Effects of aromatase inhibitors on reproduction in male three-spined sticklebacks, Gasterosteus aculeatus, exposed to long and short photoperiods

Three-spined stickleback Gasterosteus aculeatus, males were implanted with Silastic capsules filled with different aromatase inhibitors; 1,4,6-androstatriene-3,17-dione or the non-steroidal CGS16949 A, 4-(5,6,7,8-tetrahydrimidazol [1,5-a]pyridin-5-yl) benzonitrile monohydrochloride or empty capsules. The fish were then exposed to long or short photoperiod. Under the long photoperiod most fish in all treatments displayed a hypertrophied kidney (a secondary sexual character in sticklebacks) and completed, quiescent spermatogenesis, similar as in the natural spawning period. Under the short photoperiod the controls had unstimulated kidneys and an active spermatogenesis, whereas the males implanted with both aromatase inhibitors had stimulated kidneys, though not to the extent as in the long photoperiod, and completed, quiescent spermatogenesis. These findings suggest that aromatization is of importance for the inhibitory effects of short photoperiod on reproduction in the stickleback. This revised version was published online in July 2006 with corrections to the Cover Date.     

Sex differences in circulating steroid hormone levels in the red drum, Sciaenops ocellatus L.

Steroid profiles of cultured and captive red drum (Sciaenops ocellatus L.) were investigated to evaluate the potential use of circulating sex steroid levels as a tool for gender identification in this species. Cultured 18‐month‐old fish were maintained on a 120‐day shortened photothermal cycle to induce precocious maturation. Additionally, wild‐caught fish were maintained in captivity under simulated natural photothermal conditions from late spring to early fall. Circulating 11‐ketotestosterone (11‐KT) levels were significantly higher in males compared with females during the early stages of gonadal growth in both cultured and captive fish. Plasma testosterone (T) levels showed a similar trend; however, the differences were significant only when males were already producing sperm. 17β‐estradiol (E₂) concentrations were low in males and females before gonadal recrudescence but increased significantly with the progression of vitellogenesis in females. These results show that a test using a minimum concentration of circulating 11‐KT could be developed to differentiate between sexes in the early stages of gonadal maturation in red drum. Moreover, plasma E₂ concentrations could be used to identify vitellogenic females. The two steroids considered together could help avoid possible error in gender identification due to unusually high levels of certain steroids encountered in some individuals.     

Effect of Human Menopausal Gonadotropin,17β-estradiol and Epidermal Growth Factor on the Quality of in vitro Maturation Bovine Oocytes

For optimizing in vitro maturation system of bovine oocytes,we firstly examined the influence of four different hormonal regimes(FSH+LH,HMG,FSH+LH+E₂ and HMG+E₂) on oocyte maturation rates.Then we studied the effects of epidermal growth factor (EGF) in the above defined medium on bovine oocyte maturation,in vitro development and quality of parthenogenetic embryos.The cell apoptotic index of parthenogenetic blastocysts was detected by TUNEL.No significant difference was observed in maturation rates in four groups supplemented with different hormones.However,human menopausal gonadotropin (HMG) provided steady maturation results in replicates.Maturation of oocytes was promoted by supplementation with 17β-estradiol (E₂).Combination of HMG and E₂ gave rise to steady and efficient mature results.The presence of EGF at 30 ng/mL concentration significantly increased maturation rate and blastocyst rate and reduced apoptotic cells in parthenogenetic blastocysts.Therefore,the optimal oocyte maturation solution could be supplemented with 0.075 IU/mL HMG,1 μg/mL E₂ and 30 ng/mL EGF.     

山羊卵母细胞无血清体外成熟培养

为研究无血清培养系统对山羊卵母细胞体外成熟及孤雌激活胚胎早期发育力的影响,分别以0.3%牛血清蛋白(BSA)(mV)、1%聚乙烯醇(PVA)(mV)和1%胰岛素转铁蛋白亚硒酸钠(ITS)(VV)代替成熟培养液中胎牛血清(FBS),以添加5?S的OM为对照组,对来源于屠宰场的卵巢卵母细胞复合体(COCs)进行体外培养。成熟率PVA添加组(50.00%)显著低于FBS组(83.23%)(P<0.01)、BSA组(71.19%)和ITS组(76.79%),与FBS组无显著差异(P>0.05);成熟卵母细胞用离子霉素联合6-DMAP激活,在SOFaa中进行孤雌胚培养,BSA、PVA、ITS及FBS各组的卵裂率依次为61.90%、46.97%、79.07%和83.58%,囊胚率分别为25.00%、19.70%、55.81%和60.45%。结果表明:在OM基础培养液中添加BSA或PVA,卵母细胞的成熟率明显低于添加FBS,且卵裂率和囊胚发育率也低于对照组(P<0.05或P<0.01);添加ITS,卵母细胞成熟率虽低于对照组,但孤雌胚卵裂率和囊胚发育率与对照组接近(P>0.05)。表明以ITS代替FBS,可用于山羊卵母细胞体外成熟培养。     

Distribution of protein disulfide isomerase during maturation of pig oocytes

Oocyte maturation in mammals is characterized by a dramatic reorganization of the endoplasmic reticulum (ER). In mice, the ER forms accumulations in the germinal vesicle (GV) stage and distinctive cortical clusters in metaphase II (MII) of the oocyte. Multiple evidence suggests that this ER distribution is important in preparing the oocyte for Ca²⁺ oscillations, which trigger oocyte activation at fertilization. In this study, we investigated the time course and illustrated the possible functional role of ER distribution during maturation of porcine oocytes by immunostaining with protein disulfide isomerase (PDI). PDI forms clusters in the cytoplasm of oocytes. After immunostaining, PDI clusters were identified throughout the cytoplasm from the GV to metaphase I (MI) stage; however, at the MII stage, the PDI formed large clusters (1–2 µm) in the animal pole around the first polar body. PDI distribution was prevented by bacitracin, a PDI inhibitor. Our experiments indicated that, during porcine oocyte maturation, PDI undergoes a dramatic reorganization. This characteristic distribution is different from that in the mouse oocyte. Moreover, our study suggested that formation of PDI clusters in the animal pole is a specific characteristic of matured porcine oocytes.     

