• Journal of Embryo Transfer
• /
• v.18 no.2
• /
• pp.151-156
• /
• 2003

The objective of this study was to investigate response of corpus luteum, recovery rate of embryos and production of transferable embryos according to superovulation by PEG 30% FSH in Hanwoo. Cows were selected as recipient, subsequent were superovulated with a total of 400 mg NIH-FSH-P1(Folltropin-V, Canada) given by one shot subcutaneously. At the time of five days after Folltropin injection, 25mg of a PGF$_2$a was injected and cows were inseminated 12 and 24h after the onset of estrus. Seven days after insemination, embryos were collected non-surgically and were cryopreserved by direct transfer methods. The results are summarized as follows : 1. Response of corpus luteum following the superovulation in Hanwoo, right ovary were 52.8%(271/513) and left ovary were 47.1%(242/513), respectively (P<0.05). 2. Recovery rate of embryos following the number of corpus luteum were 83.0%(426/513). 3. Mean number of embryos recovered and transferable embryos were 7.74 and 6.43, respectively (P<0.05). 4. In the total number of transferable embryos per flush were collected 6.4 and all saved transferable embryos were 355.

• Journal of Embryo Transfer
• /
• v.27 no.3
• /
• pp.141-147
• /
• 2012

This study was operated to establish induction using ultrasonography by estimating the relation of follicle size and estrus manifestation. Clinical estrus symptoms were observed 97.4% in cows and 87.5% in heifers when overall 55 cows were induced to estrus in a single dose of $PGF_{2{\alpha}}$ after verifying CL through ultrasonography, which means estrus hours among those 52 cows showing the clinical estrus symptoms were estimated 2.39 days on cows and for 2.37 days on heifers which showed no differences (p>0.05). The estrus manifestation hours according to the follicle size in cows didn't have any significance each other (p>0.05), though estrus hours was 54 hours (the shortest) with follicle size bigger than 10 mm and were made up within 69 hours. The estrus manifestation hours according to the follicle size in heifers didn't have any significance each other (p>0.05) and took around 42 hours (the shortest) with follicle size of 5mm (the smallest) and were made up within 66 hours. Follicles after $PGF_{2{\alpha}}$ injection were ovulated and assigned to many phases as follows; Group 1 (growing phase) - continuously growing into ovulation, Group 2 (growing and static phase) - delaying in growth after the growth of follicles, Group 3 (static and growing phase) - growing after growth delay, Group 4 (regressing and new growing phase) - the follicle is closed and a new follicle grows. In addition, the process of follicle development and estrus hours had no significance each other (p>0.05), though estrus manifestation hours in Group 1 and 2 was relatively short, and in Group 3 and 4 for a relatively long time. In the result of all above, the estrus manifestation hours after $PGF_{2{\alpha}}$ injection has no differences accoring to the follicle size in cows and heifers. Therefore, High pregnancy rate is obtained when practicing artificial insemination within 3 days in estrus or TAI in 72 to 80 hours after adminitrating $PGF_{2{\alpha}}$.

• Applied Microscopy
• /
• v.33 no.2
• /
• pp.105-116
• /
• 2003

In Vitro fertilization of U. unicinctus eggs observed by immunofluorescence and electron microscopes revealed an overview of the meiotic pattern of the tide animals. The eggs have been fertilized early at germinal vesicle stage, followed by germinal vesicle break down (GVBD), but pre-mitotic aster like structure could not be resolved by the methods employed in this work. The meiotic features, such as rotation-shift movement of spindle fibers, behavior of spermatozoonmonaster in the egg cytoplasm and active spindle fiber of the 1st polar body, have been observed. The antitubulin-FITC fluorescence show the 2nd meiotic apparatus appeared firstly parallel to the tangential line of the oolemma, proceeding the meiosis, its bipolarity is rotated and shifted towards the oolemma. The polar bodysite of the oolemma was not amorphous, but active in a sense of anti-tubulin-FITC reactions during the extrusions of the polar bodies. The immunofluorescence reactions of the spermatozoon centriole appeared at a later stage of the 2nd meiosis. During the time periods, the fertilized spermatozoon resided in the egg cytoplasm. Activating the centrioles, spermatozoon approaches towards the chromosomal materials of the 2nd oocyte. This suggests that spermatozoon centrioles initiate and play a roll to fuse male and female pronuclei.

