Journal of Reproduction and Development
Online ISSN : 1348-4400
Print ISSN : 0916-8818
ISSN-L : 0916-8818
Volume 41, Issue 2
May
Displaying 1-11 of 11 articles from this issue
Original Articles
  • Hiroshi HARAYAMA, Takashi MIYANO, Hiroshi MASUDA, Masashi MIYAME, Seis ...
    1995 Volume 41 Issue 2 Pages 113-121
    Published: 1995
    Released on J-STAGE: November 30, 2001
    JOURNAL FREE ACCESS
    Distribution of the 25-kDa epididymal anti-agglutinin for spermatozoa was examined by the Western blotting technique in fluid collected from boar rete testes and epididymides. Fluid was collected from the rete testes and various regions of the epididymis, and then plasma and sperm extracts were prepared. Antiserum was raised against the anti-agglutinin, and specific antibody was purified from it. In the plasma, a reaction between the anti-agglutinin and purified antibody was first observed in the proximal corpus epididymidis and became stronger in the more distal regions. In extracts from washed spermatozoa using the original sample buffer, anti-agglutinin was detected between the proximal corpus and the distal cauda and more strongly in the distal corpus. A thorough washing of spermatozoa on a discontinuous Percoll gradient brought about only minor changes in the detection pattern of anti-agglutinin. However, in samples extracted from washed spermatozoa using a modified sample buffer containing twofold detergent and twofold 2-mercaptoethanol, a stronger reaction was observed between the anti-agglutinin and purified antibody in the distal cauda than in the distal corpus. These results indicate that free or loosely sperm-bound anti-agglutinin increases in the corpus and cauda epididymides. It is also suggested that solubility of the anti-agglutinin associated with spermatozoa in the original sample buffer decreases in the cauda epididymidis.
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  • Maki MORITA, Hajime MIYAMOTO, Takashi ISHII, Miki SUGIMOTO
    1995 Volume 41 Issue 2 Pages 123-128
    Published: 1995
    Released on J-STAGE: November 30, 2001
    JOURNAL FREE ACCESS
    Ultrastructural features of the secretory cells of the goat's oviduct were studied at the early follicular phase, the follicular phase, the early luteal phase, the mid-luteal phase, and the late luteal phase. The secretory cells began to regenerate at the early follicular phase. At the follicular phase, numerous secretory granules of various size and electron density were seen in the secretory cells. At the early luteal phase, maximum secretory activity by exocytosis was apparent especially in the ampulla. At the mid-luteal and the late luteal phase, many ampullar secretory cells had large cytoplasmic protrusions which often contained nuclei. The pinching-off of protrusion was observed at these phases. Apoptosis-like fragments were seen in the ampullar epithelium at the late luteal phase, when the secretory cells were atrophied.
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  • Satoru YAMAZAKI, Kyoko KAWAHATA, Taichi GOTO, Jutaro TAKAHASHI, Yasuhi ...
    1995 Volume 41 Issue 2 Pages 129-132
    Published: 1995
    Released on J-STAGE: November 30, 2001
    JOURNAL FREE ACCESS
    Early pregnancy factor (EPF) is detectable in sera of pregnant cattle within 24 to 48 h after fertilization. It has been considered that the repeat breeder is relatively sterile as a result of fertilization failure and the early embryonic loss. This study used the rossete inhibition test to clarify whether repeat breeding is a result of early embyronic loss. We measured serum EPF in cattle that were diagnosed as repeat breeders to examine the incidence of early embryonic losses after artificial insemination (AI) (n=20). There were no significant differences in the presence of EPF on day 3 after AI for 20 repeat breeders and controls. Furthermore, EPF-positive rates decreased about 60% by day 21 after AI for both repeat breeders and a control group. It is suggested that a high incidence of early embryonic losses takes place in cattle, and we attempted embryo transfer (ET) as the treatment of choice.
