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Alzheimer's disease (AD) is the most common chronic progressive neurodegenerative disease in the elderly. It has an increasing prevalence and a growing health burden. One of the limitations in studying AD is the lack of animal models that show features of Alzheimer's pathogenesis. The tree shrew has a much closer genetic affinity to primates than to rodents and has great potential to be used for research into aging and AD. In this study, we aimed to investigate whether tree shrews naturally develop cognitive impairment and major AD-like pathologies with increasing age. Pole-board and novel object recognition tests were used to assess the cognitive performance of adult (about 1 year old) and aged (6 years old or older) tree shrews. The main AD-like pathologies were assessed by Western blotting, immunohistochemical staining, immunofluorescence staining, and Nissl staining. Our results showed that the aged tree shrews developed an impaired cognitive performance compared to the adult tree shrews. Moreover, the aged tree shrews exhibited several age-related phenotypes that are associated with AD, including increased levels of amyloid-β (Aβ) accumulation and phosphorylated tau protein, synaptic and neuronal loss, and reactive gliosis in the cortex and the hippocampal tissues. Our study provides further evidence that the tree shrew is a promising model for the study of aging and AD., (© 2023. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

Tree shrews are a nonprimate species used in a range of biomedical studies. Recent genome analysis of tree shrews found that the sequence identities and the numbers of genes of cytochrome P450 (CYP or P450), an important family of drug-metabolizing enzymes, are similar to those of humans. However, tree shrew P450s have not yet been sufficiently identified and analyzed. In this study, novel CYP2D8a and CYP2D8b cDNAs were isolated from tree shrew liver and were characterized, along with human CYP2D6, dog CYP2D15, and pig CYP2D25. The amino acid sequences of these tree shrew CYP2Ds were 75%-78% identical to human CYP2D6, and phylogenetic analysis showed that they were more closely related to human CYP2D6 than rat CYP2Ds, similar to dog and pig CYP2Ds. For tree shrew CYP2D8b, two additional transcripts were isolated that contained different patterns of deletion. The gene and genome structures of CYP2Ds are generally similar in dogs, humans, pigs, and tree shrews. Tree shrew CYP2D8a mRNA was most abundantly expressed in liver, among the tissue types analyzed, similar to dog CYP2D15 and pig CYP2D25 mRNAs. Tree shrew CYP2D8b mRNA was also expressed in liver, but at a level 7.3-fold lower than CYP2D8a mRNA. Liver microsomes and recombinant protein of both tree shrew CYP2Ds metabolized bufuralol and dextromethorphan, selective substrates of human CYP2D6, but the activity level of CYP2D8a greatly exceeded that of CYP2D8b. These results suggest that tree shrew CYP2D8a and CYP2D8b are functional drug-metabolizing enzymes, of which CYP2D8a is the major CYP2D in liver. SIGNIFICANCE STATEMENT: Novel tree shrew CYP2D8a and CYP2D8b cDNAs were isolated from liver. Their amino acid sequences were 75%-78% identical to human CYP2D6. For CYP2D8b, two additional transcripts contained different patterns of deletion. Tree shrew CYP2D8a mRNA was abundantly expressed in liver, similar to dog CYP2D15 and pig CYP2D25 mRNAs. Recombinant tree shrew CYP2Ds catalyzed the oxidation of bufuralol and dextromethorphan. Tree shrew CYP2D8a and CYP2D8b are functional drug-metabolizing enzymes, of which CYP2D8a is the major CYP2D in liver., (Copyright © 2024 by The American Society for Pharmacology and Experimental Therapeutics.)

Motilin is an important hormonal regulator in the migrating motor complex (MMC). Free fatty acid receptor-1 (FFAR1, also known as GPR40) has been reported to stimulate motilin release in human duodenal organoids. However, how FFAR1 regulates gastric motility in vivo is unclear. This study investigated the role of FFAR1 in the regulation of gastric contractions and its possible mechanism of action using Suncus murinus . Firstly, intragastric administration of oleic acid (C18:1, OA), a natural ligand for FFAR1, stimulated phase II-like contractions, followed by phase III-like contractions in the fasted state, and the gastric emptying rate was accelerated. The administration of GW1100, an FFAR1 antagonist, inhibited the effects of OA-induced gastric contractions. Intravenous infusion of a ghrelin receptor antagonist (DLS) or serotonin 4 (5-HT4) receptor antagonist (GR125487) inhibited phase II-like contractions and prolonged the onset of phase III-like contractions induced by OA. MA-2029, a motilin receptor antagonist, delayed the occurrence of phase III-like contractions. In vagotomized suncus, OA did not induce phase II-like contractions. In addition, OA promoted gastric emptying through a vagal pathway during the postprandial period. However, OA did not directly act on the gastric body to induce contractions in vitro . In summary, this study indicates that ghrelin, motilin, 5-HT, and the vagus nerve are involved in the role of FFAR1 regulating MMC. Our findings provide novel evidence for the involvement of nutritional factors in the regulation of gastric motility.

Cytochromes P450 (CYPs or P450s) are important enzymes for drug metabolism. Tree shrews are non-primate animal species used in various fields of biomedical research, including infection (especially hepatitis viruses), depression, and myopia. A recent tree shrew genome analysis indicated that the sequences and the numbers of P450 genes are similar to those of humans; however, P450s have not been adequately identified and analysed in this species.In this study, a novel CYP2E1 was isolated from tree shrew liver and was characterised in comparison with human, dog, and pig CYP2E1. Tree shrew CYP2E1 and human CYP2E1 showed high amino acid sequence identity (83%) and were closely related in a phylogenetic tree.Gene and genome structures of CYP2E1 were generally similar in humans, dogs, pigs, and tree shrews. Tissue expression patterns showed that tree shrew CYP2E1 mRNA was predominantly expressed in liver, just as for dog and pig CYP2E1 mRNAs. In tree shrews, recombinant CYP2E1 protein and liver microsomes metabolised chlorzoxazone and p -nitrophenol, probe substrates of human CYP2E1, just as they do in dogs and pigs.These results suggest that tree shrew CYP2E1 encodes a functional drug-metabolising enzyme that plays a role in the liver, similar to human CYP2E1.

Among the rare venomous mammals, the short-tailed shrew Blarina brevicauda has been suggested to produce potent neurotoxins in its saliva to effectively capture prey. Several kallikrein-like lethal proteases have been identified, but the active substances of B. brevicauda remained unclear. Here, we report Blarina paralytic peptides (BPPs) 1 and 2 isolated from its submaxillary glands. Synthetic BPP2 showed mealworm paralysis and a hyperpolarization shift (-11 mV) of a human T-type Ca 2+ channel (hCa v 3.2) activation. The amino acid sequences of BPPs were similar to those of synenkephalins, which are precursors of brain opioid peptide hormones that are highly conserved among mammals. However, BPPs rather resembled centipede neurotoxic peptides SLPTXs in terms of disulfide bond connectivity and stereostructure. Our results suggested that the neurotoxin BPPs were the result of convergent evolution as homologs of nontoxic endogenous peptides that are widely conserved in mammals. This finding is of great interest from the viewpoint of the chemical evolution of vertebrate venoms., Competing Interests: Conflict of interest The authors declare that they have no conflicts of interest with the contents of this article., (Copyright © 2023 The Authors. Published by Elsevier Inc. All rights reserved.)

