Animal disease models contribute to a better understanding of the pathogenic mechanisms of human and animal diseases and help develop treatments for them. Ligation of the rat iliac arteries is performed to reproduce erectile dysfunction and peripheral arterial disease. Although knowledge of the ramification of branches from the rat iliac artery is important to perform such surgery, descriptions in previous studies are insufficient. Therefore, 17 male and 18 female Wistar rats were observed to elucidate the detailed ramification patterns of branches from the iliac arteries with the latex injection method. The iliac arteries branched off the umbilical, cranial gluteal, lateral and medial circumflex femoral, external pudendal, and caudal epigastric arteries, and the common trunk of the caudal gluteal and internal pudendal arteries. The branching pattern of the umbilical, cranial and caudal gluteal, and internal pudendal arteries varied greatly and was categorized as Types 1 to 3 based on the number of branching levels along the proximodistal axis of the iliac arteries. Based on the same criteria, the ramification patterns of the lateral and medial circumflex femoral arteries were also divided into Groups 1 and 2. The external pudendal and caudal epigastric arteries originated from the external iliac artery mainly as a common trunk or less frequently as independent arteries in this order. The detailed branching patterns of the rat iliac arteries elucidated in the present study are beneficial for the refinement of surgical procedures.

The midcingulate cortex (MCC; area 24’) resides in the mid-rostrocaudal part of the cingulate gyrus, and it plays important roles in nociceptive, cognitive and skeletomotor functions. The MCC has recently been shown to consist of four cortical areas (areas a24a’, a24b’, p24a’ and p24b’) in the rabbit, based on immunohistochemistry. To further characterize the organization of these areas, here we immunohistochemically identified structures immunopositive (+) for calretinin (CR) as a marker of a subpopulation of inhibitory neurons. CR+ somata were identified as multipolar and bipolar neurons. The multipolar neurons were predominant throughout the MCC. CR+ somata were present mainly in layer (L) 2/3 and L6, and CR+ fibers occurred mainly in L1, L2/3 and L6. However, there were differences in the distribution of CR+ structures in each area. CR+ somata tended to be most densely distributed in area a24a’, followed by area p24a’, area a24b’ and area p24b’. CR+ fibers were most densely distributed in area p24a’, followed by area p24b’, area a24a’ and area a24b’. In addition, only areas p24a’ and p24b’ enclosed patchy CR+ fibers and terminals in deep L2/3. These results show the distinct distribution of CR+ structures in each area of the MCC in the rabbit, suggesting that CR+ neurons may contribute to information processing for cognitive functions in somewhat different manners in each area of the MCC.

In dogs, variation in the branching pattern of renal veins is rare with only few patterns reported. This report describes two unusual anomalies of the renal vein branching patterns in two dogs. In dog 1, a common renal trunk drained both kidneys, in a T-shape pattern, in the caudal vena cava after a long right perirenal course. In dog 2, a common venous trunk branched cranially from the pre-renal segment of an azygos-caudal vena cava venous trunk and divided into the renal veins in a Y-shape pattern. Proper knowledge of the possible anatomical variations in renal venous drainage may be helpful during imaging assessment and surgical planning of several canine diseases involving the abdominal vasculature and retroperitoneal space.

A reverse transcription-polymerase chain reaction (RT-PCR) method was developed for broadly detecting the avian encephalomyelitis virus (AEV). The new primers were based on conserved sequences of the 5’-untranslated region of AEV, because the virus was not detected using previous reported RT-PCR. By applying this method to the chicken samples with suspected AEV infection in Japan, we successfully obtained PCR products of the predicted size from all samples, and we confirmed the presence of AEV via sequence analysis.

We evaluated the relationship between the severity of coliform mastitis and bacterial load in 106 quarter milk samples. We found no significant relationship between somatic cell count and coliform bacterial load in milk in bovine clinical coliform mastitis. Results of the Cochran-Armitage test for trend in milk bacterial load proportions indicated a significant decreasing low group (P<0.001), increasing medium group (P<0.002) and increasing high group (P<0.02) with increasing clinical grade. The present study indicates that the coliform bacterial load in milk is significantly associated with clinical severity states in cases of bovine coliform mastitis, and can be a useful indicator for optimal management of this disease.

