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Pheromonal communication plays a key role in the sociosexual behavior of rodents. The coadaptation between pheromones and chemosensory systems has been well illustrated in insects but poorly investigated in rodents and other mammals. We aimed to investigate whether coadaptation between male pheromones and female reception might have occurred in brown rats Rattus norvegicus. We recently reported that major urinary protein (MUP) pheromones are associated with male mating success in a brown rat subspecies, R. n. humiliatus (Rnh). Here, we discovered that MUPs were less polymorphic and occurred at much lower concentrations in males of a parapatric subspecies, R. n. caraco (Rnc), than in Rnh males, and found no association between pheromones and paternity success. Moreover, the observation of Rnc males that experienced chronic dyadic encounters and established dominance–submission relationships revealed that the dominant males achieved greater mating success than the subordinate males, but their MUP levels did not differ by social status. These findings suggest that male mating success in Rnc rats is related to social rank rather than to pheromone levels and that low concentration of MUPs might not be a reliable signal for mate choice in Rnc rats, which is different from the findings obtained in Rnh rats. In addition, compared with Rnh females, Rnc females exhibited reduced expression of pheromone receptor genes, and a lower number of vomeronasal receptor neurons were activated by MUP pheromones, which imply that the female chemosensory reception of pheromones might be structurally and functionally coadapted with male pheromone signals in brown rats.

Background Invasive species bring a serious effect on local biodiversity, ecosystems, and even human health and safety. Although the genetic signatures of historical range expansions have been explored in an array of species, the genetic consequences of contemporary range expansions have received little attention, especially in mammal species. In this study, we used whole-genome sequencing to explore the rapid genetic change and introduction history of a newly invasive brown rat (Rattus norvegicus) population which invaded Xinjiang Province, China in the late 1970s. Results Bayesian clustering analysis, principal components analysis, and phylogenetic analysis all showed clear genetic differentiation between newly introduced and native rat populations. Reduced genetic diversity and high linkage disequilibrium suggested a severe population bottleneck in this colonization event. Results of TreeMix analyses revealed that the introduced rats were derived from an adjacent population in geographic region (Northwest China). Demographic analysis indicated that a severe bottleneck occurred in XJ population after the split off from the source population, and the divergence of XJ population might have started before the invasion of XJ. Moreover, we detected 42 protein-coding genes with allele frequency shifts throughout the genome for XJ rats and they were mainly associated with lipid metabolism and immunity, which could be seen as a prelude to future selection analyses in the novel environment of XJ. Conclusions This study presents the first genomic evidence on genetic differentiation which developed rapidly, and deepens the understanding of invasion history and evolutionary processes of this newly introduced rat population. This would add to our understanding of how invasive species become established and aid strategies aimed at the management of this notorious pest that have spread around the world with humans.

Murine rodents are excellent models for study of adaptive radiations and speciation. Brown Norway rats (Rattus norvegicus) are successful global colonizers and the contributions of their domesticated laboratory strains to biomedical research are well established. To identify nucleotide-based speciation timing of the rat and genomic information contributing to its colonization capabilities, we analyzed 51 whole-genome sequences of wild-derived Brown Norway rats and their sibling species, R. nitidus, and identified over 20 million genetic variants in the wild Brown Norway rats that were absent in the laboratory strains, which substantially expand the reservoir of rat genetic diversity. We showed that divergence of the rat and its siblings coincided with drastic climatic changes that occurred during the Middle Pleistocene. Further, we revealed that there was a geographically widespread influx of genes between Brown Norway rats and the sibling species following the divergence, resulting in numerous introgressed regions in the genomes of admixed Brown Norway rats. Intriguing, genes related to chemical communications among these introgressed regions appeared to contribute to the population-specific adaptations of the admixed Brown Norway rats. Our data reveals evolutionary history of the Brown Norway rat, and offers new insights into the role of climatic changes in speciation of animals and the effect of interspecies introgression on animal adaptation.

