• Korean Journal of Animal Reproduction
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• v.26 no.3
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• pp.253-261
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• 2002

This study was carried out to investigate the semen characteristic, motility and viability and sperm motion characteristic by CASA test for establishing the Jindo-dog's semen freezing system. The results obtained are as follow: 1. The semen was collected 63 times. Average volume of semen, concentration of sperm, total number of sperm, progressive motility and viability were 3.8 $m\ell$ 145.6$\times$10$^{6}$ cells/$m\ell$, 396.2 x10$^{8}$ cells, 79.7% and 89.5%, respectively. Also, Fawn (Yellow) Jindo-dog comparing with White Jindo-dog showed better concentration of sperm, total number of sperm, progressive motility and viability. Among all dogs, the results of No. 2 Fawn Jindo-dog were the best. 2. The average progressive motility and viability of semen from 46 times were 73.5%, 82.3% before freezing and 51.1%, 64.9% after freezing. So, the freezing of semen has affected the progressive motility and viability. The progressive motility and viability of Fawn Jindo-dog's semen, before and after freezing, were better than White Jindo-dog. And No. 2 Fawn Jindo-dog showed the best results and showed significantly different among all dogs (P<0.05). 3. The 44 times-tested .esults by CASA system were as follow; MOT (motility) 65.6%. PROG (progressive motility) 54.8%, VAP (average path velocity) 75.3 ${\mu}{\textrm}{m}$/sec, VCL (curve linear velocity) 90.0 ${\mu}{\textrm}{m}$/sec, VSL (straight-line velocity) 69.4 ${\mu}{\textrm}{m}$/sec and ALH (amplitude of lateral head displacement) 4.4 ${\mu}{\textrm}{m}$. Although the motion characteristic of frozen semen were not significantly different between White and Fawn Jindo-dog, No. 2 Fawn Jindo-dog showed the best results and was significantly different among all dogs (P<0.05). 4. The success rate of frozen semen production between White and Fawn Jindo-dog were 43% (13/28), 94% (33/35), respectively, and the total success rate was 73% (46/63). The freezing-ability of Fawn Jindo-dog's semen was better than the other. Conclusively, the present results indicated that the characteristic and motility of Jindo-dog') semen were suitable for processing frozen semen, artificial insemination and mass production system. Also, the selection of suitable dog-breed was so important because the characteristic and freezing-ability of semen were significantly different between White and Fawn Jindo-dogs and among all individual dogs.

• Korean Journal of Veterinary Research
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• v.47 no.1
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• pp.135-138
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• 2007

A 3 year-old female Afghan hound came to the Veterinary Referral Hospital of the College of Veterinary Medicine, Seoul National University for artificial insemination (AI) with frozen semen. In order to inseminate, semen was frozen in USA 3 years ago. Frozen semen was sent by air from Santiago to Seoul for AI. The stud died 2 years ago, so we could only use a limited amount of frozen semen in that estrus cycle. The number of total motile spermatozoa was $59.4{\times}10^6$and the total volume was 1.2 ml. The frozen spermatozoa were thawed in $70^{\circ}C$ water for 8 sec, which were then deposited at the bilateral uterine horns by a surgical method. The number of corpus luteum was 6. Sixty days after artificial insemination resulted in the birth of 4 puppies, all of which are alive and healthy.

• Proceedings of the Korean Society of Embryo Transfer Conference
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• 2002.11a
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• pp.92-92
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• 2002

