吉村 文

Several lines of evidence suggest that stress induces the neurovascular dysfunction associated with increased blood-brain barrier (BBB) permeability, which could be an important pathology linking stress and psychiatric disorders, including major depressive disorder (MDD). However, the detailed mechanism resulting in BBB dysfunction associated in the pathophysiology of MDD still remains unclear. Herein, we demonstrate the role of vascular endothelial growth factor (VEGF), a key mediator of vascular angiogenesis and BBB permeability, in stress-induced BBB dysfunction and depressive-like behavior development. We implemented an animal model of depression, chronic restraint stress (RS) in BALB/c mice, and found that the BBB permeability was significantly increased in chronically stressed mice. Immunohistochemical and electron microscopic observations revealed that increased BBB permeability was associated with both paracellular and transcellular barrier alterations in the brain endothelial cells. Pharmacological inhibition of VEGF receptor 2 (VEGFR2) using a specific monoclonal antibody (DC101) prevented chronic RS-induced BBB permeability and anhedonic behavior. Considered together, these results indicate that VEGF/VEGFR2 plays a crucial role in the pathogenesis of depression by increasing the BBB permeability, and suggest that VEGFR2 inhibition could be a potential therapeutic strategy for the MDD subtype associated with BBB dysfunction.

<p>Objectives: Proximal stoma creation in neonates results in growth failure and distal intestinal atrophy. "Recycling stool" consists of stool injection from the proximal limb to the distal limb of a stoma. Because this method may prevent distal bowel atrophy and increase body weight, we investigated the effects of recycling stool upon distal intestinal mucosa by generating an ileostomy model in rats.</p><p>Methods: An ileostomy was created 5 cm proximal to the cecum in male Wistar/ST rats. Discharged stool or saline was injected into the distal limb, twice per day for 7 days. The intestinal adaptation was assessed by measuring the villus height and counting goblet cell number. Proliferation and apoptosis were analyzed by Ki67 and TUNEL immunostaining.</p><p>Results: The ratios of the height of the distal villi (D) to the that of proximal villi (P) were 0.97 (median [range] of D and P length: 421 [240–729] μm and 436 [294–638] μm, P<0.05) in the stool-injected group and 0.81 in the saline-injected group (442 [315–641] μm and 548 [236–776] μm, P<0.05). Compared with the saline-injected group, the stool-injected group showed elevated numbers of goblet cells (3.6 [2.0–7.6] vs. 4.9 [2.4–7.5] cells/100-μm villus length) and Ki67-positive cells (26.8% [13.8%–35.4%] vs. 40.1% [31.2%–45.7%]), along with a reduced number of apoptotic cells (5.0 [2.0–14.0] vs. 4.0 [1.0–9.0] cells/100-μm villus length).</p><p>Conclusions: Recycling stool prevented distal intestinal atrophy; this experimental design may facilitate further studies concerning alternative methods to prevent intestinal atrophy and growth failure.</p>

Daily fat and sugar intake has increased in Japan, while total energy intake has decreased. However, the number of type 2 diabetes mellitus patients has increased, and this often causes renal injury characterized by autophagic vacuoles. Although many studies with comparisons of high fat or sugar versus a normal macronutrient balanced diet have been reported, there are few studies that equalized calorie intake and body weights. In the current study, AIN93M diets (CONT group) with matching energy content with lard derived high saturated fat (LARD group), soybean oil derived unsaturated fat (SOY OIL group) and sucrose (SUCROSE group) were provided to compare their effects on renal morphology in streptozotocin-injected CD-1 mice without causing obesity. The number of renal tubular vacuoles was higher in SUCROSE and slightly higher in LARD compared with CONT mice, and was higher in LARD and SUCROSE compared with SOY OIL mice. Most of those vacuoles were LAMP1-positive, a marker of lysosomal autophagy. These results suggest that despite identical energy contents, diets with high sucrose or saturated fat compared to unsaturated fat may aggravate lysosomal renal injury in a non-obese, streptozotocin-induced model of diabetes mellitus.