Two parapatric Rattus norvegicus subspecies, R. n. humiliatus (RNH) and R. n. caraco (RNC), are classified according to morphological divergence and are mainly distributed in North and Northeast China. Here, we aimed to explore the population genetic structure, genetic boundary, and gene flow in these rats using 16 microsatellite loci. Structure analysis and principal component analysis revealed 3 ancestral clusters. We found that the intermediate cluster exhibited higher genetic diversity and a lower inbreeding coefficient than the other 2 clusters. The genetic differentiation between the 3 clusters was significant but weak, with a higher FST value being observed between the clusters on both sides. The subspecies boundary inferred from microsatellite markers may indicate the existence of an admixture or hybridization area covering Liaoning, Inner Mongolia, and Jilin Provinces, rather than corresponding to the administrative provincial boundaries between Liaoning and Jilin. The RNH and RNC subspecies presented moderate gene exchange and an asymmetric bidirectional gene flow pattern, with higher gene flow from the RNH subspecies to the RNC subspecies, constraining speciation. Such genetic characteristics might be explained by biological processes such as dispersal ability, mate choice, and dynamic lineage boundaries.

Hexadecanol (16OH) and hexadecyl acetate (16Ac) are two pheromones secreted in a large quantity by mouse preputial glands and act on male and female mice differentially. Yet the underlying molecular and cellular mechanisms remain to be elucidated. In this study, we examined the activation of vomeronasal sensory neurons (VSNs) by these two pheromones and mapped the downstream neural circuits that process and relay their chemosignals. Using the calcium imaging method and immunohistochemistry, we found that a small number of VSNs were activated by 16OH, 16AC, or both in the male and female mice, most of which were located apically in the vomeronasal epithelium, and their numbers did not increase when the concentrations of 16OH and 16Ac were raised by 10,000-fold except that of female VSNs in response to 16OH. In the accessory olfactory bulb (AOB), the two pheromones evoked more c-Fos+ neurons in the anterior AOB (aAOB) than in the posterior AOB (pAOB); and the increases in the number of c-Fos+ neurons in both aAOB and pAOB were dose-dependent; and between sexes, the female AOB responded more strongly to 16OH than to 16Ac whereas the male AOB had the opposite response pattern. This sexual dimorphism was largely retained in the downstream brain regions, including the bed nucleus of the stria terminalis (BNST), the medial amygdaloid nucleus (MeA), the posteromedial cortical amygdaloid nucleus (PMCo), the medial preoptic area (MPA), and the ventromedial hypothalamic nucleus (VmH). Taken together, out data indicate that there is one V1r receptor each for 16OH, 16Ac, or both, and that activation of these receptors evokes sexually dimorphic neural circuits, directing different behavioral outputs and possibly modulating other pheromone-induced responses.

Phenotypic variation and its epigenetic regulations within the inbred isogenic mice have long intrigued biologists. Here, we used inbred C57BL/6 mice to examine the individual differences and the inheritance of social dominance and male pheromones, expecting to create a model for studying the underlying epigenetic mechanisms for the evolution of these traits. We used a repeated male-male contest paradigm to form stable dominance-submission relationships between paired males and make superior or inferior quality manifest. Females showed olfactory preferences for the urine of dominant males to that of subordinate opponents. Gas chromatography-mass spectrometer analysis revealed that dominance-related or superior quality related pheromones were actually exaggerated male pheromone components (e.g., E-β-farnesene, hexadecanol, and 1-hexadecanol acetate) of preputial gland origin. Although the socially naive sons of both dominant and subordinate males elicited the same female attraction when reaching adulthood, the former could dominated over the latter during undergoing the male-male competition and then gained more attraction of females. Our results demonstrated that social dominance or superior quality and the related pheromones were heritable and could be expressed through the interaction between aggression-related epigenotypes and male-male contests. It suggested that the evolution of sexually selected traits could be epigenetically determined and promoted through female mate choice. The epigenetic mechanisms driving the individual differences in behavior and male pheromones deserve further studies.

