Journal of Reproduction and Development
Online ISSN : 1348-4400
Print ISSN : 0916-8818
ISSN-L : 0916-8818
Volume 39, Issue 1
Displaying 1-12 of 12 articles from this issue
  • Yutaka FUKUI, Hidekuni HIRAI, Koji HONDA, Kenichi HAYASHI
    1993 Volume 39 Issue 1 Pages 1-5
    Published: 1993
    Released on J-STAGE: May 15, 2008
    JOURNAL FREE ACCESS
    During the non-breeding season (April to July), 187 ewes treated wth either an intravaginal sponge impregnated with medroxyprogesterone acetate (MAP) or a controlled internal drug release (CIDR®) device for 9 days and an injection of pregnant mare serum gonadotropin (PMSG) at one day before the cessation of progestogen treatment were inseminated into the uterus by laparoscopy with frozen-thawed semen at fixed-time basis (36, 48 or 60h after the removal of sponge or device). Insemination was also performed at 18 h after estrus detection as a control. The time to the onset of estrus after CIDR® treatment was significantly (P<0.05) earlier (mean: 24.9h) than that in ewes treated with MAP sponge (mean: 30.0h). The percentages of ewes lambing and lambs born per ewe lambing for inseminations at 36, 48 and 60 h were 66.7% and 1.69; 52.0% and 1.31; 46.2% and 1.75 for MAP-treated ewes, and 33.3% and 1.50; 73.9% and 1.53; 40.9% and 1.56 for CIDR®-treated ewes. For control ewes, they were 59.1% and 1.62; and 47.6% and 1.90 for MAP and CIDR® treatments, respectively. There was no significant differences in the lambing rates and prolificacy between MAP and CIDR® treatments and among the insemination times. An optimum time for an intrauterine insemination with frozen-thawed semen appears to be different in the use of MAP sponge (range: 36 to 60h) and CIDR® (48h). A significant difference in the lambing rate was found among the sheep farms conducting the present study.
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  • Katsumi ISHIDA, Hiroshi AMEMIYA, Takashi UMEDA, Hideo MOHRI, Makoto OK ...
    1993 Volume 39 Issue 1 Pages 7-12
    Published: 1993
    Released on J-STAGE: May 15, 2008
    JOURNAL FREE ACCESS
    A new microflushing analysis method for sperm motility is proposed. Conventionally, spermatozoa have been immersed in mineral oil and manipulated without exposure to the air. In this method, since the spermatozoa scarcely adhere to the surface of the slide glass during microscopic observation of sperm motility, usually over 90% of the motility is maintained. Furthermore, it became possible to detect and characterize the flagellar bend newly formed in a different test reagent. Because this method requires only an extremely small amount of spermatozoa for assay, repeated assay even for small animals is possible. Therefore, the microflushing method is considered to be one of the best method for assaying sperm motility.
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  • Yuji HIRAO, Junko KIMURA, Takashi MIYANO, Seishiro KATO
    1993 Volume 39 Issue 1 Pages 13-17
    Published: 1993
    Released on J-STAGE: May 15, 2008
    JOURNAL FREE ACCESS
    Ovaries from newborn female mice were cultured for 4 or 8 days in Eagle's minimum essential medium supplemented with pregnant mare's serum gonadotrophin (PMSG) at a concentra-tion of 0, 2, 10 or 50IU/ml. At the beginning of culture, most oocytes in the ovaries were smaller than 20μm in diameter and no zona pellucida was formed around them. During 4 days of culture, promotion of oocyte growth by PMSG (10 and 50IU/ml) was observed. After 8 days, proportions of oocytes with a continuous zona pellucida were significantly higher in PMSG-supplementation groups. In the cultured ovaries, however, the multilayered granulosa cells, that could be often seen around growing oocytes in 8-day-old mouse ovaries, were not observed at all irrespective of the presence of PMSG. These results suggest that PMSG promotes early oocyte growth and zona pellucida formation of oocytes in neonatal mice, although it has a little effect on the proliferation of granulosa cells.