Considerable attention has been focused on the cryopreservation of semen and estrus induction in dog, as consequence of poor productivity caused by long anestrus period, in order to enhance the productivity of youngs and to preserve the breeds. The objectives of this study were to evaluate semen quality after cryopreservation and to evaluate the Pregnancy rate after insemination (AI). Fifty infertilie dogs (age 2∼3 years) were selected for the study and divided into three different estrus induction treatment groups. Group 1: dogs (n=15) were given clomifene (0.1 mg/kg) orally for five days at 12 hr intervals. Group 2: dogs (n=15) were given bromocriptine (50 $\mu\textrm{g}$/kg) orally for five days at 12 hr intervals, followed by single injection intravenously of 500 IU GnRH (Group 3, n=20) when pro-estrus occurred. The rates of pregnancy in estrus inducted dogs mated naturally compared to those inseminated artificially with ejaculated fresh semen and frozen-thawed semen. Estrus detection was performed using the method of vaginal smear and confirmed by the plasma progesterone assay. The ejaculated semen to freeze was exposed to a mixture of Tris extender with cryoprotectant (Trisma, 81 mM: TES, 209 mM: citric acid, 6 mM; glucose, 5 mM; glycerol, 8%) and cryopreserved gradually by slow-cooling at 17 cm above the surface of liquid nitrogen (LN$_2$) for 23 min. The motility of frozen-thawed spermatozoa was assessed by phase-contrast microscopy. To assess their viability and acrosome content, spermatozoa were stained with a vital stain and Fluorescence conjugated lectin Pisum Savitum Agglutinin (FITC/PAS), respectively. Pregnancy was confirmed by ultrasonograpy on day 25, 35 and 55 post insemination. The use of fresh semen, the pregnancy rates were observed 66.6, 66.6, 75.0 and 83.3% in natural estrus, clomifene induced, bromocriptine induced and a combination of GnRH and bromocriptine, respectively. The use of frozen-thawed semen, the pregnancy rates were observed 66.6, 33.3, 50.0 and 60.0% in natural estrus, clomifene induced, bromocriptine induced and a combination of GnRH and bromocriptine, respectively. No difference was observed in the number of offspring produced among natural estrus and treated groups inseminated with fresh or frozen-thawed semen. In conclusion, the pregnancy rate of dogs treated with a combination of GnRH and bromocriptine was more effective than use of clomifene or bromocriptine only. In addition, frozen-thawed semen can be used successfully far artificial insemination in dog.

• Journal of Veterinary Clinics
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• v.18 no.1
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• pp.44-47
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• 2001

Semen were collected from 9 male dogs and frozen by liquid nitrogengas. Frozen semen were thawed at 7$0^{\circ}C$ for 8 seconds. About $2{\timss}10^8$ sperm per insemination were inseminated to 10 bitches (3 Retrievers, 4 Chihuahuas, 1 Yorkshire Terriers, 1 Maltese, and 1 Poodle) at three and six days after the estimated peak of luteinizing hormone. For small breed dogs, uretero-renoscope (Kahl Storz, Germany, 12.5 Fr) was used for trans-cervical insemination, whereas cystoscope(Kahl Storz, Germany, 22Fr) was used for large breeds (Retrievers). Pregnancy was diagnosed by ultrasonography at 30 days after insemination. All of 3 Retrievers (100.0%) and 3 bitches of 7 small breed dogs (42.9%) were conceived (60.0% in total). This result indicated that trans-cervical insemination using endoscope is an effective method for AI with frozen semen not only for large breed dogs such as Retriever but also for small breeds.

• Asian-Australasian Journal of Animal Sciences
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• v.16 no.2
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• pp.180-184
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• 2003

The intrauterine inseminator (IUI) was developed to provide the method of depositing dog semen into the uterine body instead of the vagina. The IUI consists of a vaginal endoscope, a balloon sheath, and injection catheter. When the endoscope is inserted into the vagina and the balloon expanded with air, the cervical os becomes visible so a injection catheter can be inserted through the cervix for deposition of the frozen-thawed semen. The efficacy of the IUI device was compared to intra-vaginal artificial insemination using semen that had been collected and frozen from pooled sperm-rich fraction of ejaculates collected from two Jindo dog donors. Aliquots of semen were extended with a Tris-egg yolk diluent, centrifuged, the seminal plasma removed, the pellet resuspended with the same diluent, and cooled to $5^{\circ}C$ over a 2 h period. A Tris-egg yolk-glycerol extender was added at $5^{\circ}C$; after 1 h, semen was loaded into 0.5 ml straws, and straws were frozen in LN vapor for 5 min, and immersed in LN for storage. The final sperm concentration for freezing was approximately $100{\times}10^{6}cells/ml$. The straws were thawed at $70^{\circ}C$ for precisely 6 sec, 1.5 ml Tris-egg yolk buffer at $38^{\circ}C$ added, and the 2 ml of thawed semen was used for a single insemination using the IUI device. Each bitch was inseminated at optimal insemination point, which was estimated by vaginal epithelial cells staining and progesterone concentration analysis. Use of the IUI device resulted in 21 of 26 females giving birth to 89 pups ($4.2{\pm}1.6$ pups per litter), while intra-vaginal AI resulted in 6 of 15 females whelping a total of 17 pups ($2.8{\pm}1.2$ pups per litter). We believe the IUI device is easier to use than previously described devices used for intrauterine insemination. In our experience the expansion of the balloon has a calming effect on the bitch that aids the inseminator. These results indicate that the IUI device was able to provide high fertility with 50 million frozen sperm per insemination and two inseminations.