Tree shrews display obvious reproductive cycles, and sexually mature male tree shrews produce little or no sperm with extremely low motility during the nonreproductive season; the mechanism underlying this phenomenon remains unknown. Because testis-specific serine/threonine kinases (TSSK) are specifically expressed in the testis and male germ cells of mammals, we hypothesized that they may have an important role in spermatogenesis or sperm function regulation in tree shrews. In addition, the expression, distribution, subcellular localization, and dynamic changes of TSSK in tree shrew sperm are unclear. Here we show that during the reproductive season, the seminiferous tubules were significantly larger as compared with the nonreproductive season and contained mature sperm and other germ cells. The mRNA expression of Tssk genes in testis was significantly higher than that in other tissues, and the mRNA level in the testis during the reproductive season was significantly higher than that in nonreproductive season. In addition, the mRNA level of Tssk3 in the testis and sperm was significantly higher than that of other members. Specifically, Tssk1 mRNA was distributed in the acrosome and throughout the flagellum of tree shrew sperm, Tssk2 was present in the acrosome, Tssk3 was localized to postacrosomal region and relocated to the main part of the flagellum after capacitation, and Tssk6 was distributed in the acrosome and postacrosomal region. These results indicate that the TSSK are important regulating reproductive function in tree shrews.

The greater white-toothed shrew Crocidura russula has been used as a sentinel species for estimating environmental risks to human populations. Previous studies in mining areas have focused on the liver of shrews as the primary target of physiological and metabolic changes due to heavy metal pollution. However, populations persist even when detoxification by the liver seems to be compromised and damage is observed. These pollutant-adapted individuals inhabiting contaminated sites may exhibit altered biochemical parameters that confer increased tolerance in various tissues other than the liver. The skeletal muscle tissue of C. russula might be an alternative tissue that allows the survival of organisms inhabiting historically polluted sites due to the detoxification of redistributed metals. Organisms from two heavy metal mine populations and one population derived from an unpolluted site were used to determine the detoxification activities, antioxidant capacity, and oxidative damage, as well as cellular energy allocation parameters and acetylcholinesterase activity (a biomarker of neurotoxicity). Muscle biomarkers differ between shrews from polluted sites and shrews from the unpolluted location, with the mine animals showing: (1) a decreased energy consumption concomitant with increased energy reserves and total available energy; (2) reduced cholinergic activity, suggesting an impairment of neurotransmission at the neuromuscular junction; (3) an overall decrease in detoxification capacity and enzymatic antioxidant response and a higher level of lipid damage. Also, some of these markers differed between females and males. These changes may have resulted from a decreased detoxifying capacity of the liver and could potentially bring about significant ecological effects for this highly active species. Heavy metal pollution induced physiological changes in Crocidura russula showing that skeletal muscle may serve as a backup sink organ allowing rapid species adaptation and evolution., Competing Interests: Declaration of competing interest The authors declare no conflict of interest., (Copyright © 2023 The Authors. Published by Elsevier B.V. All rights reserved.)

Tree shrews (Tupaia belangeri) are a non-rodent primate-like species sometimes used for biomedical research involving hepatitis virus infections and toxicology. Genome analysis has indicated similarities between tree shrews and humans in the numbers of cytochromes P450 (P450 or CYP), which constitute a family of important drug-metabolizing enzymes; however, P450s have not been fully investigated in tree shrews. In this study, we identified CYP1A1, CYP1A2, CYP1B1, and CYP1D1 cDNAs from tree shrew liver and compared their characteristics with dog, pig, and human CYP1As. The deduced amino acid sequences of tree shrew CYP1s were highly identical (82-87 %) to human CYP1s. In tree shrews, CYP1A1 and CYP1A2 mRNAs were preferentially expressed in liver, whereas CYP1D1 mRNA was preferentially expressed in kidney and lung. In contrast, CYP1B1 mRNA was expressed in various tissues, with the most abundant expression in spleen. Among the tree shrew CYP1 mRNAs, CYP1A2 mRNA was most abundant in liver, and CYP1B1 mRNA was most abundant in kidney, small intestine, and lung. All tree shrew CYP1 proteins heterologously expressed in Escherichia coli catalyzed caffeine and estradiol in a similar manner to tree shrew liver microsomes and human, dog, and pig CYP1 proteins. These results suggest that tree shrew CYP1A1, CYP1A2, CYP1B1, and CYP1D1 genes, different form human pseudogene CYP1D1P, are expressed in liver, small intestine, lung, and/or kidney and encode functional drug-metabolizing enzymes important in toxicology., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could influence or appear to influence the work reported in this article., (Copyright © 2022 Elsevier Inc. All rights reserved.)

Soricidae spp. (shrews) play an essential role in soil ecosystems and, due to their habitat and behavior, are exposed to soil pollutants, such as pesticides. Still, toxicity risk in Soricidae spp. has not been appropriately assessed. In this study, the musk shrew (Suncus murinus) was used as a model organism for toxicity assessment in Soricidae. Considering their carnivorous diet, it is reasonable to assume that the musk shrew has unique metabolic traits that are different from those of other common experimental models. This study describes the cytochrome P450 (CYP)-dependent metabolism affected by acetamiprid (ACP), a neonicotinoid insecticide. Pharmacokinetics analysis, an in vitro metabolic assay, and genetic analysis of CYP were performed and compared with data from mice and rats. Through phylogenetic and syntenic analyses, three families of CYP were identified in the musk shrew. Pharmacokinetic analysis showed that the blood concentration of ACP decreased more quickly in musk shrews than in mice. Moreover, the in vitro metabolic assay suggested more efficient metabolic responses toward ACP in musk shrews than in mice or rats. One of the CYP2A isoforms in musk shrews might be linked to a better ACP metabolism. From the results above, we describe novel metabolic traits of the musk shrew. Future research on recombinant CYP enzymes is necessary to fully understand CYP-dependent metabolism of xenobiotics in musk shrews., Competing Interests: Declaration of competing interest The authors declare no conflicts of interest., (Copyright © 2022 Elsevier Inc. All rights reserved.)

Multiple representatives of eulipotyphlan mammals such as shrews have oral venom systems. Venom facilitates shrews to hunt and/or hoard preys. However, little is known about their venom composition, and especially the mechanism to hoard prey in comatose states for meeting their extremely high metabolic rates. A toxin (BQTX) was identified from venomous submaxillary glands of the shrew Blarinella quadraticauda. BQTX is specifically distributed and highly concentrated (~ 1% total protein) in the organs. BQTX shares structural and functional similarities to toxins from snakes, wasps and snails, suggesting an evolutional relevancy of venoms from mammalians and non-mammalians. By potentiating thrombin and factor-XIIa and inhibiting plasmin, BQTX induces acute hypertension, blood coagulation and hypokinesia. It also shows strong analgesic function by inhibiting elastase. Notably, the toxin keeps high plasma stability with a 16-h half-life in-vivo, which likely extends intoxication to paralyze or immobilize prey hoarded fresh for later consumption and maximize foraging profit., (© 2022. The Author(s), under exclusive licence to Springer Nature Switzerland AG.)