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  • Keiji TANIMOTO, Kouichi TAMURA, Fumihiro SUGIYAMA, Kazuo MURAKAMI, Aki ...
    1993 Volume 39 Issue 1 Pages 19-24
    Published: 1993
    Released on J-STAGE: May 15, 2008
    JOURNAL FREE ACCESS
    Renin is the rate-limiting enzyme in the renin-angiotensin system, the only known biochemical cascade that produces the potent octapeptide effector molecule angiotensin II that controls, to a large degree, cardiovascular homeostasis and reproductive function in mammals. In the present study, we have cloned the Ren-1C gene from the C57BL/6 mouse and determined its genomic structure. In order to provide the molecular basis for the application of the gene targeting method to the renin gene, we also characterized the structure and expression of the renin gene in embryonic stem D-3 (ES-D3) cells and in its parental mouse strains, 129/Sv mouse. The Ren-1C gene is 12 kilobases long and is composed of 9 exons interrupted by 8 introns. Renin mRNA was undetectable in ES-D3 cells but was found to be highly expressed in the submandibular gland of the 129/Sv mouse, in addition to the kidney. Southern blot analysis reveals that ES-D3 cells carry the additional duplicated renin gene, Ren-2, consistent with the classification of the 129/Sv mouse as a two-renin gene strain.
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  • Takayuki KAMEI, Kunio SHIOTA, Tomoya OGAWA, Michio TAKAHASHI, Katsushi ...
    1993 Volume 39 Issue 1 Pages 25-31
    Published: 1993
    Released on J-STAGE: May 15, 2008
    JOURNAL FREE ACCESS
    Our previous work has demonstrated that immunoreactive basic fibroblast growth factor (bFGF)-like substance presents in germ cell nuclei of adult rat testis. In the present study, interactions of bFGF-like substance with a sub-uclear material were investigated. When nuclear extract from adult rat germ cell was analysed by heparin-Sepharose affinity chromatography and Western blot, germ cell nuclei contained 2 forms of bFGF-like substances with molecular weights of 16.5 and 24.5 kDa. The immunoreactivity for bFGF-like substance of germ cell nuclei decreased after treatment with either micrococcal nuclease or DNase l. Furthermore, South-Western blots analysis revealed these 2 forms of bFGF-like substance bind with high affinity to the exogenous salmon testis DNA. Thus, these results suggest that the bFGF-like substances are associated with DNA in the germ cell.
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  • Takayuki KAMEI, Kunio SHIOTA, Michio TAKAHASHI, Tomoya OGAWA, Katsushi ...
    1993 Volume 39 Issue 1 Pages 33-39
    Published: 1993
    Released on J-STAGE: May 15, 2008
    JOURNAL FREE ACCESS
    We have previously described immunoreactive basic fibroblast growth factor (bFGF)-like substance in adult rat testis and have shown that this bFGF-like substance presents in the nucleus of germ cells. In the present study, the distribution of bFGF was examined immunohistochemically in 3 or 12-day-old rat testes. In 3-day-old, immunoreactivity for bFGF was detected in the cytoplasm of gonocytes (prespermatogonia) and fetal Leydig cells. In contrast, in 12-day-old, immunoreactivity for bFGF was detected exclusively in the nucleus of germ cells. Thus, bFGF seems to translocate from the cytoplasm to the nucleus during formation of the first wave of spermatogenesis. Using reverse transcription-polymerase chain reaction (RT-PCR), bFGF mRNA was detected in the 3-day-old rat whole testis. Moreover, in situ hybridization analysis revealed gonocytes and fetal Leydig cells contain mRNA for bFGF. These results indicate that bFGF is synthesized in gonocytes and fetal Leydig cells before the formation of the first wave of spermatogenesis.