Male animals with more conspicuous visual and acoustic signals increase their mating success, but also increase the risk of being attacked by eavesdropping predators. In rodents, males with richer sex pheromones often have higher attractiveness to females, but whether or not the males are also at higher predation risk is poorly known. Here, we used 2 laboratory inbred strains of the rat Rattus norvegicus, Brown Norway (BN) and Lewis (LEW), and wild-captured rats as odor donors to assess the relationship between the pheromone levels in male rats and attractiveness to domestic cats Felis catus. LEW rats had significantly higher levels of male pheromones (e.g., 4-heptanone, 2-heptanone, and 9-hydroxy-2-nonanone) than BN rats. Simultaneously, wild-captured male rats were selectively assigned to 2 groups (HIGH or LOW) based on pheromone content as determined by gas chromatography–mass spectrometry (GC-MS). Binary choice tests were carried out during the night in the test room. We found that cats spent more time investigating male bedding and urine of LEW rats than the counterpart of BN rats. Likewise, cats also preferred bedding and urine odor of the HIGH wild rats compared with the counterparts of LOW wild rats. Adding synthetic analogs of the 3 pheromone ketones into the urine of either BN rats or LOW wild rats significantly increased their attractiveness to cats. Our data suggest that the rats with exaggerated male pheromones more strongly attracted predators and thus as a consequence may suffer from elevated predation risk.

Whole-genome sequencing of wild-derived rat species can provide novel genomic resources, which may help decipher the genetics underlying complex phenotypes. As a notorious pest, reservoir of human pathogens, and colonizer, the Asian house rat, Rattus tanezumi, is successfully adapted to its habitat. However, little is known regarding genetic variation in this species. In this study, we identified over 41,000,000 single-nucleotide polymorphisms, plus insertions and deletions, through whole-genome sequencing and bioinformatics analyses. Moreover, we identified over 12,000 structural variants, including 143 chromosomal inversions. Further functional analyses revealed several fixed nonsense mutations associated with infection and immunity-related adaptations, and a number of fixed missense mutations that may be related to anticoagulant resistance. A genome-wide scan for loci under selection identified various genes related to neural activity. Our whole-genome sequencing data provide a genomic resource for future genetic studies of the Asian house rat species and have the potential to facilitate understanding of the molecular adaptations of rats to their ecological niches.

Background In rats, urine-borne male pheromones comprise organic volatile compounds and major urinary proteins (MUPs). A number of volatile pheromones have been reported, but no MUP pheromones have been identified in rat urine. Results We used sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE), isoelectric focusing electrophoresis (IEF), nano-liquid chromatography-tandem mass spectrometry (nLC-MS/MS) after in gel digestion of the proteins and quantitative real-time PCR (qRT-PCR) and showed that the levels of two MUPs, odorant-binding protein 3 (OBP3) (i.e. PGCL4) and MUP13 (i.e. PGCL1), in urine and their mRNAs in liver were higher in males than in females and were suppressed by orchidectomy and restored by testosterone treatment (T treatment). We then generated recombinant MUPs (rMUPs) and found that the sexual attractiveness of urine from castrated males to females significantly increased after the addition of either recombinant OBP3 (rOBP3) or recombinant MUP13 (rMUP13). Using c-Fos immunohistochemistry, we further examined neuronal activation in the brains of female rats after they sniffed rOBP3 or rMUP13. Both rOBP3 and rMUP13 activated the accessory olfactory bulb (AOB), medial preoptic area (MPA), bed nucleus of the stria terminalis (BST), medial amygdala (MeA), posteromedial cortical amygdala (PMCo) and ventromedial nucleus of the hypothalamus (VMH), which participate in the neural circuits responsible for pheromone-induced sexual behaviours. In particular, more c-Fos-immunopositive (c-Fos-ir) cells were observed in the posterior AOB than in the anterior AOB. Conclusions The expression of OBP3 and MUP13 was male-biased and androgen-dependent. They attracted females and activated brain areas related to sexual behaviours in female rats, suggesting that both OBP3 and MUP13 are male pheromones in rats. Particularly, an OBP excreted into urine was exemplified to be a chemical signal.

The central 5-hydroxytryptamine system impairs sociosexual behaviors and olfaction preferences in sexually naive mice. However, it remains unknown whether reproductive experiences impart an effect on the sexual olfactory preferences of female mice lacking central serotonin. Here, we aimed at examining such effects and the underlying mechanisms using Tph2 knockout female mice. Sexually naive Tph2-/- female mice failed to recognize olfactory cues regarding sex, genetic relatedness, and social hierarchy despite exhibiting normal olfactory discrimination. However, reproduction-experienced Tph2-/- female mice recovered sexual olfactory preferences, as did sexually naive Tph2+/+ females. Meanwhile, both the estrogen receptor α and oxytocin receptor in the amygdala of reproduction-experienced Tph2-/- females presented upregulated expression at the mRNA level and an upward tendency at the protein level vs. sexually naive Tph2-/- females. Intracerebroventricular administration of a combination of estrogen receptor α and oxytocin receptor agonists, but not either agent alone, could restore the sexual olfactory preferences of sexually naive Tph2-/- female mice to some degree. We speculate that estrogen receptor α and oxytocin receptor activation in the amygdala after reproductive experiences restores sexual olfactory recognition in Tph2-/- female mice.