Heavy metals accumulated in the environment due to the mining industry may impact on the health of exposed wild animals with consequences at the population level via survival and selection of the most resistant individuals. The detection and quantification of shifts in gene frequencies or in the genetic structure in populations inhabiting polluted sites may be used as early indicators of environmental stress and reveal potential 'candidate gene biomarkers' for environmental health assessment. We had previously observed that specimens of the Greater white-toothed shrew (Crocidura russula) from two heavy metal mines in Southern Portugal (the Aljustrel and the Preguiça mines) carried physiological alterations compared to shrews from an unpolluted site. Here, we further investigated whether these populations showed genetic differences in genes relevant for physiological homeostasis and/or that are associated with pathways altered in animals living under chronic exposure to pollution, and which could be used as biomarkers. We analysed the mitochondrial cytochrome b (Cytb) gene and intronic and/or exonic regions of four nuclear genes: CYP1A1, LCAT, PRPF31, and p53. We observed (1) population differences in allele frequencies, types of variation, and diversity parameters in the Cytb, CYP1A1, and p53 genes; (2) purifying selection of Cytb in the mine populations; (3) genetic differentiation of the two mine populations from the reference by the p53 gene. Adding to our previous observations with Mus spretus, we provide unequivocal evidence of a population effect exerted by the contaminated environment of the mines on the local species of small mammals., (© 2021. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

Venomousness is a complex functional trait that has evolved independently many times in the animal kingdom, although it is rare among mammals. Intriguingly, most venomous mammal species belong to Eulipotyphla (solenodons, shrews). This fact may be linked to their high metabolic rate and a nearly continuous demand of nutritious food, and thus it relates the venom functions to facilitation of their efficient foraging. While mammalian venoms have been investigated using biochemical and molecular assays, studies of their ecological functions have been neglected for a long time. Therefore, we provide here an overview of what is currently known about eulipotyphlan venoms, followed by a discussion of how these venoms might have evolved under ecological pressures related to food acquisition, ecological interactions, and defense and protection. We delineate six mutually nonexclusive functions of venom (prey hunting, food hoarding, food digestion, reducing intra- and interspecific conflicts, avoidance of predation risk, weapons in intraspecific competition) and a number of different subfunctions for eulipotyphlans, among which some are so far only hypothetical while others have some empirical confirmation. The functions resulting from the need for food acquisition seem to be the most important for solenodons and especially for shrews. We also present several hypotheses explaining why, despite so many potentially beneficial functions, venomousness is rare even among eulipotyphlans. The tentativeness of many of the arguments presented in this review highlights our main conclusion, i.e., insights regarding the functions of eulipotyphlan venoms merit additional study.

Afrotheria is a clade of African-origin species with striking dissimilarities in appearance and habitat. In this study, we compared whole proteome sequences of six Afrotherian species to obtain a broad viewpoint of their underlying molecular make-up, to recognize potentially unique proteomic signatures. We find that 62% of the proteomes studied here, predominantly involved in metabolism, are orthologous, while the number of homologous proteins between individual species is as high as 99.5%. Further, we find that among Afrotheria, L. africana has several orphan proteins with 112 proteins showing < 30% sequence identity with their homologues. Rigorous sequence searches and complementary approaches were employed to annotate 156 uncharacterized protein sequences and 28 species-specific proteins. For 122 proteins we predicted potential functional roles, 43 of which we associated with protein- and nucleic-acid binding roles. Further, we analysed domain content and variations in their combinations within Afrotheria and identified 141 unique functional domain architectures, highlighting proteins with potential for specialized functions. Finally, we discuss the potential relevance of highly represented protein families such as MAGE-B2, olfactory receptor and ribosomal proteins in L. africana and E. edwardii, respectively. Taken together, our study reports the first comparative study of the Afrotherian proteomes and highlights salient molecular features.

Animals that use venom to feed on a wide diversity of prey may evolve a complex mixture of toxins to target a variety of physiological processes and prey-defense mechanisms. Blarina brevicauda, the northern short-tailed shrew, is one of few venomous mammals, and is also known to eat evolutionarily divergent prey. Despite their complex diet, earlier proteomic and transcriptomic studies of this shrew's venom have only identified two venom proteins. Here, we investigated with comprehensive molecular approaches whether B. brevicauda venom is more complex than previously understood. We generated de novo assemblies of a B. brevicauda genome and submaxillary-gland transcriptome, as well as sequenced the salivary proteome. Our findings show that B. brevicauda's venom composition is simple relative to their broad diet and is likely limited to seven proteins from six gene families. Additionally, we explored expression levels and rate of evolution of these venom genes and the origins of key duplications that led to toxin neofunctionalization. We also found three proteins that may be involved in endogenous self-defense. The possible synergism of the toxins suggests that vertebrate prey may be the main target of the venom. Further functional assays for all venom proteins on both vertebrate and invertebrate prey would provide further insight into the ecological relevance of venom in this species., (© The Author(s) 2020. Published by Oxford University Press on behalf of the Society for Molecular Biology and Evolution.)

The intestinal epithelial monolayer forms a mucosal barrier between the gut microbes and the host tissue. The mucosal barrier is composed of mucins and antimicrobial peptides and proteins (AMPs). Several animal studies have reported that Paneth cells, which occupy the base of intestinal crypts, play an important role in the intestinal innate immunity by producing AMPs, such as lysozyme, Reg3 lectins, α-defensins, and group IIA secretory phospholipase A2 (GIIA sPLA2). The house musk shrew (Suncus murinus) has only a few intestinal commensal bacteria and is reported to lack Paneth cells in the intestine. Although the expression of lysozyme was reported in the suncus intestine, the expression of other AMPs has not yet been reported. Therefore, the current study was focused on GIIA sPLA2 expression in Suncus murinus. GIIA sPLA2 mRNA was found to be most abundant in the spleen and also highly expressed in the intestine. Cells expressing GIIA sPLA2 mRNA were distributed not only in the crypt, but also in the villi. In addition, intragastric injection of lipopolysaccharide increased GIIA sPLA2 expression in the small intestine of suncus. These results suggest that suncus may host unique AMP-secreting cells in the intestine., (Copyright © 2019 Elsevier Ltd. All rights reserved.)

Lysozyme is one of the most prominent antimicrobial peptides and has been identified from many mammalian species. However, this enzyme has not been studied in the order Insectivora, which includes the most primitive placental mammals. Here, we done the lysozyme cDNA from Suncus murinus (referred to as suncus, its laboratory name) and compare the predicted amino acid sequence to those from other mammalian species. Quantitative PCR analysis revealed a relatively higher expression of this gene in the spleen and gastrointestinal tract of suncus. The lysozyme-immunopositive (ip) cells were found mainly in the red pulp of the spleen and in the mucosa of the whole small intestine, including the follicle-associated epithelium and subepithelial dome of Peyer's patches. The lysozyme-ip cells in the small intestine were mostly distributed in the intestinal crypt, although lysozyme-expressing cells were found not only in the crypt but also in the villi. On the other hand, only a few lysozyme-ip cells were found in the villi and some granules showing intense fluorescence were located toward the lumen. As reported for other mammals, Ki67-ip cells were localized in the crypt and did not co-localize with the lysozyme-ip cells. Moreover, fasting induced a decrease in the mRNA levels of lysozyme in the intestine of suncus. In conclusion, we firstly identified the lysozyme mRNA sequence, clarified expression profile of lysozyme transcripts in suncus and found a unique distribution of lysozyme-producing cells in the suncus intestine.

The pineal gland structure and ultrastructure in the Northern (Blarina brevicauda) and Southern short-tailed shrew (Blarina carolinensis) are described by light and electron microscopy. Results observed were similar to other mammals of Insectivora described previously, specifically, the hedgehog (Erinaceus europaeus) and the Old World mole (Talpa europea). Two different types of pinealocytes were noticed by electron microscopy, in addition to relatively few glial cells. Granular vesicles were not noticed in abundance. The granular endoplasmic reticulum was observed and studded with vesicles. The golgi apparatus was well developed and appeared often. Synaptic ribbons were observed in several different formations consisting of ribbons and/or rods. The ciliary derivative, the rudimentary photoreceptor structures found in the pinealocytes of population I, was noticed in a 9 + 0 tubular pattern. Within these semifossorial shrews, the relationship between specific intracellular organelles and their function was discussed.