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  • Hiroshi HARAYAMA, Hiroshi KUSUNOKI, Seishiro KATO
    1993 Volume 39 Issue 1 Pages 41-45
    Published: 1993
    Released on J-STAGE: May 15, 2008
    JOURNAL FREE ACCESS
    The objectives of the present study are to characterize the motility and penetrability into zona-free hamster eggs of boar spermatozoa from 7 regions of the epididymis. Spermatozoa from the caput epididymidis were immotile or exhibited a flagellant or circular type of movement. However, 80% of the cells from the proximal cauda showed an intensive, forward movement. When the spermatozoa from each region of the epididymis were coincubated with zona-free hamster eggs after preincubation for inducing the acrosome reaction, they exhibited high penetration rates (95-100%) and average number of spermatozoa (3.4-5.8) in penetrated eggs irrespective of their origin. These findings indicate that most of the spermatozoa acquire the ability to exhibit progressive motility during their transit through the corpus, and that at least some of the cells located in the proximal caput have the ability to fuse with egg plasma membrane.
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  • Junko YOSHIDA, Junpei KIMURA, Azuma TSUKISE, Masaomi OKANO
    1993 Volume 39 Issue 1 Pages 47-54
    Published: 1993
    Released on J-STAGE: May 15, 2008
    JOURNAL FREE ACCESS
    The vomeronasal organ of the rat fetuses at 10, 11, 12, 13, 14, 16 and 18 days of gestation was studied by light and scanning electron microscopy. The primordium of the vomeronasal organ was observed first on 11 days of gestation as a thickening of the ectodermal layer on the mediorostral wall of the olfactory pit, whereas the olfactory placode appeared on the 10th day. The ectodermal thickening, which was identified as the vomeronasal placode, was situated on the separate geometric territory lying apart from the olfactory epithelium, moreover the vomeronasal placode appeared one day later than the olfactory placode did. The vomeronasal nerves and intraepithelial capillaries were observed on 13 days of gestation and immature vomeronasal cartilage cells accumulated around the vomeronasal duct on 14-days. The vomeronasal complex appeared 1 or 2 days later than the structures in the olfactory organ. These findings were similar to those reported in other rodents such as mice and golden hamsters, however in the hamster the vomeronasal duct, cartilage and gland, and in the mouse the vomeronasal nerve, appeared earlier respectively than those in the rat.
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  • Toshiyuki KUDO, Seiji SATO, Shizuyo SUTOU
    1993 Volume 39 Issue 1 Pages 55-63
    Published: 1993
    Released on J-STAGE: May 15, 2008
    JOURNAL FREE ACCESS
    Bovine male-specific repetitive DNA's were cloned for the sexing of bovine embryos by means of the polymerase chain reaction (PCR). The cloning method used was PERT (phenol emulsion reassociation technique), in which most male DNA's were absorbed by an excess amount of female DNA's to enrich Y-specific DNA's. Among 20 clones examined, one clone was male-specific. The others were gender-neutral; three of these were used as probes for internal controls. The male-specific clone was used as a probe to isolate other male-specific clones from a bovine male genomic DNA library. A clone thus obtained contained at least 3 male-specific EcoRI fragments. The original and the secondarily cloned DNA's were all sequenced. They were repetitive in nature. Oligonucleotides (20 mers) were synthesized on the basis of both the male-specific and gender-neutral DNA sequences as primers for PCR. A combination of 2 pairs of primers, i.e., male-specific and gender-neutral, gave discrete PCR products allowing discrimination between male and female DNA. The detection limit of the template DNA was 10 pg, equivalent to the DNA from 3 cells.
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  • Yoshiaki ITAGAKI, Seiji SATO, Yuji SHITANAKA, Toshiyuki KUDO, Yasumasa ...
