• Applied Microscopy
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• v.31 no.3
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• pp.223-233
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• 2001

The spermiogenesis of a Korean octopus, Octopus minor, inhabiting western of Korea Sea was observed by electron microscopy . The obtained results are as follows: The spermiogenesis of Octopus miner proceeds through four stages; early- , mid- , and late-spermatid, and mature sperm. An early spermatid is a spherical cell looking light due to the low electron density. The acrosome formed from Golgi complex of the upper nucleus looks dark due to the high electron density. The extra-nuclear rod (enr) stemming from proximal centriole is transformed from round shape into oval shape, elongating to the upper nucleus. In our observation, the axoneme was being formed from distal centriole, and the manchette composed of a number of microtubules is also found around nuclear membrane. In a mid-spermatid, chromatins in the nucleus contract shaping fine threads, and the manchette is also observed around nuclear membrane. Especially, the spherical acrosome is transformed into long oval one which is tinged with a number of horizontal stripes and has the middle electron density. In a late-spermatid, chromatins in the nucleus contract thick and short. Furthermore, the mitochondrial sleeve, in which the axoneme is surrounded with mitochondria, is observed at middle piece. The axoneme has a typical structure of 9+2 and around it, 9 coarse fibers are observed. Also in the acrosome cavity of mature sperm, horizontal striation is found. However, regularly spaced processes are peculiarly observed in there. A sperm is about 390 fm long, whose head is bent a little like a banana while the acrosome region is helical. In the middle piece of sperm, $11\sim12$ mitochondria are surrounding coarse fibers that reach the main piece of tail, while nothing but 9+2 structured axoneme is found in the end piece.

• Journal of Animal Science and Technology
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• v.63 no.1
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• pp.58-68
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• 2021

Several herbs including Artemisia are known to possess conceptive property. In the present study, mouse spermatozoa were incubated with ethanol extract of Artemisia vulgaris leaves. The effect of extract on acrosome exocytosis was studied by labeling spermatozoa with fluorescein isothiocyanate (FITC) peanut agglutinin and by staining with Coomassie blue. Viability and membrane integrity were studied by Trypan-blue staining and hypo-osmotic swelling test. Artemisia extract at very low concentration caused precocious acrosome reaction and loss of sperm viability. Acrosome reaction increased remarkably from 22.63% to 88.42% with increasing extract concentration from 0 to 2,000 ㎍/mL. However, the viability loss of spermatozoa was increased from 11.71% in control to 63.73% in samples treated, evaluated by Trypan-blue staining method. Membrane damage caused by the extract, evaluated by hypo-osmotic swelling test was even low, ranging from 2.27% to only 24.23%. These results indicate that Artemisia extract might block fertilization by causing precocious acrosome exocytosis in spermatozoa. A direct contraceptive effect was tested by injecting the plant extract into the vagina of female mice and then allowing them to mate with normal males. The treated female mice delivered significantly fewer litters in comparison to the control.

• Proceedings of the Korean Society of Toxicology Conference
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• 2001.05a
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• pp.162-162
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• 2001

Selenium is an essential micronutrient for mammals and its biological functions are mediated by a selenoprotein. The mitochondrial capsule selenoprotein (MCS) is a selenoprotein that is necessary for the maintenance and stabilization of the sperm mitochondrial membrane.(omitted)

• Asian-Australasian Journal of Animal Sciences
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• v.22 no.12
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• pp.1614-1619
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• 2009

The present study was conducted to observe the effect of increased osmolality of basic tris extender supplemented with trehalose and sucrose on post-thawing quality (motility, progressive motility, viability, the rate of acrosome abnormality, total abnormality and membrane integrity) of Markhoz goat spermatozoa. Fresh semen samples were evaluated for motility and sperm concentration. Only semen samples with motility more than 70% and sperm concentration higher than $3{\times}10^{9}$ sperm/ml were used for cryopreservation. In Exp. 1, trehalose (50, 75 or 100 mM) and sucrose (40, 60 or 80 mM) were added to a basic tris diluent. Based on the results of experiment 1, the goal of Exp. 2 was to investigate the combinational effects of the highest and lowest concentrations ($T_{100}+S_{80}$ or $T_{50}+S_{40}$) of trehalose and sucrose. As the control, semen was diluted and frozen in the tris diluent without trehalose or sucrose. The results in Exp. 1 showed that all evaluated spermatozoa characteristics improved significantly after freezing and thawing (p<0.05) and at the same time the increase of trehalose and sucrose concentrations in basic extenders was seen, with the best results obtained for extenders containing 70 and 100 mM trehalose and 80 mM sucrose. Comparing these results with those of control diluents, the effects of supplementation were significantly (p<0.05) better. In Exp. 2, the results showed no significant differences (p>0.05) between $T_{100}+S_{80}$ and $T_{50}+S_{40}$ extenders, but the results of $T_{50}+S_{40}$were slightly better than obtained with $T_{100}+S_{80}$ diluents. Furthermore, the results of this experiment indicated that the sperm characteristics in the isotonic control extender were significantly (p<0.05) lower than examined extenders. In conclusion, the results of this study indicated that goat sperm can tolerate hypertonic trehalose and sucrose solutions better than isotonic control diluents in the freezing period. In particular, these positive effects have been shown for acrosome integrity, which is very important for the fertilization capacity of sperm. The data indicated that addition of trehalose plus sucrose to the freezing extender can be recommended for cryopreservation of goat spermatozoa, but more data is needed on pregnancy rate, acrosome reaction and IVF to ascertain the real effect.