© 2020 Elsevier B.V. The increasing Yezo sika deer (Cervus nippon yesoensis) population is creating a large problem. Yezo sika deer are an important blood meal source, and these deer contribute to the maintenance of tick populations. Theileria spp. infections in Yezo sika deer and T. orientalis infections in cows occur at high frequencies, and the same tick species infests both deer and cows. Therefore, a specific detection method to identify deer Theileria spp. is important. In this study, we establish a novel molecular detection method for identifying Theileria spp. from deer and tick samples using loop-mediated isothermal amplification (LAMP). This method targets a metalloprotease/cell division cycle protein gene homologue. Our LAMP protocol was able to detect deer Theileria and did not show cross reactivity with other closely related protozoan parasites, including T. orientalis. The LAMP method showed sensitivity and specificity equivalent to those of nested PCR performed on the same field samples from deer and ticks. These results demonstrate the applicability of LAMP to field surveys in which the detection of deer Theileria spp. is required. In conclusion, due to its simplicity, specificity, and reliability, we suggest our LAMP protocol as an appropriate method for routine surveys to detect Yezo sika deer and ticks infected with deer Theileria spp. parasites. Additionally, this LAMP method offers great promise as a useful tool to distinguish Yezo sika deer Theileria from related Theileria parasites present in livestock.

Rodent models of chronic restraint stress (CRS) are often used as simple models of depressive disorder. However, these models of stress have been mainly developed in rats, and the behavioral phenotypes of CRS models are still controversial. In this study, we compared the physiological and behavioral responses of C57BL/6J (B6) and BALB/c mice, which are commonly used in genetic and behavioral studies, to CRS. In addition to measuring physiological parameters and the levels of corticosterone (a stress hormone) in response to stress, we also examined changes in the levels of testosterone (an anti-stress hormone), which have rarely been studied in stressed mice. The mice were exposed to CRS for 6 h a day for 21 days. In both B6 and BALB/c mice, CRS elicited several physiological stress responses, including decreased body weight gain and changes in the tissue weights of stress-related organs. Accumulated corticosterone in the hair was measured, and BALB/c mice had significantly greater levels than control mice and B6 mice after CRS. On the other hand, in the case of accumulated testosterone in the hair, both B6 mice and BALB/c mice showed significantly higher concentrations than control mice, but the degree of change was not different between the two strains. In the sucrose preference test, BALB/c mice, but not B6 mice, showed anhedonia-like behavior after CRS. However, neither strain showed depressive-like behavior in the forced swim or tail suspension test. Our results show that the physiological and behavioral stress responses of BALB/c mice are greater than those of B6 mice, although anti-stress responses to CRS are similar in both strains. This suggests that BALB/c mice are likely to be advantageous for use as a CRS-induced depression model.

Prolonged and intense stress chronically increases blood concentration of glucocorticoids, which in turn causes downregulation of glucocorticoid receptor (GR) in the central nervous system (CNS). This process has been suggested to be involved in the pathogenesis of major depressive disorder (MDD). Here, we found that basic fibroblast growth factor (bFGF) increased the expression of GR in the rat cerebral cortex and cultured cortical neurons and restored the reduced GR expression caused by glucocorticoid exposure. Among intracellular signaling pathways stimulated by bFGF, extracellular signal-regulated kinase/mitogen-activated protein kinase (ERK/MAPK) pathway was responsible for the upregulation of GR. The bFGF-induced GR was functional as a transcription factor to enhance transcription of a target gene. Because high stress augments bFGF levels in the brain, it is likely that bFGF plays a compensating role for reduced GR expression after stress and thus should be studied as a therapeutic target for the treatment of MDD.