• Reproductive and Developmental Biology
• /
• v.36 no.2
• /
• pp.115-120
• /
• 2012

This study was conducted to examine the optimal concentration and treatment time of antioxidants for inhibition of the ROS generation in bovine somatic cell nuclear transfer (SCNT) embryos. Bovine oocytes were activated parthenogenetically, during which oocytes were treated with various antioxidants to determine the optimal concentrations and kind of antioxidants. Determined antioxidants were applied to oocytes during in vitro maturation (IVM) and/or SCNT procedures. Finally, antioxidant-treated SCNT embryos were compared with in vitro fertilized (IVF) embryos. $H_2O_2$ levels were analyzed in embryos at 20 h of activation, fusion or insemination by staining of embryos in $10{\mu}M$ 2'7'-dichlorodihydrofluorescein diacetate (H2DCFDA) dye, followed by fluorescence microscopy. $H_2O_2$ levels of parthenogenetic embryos were significantly lower in $25{\mu}M$ ${\beta}$-mercaptoethanol (${\beta}$-ME), $50{\mu}M$ L-ascorbic acid (Vit. C), and $50{\mu}M$ L-glutathione (GSH) treatment groups than each control group ($24.0{\pm}1.5$ vs $39.0{\pm}1.1$, $29.7{\pm}1.0$ vs $37.0{\pm}1.2$, and $32.9{\pm}0.8$ vs $36.3{\pm}0.8$ pixels/embryo, p<0.05). There were no differences among above concentration of antioxidants in direct comparison ($33.6{\pm}0.9{\sim}35.2{\pm}1.1$ pixels/embryo). Thus, an antioxidant of $50{\mu}M$ Vit. C was selected for SCNT. $H_2O_2$ levels of bovine SCNT embryos were significantly lower in embryos treated with Vit. C during only SCNT procedure ($26.4{\pm}1.1$ pixels/embryo, p<0.05) than the treatment group during IVM ($29.9{\pm}1.1$ pixels/embryo) and non-treated control ($34.3{\pm}1.0$ pixels/embryo). Moreover, $H_2O_2$ level of SCNT embryos treated with Vit. C during SCNT procedure was similar to that of IVF embryos. These results suggest that the antioxidant treatment during SCNT procedures can reduce the ROS generation level of SCNT bovine embryos.

• Journal of Embryo Transfer
• /
• v.25 no.4
• /
• pp.263-266
• /
• 2010

Prediction of semen's fertilizing ability used in artificial insemination (AI) is one of very important factors on pig reproductive performance. In vitro fertilization (IVF) has been used for indirect evaluation of sperm's fertilizing ability and it has been showed as highly correlated index. In swine industry, increasing interest in preservation of boar semen raises questions on the sperm motility from semen used in commercial AI centers. Mitochondria in sperm mid-piece generate the energy to support motility and could be an explanation of impaired fertility. Objective of this study was to suggest usable sperm motility to farms in measuring the effect of sperm motility and sperm abnormality on in vitro production of embryo in which sperm's fertilizing ability can be determined indirectly. Semen samples were provided from local AI center and used within 3 days after collection. Semen samples were divided by 4 different motile groups (>70%; 61~70%; 51~60%; <50%) using CASA (computer-assisted sperm analysis) on the days of IVF. Developmental rate to the blastocyst stage from over 61% motile sperm group showed significantly higher rate than below 60% motile sperm group ($16.5{\pm}0.7{\sim}18.4{\pm}0.8%$ vs $6.3{\pm}0.8{\sim}11.5{\pm}0.7%$, p<0.05). In experiment to determine the relationship between sperm motility and viability and abnormality, over 61% motile sperm groups showed significantly higher viability rate compared to below 60% motile sperm groups ($84.8{\pm}4.0{\sim}88.1{\pm}4.0%$ vs $69.1{\pm}4.0{\sim}74.2{\pm}4.0%$, p<0.05). On the other hand, morphological sperm abnormality showed significantly higher in over 70% motile sperm group ($10.2{\pm}2.2$ vs $16.0{\pm}2.2{\sim}21.0{\pm}2.2%$, p<0.05). In experiment to find the correlation between sperm motility of 4 different motile groups and amount of mitochondria, lower motility group also showed lower level of mitochondria (p<0.05). The mitochondria parameter used in this study showed another possibility to differentiate the sperm motility. Taken together, because below 60% motile semen used in AI reduce the fertility, AI centers should provide the over 60% motile sperm to the farms at the time of AI.