• Journal of Life Science
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• v.11 no.1
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• pp.54-61
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• 2001

The comparative morphology of epididymal spermatozoa in the lesser white-toothed shrew, Crocidura suaveolens, the Japanese white-toothed shrew, C. dsinezumi and the big white-toothed shrew, C. lasiura, belonging to the subfamily Crocidurinae was studied with the light and electron microscopes. The spermatozoa of C. lasiura and C. dsinezumi were characterized by the large acrosome, serrated inner acrosomal membrane, common apical body and fistulous proximal centriole with slightly dense electron granular materials, which are the charateristics of the Crocidurinae. The C. suaveolens, however, is distinguished from the two species mentioned above in the sperm morphology. That is, the spermatozoa possess not only the charateristic of the Crocidurinae such aw the large acrosome, but also those of the Soricinae, i.e. the smooth inner acrosomal membrane, wavy, finger-like and electron-dense apical body, and the solid proximal centriole filled with electron-dense materials. The results suggest that C. suaveolens has conserved characteristics of the Soricinae.

• Journal of Animal Reproduction and Biotechnology
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• v.39 no.3
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• pp.179-193
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• 2024

Background: Because oxidative stress can induce decreased quality of caprine semen during the storage, there has been limitation for the use of stored semen in the assisted reproductive technologies. The present study, therefore, assesses the potential of Annona muricata (A. muricata) to reduce semen storage associated-damages. Methods: Semen was collected by electro-ejaculation from ten bucks, and extended with Tris-egg yolk (TEY) supplemented with A. muricata leaf aqueous extract (SAE) at 20 (SAE20), 40 (SAE40), and 80 (SAE80) ㎍/mL. Sperm variables including motility, motion characteristics, viability, membrane functionality, and DNA integrity were assessed at different storage periods (6, 24, 48, and 72 hr). In addition, oxidative stress indicators in the extender supplemted with SAE were also assessed for each group. Results: By adding SAE at 80 ㎍/mL in TEY, the storage of goat buck semen was improved, resulting in reduced loss of sperm motility, viability, DNA fragmentation, and membrane integrity during chilled storage at 4℃ for up to 72 hr. In addition, enrichment of TEY extender with SAE significantly (p < 0.05) reduced malondialdehyde, an indicator of oxidative stress, compared to the negative control. Conclusions: Supplementation of SAE in TEY extender can reduce buck spermatozoa liquid storage associated damages due to oxidative stress.

• Reproductive and Developmental Biology
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• v.31 no.2
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• pp.121-126
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• 2007

The objective of this study was to establish the optimal conditions for hypoosmotic swelling (HOS) test to assess the functional integrity of the membranes of boar fresh or frozen/thawed spermatozoa. When pooled semen sample was incubated for 30 min at $37^{\circ}C$ with different test solution of varied osmolarity, the highest percentage of HOS positive spermatozoa was observed in a 150 mOsmol fructose/Na-citrate solution (33.6%). Incubation time did not affect significantly the score of HOS positive spermatozoa observed in a 150 mOsmol fructose/Na-citrate solution at $37^{\circ}C$, but the osmolarity affected the score of HOS positive spermatozoa under the same condition above. Fresh semen was significantly better than frozen/thawed semen in semen parameters evaluated such as motility, viability, membrane integrity and lipid peroxidation (p<005). In the relationships of sperm parameters, motility vs viability, motility vs membrane integrity and viability vs membrane integrity were positively correlated ($0.82{\sim}0.94$) but lipid peroxidation vs other estimated factors was negatively correlated ($- 0.90{\sim}- 0.98$). Among the evaluation methods, motility vs Viability, motility vs membrane integrity and lipid peroxidation vs other estimated factors were significantly correlated (p<0.05). These results of this. study indicate that the optimal condition of HOST in boar spermatozoa is a 150 mOsmol fructose/Na-citrate solution for 30 min incubation at $37^{\circ}C$ and HOST can substitute the examination of motility, viability and lipid peroxidation.

