ,用激光扫描共聚焦显微镜监测 ,以 fluo-3荧光相对值指示毛细胞 [Ca2 + ] i的高低。 3 0 nmol/L thapsigargin使外毛细胞 [Ca2 + ] i由静态值 1.0增至 1.64± 0 .76,再加入 10mmol/L caffeine后更增至 2 .45± 1.5 9(x± s,n=11,F=7.90 ,P<0 .0 1)。Q值检验示两种试剂引起外毛细胞 [Ca2 + ] i增高程度的差别有极显著意义 (P<0 .0 1) ,表明毛细胞内有对三磷酸肌醇敏感和对 ryanodine敏感的两种钙库参与了胞内 Ca2 + 释放机制     

GnRH激动剂主动免疫母羊对生殖激素分泌的作用

目的:探讨GnRH激动剂(GnRHa)主动免疫对绵羊生殖激素合成与分泌的作用,并深入研究GnRH-A免疫调节动物生殖功能的机理。方法:42只5～6月龄母绵羊(Ovis aries)随机分为6组(n=7),EG-Ⅰ、EG-Ⅱ和EG-Ⅲ分别于0和14天皮下注射阿拉瑞林抗原200、300和400μg;EG-Ⅳ和EG-Ⅴ分别皮下注射阿拉瑞林抗原200、300、0、7、14和21天各一次,共4次;对照组在0和14天皮下注射抗原溶媒(除不用阿拉瑞林外,其余成分和制备方法与阿拉瑞林抗原相同)2.0 ml。无菌采集不同时段的血液,分离血清。以ELISA测定血清GnRH抗体浓度,用激素检测试剂盒(ELISA)分别测定血清GnRH、FSH、LH和E2浓度。结果:①阿拉瑞林首次免疫7天后,各实验组的抗体浓度逐渐升高,EG-Ⅰ、EG-Ⅱ和EG-Ⅲ分别在28、28和35天达到峰值(P<0.05),EG-IV和EG-V则在在45天达到峰值(P<0.01),至60天时仍明显高于对照组(P<0.05)。14～60天间EG-IV和EG-V抗体浓度均高于EG-Ⅰ、EG-Ⅱ和EG-Ⅲ(P<0.05)。②EG-Ⅰ、EG-Ⅱ的GnRH在21和28天抵谷值(P<0.05),EG-Ⅲ、EG-Ⅳ和EG-Ⅴ则在45天抵谷值(P<0.01),且以EG-Ⅴ为最低。谷值之后逐渐上升趋势,70天时达到免疫注射前水平。③实验组血清FSH浓度始终高于对照组(P<0.05)。EG-Ⅰ、EG-Ⅱ和EG-Ⅲ于28、28和35天达到峰值(P<0.05),而EG-IV和EG-V在60天达到高峰值(P<0.01)。④实验组绵羊血清LH呈下降趋势,EG-Ⅰ、EG-Ⅱ和EG-Ⅲ分别在21、21和28天达到谷值(P<0.01),EG-Ⅳ和EG-Ⅴ在35天达到谷值(P<0.01)。35天时EG-Ⅳ和EG-Ⅴ低于EG-Ⅰ、EG-Ⅱ和EG-Ⅲ。⑤各组的血清E2含量无显著差异。结论:GnRH激动剂(阿拉瑞林)抗原主动免疫可促进GnRH抗体的生成,抑制母羊GnRH和LH的合成与分泌,增强FSH的合成与分泌,且随着注射剂量和注射次数的增加,这种作用更加明显,而对血清E2无明显影响。     

Embryo implantation and GnRH antagonists: GnRH antagonists do not activate the GnRH receptor

Recent suggestions that gonadotrophin-releasing hormone (GnRH) antagonists activate the GnRH receptor are discussed. Most of the studies cited in support of this suggestion are in-vitro studies, testing supra-pharmacological doses of GnRH analogues in cancer cell lines, whereas GnRH antagonists, e.g. ganirelix or cetrorelix, do not affect the steroidogenesis of human granulosa cells in vitro. In patients treated with GnRH antagonists prior to IVF or intracytoplasmic sperm injection (ICSI), oocyte maturity and fertilization rates are equal to those achieved following a long protocol of GnRH agonists. Although there is a tendency towards a lower pregnancy rate (not statistically significant) in the initial trials using GnRH antagonist with either recombinant FSH or human menopausal gonadotrophin (HMG) for ovarian stimulation, this new treatment option of GnRH antagonists facilitates short and simple treatment and improves the convenience and safety for the patient. As with GnRH agonists in the past, the clinical outcome of GnRH antagonist treatment will improve with time as more clinical experience is gained (learning curve) and the treatment protocol is optimized. Moreover, a GnRH agonist instead of human chorionic gonadotrophin (HCG) may be used for triggering ovulation and will decrease the cancellation rate and minimize the risk for developing ovarian hyperstimulation syndrome (OHSS).     