• Clinical and Experimental Reproductive Medicine
• /
• v.27 no.3
• /
• pp.267-273
• /
• 2000

Objectives: We have previous reported that thawed testicular sperm and sperm extracted from seminiferous tubule could achieved optimal fertilization and pregnancy in azoospermic patients. However, thawed testicular sperm did not show motility in many cases. Therefore we studied viability of immotile sperm extracted from frozen-thawed seminiferous tubule using hypo-osmotic swelling (HOS) test and eosin-Y test. Materials and Methods: After sperm extraction using for ICSI, the remained sections of seminiferous tubules were frozen with a computerized freezer. For thawing and preparation of testicular sperm, the seminiferous tubules were thawed by removing from $LN_2$ and letting them at room temperature for 10 min followed by %37^{\circ}C$ water bath for 10 min. The prepared samples were washed for free of preservation medium and sperm preparation method described previous. Sperm was suspended in 0.1 ml hypoosmotic solution. After 30 minutes, the type of distally coiled sperm were assessed. Results: In 44 cases of cryopreservation of seminiferous tubules in obstructive azoospennic patients, the fertilization rates with 2PN were 71.4% and pregnancy rates were 34.1%. The presence of motile spermatozoa on subsequent post-thaw testicular sperm remarked 15.1% and were increased to 77.3% just before ICSI. After sperm extracted from frozen-thawed seminiferous tubule, 3 hrs later in in vitro culture, the cases of presence of motile sperm, reaction of hypo-osmotic swelling test and viable sperm were 63.6% (28/44), 93.2% (41/44), and 77.3% (34/44), respectively. Conclusions: Just after post-thawed testicular sperm did not showed motility. Although motility was gained after in vitro culture, many cases showed non-motile sperm until optimal insemination time. However, HOS test showed positive reaction in non-motile sperm. Therefore, HOS test is an alternative method for the selection of viable sperm for ICSI.

• Korean Journal of Ichthyology
• /
• v.34 no.4
• /
• pp.244-252
• /
• 2022

The aim of this study was to investigate the early life history of Squalidus gracilis majimae by observing the development of the eggs and morphology of juvenile fish. Squalidus gracilis majimae used in this study were captured in 2021 June from a tributary flowing into Wicheon, located in Uiseong-gun, Gyeongsangbuk-do. For artificial insemination, ovaprim was injected at 0.5 mL/kg. Egg development occurred 12 h after injection. The size of mature eggs of S. gracilis majimae was 0.80~0.85 mm (0.83±0.01 mm, n=30). Incubation time at 22±1℃ ranged from 62.5 to 69 h. The length of larvae immediately after hatching was 2.39~2.53 mm (2.44±0.04 mm, n=30). On the 18th day after hatching, the larvae had a total length of 8.51~9.84 mm (8.95±0.42 mm, n=30), and the tip of the spine was completely curved. On the 22nd day after hatching, the juvenile fish had a total length of 12.6~13.6 mm (13.0±0.36 mm, n=30), with dorsal fins at iii7 and anal fins at i7. Egg size and hatched larvae size were similar for S. gracilis majimae and S. multimaculatus, and cupulae showed a singularity as it disappeared last among related species.