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  • Yukio KANAI, Noritoshi YAGYU, Takashi SHIMIZU
    1995 Volume 41 Issue 2 Pages 133-139
    Published: 1995
    Released on J-STAGE: November 30, 2001
    JOURNAL FREE ACCESS
    An experiment was designed to determine if hypogonadism in heat stressed animals is associated with reduced endocrine function of the pituitary or the ovary. A long day-induced anestrous goat model in which ovulatory follicular growth is known to be induced by hourly injections of a small dose of GnRH was used. Ten anestrous animals, divided into two groups of 5 each, were treated with GnRH (3 ng/kg body weight) for 48 h under two temperature conditions, 25 C (Control) or 38 C (Heat stress). Changes in plasma concentrations of LH and estradiol were monitored at 2 to 4 h-intervals. In addition LH was monitored every 15 min for 16 h starting at 4 h before the first GnRH injection. GnRH treatment resulted in a prompt and sustained increase in plasma estradiol concentrations accompanied by a preovulatory LH surge within 24 h of GnRH treatment in all control animals. Heat stress abolished such ovarian responses in 4 out of 5 animals with the estradiol concentrations remained unchanged or increased slightly (7.7 pg/ml vs 2.0 pg/ml, P<0.01). Before GnRH treatment, the frequency but not the amplitude of the LH pulse was higher in the heat stressed group (0.75/4 h vs 2.75/4 h, P<0.05). The active pulsatile secretion was associated with slightly lowered estradiol concentration during this period (2.4 pg/ml vs 1.4 pg/ml P<0.05). GnRH-treatment resulted in a synchronized pulsatile secretion of LH in both groups with the frequency of LH pulses being controlled at once every hour (3.67/4 h vs 4.17/4 h). The amplitude of GnRH-induced LH pulse (0.81 ng/ml vs 1.62 ng/ml, P<0.01) as well as the baseline (0.73 ng/ml vs 2.02 ng/ml, P<0.05) and mean concentrations (1.19 ng/ml vs 2.81 ng/ml, P<0.05) of LH were significantly higher in the heat stressed group. These results demonstrate that heat stress abolishes GnRH-induced ovulatory responses in anestrous goats not mediated by a decreases in pituitary LH secretion, but rather by reducing responsiveness of the ovary to LH.
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  • Shiro KURUSU, Satoshi SHINGAKI, Yoshiko MUNEKATA, Mitsumori KAWAMINAMI ...
    1995 Volume 41 Issue 2 Pages 141-147
    Published: 1995
    Released on J-STAGE: November 30, 2001
    JOURNAL FREE ACCESS
    To investigate the presence and possible physiological role of cytosolic phospholipase A2 (cPLA2) in corpora lutea (CL), immunoblot analysis and activity measurements of the enzyme were performed on the cytosol of rat CL. The mouse monoclonal antibody against human cPLA2 immunoprecipitated a single protein (ca 110 kDa Mr) in several rat tissues, and the immunoreaction decreased in the absence of EGTA during homogenization. These results suggest that the antibody can recognize rat cPLA2. Subsequent immunoblot analysis detected little cPLA2 in adult rat CL on days 6 and 12 of pregnancy. In both adult and immature rats, the enzyme protein was not detectable in CL on day 6 of pseudopregnancy (PSP6); however, it became detectable in CL of PSP12. In these models, luteal cytosolic PLA2 activity, as determined by the release of fatty acid from liposomes of phosphatidylcholine, clearly increased from PSP6 to PSP12, and the increase persisted during treatment with the secreted PLA2 inactivator ditiothreitol. The present results suggest a possible involvement of cPLA2 in luteal eicosanoids production and luteolytic processes.