Brain-derived neurotrophic factor (BDNF) critically controls the fate and function of the neuronal network and has received much attention as a target of many brain diseases. Dopaminergic system dysfunction has also been implicated in a variety of neuropsychiatric diseases. Rotigotine, a non-ergot dopamine receptor agonist, is used in the treatment of Parkinson's disease and restless legs syndrome. To investigate the effects of rotigotine on neuronal functions both in vivo and in vitro, rats and primary cortical neurons were administered rotigotine, and the mRNA and protein expression levels of BDNF, its receptor TrkB and downstream signaling molecules, and synaptic proteins were determined. We found that BDNF protein was increased in the cortex and hippocampus of rats after 7 days of rotigotine treatment. In contrast, BDNF mRNAs were reduced 6 h after rotigotine treatment in cultured neurons presumably through the transient suppression of neuronal activity. We identified differential expression of D1, D2, and D3 receptors in the rat brain and cultured neurons. The observed increase in the expression of BDNF protein in the cortex and hippocampus after subchronic administration of rotigotine suggests that it may exert its medical effect in part through improving BDNF function in the brain. In addition, our results highlight the complex relationships between rotigotine and BDNF expression, which depend on the brain region, time course, and dose of the drug.

Extracellular vesicles (EVs) can modulate microenvironments by transferring biomolecules, including RNAs and proteins derived from releasing cells, to target cells. To understand the molecular mechanisms maintaining the neural stem cell (NSC) niche through EVs, a new transgenic (Tg) rat strain that can release human CD63-GFP-expressing EVs from the NSCs was established. Human CD63-GFP expression was controlled under the rat Sox2 promoter (Sox2/ human CD63-GFP), and it was expressed in undifferentiated fetal brains. GFP signals were specifically observed in in vitro cultured NSCs obtained from embryonic brains of the Tg rats. We also demonstrated that embryonic NSC (eNSC)-derived EVs were labelled by human CD63-GFP. Furthermore, when we examined the transfer of EVs, eNSC-derived EVs were found to be incorporated into astrocytes and eNSCs, thus implying an EV-mediated communication between different cell types around NSCs. This new Sox2/human CD63-GFP Tg rat strain should provide resources to analyse the cell-to-cell communication via EVs in NSC microenvironments.

Growing evidence suggests that excess glucocorticoids (GCs) exposure during the pregnancy results in behavioral abnormality in offspring. Although research using animal models has demonstrated that systemic GCs treatment impairs development of fetal brain, direct impact of GCs on the phenotype of embryonic neural stem/progenitor cells (eNSPCs) and its mechanism has not been fully understood. Here, we investigated the effect of chronic GCs exposure on cell proliferation, differentiation, and survival of eNSPCs in vitro. Corticosterone (CORT, a murine GC) treatment did not affect the proliferation of eNSPCs. On the other hand, decreased expression of neuronal, synaptic, and astroglial marker proteins were observed when the differentiation of eNSPCs was induced in the presence of CORT. CORT also reduced the survival rate of eNSPCs after the differentiation. Moreover, CORT inhibited extracellular signal-regulated kinase (ERK) and phosphatidylinosito13-kinase/Akt (PI3K/Akt) signaling pathways, which were activated during cell differentiation of eNSPCs. Inhibiting these signaling pathways reduced neural differentiation and eNSPCs viability, indicating their essential roles in the eNSPCs differentiation. Furthermore, IGF-I, a potent PI3K/Akt and ERK signaling stimulator, partially restored the adverse effect of CORT on eNSPCs, suggesting a possible involvement of the repression of these intracellular signaling in the GCs-caused eNSPCs impairment. (C) 2016 Elsevier Ireland Ltd and Japan Neuroscience Society. All rights reserved.