    1993 Volume 39 Issue 1 Pages 65-72
    Published: 1993
    Released on J-STAGE: May 15, 2008
    JOURNAL FREE ACCESS
    We have previously cloned and characterized both bovine male-specific and genderneutral DNA's. Primers for the polymerase chain reaction (PCR) were synthesized on the basis of these DNA's to give differential PCR products between males and females. Serial dilution of bovine somatic cells of both sexes revealed that 10 cells were sufficient to discriminate males from females under our conditions. The usefulness of our primers to sex bovine embryos was confirmed by using purified control DNA's, cytogenetic analysis of demi-embryos, coincidence of a pair of asymmetrical biopsy samples and embryo transfer. All the embryos used here were produced by in vitro fertilization and culture methods. A successful determination of sex was made for 83 out of 84 embryos examined (98.8%). The sex ratio (54.2% male) did not differ significantly from the expected 1:1 ratio. The viability of biopsied embryos (86.3%) after 24 h culture in vitro was not significantly different from that of intact embryos (92.3%). Also, the mean numbers of cells of the biopsied (153.1, n=42) and intact (170.7, n=39) embryos were not significantly different. Out of seven recipients received the embryos with predicted sex, three went to term, producing one male and 2 female calves in accordance with the sex prediction before transfer. In conclusion, our method appears to be an efficient and reliable one to determine the sex of bovine preimplantation embryos.
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  • Hiroshi TORIUMI, Shigeo OHBA, Yasushi KUWABARA, Kaoru TAKAGI, Shigehis ...
    1993 Volume 39 Issue 1 Pages 73-77
    Published: 1993
    Released on J-STAGE: May 15, 2008
    JOURNAL FREE ACCESS
    When 19, 025 sows were administered with a calcium (Ca) and phosphorus (P) preparation containing 2.55-fold Ca and 1.54-fold P of the respective standardized mineral contents for breeders, a significantly (P<0.01) higher litter male/female ratio (secondary) was indicated in the calcium and phosphorus (Ca/P) treatment period compared to that of pre-treatment (control) interval. During the pre-treatment period, the litter sex ratio of sows indicated a value similar to previous findings. By supplementing this Ca/P preparation in breeders for a 2-year period, the male/female ratio was significantly enhanced without any apparent influences from the breed, mating pattern, seasonal variation and parity history. Further, effects on gestation period and litter size were negligible with supplementation of high Ca/P levels. On comparing the male ratio (1.395; taking the female ratio as 1) in the post-treatment (1.066) and pre-treatment (0.992) periods, the former interval manifested a significantly higher value, indicating favorable effects of the high Ca/P supplementary diet on the male/female ratio. Further, the longer the sows were supplemented with this Ca/P preparation, the higher was the male/female ratio. Terminating supplementation of the high Ca/P preparation during the post-treatment period manifested a recovery tendency of the male/female ratio to pre-treatment values.
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  • Tamotsu ISOGAI, Itsuo SHIMOHIRA, Kazuo KIMURA
    1993 Volume 39 Issue 1 Pages 79-84
    Published: 1993
    Released on J-STAGE: May 15, 2008
    JOURNAL FREE ACCESS
    The effects of the time after calving at flush of the first treatment (TIME) and the age at the last calving (AGE) on embryo production following repeated superovulation treatment were investigated for the determination of appropriate timing for starting embryo production in Holstein donors. TIME on the number of normal embryos recovered showed a linear decrease from 131-190 d to ?? 461 d, and there were more normal embryos in the classes of donors treated during 70-130 d, 131-190 d, and 191-250 d than in the class of donors treated during ?? 461 d (P<0.05). AGE described curves with the peak at 5 yr, and a significantly greater numbers of normal embryos were recovered from donors calved at 4 yr and 5 yr than from donors calved at ?? 7 yr (P<0.05). However, there was no statistically significant difference among donors calved at 3 to 6 yr. It is desirable that donor selection be made as early as possible for genetic improvement, and these results suggest that it is possible to select donor by records of the first lactation and to start embryo production about 70 d after the second calving.
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