Embryo implantation and GnRH antagonists: embryo implantation: the Rubicon for GnRH antagonists

When gonadotrophin-releasing hormone (GnRH) was discovered, the agonist and antagonist of GnRH were developed to control the release of FSH and LH by the gonadotrophs. More than 10 years of research were needed to develop a GnRH antagonist free of histamine release. Recent studies have shown that these GnRH antagonists are effective in preventing a rise in LH during ovarian stimulation in IVF. However, a decrease in ongoing pregnancies seems to suggest that implantation rates per transferred embryo are reduced in GnRH antagonist-stimulated cycles. In my opinion, these data highlight an area less well known to clinicians: the role of the GnRH antagonist at the cellular level in extrapituitary tissues. There are sufficient data in the literature suggesting that GnRH antagonist is an inhibitor of the cell cycle by decreasing the synthesis of growth factors. Given that, for folliculogenesis, blastomere formation and endometrium development, mitosis is everything; the interaction between the GnRH antagonist and the GnRH receptor (present in all these cells and tissues) may compromise the mitotic programme of these cells. This is the Rubicon for the GnRH antagonist: to demonstrate irrevocably that, at the minimal doses necessary to suppress LH release, it does not affect processes such as implantation, embryo development and folliculogenesis.     

GnRH拮抗剂与GnRH激动剂短方案IVF-ET结局的比较

目的比较GnRH antagonist与GnR Hagonist短方案的IVF-ET结局。方法2006年8月至2007年8月GnR Hantagonist治疗组54人和GnR Hagonist短方案对照组132人,记录促性腺激素的用量及其用药天数、hCG日子宫内膜厚度和激素水平、获卵数、受精率、卵裂率、优胚率、妊娠率和OHSS发生率等指标。结果两组促性腺激素的用量及其用药天数、获卵数、受精率、卵裂率、着床率和妊娠率相比较均无显著差异（P〉0.05）。GnR Hantagonist组在hCG日激素水平低,与对照组比较其差异有统计学意义。结论行GnR Hantagonist方案IVF-ET助孕治疗与传统的GnR Hagonist短方案比较,其hCG日雌激素水平下降可能是OHSS发生率显著下降的主要因素;但卵泡的发育、卵母细胞的受精率、卵裂率及妊娠率和着床率均不受影响。GnR Hantagonist的使用为IVF-ET助孕药物提供了一种新的选择。     

GnRH agonist versus GnRH antagonist in oocyte donation cycles: a prospective randomized study

BACKGROUND: The specific role of LH in folliculogenesis and oocyte maturation is unclear. GnRH antagonists, when administered in the late follicular phase, induce a sharp decrease in serum LH which may be detrimental for IVF outcome. This study was performed to evaluate whether the replacement of GnRH agonist (triptorelin) by a GnRH antagonist (ganirelix; NV Organon) in oocyte donation cycles has any impact on pregnancy and implantation rates. METHODS: A total of 148 donor IVF cycles was randomly assigned to use either a GnRH antagonist daily administered from the 8th day of stimulation (group I) or a GnRH agonist long protocol (group II) for the ovarian stimulation of their donors. The primary endpoints were the pregnancy and the implantation rates. RESULTS: The clinical pregnancy rate per transfer (39.72%, 29/73 versus 41.33%, 31/75) based on transvaginal scan findings at 7 weeks of gestation, the implantation rate (23.9 versus 25.4%) and the first trimester abortion rate (10.34 versus 12.90%) were similar in the two groups. CONCLUSION: In oocyte donation cycles the replacement of GnRH agonist by a GnRH antagonist appears to have no impact on the pregnancy and implantation rates when its administration starts on day 8 of stimulation.     

大鼠颌下腺GnRH及其受体的免疫组织化学双标记和年龄变化

用免疫组织化学法，以邻片双标记技术研究了大鼠颌下腺内ＧｎＲＨ及ＧｎＲＨ受体的分布；并对不同的发育阶段大鼠颌下腺内ＧｎＲＨ及其受体进行了原位定量分析，结果表明，雄性成年大鼠颌下腺浆液性腺泡的腺上皮细胞及各级导管上皮细胞既呈ＧｎＲＨ又呈ＧｎＲＨ受体免疫反应性。     

Embryo implantation and GnRH antagonists: the search for the human placental GnRH receptor

Hypothalamic gonadotrophin-releasing hormone (GnRH) plays a major role in the endocrine control of reproduction. Acting through its high affinity receptors on pituitary gonadotrophs, it regulates the secretion of gonadotrophins. In addition, GnRH also functions as a local regulator in a number of other cell lines and tissues, including the placenta. In a manner analogous to hypothalamic GnRH stimulation of LH and FSH from the anterior pituitary, GnRH was found to cause a dose-dependent release of human chorionic gonadotrophin (HCG) from the placenta. So began the search for a putative GnRH receptor in the human placenta. Although early radio-receptor studies reported specific binding, the properties of these 'putative' GnRH binding sites were found to differ significantly from those of their pituitary counterparts in several important respects. This was followed by a series of contradictory reports that led to more questions and opened up avenues for further investigations. Even after nearly two decades of research, the human placental GnRH receptor has not been characterized beyond all reasonable doubt. This review recalls the discovery, the controversies and unanswered questions concerning the human placental GnRH receptor.     

