Jian-Xu Zhang

Social regulation of aggression by pheromonal activation of Or65a olfactory neurons in Drosophila

When two socially naive Drosophila males meet, they will fight. However, prior social grouping of males reduces their aggression. We found olfactory communication to be important for modulating Drosophila aggression. Although acute exposure to the male-specific pheromone 11-cis-vaccenyl acetate (cVA) elicited aggression through Or67d olfactory receptor neurons (ORNs), chronic cVA exposure reduced aggression through Or65a ORNs. Or65a ORNs were not acutely involved in aggression, but blockade of synaptic transmission of Or65a ORNs during social grouping or prior chronic cVA exposure eliminated social modulation of aggression. Artificial activation of Or65a ORNs by ectopic expression of the Drosophila gene TrpA1 was sufficient to reduce aggression. Social suppression of aggression requires subsets of local interneurons in the antennal lobe. Our results indicate that activation of Or65a ORNs is important for social modulation of male aggression, demonstrate that the acute and chronic effects of a single pheromone are mediated by two distinct types of ORNs, reveal a behaviorally important role for interneurons and suggest a chemical method to reduce aggression in animals.

Uropygial Gland-Secreted Alkanols Contribute to Olfactory Sex Signals in Budgerigars

The possible role of uropygial gland-secreted compounds in olfactory discrimination of sex or sex attractants in the budgerigar, Melopsittacus undulatus, was investigated using behavioral 2-choice tests and gas chromatography-mass spectrometry analysis. Our data showed that female budgerigars were capable of distinguishing males from females in a Y maze via body odor, indicating its sexual dimorphism. When we conducted a chemical assay of the uropygial preen gland secretions, we found 4 times more volatile octadecanol, nonadecanol, and eicosanol in ratios in males than in females, making them putative male pheromone candidates. Female birds also showed overt preferences for the odor of male preen gland secretions or the 3-alkanol blend equivalent preened onto the plumage of a male over that of female counterparts. Removal of any one alkanol was associated with a loss of attractiveness to the female. In another test device (a test cage) with visible male bird stimulus, females chose the male with the 3-alkanol blend of males over the other male with female preen gland secretion, whereas did not differentiate their responses between the males with either this blend or male preen gland secretions. The behavioral data robustly suggested that the 3 alkanols synergistically created a female attractant odor or male pheromone in the budgerigar and that bird uropygial glands have broader implications in sexual behavior than previously known. This is the first investigation with bioassay of components of the gland in a bird species.

Effects of weasel odor on behavior and physiology of two hamster species.

This study examined the behavioral and physiological effects of long-term exposure to overdose of aversive odor (predator odor) in two species of hamsters. About 0.05 mg of anal gland secretions of Siberian weasels (Mustela sibirica) was smeared at the oronasal groove of wild male ratlike hamsters (Cricetulus triton) (natural prey) and laboratory golden hamsters (Mesocricetus auratus) once every day for 4 weeks. After 28 days, the experimental groups of both hamster species displayed higher cortisol level, larger adrenal gland (in ratlike hamsters only), smaller thymus and flank gland, and lower aggression level than the conspecific control group (presented with water). Thus, the long-term presence of overdose of the anal gland secretion of the Siberian weasel could lower the aggression and social rank and suppress the immunity in the hamsters. The reproductive conditions of these prey species, however, seemed not to be affected. In addition, the similarities in the behavioral and physiological responses to the predator odor between the two species of hamsters showed that the responses to predator odor might be innate.

Dual Role of Preputial Gland Secretion and its Major Components in Sex Recognition of Mice

This study was aimed at validating the sexual attractiveness of hexadecanol and hexadecyl acetate, two putative pheromone compounds, from preputial gland secretion of mice. These two compounds have been reported to be among the major components of preputial gland secretion in both sexes but higher in quantity in males than females. In this study, we show that castration suppressed the production of the two compounds, further suggesting their association with maleness. Adding preputial gland secretion and the synthetic analogs of the two compounds to castrated male urine at their physiological levels in intact males increased the attractiveness of castrated male urine to female mice, showing that the two compounds were indeed male pheromones. Furthermore, their sexual attractiveness disappeared upon removing the vomeronasal organs (VNOs) from female recipients. Replenishing castrated male urine with preputial gland secretion and the two compounds at their physiological levels in females increased the attractiveness of castrated male urine to males. Such a reversal of sexual attractiveness for hexadecanol and hexadecyl acetate suggests that they had opposing dual effects in sexual attractiveness in a dosage-dependent manner.