• Journal of Practical Agriculture & Fisheries Research
• /
• v.20 no.2
• /
• pp.49-56
• /
• 2018

Recently, attempts have been made by dairy farms to shorten the time until the first estrus by feeding animals a high-energy diet during the heifer period. Energy consumption during the heifer period has been shown to be related to milk yield and reproductive performance. The present study was conducted to analyze the relationship between reproductive performance and energy intake in heifers. There were no differences in height or body weight between animals that were fed the Korean standard diet versus a high energy diet (110% of the Korean standard). However, there was a difference in the timing of the first estrus: 329.3 days under normal energy conditions versus 311 days under high energy conditions. There was no difference in the number of artificial insemination per gestation, but both the rate of female calf production and the weight of individual female calves increased in response to the high energy diet. Based on these results, it may be predicted that the reproductive performance of dairy cows will increase when the animals are fed a high energy diet (110% of the Korean standard). These results demonstrate a correlation between the amount of energy consumed and reproductive performance, and will be of great value to the dairy farming industry.

• Clinical and Experimental Reproductive Medicine
• /
• v.34 no.3
• /
• pp.159-166
• /
• 2007

Objective: To investigate whether semen parameters in infertile couples who undergone intrauterine insemination (IUI) change in the subsequent IUI cycle and the subsequent in vitro fertilization (IVF) cycle. Methods: Fifty-three infertile couples who had failed to become pregnant after the first IUI cycle with computer-assisted semen analysis (CASA) were included. After the first IUI, thirty-eight couples underwent the second IUI (Group 1), and fifteen underwent IVF-ET procedure (Group 2). All semen parameters including semen volume, concentration, motility and total motile sperm count were analyzed in the second IUI or IVF-ET procedure for comparison with the result of first IUI. Results: There were no significant differences in husband age, interval between the first and second procedure and cause of infertility. In Group 1, only sperm motility at the time of the latter IUI was significantly decreased when compared to the former IUI irrespective of the first semen parameters. In Group 2, sperm concentration, motility and total motile sperm count at the time of subsequent IVF were lower than the former IUI. By sub-analyses of Group 2, in the group of optimal semen parameter at IUI cycle, sperm concentration and total motile sperm count at the time of subsequent IVF were lower than the former IUI, while in the group of suboptimal semen parameter at IUI cycle, only sperm motility at the time of subsequent IVF were lower than the former IUI. Conclusion: The semen parameters in couples converted to IVF cycle were more adversely affected than those remained in IUI cycle. Further study on psychological stress should be necessary to explain the reason.

• Development and Reproduction
• /
• v.11 no.2
• /
• pp.127-135
• /
• 2007

In order to monitor the developmental features of embryos, larvae, and juveniles of Konoshiro Gizzard Shad Konosirus punctatus, the fertilized eggs were gotton using artificial insemination. Konosirus punctatus were caught in Mankyung-myeon, Kimjae, Jeollabuk-do at June of 2004, and experiments were carried out in Ichthyology laboratory in Chonnam National University. Konosirus punctatus spawned draft egg from March to June. The fertilized eggs were cultured in $19.0{\sim}23.0^{\circ}C$(mean, $21.2^{\circ}C)$. The eggs had spherical shape and the diameter is $1.14{\sim}1.34\;mm$(mean, 1.21 mm). The lens began to appear from 35 hr 53 min after fertilization. At the moment, the movement of larvae was more active, and the tail was separated completely from yolk, the heart had forms, and melanophore appeared. Hatching was observed from 37 hr 10 min after fertilization. The total length of the hatched larvae was $4.26{\sim}5.30\;mm$(mean, 4.96 mm), but the mouth and anus were not opened at the time when the larva had yolk sack, and had $22{\sim}27$ myometium, and the anus located just abdominal front of the tail fin, and melanophore accumulated in the eye. Post-larvae used yolk completely after 2 day of hatching, and the total length was $4.96{\sim}5.74\;mm$(mean, 5.24 mm). From 16 days after hatching, the tail had curved tail end, and appeared the stems for pectoral, dorsal, and caudal fins. At 53 days after hatching, the total length of post-larvae was $27.11{\sim}34.09\;mm$(mean, 30.11 mm), and the frontal part of head developed like an adult one. At this time, fins and body are transferred to those of adult Konosirus punctatus. Fishes have a different shape and location of melanophore even in the same family. This research was tried to elucidate the early developmental features of Konosirus punctatus, together with species-specific pattern of melanophore.