Mercury (Hg) methylation is often elevated at the terrestrial-peatland interface, but methylmercury (MeHg) production at this "hot spot" has not been linked with in situ biotic accumulation. We examined total Hg and MeHg levels in peat, invertebrates and tissues of the insectivore Sorex cinereus (masked shrew), inhabiting a terrestrial-peatland ecotone in northern Minnesota, USA. Mean MeHg concentrations in S. cinereus (71 ng g -1 ) fell between concentrations measured in spiders (mean 70-140 ng g -1 ), and ground beetles and millipedes (mean 29-42 ng g -1 ). Methylmercury concentrations in S. cinereus increased with age and differed among tissues, with highest concentrations in kidneys and muscle, followed by liver and brain. Nearly all Hg in S. cinereus was in the methylated form. Overall, the high proportional accumulation of MeHg in peat at the site (3.5% total Hg as MeHg) did not lead to particularly elevated concentrations in invertebrates or shrews, which are below values considered a toxicological risk.

Small non-migratory mammals with Northern distribution ranges apply a variety of behavioural and physiological wintering strategies. A rare energy-saving strategy is Dehnel's phenomenon, involving a reduction and later regrowth of the body size, several organs and parts of the skeleton in red-toothed shrews (Soricidae). The size extremes coincide with major life stages. However, the physiological consequences for the shrew's metabolism remain poorly understood. In keeping with the energetic limitations that may induce the size changes, we hypothesised that metabolic incorporation rates should remain the same across the shrews' lifetimes. In contrast, fat turnover rates should be faster in smaller subadults than in large juveniles and regrown adults, as the metabolic activity of fat tissue increases in winter individuals (subadults). Measuring the changes in the ratio of exhaled stable carbon isotopes, we found that the baseline diet of shrews changed across the season. A diet switch experiment showed that incorporation rates were consistently rapid ( t 50 =38.2±21.1-69.3±53.5 min) and did not change between seasons. As predicted, fat turnover rates were faster in size-reduced subadults ( t 50 =2.1±1.3 h) compared with larger juveniles ( t 50 =5.5±1.7 h) and regrown adults ( t 50 =5.0±4.4 h). In all three age/size classes, all body fat was turned over after 9-24 h. These results show that high levels of nutrient uptake are independent of body size, whereas fat turnover rates are negatively correlated with body size. Thus, the shrews might be under higher pressure to save energy in winter and this may have supported the evolution of Dehnel's phenomenon., Competing Interests: Competing interestsThe authors declare no competing or financial interests., (© 2017. Published by The Company of Biologists Ltd.)

Nicotinic acetylcholine receptors (nAChRs) are neuromuscular proteins responsible for muscle contraction upon binding with chemical stimulant acetylcholine (ACh). The α-neurotoxins of snake mimic the structure of ACh and attacks nAChRs, which block the flow of ACh and leads to numbness and paralysis. The toxin-binding site of alpha subunit in the nAChRs is highly conserved throughout chordate lineages with few exceptions in resistance organisms. In this study, we have analyzed the sequence and structures of toxin-binding/resistant nAChRs and their interaction stability with toxins through molecular docking and molecular dynamics simulation (MDS). We have reported the potential glycosylation residues within the toxin-binding cleft adding sugar moieties through N-linked glycosylation in resistant organisms. Residue variations at key positions alter the secondary structure of binding cleft, which might interfere with toxin binding and it could be one of the possible explanations for the resistance to snake venoms. Analysis of nAChR-α-neurotoxin complexes has confirmed the key interacting residues. In addition, drastic variation in the binding stability of Mongoose nAChR-α-Bungarotoxin (α-BTX) and human nAChR-α-BTX complexes were found at specific phase of MDS. Our findings suggest that specific mutations in the binding site of toxin are potentially preventing the formation of stable complex of receptor-toxin, which might lead to mechanism of resistance. This in silico study on the binding cleft of nAChR and the findings of interacting residues will assist in designing potential inhibitors as therapeutic targets.

The function of the RIG-I-like receptors (RLRs; including RIG-I, MDA5, and LGP2) as key cytoplasmic sensors of viral pathogen-associated molecular patterns (PAMPs) has been subjected to numerous pathogenic challenges and has undergone a dynamic evolution. We found evolutionary evidence that RIG-I was lost in the Chinese tree shrew lineage. Along with the loss of RIG-I, both MDA5 (tMDA5) and LGP2 (tLGP2) have undergone strong positive selection in the tree shrew. tMDA5 or tMDA5/tLGP2 could sense Sendai virus (an RNA virus posed as a RIG-I agonist) for inducing type I IFN, although conventional RIG-I and MDA5 were thought to recognize distinct RNA structures and viruses. tMDA5 interacted with adaptor tMITA (STINGTMEM173/ERIS), which was reported to bind only with RIG-I. The positively selected sites in tMDA5 endowed the substitute function for the lost RIG-I. These findings provided insights into the adaptation and functional diversity of innate antiviral activity in vertebrates., Competing Interests: The authors declare no conflict of interest.

The medial preoptic nucleus (MPN) and the bed nucleus of the stria terminalis (BNST) of mice contain sexually dimorphic nuclei (SDNs) that are larger and have more neurons expressing calbindin D-28K (CB), a calcium-binding protein, in males than females. However, it is largely unknown whether such SDNs exist in species other than rodents. In this study, we performed an immunohistochemical study of CB in the MPN and BNST of musk shrews and Japanese quails to examine the existence of homologs of SDNs in mice. Like mice, musk shrews had a SDN exhibiting male-biased sex differences in volume and CB-immunoreactive (ir) cell number in the MPN. The BNST of musk shrews also contained a male-biased SDN, but consisted of non-CB neurons. The paratenial thalamic nucleus of musk shrews, but not mice, had more CB-ir cells in males than females. In Japanese quails of both sexes, CB-ir cells in the MPN and BNST were extremely small in number and did not cluster. These results suggest that the distribution of CB neurons differs among these species. Musk shrews may have a homolog of the SDN composed of CB neurons in the MPN of mice., (Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.)

Non-essential "heavy" metals such as cadmium tend to accumulate in an organism and thus are a particular threat for long-lived animals. Here we show that two unrelated, short-lived groups of mammals (rodents and shrews, separated by 100 Mio years of evolution) each have independently acquired mutations in their metal-responsive transcription factor (MTF-1) in a domain relevant for robust transcriptional induction by zinc and cadmium. While key amino acids are mutated in rodents, in shrews an entire exon is skipped. Rodents and especially shrews are unique regarding the alterations of this region. To investigate the biological relevance of these alterations, MTF-1s from the common shrew (Sorex araneus), the mouse, humans and a bat (Myotis blythii), were tested by cotransfection with a reporter gene into cells lacking MTF-1. Whereas shrews only live for 1.5-2.5 years, bats, although living on a very similar insect diet, have a lifespan of several decades. We find that bat MTF-1 is similarly metal-responsive as its human counterpart, while shrew MTF-1 is less responsive, similar to mouse MTF-1. We propose that in comparison to most other mammals, the short-lived shrews and rodents can afford a "lower-quality" system for heavy metal homeostasis and detoxification.