Bovine tuberculosis (bTB) control programs generally rely on intradermal tuberculin tests for the antemortem diagnosis of Mycobacterium bovis infection in cattle, but these tests detect only a portion of the infected animals. The aim of the present study was to evaluate the diagnostic coverage of a combination of the bTB antemortem techniques known as the comparative intradermal tuberculin test (CITT) and an ELISA based on a recombinant chimera of ESAT-6/MPB70/MPB83 as the antigen in cattle. The results were compared to postmortem findings based on M. bovis culturing and PCR. Paired comparisons of all data (n=92) demonstrated that ELISA and LST results compared to the culturing results did not present significant differences (P=0.27 on McNemar’s test and P=0.12 on Fisher’s exact test, respectively). Using culturing as the gold standard, the sensitivity and specificity of ELISA were 79.5% (95% CI: 64.5–89.2%) and 75.5% (95% CI: 62.4–85.1%), respectively, whereas LST demonstrated 100% sensitivity (95% CI: 91.03–100%) and 92.5% specificity (95% CI: 82.1–97.0%). The ELISA results did not reveal significant differences in relation to the LST results (P>0.99 on Fisher’s exact test). Using the latter as the gold standard, the sensitivity and specificity of ELISA were 79.1% (95% CI: 64.8–88.6%) and 79.6% (95% CI: 66.4–88.5%), respectively. The use of ELISA with the recombinant chimera of ESAT-6/MPB70/MPB83 as the antigen complements the diagnostic coverage provided by CITT and increases the removal of infected animals from herds.

The aim of this study was to elucidate the bone metabolic status after taking colostrum in newborn calves. Fourteen neonatal calves were randomly allocated to two groups fed either unheated or heated (60°C, 30 min) colostrum three times on the first day (2 l every 10 hr; 6 l in total). Heat treatment on colostrum was to reduce the bone metabolic markers assumed as heat-sensitive. The concentrations of four bone metabolic markers (the enzymes from bone cells or the bone collagen fragments) and a bone protective protein, osteoprotegerin (OPG), were measured in the blood of calves during a week after the birth and in the colostrum. The colostral concentrations of four bone metabolic markers were reduced by heating. Then those circulatory markers peaked after colostrum intake in the calves fed unheated colostrum; whereas those fed heated colostrum did not show such changes. However, the plasma tartrate resistant acid phosphatase 5b (TRAP5b) activity was transiently increased after taking colostrum in both groups. Meanwhile, heating did not decrease colostral OPG and there was no significant rise in the serum OPG concentrations after the first colostrum intake in both groups. The study revealed that the blood concentrations of studied bone metabolic markers depended on those colostral values except for TRAP5b. Based on the plasma TRAP5b changes, accelerated formation of premature osteoclast cells may be induced by colostrum intake. Meanwhile, colostral OPG absorption is less likely to impact on its circulating levels.

The purpose of this study was to use magnetic resonance imaging (MRI) to compare the volumes and three-dimensional configurations of the soft palate and nasopharynx in non-brachycephalic and brachycephalic dogs with different body weights, and infer which factors influence nasopharyngeal volume. This was a retrospective observational study. The brain MRI medical records of all dogs referred to the Veterinary Medical Center, Chungbuk National University, between 2013 and 2016, for evaluation of intracranial disease were reviewed. There was a significant difference in the two-dimensional parameters including soft palate length/skull length ratio (P<0.01) and maximum soft palate thickness (P<0.01), and three-dimensional parameters which included soft palate volume (P<0.01), nasopharyngeal volume (P<0.01), soft palate/total upper airway volume ratio (P<0.01), and nasopharyngeal volume/total upper airway volume ratio (P<0.01), between brachycephalic and non-brachycephalic dog breeds. Nasopharyngeal volume correlated positively with the maximum soft palate thickness and body weight in all breeds. The three-dimensional morphologic grades of soft palate were significantly different between the two groups. In brachycephalic breeds, Grade 3 was observed in 33% of cases but was absent in non-brachycephalic breeds, where Grade 1 was present in 85% of the cases. We can conclude that three-dimensional morphology and upper airway volume are significantly different between brachycephalic and non-brachycephalic breeds, and body weight and maximum soft palate thickness are the key factors associated with a decreased nasopharyngeal volume.