Time-dependent effect of ethanol on GnRH and GnRH receptor mRNA expression in hypothalamus and testis of adult and pubertal rats

Chronic exposure to ethanol suppresses the male reproductive activity which is primarily involved in the release of hypothalamic gonadotropin-releasing hormone (GnRH). The testicular GnRH and GnRH receptors (GnRH-R) are found in seminiferous tubules, which are predicted to act as a local regulator of spermatogenesis, although the function is not well known. In this study, we investigated the chronic ethanol effect on GnRH mRNA expression in hypothalamus and testis using in situ hybridization and RNase protection assay (RPA). The effect of ethanol on expressional changes of GnRH and GnRH-R mRNA was observed in adult and pubertal rats according to age and time from 2 weeks (short term) and 4 weeks (long term) periods. The results showed that GnRH mRNA expression in adult and pubertal rats was dramatically decreased in the testis while no significant change was observed in hypothalamus after both short and long term exposure to ethanol. The pubertal rats showed decrease in testicular GnRH and GnRH-R mRNA expression, whereas GnRH mRNA was increased significantly, while GnRH-R mRNA was further decreased after long term exposure in adults. This study suggested that chronic ethanol administration is more effective to testicular GnRH and GnRH-R mRNA expression than hypothalamus and causes a negative effect on the spermatogenesis process. Furthermore, our finding suggests that the deteriorative effects of ethanol on gonadal activity are more lethal in puberty than adults.     

GnRH antagonist in assisted reproduction: a Cochrane review

BACKGROUND: In the present systematic review, we wished to compare the efficacy of gonadotrophin-releasing hormone (GnRH) antagonist and GnRH agonist administration for controlled ovarian hyperstimulation in assisted conception. METHODS: Five randomized controlled trials fulfilled the inclusion criteria. In four studies, the multiple low-dose (0.25 mg) antagonist regimen was applied and, in one study, the single high-dose (3 mg) antagonist regimen was investigated. In all trials, reference treatment included a long protocol of GnRH agonist (buserelin, leuprorelin or triptorelin) starting in the mid-luteal phase of the preceding cycle. RESULTS: In comparison with the long protocol of GnRH agonist, the overall odds ratio for the prevention of premature LH surges was 1.76 [95% confidence interval (CI) 0.75-4.16], which was not statistically significant. There were significantly fewer clinical pregnancies in those treated with GnRH antagonists (OR 0.79; 95% CI 0.63-0.99). There was no statistically significant reduction in incidence of severe ovarian hyperstimulation syndrome between the two regimens (relative risk 0.51; OR 0.79; 95% CI 0.22-1.18). CONCLUSIONS: We concluded that the fixed GnRH antagonist protocol is a short and simple protocol with good clinical outcome, but the lower pregnancy rate compared with the GnRH agonist long protocol and the non-significant difference between both protocols regarding prevention of premature LH surge and prevention of severe ovarian hyperstimulation syndrome necessitates counselling subfertile couples before recommending change from GnRH agonist to antagonist. The clinical outcome may be further improved by developing more flexible antagonist regimens, taking into account individual patient characteristics.     

GnRH antagonist versus long GnRH agonist protocol in poor responders undergoing IVF: a randomized controlled trial

BACKGROUND. This is the first published report of a prospective, randomized, controlled trial comparing a fixed, multi-dose GnRH antagonist protocol with a long GnRH agonist protocol in poor responders undergoing IVF. METHODS. Sixty-six poor responders were randomized into two groups: the study group received 0.25 mg of cetrorelix daily starting on day 6 of stimulation; the control group received 600 microg of buserelin acetate daily starting in the mid-luteal phase of the preceding cycle. Both groups were given a fixed dose of recombinant FSH (300 IU daily) for stimulation. RESULTS. There were no significant differences in the cycle cancellation rates, duration of stimulation, consumption of gonadotrophins, and mean numbers of mature follicles, oocytes and embryos obtained. The implantation rates were similar, but the number of embryos transferred was significantly higher for the antagonist group (2.32 +/- 0.58 versus 1.50 +/- 0.83; P = 0.01). The pregnancy rates were also higher in the antagonist group, but the difference was not statistically significant. CONCLUSION. A fixed multi-dose GnRH antagonist protocol is feasible for patients who are poor responders on a long agonist protocol; however, our study failed to demonstrate an overall improvement in ovarian responsiveness. Clinical outcomes may be improved by developing more flexible antagonist regimens, an approach that requires further evaluation.