Extracellular vesicles (EVs) play an important role in the transfer of biomolecules between cells. To elucidate the intercellular transfer fate of EVs in vivo, we generated a new transgenic (Tg) rat model using green fluorescent protein (GFP)-tagged human CD63. CD63 protein is highly enriched on EV membranes via trafficking into late endosomes and is often used as an EV marker. The new Tg rat line in which human CD63-GFP is under control of the CAG promoter exhibited high expression of GFP in various body tissues. Exogenous human CD63-GFP was detected on EVs isolated from three body fluids of the Tg rats: blood serum, breast milk and amniotic fluid. In vitro culture allowed transfer of serum-derived CD63-GFP EVs into recipient rat embryonic fibroblasts, where the EVs localized in endocytic organelles. These results suggested that this Tg rat model should provide significant information for understanding the intercellular transfer and/or mother-child transfer of EVs in vivo.

MicroRNAs (miRs) play important roles in neuronal differentiation, maturation, and synaptic function in the central nervous system. They have also been suggested to be implicated in the pathogenesis of neurodegenerative and psychiatric diseases. Although miR-132 is one of the well-studied brain-specific miRs, which regulates synaptic structure and function in the postnatal brain, its function, in the prenatal brain is still unclear. Here, we investigated miR-132 function during differentiation of rat embryonic neural stem cells (eNSCs). We found that miR-132 expression significantly increased during the fetal rat brain development and neural differentiation of eNSCs in vitro. Furthermore, miR-132 expression was increased during differentiation through MAPK/ERK1/2 pathway. Inhibition of ERK1/2 activation resulted in increased levels of synaptic proteins including PSD-95, GluR1 and synapsin I. Silencing of miR-132 also increased PSD-95 and GluR1. Considering that miR-132 increases synaptic proteins in differentiated cortical neurons, our result shows a novel function of miR-132 as a negative regulator for synaptic maturation in the neuronal differentiation of eNSCs. (C) 2016 Elsevier Ltd. All rights reserved.

Neuronal cell survival and synaptic plasticity are controlled through expression of various neurotrophic factors including brain-derived neurotrophic factor (BDNF). In the present study, we examined the mechanism behind BDNF-induced Bdnf mRNA production and the physiological role of its amplification system using cortical neurons. Exogenous BDNF was applied to the cultured cortical neurons at days in vitro (DIV) 3 and DIV 7 with or without inhibitors for intracellular signaling. Expression levels of total Bdnf and Bdnf variants (exon I, exon IV, and exon VI) were biphasically increased after the BDNF application in different developing stage of neurons. Inhibitor for extracellular signal-regulated kinase, calmodulin dependent protein kinase II, or protein kinase A repressed the BDNF-induced Bdnf mRNA expression. Furthermore, we found that application of TrkB-Fc, which scavenges produced endogenous BDNF, resulted in weakened BDNF/TrkB signaling and decreased expression of postsynaptic proteins, suggesting that newly synthesized BDNF induced by the self-amplification system contributes to the synaptic maturation and function. (C) 2015 Elsevier Ltd. All rights reserved.

The prevalence of Campylobacter jejuni in wild birds is a potential hazard for human and animal health. The aim of this study was to establish the prevalence of C. jejuni in wild birds in Tokachi area, Hokkaido, Japan and investigate their virulence in vitro. In total, 173 cloacal swabs from individual wild birds were collected for the detection of Campylobacter spp. Thirty four samples (19.7%) were positive for Campylobacter of which 94.1% (32/34 samples) were C. jejuni. Additionally, one C. coli and one C. fetus were isolated. Seven C. jejuni isolates (one from crows and the other from pigeons) had important virulence genes including all three CDT genes (cdtA, cdtB and cdtC) and fiaA,flaB, ciaB and cadF, and the other isolates were lacking cdtA gene. Further studies on in vitro virulence-associated phenotypes, such as motility assay on soft agar and invasion assay in Caco-2 cells, were performed. The wild bird C. jejuni isolates adhered and invaded human cells. Although the numbers of viable intracellular bacteria of wild bird isolates were lower than a type strain NCTC11168, they persisted at 48-hr and underwent replication in host cells.