This study proposed a novel radiographic positioning in order to image the cranioventral lung region using a portable X-ray unit and digital radiography system. In the novel position, calves were restrained in a chute and a unilateral forelimb was pulled cranially with the contralateral forelimb tied to the chute; the forelimbs were then spread cranio-caudally as in a scissor position (Three-legged view: TL view). In a preliminary study, we applied the TL view for imaging of 14 clinically healthy calves. In a clinical study, accuracy in detecting cranioventral lung lesions was compared between the standard standing view and the TL view for 19 calves, which were culled from herd; the results of postmortem examination were used as gold standard. Seven evaluators independently interpreted the images. The median (range) number of trials and the time for obtaining optimal position were 2 (1–7) and 263 sec (105–488), respectively in 14 healthy calves. Calves thicker than approximately 40 cm were not considered candidates for TL view in this setting because of difficulty in restraint and the low output of the portable X-ray unit. The TL view improved the detection of consolidation in the cranioventral lung region, compared with the standard view. The TL view was considered an optional view when the cranioventral lung region was an area of interest, because this view was relatively easy to perform and required a small number of personnel, even for large calves.

We investigated the effects of an injectable pimobendan solution (0.15 mg/kg) on cardiac function in healthy dogs. Fifteen dogs were divided into placebo, intravenous pimobendan injection, and subcutaneous pimobendan injection groups. In the placebo, the heart rate, systolic and end-diastolic left ventricular pressure (LVPs and LVEDP), and peak positive (max dP/dt) and negative (min dP/dt) first derivatives of the left ventricular pressure did not change for 60 min. After the intravenous pimobendan injection, LVEDP decreased significantly within 5 min, while the max dP/dt increased, and the effects continued until 60 min. In comparison, there were no hemodynamic changes after the subcutaneous pimobendan injection. This study demonstrates that injectable pimobendan induced a rapid inotropic effect and decreased the LVEDP in dogs.

To identify reference values for the Schirmer tear test I (STT I) in Japanese black cattle, investigate the effects of variables (sex, age, time of day, and environment in the barn including environmental temperature, humidity, illuminance, and ammonia concentration) on tear production, and determine total tear protein concentration. One-hundred-and-thirty-seven Japanese black cattle (67 females and 70 males, age 3–90 days) were evaluated. The mean STT I values was 18.9 ± 2.9 mm/min (n=263) and significant effects were age and ammonia concentration in the barn. Mean total tear protein concentration was 1.18 ± 0.30 mg/ml in healthy cattle older than 15 days (n=38). It was suggested that age and ammonia concentration are related to fluctuation of tear volume.

A 12-year-old, castrated male cat with diabetes mellitus was diagnosed with acromegaly and examined with magnetic resonance imaging (enlarged pituitary gland, 8 mm); serum hormone concentrations were measured. After the first course of radiation therapy (4 Gy, 12 fractions), insulin administration was not required from day 420 after diagnosis. Enlarged pituitary tumor (8 mm) recurred, and insulin dosage amount of the cat was increased on day 1,065. The second course of radiation therapy (6 Gy, 4 fractions) was performed on day 1,201 and insulin administration was again discontinued. However, the cat died from lymphoma on day 1,397. Postmortem examination revealed pituitary adenoma. Most tumor cells were positive for chromogranin A, synaptophysin, and growth hormone immunohistochemistry. The pancreatic islet cells revealed diffuse hyperplasia. We achieved long-term successful management of an acromegalic cat with two courses of RT. However, a protocol for a second course of RT for feline recurrent pituitary tumor should be further discussed.

Fibrotic lesion is formed by myofibroblasts capable of producing collagens. The myofibroblasts are characterized by immunoexpressions of vimentin, desmin and α-smooth muscle actin (α-SMA) in varying degrees. The cellular characteristics remain investigated in myocardial fibrosis. We analyzed immunophenotypes of myofibroblasts appearing in isoproterenol-induced myocardial fibrosis in rats until 28 days after injection (10 mg/kg body weight); the lesions developed as interstitial edema and inflammatory cell reaction on 8 hr and days 1 and 3, and fibrosis occurred on days 1, 3, 7, 14, and 21 by gradual deposition of collagens, showing the greatest grade on day 14; the lesions gradually reduced with sporadic scar until day 28. Myofibroblasts expressing vimentin and α-SMA increased with a peak on day 3, and then, gradually decreased onwards. Interestingly, Thy-1 expressing cells appeared in the affected areas, apparently being corresponding to the grade similar to vimentin- and α-SMA-positive cells. Thy-1 is expressed in immature mesenchymal cells such as pericytes with pluripotent nature. The immunoreactivity for A3-antigen, a marker for immature mesenchymal cells, was seen in some surrounding cells. There were no cells reacting with antibodies to nestin or glial fibrillary acidic protein, although hepatic myofibroblats have been reported to react with these antibodies. Collectively, myofibroblasts appearing in rat myocardial fibrosis may have been derived from immature mesenchymal cells positive for Thy-1 or A3-antigen, with thereafter showing expressions of vimentin and α-SMA in differentiation.