Animal models are indispensible to investigate the pathogenesis and treatments of non-alcoholic fatty liver diseases (NAFLD). Altered cholesterol metabolism has been implicated into the pathogenesis of NAFLD. Here, using high fat, cholesterol and cholate diet (HFHC), we generated a novel tree shrew (Tupaia belangeri chinensis) model of NAFLD, which displayed dyslipidemia with increased levels of plasma alanine aminotransferase (ALT) and aspartate aminotransferase (AST), total cholesterol (TC), low density lipoprotein-cholesterol (LDL-c) and high density lipoprotein-cholesterol (HDL-c), but decreased level of triglycerides (TG). Liver histopathology and genes expression indicated that HFHC diet successfully induced liver steatosis to inflammation and fibrosis progressively within 10 weeks. Moreover, HFHC induced the transcriptional expression of lipoprotein lipase (lpl) in the liver, but repressed the expression of LDL receptor, and the endogenous synthesis pathway and excretion of cholesterol. Notably, Poloxamer 407 (P-407) inhibition of LPL improved the severity of steatosis and reduced inflammation. These results illustrated that LPL plays an important role in cholesterol metabolism in NAFLD, and the tree shrew may be a valuable animal model for further research into NAFLD.

Many small mammals bask in the sun during rewarming from heterothermy, but the implications of this behaviour for their energy balance remain little understood. Specifically, it remains unclear whether solar radiation supplements endogenous metabolic thermogenesis (i.e., rewarming occurs through the additive effects of internally-produced and external heat), or whether solar radiation reduces the energy required to rewarm by substituting (i.e, replacing) metabolic heat production. To address this question, we examined patterns of torpor and rewarming rates in eastern rock elephant shrews (Elephantulus myurus) housed in outdoor cages with access to either natural levels of solar radiation or levels that were experimentally reduced by means of shade cloth. We also tested whether acclimation to solar radiation availability was manifested via phenotypic flexibility in basal metabolic rate (BMR), non-shivering thermogenesis (NST) capacity and/or summit metabolism (Msum). Rewarming rates varied significantly among treatments, with elephant shrews experiencing natural solar radiation levels rewarming faster than conspecifics experiencing solar radiation levels equivalent to approximately 20% or 40% of natural levels. BMR differed significantly between individuals experiencing natural levels of solar radiation and conspecifics experiencing approximately 20% of natural levels, but no between-treatment difference was evident for NST capacity or Msum. The positive relationship between solar radiation availability and rewarming rate, together with the absence of acclimation in maximum non-shivering and total heat production capacities, suggests that under the conditions of this study solar radiation supplemented rather than substituted metabolic thermogenesis as a source of heat during rewarming from heterothermy.

The telomere structure in the Iberian shrew Sorex granarius is characterized by unique, striking features, with short arms of acrocentric chromosomes carrying extremely long telomeres (up to 300 kb) with interspersed ribosomal DNA (rDNA) repeat blocks. In this work, we investigated the telomere physiology of S. granarius fibroblast cells and found that telomere repeats are transcribed on both strands and that there is no telomere-dependent senescence mechanism. Although telomerase activity is detectable throughout cell culture and appears to act on both short and long telomeres, we also discovered that signatures of a recombinogenic activity are omnipresent, including telomere-sister chromatid exchanges, formation of alternative lengthening of telomeres (ALT)-associated PML-like bodies, production of telomere circles, and a high frequency of telomeres carrying marks of a DNA damage response. Our results suggest that recombination participates in the maintenance of the very long telomeres in normal S. granarius fibroblasts. We discuss the possible interplay between the interspersed telomere and rDNA repeats in the stabilization of the very long telomeres in this organism., (Copyright © 2014, American Society for Microbiology. All Rights Reserved.)

Based on the results of a study conducted in 2006-2010 in the Buyunda River basin (a feeder of the Kolyma River), the influence of the population density of common shrews (Sorex) on some of the physiological and biochemical parameters (glycogen and lipids in the liver, the relative weight of the spleen, the white and brown adipose tissue cellularity of bone and brain tissue) was investigated. The content of energy reserve substances was correlated with the number of animals (fat deposits had a negative correlation; the glycogen content in the liver had a direct correlation). For the rest of the physiological-biochemical parameters, no significant correlation with the population density was detected, although for the content of brown fat and cellularity of bone marrow tissue in Sorex isodon, as well as the relative weight of the spleen in both species of shrews, a trend was observed. We suggest that the identified physiological changes indicate irregular feeding of animals in years with higher population densities.

The distribution of mucous secreting goblet cells was examined in the gastrointestinal tracts of three insectivores namely: Acomys spinosissimus (Southern African spiny mouse), Crocidura cyanea (Reddish gray musk shrew) and Amblysomus hottentotus (Hottentot golden mole) in order to improve our understanding of the quality and composition of the protective intestinal biofilm. Intestinal tracts were fixed and processed to wax for histology. Serial transverse sections were stained using alcian blue-periodic acid Schiff, alcian blue-aldehyde fuchsin and alcian blue-high iron diamine techniques. Photomicrographs of the stained sections were analyzed by quantifying the number of goblet cells containing mucins per mm(2) in the surface epithelial or crypt areas. Neutral mucins predominated in the gastric epithelium of all three insectivores, while sialomucins were absent in the stomach of C. cyanea. In all three species, goblet cells producing a mixture of neutral and acid mucins were most abundant throughout the intestinal tract as were cells secreting a mixture of sulfomucins and sialomucins. However, differences between the insectivore species were observed in the qualitative expression and distribution of mucins throughout the intestinal tract. Similarities between the insectivores of this study and other distantly related species suggest that mixed mucin goblet cells are essential for the formation of the biofilm, irrespective of their diet or taxonomy., (Copyright © 2013 Elsevier GmbH. All rights reserved.)

The present study describes the organisation of the cholinergic, catecholaminergic, and serotonergic neurons in the brains of the giant otter shrew, the Hottentot golden mole and the four-toed sengi, and the orexinergic (hypocretinergic) system in the giant otter shrew and four-toed sengi. The aim of the present study was to investigate the possible differences in the nuclear complement of these neural systems in comparison to previous studies on other Afrotherian species and mammalian species in general. Brains of the golden mole, sengi and giant otter shrew were coronally sectioned and immunohistochemically stained with antibodies against cholineacetyl-transferase, tyrosine hydroxylase, serotonin and orexin-A. The majority of nuclei revealed in the current study were similar among the species investigated, to other Afrotherian species, and to mammals generally, but certain differences in the nuclear complement highlighted phylogenetic interrelationships. The golden mole was observed to have cholinergic interneurons in the cerebral cortex, hippocampus, olfactory bulb and amygdala. The four-toed sengi had cholinergic neurons in both colliculi and in the cochlear nucleus, but lacked the catecholaminergic A15d group in the hypothalamus. In both the golden mole and the four-toed sengi, the locus coeruleus (A6d group) was made up of few neurons. The golden mole also exhibited an unusual foreshortening of the brain, such that a major (mesencephalic?) flexure in the brainstem was evident., (Copyright © 2013 Elsevier B.V. All rights reserved.)