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  • Seizo HAMANO, Akiko KOIKEDA
    1995 Volume 41 Issue 2 Pages 149-152
    Published: 1995
    Released on J-STAGE: November 30, 2001
    JOURNAL FREE ACCESS
    The objective of this study was to compare in vitro development to the blastocyst stage of bovine embryos obtained by in vitro fertilization within a breed and between different breeds of cattle. The timing of maturaion division did not differ among oocytes of Japanese Black (JB), Japanese Brown (JBr) and Japanese Shorthorn (JS); 91.5 to 97.5% of the oocytes were at the second metaphase after 21 h of culture. The rate of development to the blastocyst stage was also similar (34.6 to 37.5%) among three kinds of embryos (JB, JBr and JS) that were fertilized in vitro and cultured for 6 to 8 days after insemination. However, significantly higher rates of development to the blastocyst stage were observed (55.0 to 67.2%) when spermatozoa from a JB bull were used for in vitro fertilization of JBr, JS and JP (Japanese Polled) oocytes as compared to the rate of development in control JB embryos (36.7%). Development to the 2- to 4-cell stage did not differ significantly. These results suggest that the effect of heterosis becomes apparent at the blastocyst stage, which might be under the control of zygotic gene activity.
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  • Naoki AOYAMA, Osamu TOYAMA, Yoshito YAMASAKI, Yoshinori MIZOGUCHI, Kao ...
    1995 Volume 41 Issue 2 Pages 153-158
    Published: 1995
    Released on J-STAGE: November 30, 2001
    JOURNAL FREE ACCESS
    Using a perfused rabbit ovary, we studied the effect of luteinizing hormone (LH) secretion on the ovulation time and the maturity of ova with 2 modes of perfusion. One mode consisted of a constant level of human chorionic gonadotropin (hCG) in the medium (the constant mode of perfusion), and the other mode mimicked the profile of LH surge in the rabbit (the surge mode of perfusion). The mean numbers of ova ovulated in the 2 modes were almost equal. The mean ovulation time in the constant mode was shorter than that in the surge mode. In the constant mode, the percentage of ova in germinal vesicle stage was larger, and in metaphase II was smaller, than those in the surge mode, while the difference in metaphase I was insignificant. The results indicate that the maturities of the ova in the surge mode were more advanced than those in the constant mode. A reason for the occurrence of the difference might be the early occurrence of ovulation in the constant mode due to excessive action on the follicular wall by various proteolytic enzymes, which are activated by the higher concentration of hCG, when compared with that of the surge mode, in the latter half of the perfusion.
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Research Notes
  • Iwao HONTA, Kazuei ITO, Jutaro TAKAHASHI, Yasuhisa YASUDA
    1995 Volume 41 Issue 2 Pages 159-163
    Published: 1995
    Released on J-STAGE: November 30, 2001
    JOURNAL FREE ACCESS
    To investigate whether early pregnancy factor (EPF) is necessary for embryonic development and implantation, anti-bovine EPF antibody was injected (2.5 mg/rat per injection) into pregnant rats intraperitoneally at 12, 36, 60 and 84 h after mating (total dose of the antibody; 10 mg). By 4.2 days post coitum (p.c.), 47.3% of embryos in the group of animals treated with anti-EPF antibody had developed to the blastocyst stage, compared with 97.2% and 95.4% in the control groups (injected with anti-nonpregnancy antibody and saline, respectively). Rats were administered with 2.5 mg of anti-EPF antibody on 0.5 and 1.5 day p.c.(postfertilization period: group 1), on 3.5 and 4.5 day p.c. (preimplantation period: group 2), and on 6.5 and 7.5 day p.c. (postimplantation period: group 3). When the animals were examined on 10 day p.c., the implantation rates in group 1 and group 2 had significantly decreased by 49.5% and 47.5%, respectively, compared with the control groups which were given injections of either anti-nonpregnancy antibody or saline. In group 3, however, the implantation rates had not been affected. Our data essentially agreed with the results of Igarashi [1] and Athanasas-Platsis et al. [2, 3] who investigated effects of anti-mouse or human EPF antibody in pregnant mice, and showed that anti-bovine EPF antibody crossreacted with rat EPF in vivo.