We analyzed blood samples of resident and migratory Japanese birds to evaluate the prevalence and genetic background of avian blood parasites in northern Japan. We used PCR targeting the mitochondrial cytochrome b gene to examine infections of Leucocytozoon, Haemoproteus, and Plasmodium parasites in blood samples from 243 birds of 14 species in three orders (Passeriformes, Columbiformes, and Anseriformes). Sequences were subjected to phylogenetic analysis. The infection rate was 21% in pigeons (Columbiformes) and 17% in Anseriformes. A high infection rate of 93.8% was found in crow species (Passeriformes). Haemoproteus and Plasmodium parasites were detected in only two species. Infected blood samples obtained from seven bird species involved two major clades of Leucocytozoon, which were divided between resident and migratory birds. The parasites, which are genetically distinct from parasites in Japanese resident birds, may have been introduced to Japan by migratory bird species.

Regulated degradation of cellular components by lysosomes is essential to maintain biological homeostasis. In mammals, three forms of autophagy, macroautophagy, microautophagy and chaperone-mediated autophagy (CMA), have been identified. Here, we showed a novel type of autophagy, in which RNA is taken up directly into lysosomes for degradation. This pathway, which we term 'RNautophagy,' is ATP-dependent, and unlike CMA, is independent of HSPA8/ Hsc70. LAMP2C, a lysosomal membrane protein, serves as a receptor for this pathway. The cytosolic tail of LAMP2C specifically binds to almost all total RNA derived from mouse brain. The cytosolic sequence of LAMP2C and its affinity for RNA are evolutionarily conserved from nematodes to humans. Our findings shed light on the mechanisms underlying RNA homeostasis in higher eukaryotes. © 2013 Landes Bioscience.

Background: The proboscis is an essential head appendage in insects that processes gustatory code during food intake, particularly useful considering that blood-sucking arthropods routinely reach vessels under the host skin using this proboscis as a probe. Results: Here, using an automated device able to quantify CO2-activated thermo (35 degrees C)-sensing behavior of the malaria vector Anopheles stephensi, we uncovered that the protruding proboscis of mosquitoes contributes unexpectedly to host identification from a distance. Ablation experiments indicated that not only antennae and maxillary palps, but also proboscis were required for the identification of pseudo-thermo targets. Furthermore, the function of the proboscis during this behavior can be segregated from CO2 detection required to evoke mosquito activation, suggesting that the proboscis of mosquitoes divide the proboscis into a "thermo-antenna" in addition to a "thermo-probe". Conclusions: Our findings support an emerging view with a possible role of proboscis as important equipment during host-seeking, and give us an insight into how these appendages likely evolved from a common origin in order to function as antenna organs.

Vector-borne diseases, such as malaria and lymphatic filariasis, are co-endemic in large parts of the world. To develop a multiplex amplification method for the simultaneous detection of multiple insect-borne infectious diseases, we used LAMP with fluorescently labeled primers to identify the SPECT2 gene of Plasmodium berghei and the cytochrome oxidase subunit I gene of Dirofilaria immitis in mosquitoes. This technique could detect as few as 100 P. berghei-infected red blood cell-equivalents or one D. immitis microfilaria. Moreover, individual species of parasites in mosquitoes could be identified when a mixture of fluorescently labeled primer sets was used. These findings suggest that the multiplex LAMP assay is sensitive and specific enough to identify parasite-bearing mosquitoes in areas where several diseases occur simultaneously. This procedure could increase the efficiency and effectiveness of arthropod-borne disease elimination programs. (C) 2010 Elsevier Inc. All rights reserved.