In this study, we examined how urine-borne volatile compounds (UVCs) and darcin of male mice are inherited from parents and interact to modulate the olfactory preferences of females using two inbred strains of mice, C57Bl/6 (C57) and BALB/c (BALB), and their reciprocal hybrids (BC = BALB♀× C57♂; CB = C57♀ × BALB♂). Chemical analysis revealed that the UVCs of C57BL/6 males were quantitatively distinguishable from those of BALB/c males. Darcin was detected in C57 urine, but not in BALB urine. The levels of UVCs and darcin in both BC and CB were intermediate between those of C57 and BALB. Behaviourally, C57 females consistently preferred BALB male urine over C57 or CB males despite that there are trace amounts of darcin in BALB urine. However, the preference for BALB urine disappeared in contact two-choice tests of BALB vs. BC pairs, and restored when recombinant darcin was added to BALB male urine. Our results suggested that both UVCs and darcin in male mice are quantitatively inherited and interact to affect the olfactory preferences of females.

Rats are predators of mice in nature. Nevertheless, it is a common practice to house mice and rats in a same room in some laboratories. In this study, we investigated the behavioral and physiological responsively of mice in long-term co-species housing conditions. Twenty-four male mice were randomly assigned to their original raising room (control) or a rat room (co-species-housed) for more than 6 weeks. In the open-field and light-dark box tests, the behaviors of the co-species-housed mice and controls were not different. In a 2-choice test of paired urine odors [rabbit urine (as a novel odor) vs. rat urine, cat urine (as a natural predator-scent) vs. rabbit urine, and cat urine vs. rat urine], the co-species-housed mice were more ready to investigate the rat urine odor compared with the controls and may have adapted to it. In an encounter test, the rat-room-exposed mice exhibited increased aggression levels, and their urines were more attractive to females. Correspondingly, the levels of major urinary proteins were increased in the co-species-housed mouse urine, along with some volatile pheromones. The serum testosterone levels were also enhanced in the co-species-housed mice, whereas the corticosterone levels were not different. The norepinephrine, dopamine, and 5-HT levels in the right hippocampus and striatum were not different between the 2. Our findings indicate that chronic co-species housing results in adaptation in male mice; furthermore, it appears that long-term rat-odor stimuli enhance the competitiveness of mice, which suggests that appropriate predator-odor stimuli may be important to the fitness of prey animals.

Background The Asian house rat (Rattus tanezumi) and the brown rat (Rattus norvegicus) are closely related species and are partially sympatric in southern China. Over the past 20 years, R. tanezumi has significantly expanded northward in China and partially replaced the native brown rat subspecies, R. n. humiliatus. Although invasive species are often more aggressive than native species, we did not observe interspecific physical aggression between R. tanezumi and R. n. humiliatus. Here, we focused on whether or not R. tanezumi was superior to R. n. humiliatus in terms of nonphysical competition, which is primarily mediated by chemical signals. Results We performed two laboratory experiments to test different paradigms in domesticated R. tanezumi and R. n. humiliatus. In Experiment 1, we caged adult male rats of each species for 2 months in heterospecific or conspecific pairs, partitioned by perforated galvanized iron sheets, allowing exchange of chemical stimuli and ultrasonic vocalization. The sexual attractiveness of male urine odor showed a tendency (marginal significance) to increase in R. tanezumi caged with R. n. humiliatus, compared with those in conspecific pairs. Hippocampal glucocorticoid receptor (GR) and brain-derived nutrition factor (BDNF) mRNA were upregulated in R. n. humiliatus and R. tanezumi, respectively, when the rats were caged in heterospecific pairs. In Experiment 2, we kept juvenile male rats in individual cages in rooms with either the same or the different species for 2 months, allowing chemical interaction. The sexual attractiveness of male urine was significantly enhanced in R. tanezumi, but reduced in R. n. humiliatus by heterospecific cues and mRNA expression of hippocampal GR and BDNF were upregulated by heterospecific cues in R. n. humiliatus and R. tanezumi, respectively. Although not identical, the results from Experiments 1 and 2 were generally consistent. Conclusions The results of both experiments indicate that nonphysical/chronic interspecific stimuli, particularly scent signals, between R. n. humiliatus and R. tanezumi may negatively affect R. n. humiliatus and positively affect R. tanezumi. We infer that chronic interspecific interactions may have contributed to the invasion of R. tanezumi into the range of R. n. humiliatus in natural habitats.