An 8-year-old castrated Toy poodle presented with swelling and proptosis of the right third eyelid caused by an exophytic mass on the bulbar surface. Histologically, the mass was composed of stratified neoplastic basaloid cells, arranged in nests and interconnecting islands, which were mixed with tubular structures. Immunohistochemically, the basaloid cells were positive for p63 and cytokeratin (CK) 14, and the inner epithelial cells of the tubular structures were positive for CK7, CK8, and CK19. According to these findings, the mass was diagnosed as a basal cell adenocarcinoma. Although basal cell adenocarcinoma is rare in animals, it should be included in the list of differential diagnoses for superficial tumors of bulbar conjunctiva of third eyelid in dogs.

Erythroblastic sarcoma in a 10-year-old Japanese Black cow with anemia is described. Tumor masses or nodules were located mainly in the thoracic cavity, and some lymph nodes were slightly enlarged. Although neoplastic involvement of the bone marrow was detected, the cow was not leukemic. The diagnosis was made based on the localized distribution of neoplastic lesions, no increase of intravascular nucleated cell number, deeply eosinophilic cytoplasm in some tumor cells, and frequent immunoreactivity of the tumor cells for hemoglobin. The tumor cells were characterized by marked pleomorphism and atypia; such morphological deviation from their normal counterparts may be connected with functional deviation resulting in the sarcomatous growth of these erythroid cells.

It has been well known in humans that eosinophil infiltration into the site of inflammation and eosinophilia occur in mycobacterial infections. However, the role of eosinophils against the mycobacterium is unclear. We showed in previous study that in situ mouse eosinophils infiltrated into tissues produce α-defensin, an anti-bacterial peptide. We investigated in this study whether eosinophils reacting to mycobacteria produce α-defensin in mice and whether it can be used as a model. We showed that mycobacterial infection induced blood eosinophilia and infiltration of α-defensin producing eosinophils that to surround mycobacteria at the site of infection. These findings were usually seen during human mycobacterial infection. We established a good model to study host defense mechanism against mycobacteria through α-defensin via eosinophils.

A Japanese Black calf became dehydrated on the first day of life and died on the third day. Gross examination revealed a large amount of yellowish-brown serous fluid in the abdominal cavity and whitish-yellow fibrin in the serosa of the abdominal organs. Patchy red spots were observed throughout the peritoneum, and the outer membrane of the umbilical arteries was dark red. Bacteriologically, Mannheimia haemolytica serotype 2 was isolated from the umbilical arteries and vein, liver, and kidney. Histopathology revealed inflammation with M. haemolytica serotype 2 in the outer membrane of the umbilical arteries and in the serosa of the bladder and intestinal tract. This is the first case of bovine peritonitis with histopathologic and immunohistochemical identification of M. haemolytica.

Eukaryotic elongation factor 2 (eEF2) kinase (eEF2K) inhibits protein translation through the phosphorylation of its specific substrate, eEF2. We previously demonstrated that eEF2K expression increases in superior mesenteric artery from spontaneously hypertensive rats (SHR) and that eEF2K mediates development of hypertension in SHR. In addition, we recently revealed that A484954, a selective eEF2K inhibitor induced relaxation via opening smooth muscle inward rectifier K⁺ (K_ir) channel in rat isolated superior mesenteric artery. Here, we further examined the effects of A484954 on contractility and blood pressure (BP) in rats. Isometric contraction of rat isolated superior mesenteric artery was measured. BP was measured by a carotid cannulation method. A484954 (10 µM) inhibited noradrenaline (NA)-induced contraction in a biphasic manner (magnitude of inhibition higher at high dose NA). A484954 also inhibited an α₁-receptor agonist, phenylephrine-induced contraction, while it was not biphasic. Specifically, a β-receptor antagonist, propranolol (1 µM) prevented the A484954-mediated inhibition of NA (high-dose)-induced contraction. A484954 (10 µM) potentiated a β-receptor agonist, isoproterenol-induced relaxation, which was completely prevented by BaCl₂ (1 mM), a K_ir channel blocker. In vivo, A484954 (122 µg/kg) inhibited NA-induced increase of BP in rats. Another eEF2K inhibitor, NH125 (22 µg/kg) also inhibited the NA-induced BP increase in rats. In summary, it was concluded that A484954 lowers NA-induced BP rise perhaps through activation of β₂-receptor-K_ir channel and subsequent vasorelaxation via inhibiting eEF2K activity.