Mosquitoes are critical vectors in many arboviral transmission cycles. Considering the increasing incidence of arboviral infections throughout the world, monitoring of vector populations for the presence of an arbovirus could be considered an important initial step of risk assessment to humans and animals. In response to this need, increased efforts to develop rapid and reliable diagnostic techniques have been undertaken; a single-step reverse transcription-loop-mediated isothermal amplification (RT-LAMP) assay was developed to detect virus in vector mosquitoes (Aedes aegypti) using the Flock House Virus (FHV) as a model. The robustness of the RT-LAMP reaction was revealed by its ability to detect FHV from an "all-in-one" template using whole mosquito bodies within 30 min. Furthermore, RT-LAMP identified successfully a mosquito carrying just a single FHV particle, a level easily overlooked in conventional analysis such as plaque forming assays. These observations suggest that RT-LAMP is more reliable and useful for routine diagnosis of vector mosquitoes in regions where the prevalence of vector-borne diseases such as West Nile fever or dengue fever are common. (c) 2008 Elsevier B.V. All rights reserved.

Background: Despite recent advances in our understanding of the basic biology behind transmission of zoonotic infectious diseases harbored by arthropod vectors these diseases remain threatening public health concerns. For effective control of vector and treatment, precise sampling indicating the prevalence of such diseases is essential. With an aim to develop a quick and simple method to survey zoonotic pathogen-transmitting vectors, LAMP (loop-mediated isothermal amplification) was applied to the detection of filarial parasites using a filarial parasite-transmitting experimental model that included one of the mosquito vectors, Aedes aegypti, and the canine heartworm, Dirofilaria immitis. Results: LAMP reactions amplifying the cytochrome oxidase subunit I gene demonstrated high sensitivity when a single purified D. immitis microfilaria was detected. Importantly, the robustness of the LAMP reaction was revealed upon identification of an infected mosquito carrying just a single parasite, a level easily overlooked using conventional microscopic analysis. Furthermore, successful detection of D. immitis in wild-caught mosquitoes demonstrated its applicability to field surveys. Conclusion: Due to its simplicity, sensitivity, and reliability, LAMP is suggested as an appropriate diagnostic method for routine diagnosis of mosquito vectors carrying filarial parasites. This method can be applied to the survey of not only canine filariasis but also lymphatic filariasis, another major public health problem. Therefore, this method offers great promise as a useful diagnostic method for filarial parasite detection in endemic filariasis regions.

The genomic DNA of the grasshopper (Oxya hyla intricata) was subjected to electrophoresis after digestion with HaeIII, and the result showed two bands of highly repetitive DNA, approximately 200 and 400 bp in length. The 200-bp HaeIII-digested fragment was cloned and characterized by sequencing and fluorescence in situ hybridization (FISH). The results showed the presence of two distinct satellite DNA (stDNA) families: one consisting of a 169-bp repeated element having an A+T content of 60.9% and the other consisting of a 204-bp repeated element having an A+T content of 53.9%. No significant homology between the two stDNA families was observed. FISH showed that the chromosomal locations of these families are different from each other. The 169-bp element was located in the C-band-positive regions of the short arms of most of the chromosomes, whereas the 204-bp element was located in the centromeric regions of three chromosome pairs. These results imply that the origins of these two DNA families are different. The results of zoo-blot hybridization to the genomic DNA from four Oxya species, O. hyla intricata, O. japonica japonica, O. chinensis formosana, and O. yezoensis, suggest that the two stDNA families found in the present study are species-specific for O. hyla intricata.

The chromosomes derived from the Japanese population of Gryllus bimaculatus were characterized by C-banding and Ag-NOR staining. The chromosome number, 2n = 28 + XX ( female)/XO ( male), corresponded with that of other populations of G. bimaculatus, but the chromosome configuration in idiograms varied between the populations. NORs were carried on one pair of autosomes and appeared polymorphous. The positive C-bands located at the centromere of all chromosomes and the distal regions of many chromosome pairs, and the size and the distribution pattern of the distal C-heterochromatin showed differences among the chromosomes. In addition, this paper reports on the characteristics of HindIII satellite DNA isolated from the genome of G. bimaculatus. The HindIII repetitive fragments were about 0.54 kb long, and localized at the distal C-bands of the autosomes and the interstitial C-bands of the X chromosome. Molecular analysis showed two distinct satellite DNA sequences, named the GBH535 and GBH542 families, with high AT contents of about 67 and 66%, respectively. The two repetitive families seem to be derived from a common ancestral sequence, and both families possessed the same 13-bp palindrome sequence. The results of Southern blot hybridization suggest that the sequence of the GBH535 family is conserved in the genomic DNAs of Gryllus species, whereas the GBH542 family is a species-specific sequence.