• Journal of Embryo Transfer
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• v.23 no.2
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• pp.115-118
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• 2008

Glycerol is the cryoprotectant most frequently used to freeze semen in several of species. The objective of the present study was to compare the effect of three different glycerol concentrations (4, 6 or 8%, v/v) on frozen-thawed dog sperm survival rate. Ejaculates from 9 dogs collected by digital manipulation were pooled and assessed by macroscopic and microscopic criteria. Semen was divided into 3 aliquots, which were centrifuged and the sperm pellets rediluted with first Tris-glucose-citric acid extender. After 1 h cooling at $4^{\circ}C$, second extender containing 4, 6 or 8% glycerol was added, respectively. The semen was loaded into 0.25 ml straws and frozen and stored in liquid nitrogen and thawed. Sperm vigor, live:dead spermatozoa ratio using HOS test, and sperm morphology using $Spermac^{(R)}$ stain were evaluated. After thawing, there were no significant differences among groups in vigor, viability and morphology. In conclusion, the three glycerol concentrations (4, 6 or 8%) can be used successfully in cryopreservation of canine semen. Therefore the use of 4% glycerol in the extender has less toxic effect and reduces of freezing injuries.

• Journal of Embryo Transfer
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• v.23 no.4
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• pp.239-243
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• 2008

This study was conducted to investigate the effects of type of the sugar supplemented to the extender on the vigor, viability and intact acrosomal rates of frozen-thawed dog spermatozoa. The ejaculated semen was diluted with TRIS-citric acid extender containing 200mM TRIS, 73mM citric acid, 6% (v/v) glycerol, 20% (v/v) egg yolk, 1% (v/v) antibiotics (streptomycin/penicillin), 44 mM sugar, which was either glucose, fructose or glucose-fructose combination, and distilled water to make the final volume of 100ml. Extended semen samples were cooled at $4^{\circ}C$ for an hour, packaged in 0.25ml straws, equilibrated for 10 minutes in liquid nitrogen vapor, and frozen in liquid nitrogen. Samples were thawed by placing straws into $37^{\circ}C$ water for 120 seconds. After thawing, vigor, viability and intact acrosomal rates of frozen-thawed semen were compared according to type of sugar. No significant differences were observed between glucose and fructose groups. In addition, combination of the 2 sugars also did not show any significant differences in the vigor, viability and intact acrosomal rates. In conclusion, glucose and fructose were equally efficient as sugar supplements for freezing extender.

• Proceedings of the Korean Society of Embryo Transfer Conference
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• 2001.05a
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• pp.30-30
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• 2001

• Proceedings of the Korean Society of Veterinary Clinics Conference
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• 2009.10a
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• pp.260-260
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• 2009

• Proceedings of the KSAR Conference
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• 2001.10a
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• pp.57-57
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• 2001

This study was carried out to investigate the general characteristics such as volume, sperm concentration, sperm motility, sperm abnormality on whole semen, RSP-S and RSP-T semen and fractional semen of small size dogs, and the effect of temperature and preservation time and cryoproservation on motility of whole and RSP-S and RSP- T semen. Multiple ejaculates were collected from small dogs by the digital manipulation of penis. 1. The volume per ejaculate semen, sperm of concentration and motility and abnormal sperm rate of 1st fractional semen were 0.65±0.09㎖, 4.52±0.35×10/sup 6/ cells/㎖, 15.64±3.85% and 5.50±0.62%. Also, 2nd fractional semen were 1.25±0.20㎖, 3.35±0.48×10/sup 6/cells/㎖, 96.25±4.65% and 4.24±0.46%. And 3rd fractional semen were 1.45±0.21㎖, 3.85±0.52×10/sup 6/cell/㎖, 92.82±4.24% and 4.66±0.58%, respectively. 2. The sperm of concentration and motility and abnormal sperm rates of whole, RSP-S and RSP-T semen were 5.45±0.82×10/sup 6/ cells/㎖, 95.55±4.65%, 4.58±0.45% and 4.82±0.36×10/sup 6/cells/㎖, 90.10±3.42%, 6.48±0.68% and 4.55±0.45× 10/sup 6/cells/㎖, 93.25±3.85%, 4.82±0.58%, respectively. 3. The motility of whole, RSP-S and RSP-T semen were higher at 4℃ than at 38℃. When preservation temperature was at 4℃, survival rates of RSP-S and RSP-T sperm were 97.54%-6.25% at 1-72 hrs, 97.40%-5.62% at 1-100 hrs, respectively. 4. The survival rates of slow and rapid frozen 2nd fraction, RSP-S and RSP-T semen were 67.3±4.45%, 88.8±4.46% and 46.4±3.84%, 74.4±4.20%, respectively. Survival rates was significantly higher in frozen RSP-S and RSP-T semen than that in control group(8.5±2.12%).