High temperature induces various metabolic disturbances in animals. However, no comprehensive information is currently available on the metabolic pathway affected by high environmental temperature. The present study examined metabolite content in the serum of heat challenged quails using metabolomic analysis. In the present study, female quails with normal laying rate at 20 weeks kept in standard condition (control group) or exposed to 34°C 4 hr per day (12:00 to 16:00 hr)(heat group) for 10 consecutive days. The metabolomic analysis identified 165 metabolites in the serum, and significant differences were observed in the serum for 7 metabolites between two groups. An analysis by MetaboAnalyst, a web-based metabolome data tool, indicate that high temperature affect ketone body metabolism, butyrate metabolism, arginine and proline metabolism. Furthermore, histological examination of liver indicates a heat challenge induced abnormal lipid metabolism. Triglyceride and cholesterol level in the liver increased, however cholesterol level decreased in the serum. Genes related to lipid metabolism significantly increased in the liver after heat challenge. The present study demonstrated that high temperature cause liver damage, thus lipid metabolic was affected. Protect liver under high temperature could be one solution for coping with high temperatures in summer.

Currently, metabolic syndrome is a worldwide concern. Thus, it is imperative to understand the mechanism of metabolic syndrome by establishing various metabolic syndrome models. In this study, we used Hatano high-avoidance animals (HAA) and low-avoidance animals (LAA), both derived from Sprague–Dawley rats by selective breeding to determine high- or low-avoidance rates in shuttle-box active avoidance tests. HAA and LAA rats have some strain differences related to eating and appetite. Therefore, we determined whether Hatano rats could be used as a metabolic syndrome model. We compared food intake, body weights, blood pressure levels, plasma component levels, and fat contents between HAA and LAA rats. The HAA rats showed more active eating, higher blood pressure, higher percentage fat, and higher triglyceride levels than the LAA rats—these features correspond to some of the risk factors associated with metabolic syndrome. Our study suggests that HAA rats can be considered as a metabolic syndrome model by focusing on their feeding behavior, blood pressure levels, and percent body fat.

A 6-day-old female Holstein displayed a dome-shaped skull and cardiac murmur on physical examination. Neurological abnormalities included progressive ataxia, decreased pupillary light reflex, and blindness soon after birth. On diagnostic imaging, CT identified expanded ventricles and thyroid hypoplasia on the left side. MRI detected expanded ventricles, especially in the rostral cerebrum at the mesencephalic aqueduct, compared with normal calves, so we suspected hydrocephalus causing stenosis of the mesencephalic aqueduct. Postmortem examination revealed a structure in the mesencephalic aqueduct resembling the “web” type of aqueductal stenosis described in humans. This case report indicates the utility of describing mesencephalic aqueductal stenosis by web and detection of other malformations on CT and MRI for antemortem diagnosis in calves.

Reduction in oocyte quality is a major factor responsible for declining fertility associated with maternal aging in cows. The objective of the present study was to determine whether intracytoplasmic sperm injection (ICSI) could increase the efficiency of embryo production in older cows. We used cows aged 30 to 50 months or >120 months, which were defined as young or aged, respectively. The distribution of cortical granules in oocytes was affected by age as older cows had lower proportion of oocytes with mature cytoplasm containing evenly dispersed cortical granules compared to young cows. Although fertilization rates did not differ significantly between the two groups after in vitro fertilization (IVF), the rate of abnormal fertilization was higher, and the numbers of total and diploid blastocysts were lower for aged cows compared to young cows. However, in the embryos produced by ICSI, there was no significant difference in these parameters between young and aged cows. Although ICSI did not improve the blastocyst development rate, ICSI increased the proportion of diploid blastocysts in aged cows compared to IVF. In conclusion, maternal aging may negatively affect cytoplasmic maturation of bovine oocytes, which could be associated with abnormal fertilization or low developmental competence of oocytes. Our data also suggests beneficial effects of ICSI on the production of chromosomally normal embryos in aged cows.