Although the brown rat (Rattus norvegicus) is widely used as a model mammal throughout biological sciences, little is known about genetic variation in wild rat populations or the relationship of commonly used inbred strains to their wild relatives. We sampled wild brown rats from the species’ presumed ancestral range in NW China and from a derived population in the UK and estimated nucleotide diversity and population subdivision, based on the sequences of 30 autosomal protein-coding loci. Neutral genetic diversity was close to 0.2% in both populations, which is about five times lower than diversity at the orthologous sites in a population of wild house mice from the species’ putative ancestral range in India. We found significant population differentiation between UK and Chinese populations, as assessed by Fst and the program STRUCTURE. Based on synonymous diversity and divergence between the brown rat and house mouse, we estimate that the recent effective population size in brown rats is approximately 130,000 (approximate 95% confidence interval 85,000-184,000), about fivefold lower than wild house mice.

The maternal environment has been shown to influence female olfactory preferences through early chemosensory experience. However, little is known about the influence of the maternal environment on chemosignals. In this study, we used two inbred mouse strains, C57BL/6 (C57) and BALB/c (BALB), and explored whether adoption could alter male chemosignals and thus influence female olfactory preferences. In Experiment 1, C57 pups were placed with BALB dams. Adult BALB females then served as the subjects in binary choice tests between paired male urine odours (BALB vs. C57, BALB vs. adopted C57 and C57 vs. adopted C57). In Experiment 2, BALB pups were placed with C57 dams, and C57 females served as the subjects in binary choice tests between paired male urine odours (C57 vs. BALB, C57 vs. adopted BALB, and BALB vs. adopted BALB). In both experiments, we found that females preferred the urine of males from different genetic backgrounds, suggesting that female olfactory preferences may be driven by genetic compatibility. Cross-fostering had subtle effects on female olfactory preferences. Although the females showed no preference between the urine odours of adopted and non-adopted males of the other strain, the BALB females preferred the urine odour of BALB males to that of adopted C57 males, whereas the C57 females showed no preference between the urine odour of C57 and adopted BALB males. Using gas chromatography-mass spectrometry (GC-MS) and stepwise discriminant analysis, we found that the ratios of volatile chemicals from urine and preputial gland secretions were altered in the fostered male mice; these changes may have resulted in the behavioural changes observed in the females. Overall, the results suggest that female mice prefer urine odours from males with different genetic backgrounds; this preference may be driven by genetic compatibility. The early maternal environment influences the chemosignals of males and thus may influence the olfactory preferences of females. Our study provides additional evidence in support of genotype-dependent maternal influences on phenotypic variability in adulthood.

The molecular evolution of DAX1, SRY, and SOX9, genes involved in mammalian sex determination, was examined in six primate species. DAX1 and SRY have been added to the X and Y chromosomes, respectively, during mammalian evolution, whereas SOX9 remains autosomal. We determined the genomic sequences of DAX1, SRY, and SOX9 in all six species, and calculated K(a), the number of nonsynonymous substitutions per nonsynonymous site, and compared this with the K(s), the number of synonymous substitutions per synonymous site. Phylogenetic trees were constructed by means of the DAX1, SRY, and SOX9 coding sequences, and phylogenetic analysis was performed using maximum likelihood. Overall measures of gene and protein similarity were closer for DAX1 and SOX9, but DAX1 exhibited nonsynonymous amino acid substitutions at an accelerated frequency relative to synonymous changes, similar to SRY and significantly higher than SOX9. We conclude that, at the protein level, DAX1 and SRY are under less selective pressure to remain conserved than SOX9, and, therefore, diverge more across species than does SOX9. These results are consistent with evolutionary stratification of the mammalian sex determination pathway, analogous to that for sex chromosomes.