The biochemical and histopathological changes in the lower esophageal sphincter (LES) in the pathogenesis of gastroesophageal reflux disease have gained interest. The least shrew is able to vomit in response to emetogens and provides a good model to study the histology of this phenomenon relative to the published reports in the commonly used but vomit-incompetent laboratory species. The LES is located at the junction of the esophagus and stomach. It typically closes at rest and opens in response to swallowing. Our findings demonstrate that the least shrew does not have a well-defined LES, lacks esophageal glands and has a mucosal valve-like projection from the terminal end of the esophagus before joining the gastric epithelium at the lesser curvature. In addition, the least shrew has thoracic and abdominal components prior to joining the gastric epithelium. The mucosal lining of the esophagus is folded, becoming clearly convoluted and forming a bucket-like structure at the level of the esophageocardiac junction (ECJ). No significant differences are to be found between the structure and thickness of the wall before and after the ECJ. Thus, the ECJ forming the LES is relatively less complex than those of other mammals including man. The distribution of enterochromaffin (EC) cells is confined to the lamina propria of the junction and is not associated with the cardiac glands, suggesting its functional involvement with the smooth muscle in and around the ECJ. In conclusion, the least shrew's anatomical sphincter appears ill-defined and is replaced by a less sturdy valve-like mucosal flap.

Red-toothed shrews (subfamily Soricinae) exhibit the highest mass-specific rates of O₂ consumption recorded among eutherian mammals, though surprisingly no data appears to be available on the functional characteristics of their hemoglobin (Hb). As a first step in addressing this shortcoming, we investigated the O₂ binding characteristics of Taiwanese brown-toothed shrew (Episoriculus fumidus) Hb and its temperature and pH dependence in the absence and presence of anionic red blood cell effectors. Although comparative data regarding the intrinsic O₂ affinity of other shrew species are currently unavailable, our data suggest that the sensitivity of this high-elevation endemic species' Hb to allosteric effector molecules is similar to that of the two lowland species of white-toothed (crocidurine) shrews examined to date. The efficient exploitation of blood O₂ reserves by E. fumidus appears to be achieved via synergistic modulation of O₂ affinity by Cl⁻ and organic phosphates that moreover dramatically lowers the overall enthalpy of oxygenation of their Hb. Oxygen unloading is presumably further enhanced by a relatively high Bohr effect (ΔLog P₅₀/ΔpH = -0.69) and marked reduction in the titratable histidine content (predicted low proton buffering value) of the component globin chains relative to human HbA. Notably, however, the limited data available suggest these latter attributes may be widespread among shrews and hence likely are not adaptations to chronic altitudinal hypoxia per se.

Shrews (Sorex cinereus) collected at a historic mine in Nova Scotia, Canada, had approximately twice the arsenic body burden and 100 times greater daily intake of arsenic compared with shrews from a nearby uncontaminated background site. Shrews store arsenic as inorganic and simple methylated arsenicals. Much of the arsenic associated with their primary food source, i.e., small invertebrates, may be soil adsorbed to their exoskeletons. A physiologically based extraction test estimated that 47 ± 2% of invertebrate arsenic is bioaccessible in the shrew gastrointestinal tract. Overall, shrews appear to be efficient at processing and excreting inorganic arsenic.

An improved and simple reversed-phase high performance liquid chromatography method with electrochemical detection for the simultaneous determination of amino acids in brain tissue of Suncus murinus was developed. Homogenates from 5 different brain areas were derivatized with o-phthalaldehyde in the presence of sodium sulphite. Subsequent separation was achieved using linear gradient elution over 30 min. The derivatives were stable for up to 20 h at 4 degrees C. The method was accurate, reproducible, and showed good linearity. The recoveries were >88% for aspartate, glutamine, glutamate, glycine and gamma-aminobutyric acid, with the limit of quantification varying from 5 to 30 pmol. The method was successfully applied for the measurement of amino acids under fed and fasted conditions., (Copyright (c) 2010 Elsevier B.V. All rights reserved.)

The organization of the nuclear subdivisions of the cholinergic, putative catecholaminergic and serotonergic systems of the brain of the elephant shrew (Elephantulus myurus) were determined following immunohistochemistry for choline acetyltransferase, tyrosine hydroxylase and serotonin, respectively. This was done in order to determine if differences in the nuclear organization of these systems in comparison to other mammals were evident and how any noted differences may relate to specialized behaviours of the elephant shrew. The elephant shrew belongs to the order Macroscelidea, and forms part of the Afrotherian mammalian cohort. In general, the organization of the nuclei of these systems resembled that described in other mammalian species. The cholinergic system showed many features in common with that seen in the rock hyrax, rodents and primates; however, specific differences include: (1) cholinergic neurons were observed in the superior and inferior colliculi, as well as the cochlear nuclei; (2) cholinergic neurons were not observed in the anterior nuclei of the dorsal thalamus as seen in the rock hyrax; and (3) cholinergic parvocellular nerve cells forming subdivisions of the laterodorsal and pedunculopontine tegmental nuclei were not observed at the midbrain/pons interface as seen in the rock hyrax. The organization of the putative catecholaminergic system was very similar to that seen in the rock hyrax and rodents except for the lack of the rodent specific C3 nucleus, the dorsal division of the anterior hypothalamic group (A15d) and the compact division of the locus coeruleus (A6c). The nuclear organization of the serotonergic system was identical to that seen in all eutherian mammals studied to date. The additional cholinergic neurons found in the cochlear nucleus and colliculi may relate to a specific acoustic signalling system observed in elephant shrews expressed when the animals are under stress or detect a predator. These neurons may then function to increase attention to this type of acoustic signal termed foot drumming., (Copyright (c) 2010 Elsevier B.V. All rights reserved.)

The diphyodont tooth replacement in mammals is characterized by a single replacement of a deciduous dentition by a permanent dentition. Despite its significance in mammalian biology and paleontology, little is known about the developmental mechanisms regulating the diphyodont replacement. Because the mouse never replaces its teeth, this study used the house shrew, Suncus murinus, as a model to investigate the control of the diphyodont replacement of a deciduous dentition by successions and additions of permanent teeth. Using morphological and gene expression analyses of serial sections, we have demonstrated the development of the upper dentition of the house shrew. In this species, the deciduous tooth germs are formed but soon become vestigial, whereas the successional and accessional (molar) germs are subsequently formed and developed. There are distinct Shh expression domains in the deciduous, successional, and accessional tooth germs, and those of the latter two germs are identified from the appearance of their primary enamel knots. The developmental sequence of tooth germs in the house shrew indicates that two adjacent primary enamel knots of the successional and accessional germs do not develop simultaneously, but with a constant time lag. We suggest that this mode of tooth succession and accession can be explained by a sequential inhibitory cascade model in which the timing of initiation and the spacing of tooth development are determined by the inhibition from the primary enamel knots of developmentally preceding adjacent tooth germs.

Genomic and biochemical analyses reveal that two independently evolved serine protease venoms in mammals and lizards have converged on nearly identical protein structures. Likewise, in two groups of frog, an identical toxin, caerulein, has arisen repeatedly from unique genes in those lineages., (Copyright 2010 Elsevier Ltd. All rights reserved.)

Objective: To investigate the distribution of ghrelin, an important appetite regulatory factor related to obesity, in the stomach of Suncus murinus, and attempted to elucidate the ghrelin-mediated regulatory effect in this animal., Methods: The stomachs of Suncus murinus were divided into 5 sections, cardia, fundus, greater curvature, lesser curvature, and pylorus, for investigating the ghrelin-producing cells by immunohistochemistry and Western blotting. Then Suncus murinus were randomized into two groups with ghrelin intraperitoneal injection (ghrelin-ip group) and saline intraperitoneal injection (control group), respectively. The effects of food intake and body weight were measured, and furthermore, the distribution of ghrelin in stomach was also investigated by immunohistochemistry and Western blotting., Results: The immunolocalization and protein levels of ghrelin differed significantly in different regions of the stomach of Suncus murinus. Furthermore, ghrelin administration did not change the rate of food intake, but resulted in an increase in body weight compared with the control group. In this study, we elucidated the distribution of ghrelin-producing cells in the stomach of Suncus murinus in detail for the first time. Ghrelin intraperitoneal administration was found to induce an increase in body weight without changing food intake in this species., Conclusion: Our study implied ghrelin showed a different regulatory function in Suncus murinus from other species. It is considered that ghrelin may be associated with obesity-resistance phenomenon in Suncus murinus.