A routine parentage test as part of a conservation program for Kiso horses identified a possible sex chromosome anomaly in a 7 months-old filly because of an aberrant result using LEX3, an X-linked marker. We then analyzed X-linked markers (LEX26, TKY38, and TKY270), Y-linked markers (Eca.YH12, Eca.YM2, Eca.YA16, and the sex-determining region Y gene), and an X/Y marker (Amelogenin gene). This analysis demonstrated that the filly had not inherited an X chromosome from her sire. A karyotyping analysis confirmed that the filly was 63,XO. As it was suspected that the horse would be sterile, we avoided using the horse as a broodmare; the information should also serve to prevent unnecessary conflict between owners transferring and receiving the horse.

The present study aimed to determine possible risk factors affecting reproductive performance, especially time taken to establish pregnancy in dairy cows, using Cox’s proportional hazard model. The data were collected from 154 Holstein Friesian cows (199 lactations). Cows diagnosed with a vaginal discharge score (VDS) of one or calving abnormality showed significantly delayed pregnancy: hazard ratio (HR)=0.654 (95% confidence interval [CI]: 0.436–0.983; P=0.041) and HR=0.457 (95% CI: 0.270–0.774; P=0.004), respectively. Our study suggested that the occurrence of VDS of one or calving abnormality might be possible risk factors increasing the number of days open and affecting reproductive performance in dairy cows.

Wild boars are a reservoir for many zoonotic pathogens and a good sentinel for surveillance of zoonotic viral infections, but collection of serum samples from wild boars in the field is sometimes difficult and requires special equipment and techniques. In this study, ELISA using meat juices extracted from the heart and diaphragm of wild boars, instead of serum samples, was performed to detect antibodies against zoonotic pathogens, Japanese encephalitis virus and hepatitis E virus. The results of ELISA using meat juice samples were significantly correlated with those using serum samples and meat juice contained one-fifth the antibodies of serum samples. As meat juice is easily collected from wild animals in the field without special equipment and techniques, ELISA using meat juice is a simple and superior method for serological survey of zoonosis among wild animals.

Forty rescued common palm civets were anesthetized. Twenty animals received intramuscular injections of alfaxalone 5 mg/kg and medetomidine 0.05 mg/kg (A-M group), whereas twenty animals received 5 mg/kg of tiletamine and zolazepam (T-Z group). The A-M group was reversed with atipamazole 0.25 mg/kg. There were no significant differences in the time from anesthetic injection to induction and intubation between the A-M and T-Z groups. The time from the injection of reversal in the A-M group and the time from cessation of isoflurane in the T-Z group to extubation, first response to recovery and ambulation were longer (P<0.05) in the T-Z group. The T-Z group recorded lower (P<0.05) rectal temperatures compared to the A-M group. This study showed that both drug combinations can be used effectively for the immobilization of civets. The A-M combination provided better anesthetic depth, but with higher incidence of bradycardia and hypoxemia. The recovery time was reduced significantly as atipamezole was used as a reversal agent in the A-M combination.

A 3-year-old male hamadryas baboon (Papio hamadryas) at the Seoul Zoo, Korea, died without any previous symptoms. Necropsy revealed severe whipworm infection in the large intestine. The animal weighed 2.6 kg and had a blood clot at the anus. Numerous whipworms were found attached to the intestinal wall, with their anterior ends embedded in the mucosa. Fecal microscopy revealed typical barrel-shaped, brown eggs of Trichuris spp., with hyaline polar plugs at each end. Histopathological examination revealed the thin anterior part of Trichuris spp. embedded in the mucosal layer and the thick posterior part at the mucosal surface or hanging freely in the intestinal lumen. This case emphasizes the importance of parasitic infection management in zoo animals.

Kiso horse is a breed of Japanese native horses. In this study, to clarify coat color gene variation in Kiso horses, we used SNaPshot™ genotyping to evaluate variation in MC1R, ASIP, and MATP genes at the Extension (E), Agouti (A), and Cream dilution (C) loci. The coat color of 149 horses was documented. The coat color of 140, 3, and 6 horses was bay, chestnut, and buckskin, respectively. Furthermore, the frequency of alleles E, e, A, a, C, and Cr was 0.80, 0.20, 0.86, 0.14, 0.98, and 0.02, respectively. Current status of coat color genes in Kiso horses was clarified, and this information will help plan further conservation of the horses.