• Journal of Animal Science and Technology
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• v.63 no.1
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• pp.46-57
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• 2021

Fruits with antioxidant enrichment can be an economically affordable supplement for mitigating oxidative damage prone spermatozoa membrane pathologies. Computer-assisted sperm analyzer and oxidative status were utilized to evaluate the impact of watermelon (Citrullus lanatus) fortification of dextrose saline as diluent for rooster semen and fertility response of hens inseminated. Watermelon juice and dextrose saline were used to formulate diluent of 7 treatments consisting of unextended semen (positive control), 10%, 20%, 30%, 40%, 50% and only dextrose saline (negative control) designated as Treatments 1-7. Pooled semen was obtained from fertile roosters and equilibrated with diluents at ratio 1:2 in the various treatments and were evaluated using computer software coupled microscope and seminal oxidative status assay. 168 laying hens randomly divided into 7 treatment of 8 replicates and 3 hen per replicate. Hen were everted, and semen (2 × 108 Spermatozoa) deposited intra-vagina and eggs collected over 8 weeks to assess fertility and hatchability of eggs laid. The result obtained revealed that watermelon-dextrose saline rooster semen diluent enhanced progressive motility, sperm kinetics and lowered non-progressive motility in T2-T6 compared to T7 over the 3 hours of evaluation. Watermelon addition to rooster semen diluent enhance the antioxidant capacity of rooster semen and lowered lipid peroxide generation. The percentage fertility was highest in T3 (81.01%) and T4 (81.24%) with lowest value obtained in T7 (73.46%). The hatchability of eggs set of hens inseminated with undiluted semen (71.46%) was lower than values for hens inseminated with watermelon inclusive extended semen (75.71%-80.39%). The optimal inclusion of 30%-40% watermelon in dextrose saline diluent enhance rooster semen kinetics, seminal oxidative stability and egg fertility.

• Journal of Animal Reproduction and Biotechnology
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• v.37 no.2
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• pp.113-120
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• 2022

Sperm cryopreservation is a fundamental process for the long-term conservation of livestock genetic resources. Yet, the packaging method has been shown, among other factors, to affect the frozen-thawed (FT) sperm quality. This study aimed to develop a new mini-straw for sperm cryopreservation. In addition, the kinematic patterns, viability, acrosome integrity, and mitochondrial membrane potential (MMP) of boar spermatozoa frozen in the developed 0.25 mL straw, 0.25 mL (minitube, Germany), or 0.5 mL (IMV technologies, France) straws were assessed. Post-thaw kinematic parameters were not different (experiment 1: total motility (33.89%, 32.42%), progressive motility (19.13%, 19.09%), curvilinear velocity (42.32, 42.86), and average path velocity (33.40, 33.62) for minitube and the developed straws, respectively. Further, the viability (38.56%, 34.03%), acrosome integrity (53.38%, 48.88%), MMP (42.32%, 36.71%) of spermatozoa frozen using both straw were not differ statistically (p > 0.05). In experiment two, the quality parameters for semen frozen in the developed straw were compared with the 0.5 mL IMV straw. The total motility (41.26%, 39.1%), progressive motility (24.62%, 23.25%), curvilinear velocity (46.44, 48.25), and average path velocity (37.98, 39.12), respectively, for IMV and the developed straw, did not differ statistically. Additionally, there was no significant difference in the viability (39.60%, 33.17%), acrosome integrity (46.23%, 43.23%), and MMP (39.66, 32.51) for IMV and the developed straw, respectively. These results validate the safety and efficiency of the developed straw and highlight its great potential for clinical application. Moreover, both 0.25 mL and 0.5 mL straws fit the present protocol for cryopreservation of boar spermatozoa.

• Korean Journal of Animal Reproduction
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• v.24 no.4
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• pp.439-449
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• 2000

This study was to investigate the expression of transgene after co-injection of spermatozoon and EGFP gene into mature oocytes in cattle. From frozen semen, spermatozoa were treated by DTT with 0.03% Tween-20, freezing and thawing or 0.02% Triton X-100 to disrupt their plasma membranes. The sperm injected oocytes were co-cultured with bovine oviduct epithelial cells in CRlaa, and expression of EGFP in embryos were observbed under epifluorescent microscope. Two pronuclei were formed in oocytes injected with sperm treated by DTT (44.6%), DTT-Tween-20 (48.4%), DTT-freezing (44.4%) and DTT-Triton X-100 (42.9%). Cleavage and blastocyst formation rates of bovine oocytes which injected with sperm treated by DTT, DTT-Tween-20, DTT-freezing, and DTT-Triton X-100 were 49.1, 58.5, 43.9, and 48.4% and 10.2, 13.0, 6.8, and 6.5%, respectively. Although the most of embryos were showing mosaicism, embryos expressing EGFP gene were observed in all treated groups. Therefore, these results indicate that membrane disrupted sperm could interact with exogenous DNA, and that this procedure may be useful to introduce foreign gene into bovine oocytes.