极早熟西瓜秀丽生育特性观察及栽培技术

极早熟西瓜秀丽系杂交一代。经系统观察探明,该品种从播种育苗至果实成熟全生育期８０ｄ左右,从雌花开放到果实成熟需２５～２６ｄ。第１雌花位于８～１１节,第２雌花位于１７～２０节。可溶性固形物达到１３．４％。叶片生长速度在新叶展开８ｄ内最快,茎蔓抽生速度在子蔓８片叶进入快速生长期,果实膨大和可溶性固形物积累速度在座果后７～２１ｄ最快。同时,研究了秀丽西瓜的关键栽培技术。     

Early metaphase II oocytes treated with dibutyryl cyclic adenosine monophosphate provide suitable recipient cytoplasm for the production of miniature pig somatic cell nuclear transfer embryos

We investigated the effects of in vitro maturation duration and treatment with dibutyryl cyclic adenosine monophosphate (dbcAMP) on the blind enucleation efficiency and developmental competence of miniature pig somatic cell nuclear transfer (SCNT) embryos. Oocytes were cultured for 22 h in NCSU-23 medium with or without 1 mM dbcAMP and then additionally cultured in dbcAMP-free NCSU-23 for 14, 18, or 22 h. Regardless of dbcAMP treatment, the rate of nuclear maturation reached a plateau at 36 and 40 h. However, mitochondrial distribution, a marker for cytoplasmic maturation, differed between the dbcAMP-untreated oocytes at 36 h and dbcAMP-treated oocytes at 40 h. The metaphase II chromosomes were adjacent to the first polar body in 68.8% and 63.5% of the dbcAMP-untreated oocytes at 36 h and dbcAMP-treated oocytes at 40 h, respectively. Furthermore, the blind enucleation efficiency by removing a small volume of cytoplasm was significantly higher in the dbcAMP-untreated oocytes at 36 h (82.9%) and dbcAMP-treated oocytes at 40 h (89.9%) than other groups. The rate of blastocyst formation was highest in the dbcAMP-treated oocytes at 40 h. Hence, this study demonstrated that dbcAMP-treated early metaphase II oocytes are suitable for the production of miniature pig SCNT embryos.     

注射含HCG和CPE的W／O／W复乳诱导日本鳗鲡性腺发育成熟的研究

用含HCG及鲤脑垂体（PG）匀浆液提取物（CPE）的W／O／W复乳，对塘养雌鳗进行注射催熟实验，雌鳗平均GSI达35．18％，最高达56．5％，其催熟效果较常规注射方法（平均GSI 27．9％，最高50．4％）好，用含LHRH－A，HCG，CPE的复乳和DOM生理盐水混悬剂对部分达到性成熟的鳗鲡进行催产，用W／O／W复乳诱导成熟的鳗鲡，排卵率为75％，用常规方法催熟的鳗鲡排卵率为55％，注射复乳制剂后，鳗鲡血清GtH含量可较长时间的保持较高水平，血清GtH的变化较常规注射方法缓和，注射乳剂还可使鳗鲡脑垂体GtH含量升高。     

HA对牛卵母细胞体外成熟和早期胚胎体外发育影响的研究

在TCM-199中添加一些确定的辅助成分,探讨在无血清培养基中添加不同浓度HA对牛卵母细胞体外成熟及早期胚胎体外发育的影响。结果表明,在卵母细胞成熟培养液(TCM-199-m)中添加4.0mg/mL的HA时,卵母细胞的成熟率和卵裂率与对照组(BSA)无显著差异(P>0.05),分别为85.14%,83.57%和89.47%,85.24%,但显著高于其他各浓度组(P<0.05),表明HA在卵母细胞无血清成熟培养中可代替BSA。在受精卵体外培养液(TCM-199-c)中添加HA时,囊胚发育率与对照组(BSA)差异不显著(P>0.05),分别为27.64%和26.51%,但显著高于TCM-199-c组(P<0.05),表明HA对牛胚胎体外发育具有明显的促进作用,在无血清培养中可以代替BSA。当在TCM-199-c+HA中添加少量的BSA时,囊胚发育率(31.19%)显著高于其他各组(P<0.05),表明HA欲完全代替BSA还有不完善之处,有待进一步研究。     

Growth of Preimplatation Bovine Embryos

Development of mammalian embryos in vitro is functionally and temporally inferior to embryo development as it occurs inside the female reproductive tract. The deficiencies of cultured embryos range from slow cleavage rates to complete developmental arrests or blocks, occurring at particular stages in many species. A variety of approaches have been used to overcome the blocks, including most extensively the coculture of preimplantation embryos with various somatic cells. However, even with coculture, development of embryos in vitro is still not equivalent to that in vivo. In most laboratories, only 25–40% of inseminated oocytes develop into morulae and blastocysts in spite of numerous variations on the basic technique. A better understanding of the factors governing embryonic growth is required before we can hope to achieve results comparable with those occurring in vivo.