Volatile Compounds in Anal Gland of Siberian Weasels (Mustela sibirica) and Steppe Polecats (M. eversmanni)

The volatile constituents in anal gland secretions of two sympatric Mustela species, the Siberian weasel (M. sibirica) and steppe polecat (M. eversmanni), were studied by the headspace technique, followed by gas chromatography–mass spectrometry (GC-MS) analysis. Nine sulfur-containing compounds were identified. They were 2,2-dimethylthietane, (Z)- or (E)-2,4-dimethylthietane, (E)-2,3-dimethylthietane, 2-ethylthietane, (E)-2-ethyl-3-methylthietane, (Z)-2-ethyl-3-methylthietane, 2-propylthietane, 3,3-dimethyl-1,2-dithiacyclopentane, and (Z)-3,4-dimethyl-2,2-dithiacyclopentane. Among them, (E)-2-ethyl-3- methylthietanes, (Z)-2-ethyl-3-methylthietanes, and (Z)-3,4-dimethyl-1,2-dithiacyclopentane were present in the polecat but not in the weasel. The predominant compound was 2,2-dimethylthietane in the weasel and (E)- or (Z)-2,4-dimethylthietane in the polecat. These differences were consistent between the two species, regardless of sex and age and, therefore, could possibly be used for species recognition. In the weasel, 2-ethylthietane was found only in the female, and the relative abundance of several compounds was significantly different between males and females. In the polecat, although no sex-specific volatile compounds were found, males and females differed in the relative abundance of several of the compounds. In both species, the relative abundance of some compounds varied with age. We conclude that these volatile compounds can be used to communicate information about species, sex, and age.

Potential Chemosignals in the Anogenital Gland Secretion of Giant Pandas, Ailuropoda melanoleuca, Associated with Sex and Individual Identity

With a combination of dichloromethane extraction and analysis by gas chromatography–mass spectrometry (GC-MS), we found 39 compounds (corresponding to 38 GC peaks) in the anogenital gland secretion (AGS) of captive adult giant pandas, Ailuropoda melanoleuca, during the non-mating season. In addition to indole, squalene, and some of the straight-chain fatty acids that had been characterized previously from the AGS of giant pandas, we identified several new compounds such as decenal, two isomers of decadienal, phenylacetic acid, 5-methylhydantoin, hydroquinone, phenylpropanoic acid, and erucic acid. Quantitative comparison of the relative abundances of the 20 main GC peaks revealed that 5-methylhydantoin, indole, and erucic acid are putative female pheromones, whereas squalene and hydroquinone are putative male pheromones. In addition to the presence of a few individual-specific compounds, the relative abundances of most of the 21 constituents varied more among individuals than within individuals. This suggests that individual identity might be coded in both digital and analog form. The chemical composition of different AGS samples from the same pandas consistently displayed a minimum cluster distance, much smaller than that between samples from different individuals in a hierarchical linkage cluster (average linkage) dendrogram. Our results indicate that the AGS might contain an “odor fingerprint.” Although putative sex pheromones such as squalene and erucic acid should be assessed further by bioassay, our study suggests that synthetic chemosignals might be useful in modulating the behavior and physiology of giant pandas.