The karyotypes of Gryllus rubens (Scudder) and Gryllus sp. collected in the US were studied using conventional Giemsa staining and two differential staining methods. Both species had a chromosome complement of 2n = 28 + XX/XO, and the X chromosome was large and metacentric. In addition, nucleolus organizer regions (NOR) were detected in the short arm of one pair in the two species, and the NOR showed variation in size. The two species had species-specific chromosome configuration and C-banding patterns. In Gryllus sp., the chromosome configuration showed polymorphism in size and type among individuals, and the distal C-bands of Gryllus sp. were larger than those of G. rubens.

The chromosomes of 4 species of grasshopper in the genus Oxya were analyzed by conventional and five differential-staining methods. All 4 species had a chromosome complement of 2n=22+XX(female)/XO(male), with very similar karyotypes. Both O. hyla intricata and O. japonica japonica had acrocentric chromosomes with minute short arms, while the chromosomes of O. chinensis formosana and O. yezoensis were almost all telocentric. In all 4 species, a secondary constriction (SC) was detected at a proximal site in chromosome 8, which had the same staining properties (positive for C- and CMA-banding negative for G- and QM-banding) in all 4 species. Our C-banding analysis revealed that the short arms of O. hyla intricata and O. japonica japonica consisted of C-heterochromatin. The 4 species yielded species-specific patterns of C-bands, and there appeared to be a close relationship between O. chinensis formosana and O. yezoensis, as suggested by the polymorphism of the distal C-bands of chromosome 2. In all 4 species of Oxya, Ag-NORs were detected on the centromeric regions of all the chromosomes, but not detected on the SC of chromosome 8. The karyosystematic relationships among the 4 species are discussed on the basis of the results of differential staining. © 2005 The Japan Mendel Society.

The karyological relationship and organization of highly repetitive DNA sequences in Japanese shrew-moles were studied by zoo-blot hybridization and fluorescence in situ hybridization (FISH). When the genomic DNA of the eastern race of Urotrichus talpoides was digested with PstI, three fragments of highly repetitive DNA sequences, approximately 0.7, 0.9, and 1.4 kb in length, were observed as distinct bands. The results of FISH in the eastern race of U. talpoides using these three fragments separately as probes showed that the 0.7-kb PstI fragment was distributed in the centromeric regions of most chromosomes, and that the 0.9- and 1.4-kb fragments were predominantly located in the C-heterochromatin region of chromosome 13p. Although the western race of U. talpoides also had three PstI fragments, 0.9- and 1.4-kb PstI fragments were more ambiguous than those of the eastern race. The PstI- digested genomic DNA in Dymecodonpilirostris produced only a faint 0.9-kb band, and its signal patterns obtained by zoo-blot hybridization were clearly different from those of U. talpoides. The 0.7-kb fragment of U. talpoides hybridized strongly with the 0.9-kb fragment of D. pilirostris. In a FISH analysis, the 0.9-kb fragment of D. pilirostris hybridized with highly repetitive DNA in the centromeric regions of most chromosomes from both D. pilirostris and U. talpoides. Zoo-blot hybridization and FISH analyses suggest that the 0.9- and 1.4-kb PstI fragments were generated specifically in the genome of U. talpoides after the common ancestor differentiated into two extant shrew-mole species. A difference in the length of the centromeric elements between U. talpoides and D. pilirostris might be observed due to certain modifications of the repeating unit.