Due to large zinc discharges into the global environment, both terrestrial and aquatic environments have been polluted with zinc. The embanked floodplains of the lower Rhine River in the Netherlands contain large amounts of heavy metals, including zinc. These large amounts of heavy metals may pose risks to flora and fauna by accumulation in food webs, and risk assessment may be required. However, toxicokinetic models for zinc metabolism in mammalian wildlife species are currently lacking. The present study describes the development of a zinc kinetics model that predicts internal zinc concentrations in small mammals by modeling zinc absorption and excretion dependent on the dietary zinc intake using adapted Michaelis-Menten equations. The equations were parameterized based on experimental data reported in the scientific literature. Within the dietary zinc range of 3 to 104 mg/kg dry weight, the model predicts internal zinc concentrations and shows that the internal zinc is regulated within this range. Outside this range, the model could not be calibrated due to a lack of data. Validation of the model with four small mammal species living in an embanked floodplain along a distributary of the Rhine River illustrated that the model predicts internal zinc concentrations differing up to a factor of 1.6 (Common shrew) to 1.7 (Common vole) from field measurements.

Historically impacted by anthropogenic activities, the nature reserve of Doñana (SW Spain) was affected by an unprecedented spillage of mud and acidic water from the Aznalcóllar pyrite mine in April 1998. Although several studies have addressed the influence of this spill on soils, water, and biota, there is little information on mammals, especially carnivorous species. We measured the concentrations of Fe, Mg, Pb, Hg, Cd, Zn, Cu, Mn, Mo, Co, and Cr in specimens of the greater white-toothed shrew, Crocidura russula, inhabiting the protected area affected by the mine spillage. We also examined other parameters to approach at the physiological effects of pollution. We found an increase in non-essential metals (Pb, Cd, and Hg), and morphometric, histological and genotoxic alterations. Age and gender were two significant factors explaining metal bioaccumulation: adults had higher Hg and Cd levels than juveniles, whereas males bioaccumulated more Pb and Co and less Mo than females. The micronucleus frequencies in blood erythrocytes were significantly higher in specimens from the polluted site than animals from the control site. Shrews from the impacted area also had hepatic alterations, namely increased liver-body ratio, focal necrosis, and signs of apoptosis in hepatocytes. Due to the relevance of small mammals in the diet of endangered species such as carnivorous birds and mammals, the findings of our study are of practical use for the management of the Doñana wildlife reserve and other protected Mediterranean wetlands.

Background: Laboratory conditions nullify the extrinsic factors that determine the wild expected lifespan and release the intrinsic or potential lifespan. Thus, wild animals reared in a laboratory often show an increased lifespan, and consequently an increased senescence phase. Senescence is associated with a broad suite of physiological changes, including a decreased responsiveness of the circadian system. The time-keeping hormone melatonin, an important chemical player in this system, is suspected to have an anti-aging role. The Greater White-toothed shrew Crocidura russula is an ideal study model to address questions related to aging and associated changes in biological functions: its lifespan is short and is substantially increased in captivity; daily and seasonal rhythms, while very marked the first year of life, are dramatically altered during the senescence process which starts during the second year. Here we report on an investigation of the effects of melatonin administration on locomotor activity of aging shrews., Methodology/principal Findings: 1) The diel fluctuations of melatonin levels in young, adult and aging shrews were quantified in the pineal gland and plasma. In both, a marked diel rhythm (low diurnal concentration; high nocturnal concentration) was present in young animals but then decreased in adults, and, as a result of a loss in the nocturnal production, was absent in old animals. 2) Daily locomotor activity rhythm was monitored in pre-senescent animals that had received either a subcutaneous melatonin implant, an empty implant or no implant at all. In non-implanted and sham-implanted shrews, the rhythm was well marked in adults. A marked degradation in both period and amplitude, however, started after the age of 14-16 months. This pattern was considerably delayed in melatonin-implanted shrews who maintained the daily rhythm for significantly longer., Conclusions: This is the first long term study (>500 days observation of the same individuals) that investigates the effects of continuous melatonin delivery. As such, it sheds new light on the putative anti-aging role of melatonin by demonstrating that continuous melatonin administration delays the onset of senescence. In addition, the shrew appears to be a promising mammalian model for elucidating the precise relationships between melatonin and aging.

Although ecotoxicological data on heavy metals are abundant, information on other potentially toxic elements with attributed deficiency and/or toxic disturbances is scarce. Here we quantify zinc, copper, iron, manganese, chromium, molybdenum, strontium, barium, and boron in bones of greater white-toothed shrews, Crocidura russula, inhabiting two protected Mediterranean coastal sites: the Ebro Delta, a wetland impacted by human activities, and the Medas Islands, a reference site. Natural and anthropogenic inputs significantly increase Fe, Mn, Mo, Sr, Ba, and B in specimens from the Ebro Delta, whereas Cu and Cr were higher in Medas' shrews. Principal component analysis allowed complete separation between sites along the first two axes in particular due to B, Sr, and Cu. This study provides metal reference values in bones of insectivores, explores their variability and bioaccumulation patterns in depth, and assesses the potential environmental risk and toxicity for biota exposed to the above elements.

Although many studies have demonstrated the action of motilin on migrating motor complex by using human subjects and relatively large animals, the precise physiological mechanisms of motilin remain obscure. One reason for the lack of progress in this research field is that large animals are generally not suitable for molecular-level study. To overcome this problem, in this study, we focused on the house musk shrew (Suncus murinus, order: Insectivora, suncus named as laboratory strain) as a small model animal, and we present here the results of motilin gene cloning and its availability for motilin study. The motilin gene has a high homology sequence with that of other mammals, including humans. Suncus motilin is predicted to exist as a 117-residue prepropeptide that undergoes proteolytic cleavage to form a 22-amino-acid mature peptide. The results of RT-PCR showed that motilin mRNA is highly expressed in the upper small intestine, and low levels of expression were found in many tissues. Morphological analysis revealed that suncus motilin-producing cells were present in the upper small intestinal mucosal layer but not in the myenteric plexus. Administration of suncus motilin to prepared muscle strips of rabbit duodenum showed almost the same contractile effect as that of human motilin. Moreover, suncus stomach preparations clearly responded to suncus or human motilin stimulation. To our knowledge, this is the first report that physiological active motilin was determined in small laboratory animals, and the results of this study suggest that suncus is a suitable model animal for studying the motilin-ghrelin family.

Some cases of Robertsonian (Rb) polymorphism in the common shrew (Sorex araneus L.) are believed not to be associated with hybrid zones. One of the hypotheses explaining the persistence of such Rb polymorphism is that they are maintained by some form of selection for Rb heterozygotes. To test this hypothesis, we compared several parameters between homozygotes and Rb heterozygotes for the mp chromosome pair. We used shrews from Jurowce population in Poland, situated within the range of the Białowieza race, where Rb polymorphism persists far from any known hybrid zone. We found no differences between the two karyotypic classes in maximum metabolic rate during running (forced activity). However, the Rb heterozygotes showed significantly higher maximum metabolic rate during swimming (forced activity combined with thermal stress). The levels of fluctuating asymmetry (FA) of homozygous and Rb heterozygous shrews were indistinguishable, indicating no effect of chromosomal heterozygosity on developmental stability of shrews. We suggest that selective advantages, such as the higher metabolic performance in activity combined with cold stress, may outweigh the expected negative effects of Rb heterozygosity upon fertility, and help to maintain huge areas of the Rb polymorphism in this species.