A Two-Component Female-Produced Pheromone of the Spider Pholcus beijingensis

Chemical signaling plays an important role in spider sexual communication, yet the chemistry of spider sex pheromones remains poorly understood. Unlike insects and mammals, the identification of spider pheromones has seldom been attempted, and no multicomponent pheromones have been found. Empty webs of sexually receptive females of Pholcus beijingensis were more attractive to male conspecifics as compared to webs of sexually unreceptive females or to mature males. Coincidently, chemical analysis revealed that (E,E)-farnesyl acetate, diisobutyl phthalate, and hexadecyl acetate of the spider webs exhibited higher relative abundances in sexually receptive females than in sexually unreceptive females or males, indicative of possible pheromone components. Two-choice behavioral assays verified that the blend of (E,E)-farnesyl acetate and hexadecyl acetate (w/w: 2:1) attracted males at a dosage equivalent to the amounts of these compounds in one spider web, whereas neither compound alone aroused males. In addition, diisobutyl phthalate (a likely contaminant from contact with plastic) alone or in combination with either of the acetates did not evoke the males’ attraction. The behavioral data suggest that (E,E)-farnesyl acetate and hexadecyl acetate comprise a two-component female-produced sex pheromone in P. beijingensis, the first multicomponent pheromone found in spiders.

Mice Respond Differently to Urine and Its Major Volatile Constituents from Male and Female Ferrets

Our previous chemical investigation showed that the concentrations of urinary volatiles from males were much higher than those from females in the ferret (Mustela furo). The current study was designed to examine the behavioral significance and ecological relevance of this difference for one of the main prey of the ferret, the house mouse (Mus musculus). Our data showed that male mice displayed no difference in their response to raw male and female ferret urine. However, they showed significantly less response to female mouse urine mixed with ferret urine than to pure female mouse urine, and to female mouse urine mixed with male ferret urine than to female mouse urine mixed with female ferret urine. Furthermore, high levels of the three major volatiles (quinoline, 2,5-dimethylpyrazine, and 4-hepatanone) in male ferret urine were as effective as raw male ferret urine was in inhibiting the response of male mice. We discuss the ecological and behavioral significance of these findings in terms of chemical mimicry and cognitive feature extraction of predator odors in mice.

Male-specific (Z)-9-tricosene stimulates female mating behaviour in the spider Pholcus beijingensis

Chemical signals play an important role in spider sexual communication, yet the chemistry of spider sex pheromones remains poorly understood. Chemical identification of male-produced pheromone-mediating sexual behaviour in spiders has also, to our knowledge, not been reported before. This study aimed to examine whether chemically mediated strategies are used by males of the spider Pholcus beijingensis for increasing the probability of copulation. Based on data from gas chromatography–mass spectrometry analysis, electroantennography assay and a series of behavioural tests, we verified that (Z)-9-tricosene is a male-specific compound in the spider P. beijingensis. This compound acts as an aphrodisiac: it increases the likelihood that a female will mate. Mate-searching males release (Z)-9-tricosene to stimulate sexual behaviour of conspecific females. In the two-choice assay, however, sexually receptive females show no preference to the chambers containing (Z)-9-tricosene. This indicates that the male pheromone of P. beijingensis is not an attractant per se to the conspecific females. This is, to our knowledge, the first identification of a male-produced aphrodisiac pheromone in spiders.

Urine-Derived Key Volatiles May Signal Genetic Relatedness in Male Rats

Olfactory cues play a vital role in kin recognition and mate choice of the rat. Here, using 2 inbred strains of rats, Brown Norway (BN) and Lewis, as models to simulate kinship via genetic distance, we examined whether urine-derived volatiles are genetically determined, and, if so, how they code for olfactory information and the degree of genetic relatedness in mate choice. Binary choice tests showed that BN females preferred the urine odor of Lewis males over that of BN males, suggesting that they avoided males genetically similar to themselves and were able to assess this olfactorily. Gas chromatography-mass spectrometry analysis revealed that the composition of urine-derived volatiles was more similar within strains than between strains and suggests that odortypes may reflect genetic relatedness. Our data further show that BN males had lower ratios of 2-heptanone and 4-heptanone and higher ratios of dimethyl sulfone and 4-ethyl phenol than Lewis males. When we supplemented BN and Lewis male urine to make each similar, the preferences of BN females were reversed. We conclude that some urine-derived volatiles covary in relative abundance with degree of genetic relatedness, and this relationship may play a key role in chemical signaling and genetic identity in this species.