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  • Hiroshi NAGASHIMA, Naomi KASHIWAZAKI, Rodney J. ASHMAN, Mark B. NOTTLE
    1995 Volume 41 Issue 2 Pages 165-170
    Published: 1995
    Released on J-STAGE: November 30, 2001
    JOURNAL FREE ACCESS
    This study aimed at increasing the efficiency of current protocols for the production of live piglets from cryopreserved embryos. Porcine early hatched blastocysts (n=163) were cryopreserved in liquid nitrogen in the presence of 1.5 M glycerol (1.5G), 1.5 M glycerol + 0.25 M sucrose (GS) or 1.8 M glycerol (1.8 G) using a conventional slow cooling method, and their post-thaw survival assessed following in vitro culture or transfer. Post-thaw survival rates of embryos were 85.1%, 91.6% and 94.1% in 1.5 G, 1.8 G and GS groups, respectively. More embryos which maintained prefreeze morphology were obtained in GS (41.7 %, P<0.001) and 1.8 G groups (29.1%, P<0.025) compared with 1.5 G group (10.0%). Transfer of 171 frozen-thawed embryos from the three groups after culture for 24 h to 4 recipients did not result in pregnancy. In contrast, transfer of 66 embryos from GS group to a recipient 3 h after thawing resulted in the birth of 4 piglets. The results demonstrate that addition of 0.25 M sucrose to the freezing media improves post-thaw survival compared with freezing with glycerol alone.
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Technical Notes
  • Kazufumi GOTO, Yasuaki TANIMOTO, Shoji OOKUTSU, Yoshihiko NAKANISHI, K ...
    1995 Volume 41 Issue 2 Pages 171-174
    Published: 1995
    Released on J-STAGE: November 30, 2001
    JOURNAL FREE ACCESS
    Follicular oocytes were collected from FSH-stimulated (n=15) and unstimulated (n=14) cows by ultlrasound-guided follicular aspiration. The mean number of aspirated follicles (≤2 mm) was significantly (P<0.05) higher in FSH-stimulated (13.7) than in unstimulated (8.5) cows. There was no significant difference in the mean number of recovered oocytes between FSH-stimulated (7.3) and unstimulated (5.6) cows. The proportions of oocytes that cleaved and developed to the morula/blastocyst stage were 53 and 23% for FSH-stimulated and 44 and 12% for unstimulated cows, respectively, indicating no significant differences between the 2 groups. Two calves were born after transfer of 8 embryos originating from FSH-stimulated cows, and one set of twins was born after the transfer of 3 embryos originating from unstimulated cows. In another experiment, oocytes were aspirated weekly for one month from 5 unstimulated cows. A total of 10 to 41 oocytes were collected from each cow, and some of them were developmentally competent in vitro after in vitro maturation and fertilization. No detrimental effects were observed in these cows by clinical and postmortem examinations.
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  • Woo-suk HWANG, Choong-ho JO, Byeong-chun LEE, Tae-young SHIN, Kye-seon ...
    1995 Volume 41 Issue 2 Pages 175-179
    Published: 1995
    Released on J-STAGE: November 30, 2001
    JOURNAL FREE ACCESS
    The objective of this study was to produce calves derived from in vitro maturation (IVM) and fertilization (IVF) of bovine follicular oocytes. Oocytes aspirated from small antral follicles (2-5 mm) of ovaries obtained at a local abattoir were matured and fertilized in vitro. At 18 h after insemination with frozen-thawed semen from Korean Native Cattle, oocytes were co-cultured for 6-7 days in tissue culture medium 199 with bovine oviduct epithelial cells. After co-culture, good quality late morulae or early blastocysts were selected by morphological criteria under stereo microscopic evaluation and were transferred to recipients on day 6 or 7 of the cycle (estrus=day 0). Recipients were monitored by observation for estrus and rectal palpation every 30 days beginning 55 days after transfer. Four of seven recipients became pregnant and one of them delivered a normal calf. This result is the first production of calf derived from IVM and IVF in Korea.
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