Haematological (WBC, RBC, Hgb and Hct) and genotoxicity (MNT) parameters, hepatic enzymatic activities (GST, GPx and GR), and a histopathological evaluation of liver, kidneys and gonads were assessed as general biomarkers of metal pollution in the shrew Crocidura russula inhabiting a pyrite mining area. Specimens exposed to metals presented a few significant alterations when compared with reference animals: GST activity decreased; micronuclei increased; and evident liver alterations related to metal exposure were observed. On the basis of all the parameters studied, age was an important factor that partly explained the observed variation, whereas sex was the least important factor. Significant correlations were also found between heavy metal concentrations and biomarkers evaluated, demonstrating the great influence of these metals in the metabolic alterations. To the best of our knowledge, these data constitute the first measurements of a battery of biomarkers in shrews from a mine site and are among the few available for insectivorous mammals.

In the present study, we determined the concentrations of Cu, Fe, Mn, and Zn in soil and several trophic compartments at a total of 16 sampling stations. The trophic compartments studied were primary producers, represented by two species of terrestrial mosses (Pseudoescleropodium purum and Hypnum cupressiforme) and oak trees (Quercus robur or Q. pyrenaica); primary consumers, represented by the wood mouse (Apodemus sylvaticus) and the yellow necked mouse (A. flavicollis); secondary consumers, represented by the shrew (Sorex granarius); and finally, detritivores, represented by slugs (Arion ater). Thirteen of the sampling stations were located in mature oak woodlands (Quercus sp.); two of the sampling stations were located in the area surrounding a restored lignite mine dump, and the other in an ultrabasic area. The analytical determinations revealed a lack of significant correlations among trophic compartments, possibly caused by effective regulation of metals by organisms and/or spatial variation in availability of metals from soil or food. Furthermore, the only element that showed a clear pattern of biomagnification was Cu; as for the other elements, there was always some divergence from such a pattern. Finally, the patterns of bioaccumulation in contaminated and woodland sampling stations were very similar, although there was enrichment of the concentrations of Cu, Mn, and Zn in the mice viscera, which, except for Mn, were related to higher edaphic concentrations.

Recent studies used Suncus murinus to investigate the anti-emetic potential of NK(1) tachykinin receptor antagonists. However, the pharmacology of tachykinin receptors in this species has not been fully characterized. In the present studies, therefore, we examined a range of tachykinin receptor agonists for a capacity to induce contractions of the isolated ileum. The tachykinin NK1 receptor preferring agonists substance P, septide and [Sar9Met(O2)11] substance P, and the tachykinin NK2 preferring agonists neurokinin A and GR 64349 (Lys-Asp-Ser-Phe-Val-Gly-R-gamma-lactam-Leu-Met-NH2) caused concentration dependent contractions with EC50 values in the nanomolar range. However, the tachykinin NK3 preferring agonists neurokinin B and senktide (1nM-1microM) induced only weak contractions. The action of senktide, but not [Sar9Met(O2)11] substance P, septide, or GR 64349, was antagonized significantly by atropine (P<0.05); tetrodotoxin and hexamethonium were inactive. The tachykinin NK1 receptor antagonist CP-99,994 ((+)-[(2S,3S)-3-(2-methoxy-benzyl-amino)-2-phenylpiperidine]) (10-100nM) inhibited substance P- and septide-induced contractions non-competitively. The pA2 value estimated for CP-99,994 against septide was 7.3+/-0.1. It also non-competitively antagonized the contractile responses induced by [Sar9Met(O2)11] substance P with a pA2 of 7.4+/-0.1. CP-99,994 also had a slight inhibitory action on neurokinin A-induced contractions, but did not modify the action of GR 64349. Conversely, the tachykinin NK2 receptor antagonist, saredutant, competitively antagonized GR 64349-induced contractions with a pA2 of 7.34+/-0.02. On the other hand, the presence of both CP-99,994 and saredutant competitively antagonized substance P-induced contraction. The present studies indicate that tachykininNK1 and NK2 receptors exist in the ileum of S. murinus and are involved in mediating contractions directly on smooth muscle, whereas tachykinin NK3 receptors may play a minor role involving a release of acetylcholine.

Neuropeptide Y (NPY) stimulates feeding, depresses sexual behavior, and its expression in the brain is modulated by energetic status. We examined the role of NPY in female musk shrews, a species with high energetic and reproductive demands; they store little fat, and small changes in energy can rapidly diminish or enhance sexual receptivity. Intracerebroventricular infusion of NPY enhanced acute food intake in shrews; however, NPY had little affect on sexual receptivity. The distribution of NPY immunoreactivity in the female musk shrew brain was unremarkable, but energy status differentially affected NPY immunoreactivity in several regions. Similar to what has been noted in other species, NPY immunoreactivity was less dense in brains of ad libitum shrews and greater in shrews subjected to food restriction. In two midbrain regions, both of which contain high levels of gonadotropin releasing hormone II (GnRH II), which has anorexigenic actions in shrews, NPY immunoreactivity was more sensitive to changes in food intake. In these regions, acute re-feeding (90-180 min) after food restriction reduced NPY immunoreactivity to levels noted in ad libitum shrews. We hypothesize that interactions between NPY and GnRH II maintain energy homeostasis and reproduction in the musk shrew.

American water shrews (Sorex palustris) are aggressive predators that feed on a variety of terrestrial and aquatic prey. They often forage at night, diving into streams and ponds in search of food. We investigated how shrews locate submerged prey using high-speed videography, infrared lighting, and stimuli designed to mimic prey. Shrews attacked brief water movements, indicating motion is an important cue used to detect active or escaping prey. They also bit, retrieved, and attempted to eat model fish made of silicone in preference to other silicone objects showing that tactile cues are important in the absence of movement. In addition, water shrews preferentially sniffed model prey fish and crickets underwater by exhaling and reinhaling air through the nostrils, suggesting olfaction plays an important role in aquatic foraging. The possibility of echolocation, sonar, or electroreception was investigated by testing for ultrasonic and audible calls above and below water and by presenting electric fields to foraging shrews. We found no evidence for these abilities. We conclude that water shrews detect motion, shape, and smell to find prey underwater. The short latency of attacks to water movements suggests shrews may use a flush-pursuit strategy to capture some prey.

Expression and localization of members of the aquaporin (AQP) family (AQP1, 2, 3, 4, and 5) in the kidney of the musk shrew (Suncus murinus) was examined by immunohistochemistry. AQP1 was expressed in the proximal tubules and in the thin limb of the loops of Henle. AQP1 was the only water channel expressed in the proximal nephron examined, indicating that AQP1 may be an independent water transporter in the proximal nephron. AQP2 and AQP5 were localized to the apical cytoplasm of the cortical to medullary collecting duct (CD) cells and AQP3 and AQP4 were localized to the basal aspect of the cortical to medullary CD cells. AQP3 expression was weaker in the cortical cells compared with the medullary cells, whereas AQP4 was strongly positive throughout the CD. These indicate that the CD is the main water reabsorption segment of the nephron and is regulated by AQPs. Indeed, apical water transport of CD cells of the musk shrew may be controlled by both AQP2 and AQP5. The characteristic expression pattern of the AQPs in this animal provides a novel animal model for elucidating the regulation of water reabsorption by AQPs in the mammalian kidney.