The ownership signature in mouse scent marks is involatile

Male house mice advertise their territory ownership through urinary scent marks and use individual-specific patterns of major urinary proteins (MUPs) to discriminate between their own scent and that of other males. It is not clear whether recognition occurs through discrimination of the non-volatile proteins or protein-ligand complexes (direct model), or by the detection of volatile ligands that are released from MUPs (indirect model). To examine the mechanism underlying individual scent mark signatures, we compared investigatory and countermarking responses of male laboratory mice presented with male scent marks from a strain with a different MUP pattern, when they could contact the scent or when contact was prevented by a porous nitrocellulose sheet to which proteins bind. Mice investigated scent marks from other males whether these were covered or not, and biochemical analysis confirmed that the porous cover did not prevent the release of volatiles from scent marks. Having gained information through investigation, mice increased their own scent marking only if they had direct contact with another male's urine, failing to do this when contact was prevented. Individual signatures in scent marks thus appear to be carried by non-volatile proteins or by non-volatile protein-ligand complexes, rather than by volatiles emanating from the scent.     

Perspectives on over-marking: is it good to be on top?

What we refer to as over-marking occurs when one individual places its scent mark on top of, touching, or adjacent to the scent mark of another individual, usually a conspecific. Over-marking frequently occurs among mammals that share common paths, trails, and runways. Despite its ubiquity among terrestrial mammals, we know little about how individuals respond to over-marks and the function(s) of over-marking. Studies on voles and golden hamsters indicate that after exploring an over-mark, individuals respond selectively to the mark of the top-scent donor relative to that of the bottom-scent donor. Thus, individuals may be able to focus their attention on a particular scent mark relevant at a particular time and in a particular context, neglecting other scent marks that are present. The function(s) of over-marking are examined within the framework of ten hypotheses. Several hypotheses are plausible. However, the bulk of the literature is consistent with hypotheses stating that over-marking serving a role in olfactory communication between opposite and same-sex conspecifics. Lastly, we postulate the costs and benefits that may be garnered by the top-scent donor of an over-mark.     

The competing countermarks hypothesis: reliable assessment of competitive ability by potential mates

Scent marking on top of (overmarking), or in the vicinity of, a scent mark already present is commonly termed countermarking. Scent marks and countermarks provide a continuous record of competitive challenges between conspecifics, thus providing a reliable advertisement of an individual's ability to dominate or defend an area to other competitors and potential mates. To test the hypothesis that females should prefer males that countermark competing scent marks in their territory over those whose own marks are partially countermarked by a competing male, we manipulated scent marks in the territories of neighbouring male house mice (captive-bred Mus domesticus). As predicted, oestrous females were more strongly attracted to approach territory owners that countermarked the scent mark challenges of competitors than those that had been countermarked, and females themselves deposited more scent marks near the scents of these males. To investigate whether female mice use scent age, overlap or intrinsic qualitative or quantitative differences between scent marks and countermarks to make this discrimination, we redeposited male scent marks artificially as marks and partially overlapping countermarks, with or without a 24-h age difference between them. While the intrinsic quality or quantity of countermarks did not affect discrimination, an age difference between the original mark and subsequent countermark was important for consistent discrimination. The ultimate function of such competitive scent signalling thus may be to provide potential mates with a reliable indicator of the competitive ability of individuals advertising their high status. Since scent marks remain in the environment and are continuously available to challenge and investigation, they may provide a particularly effective and reliable means of dominance advertisement. Copyright 1999 The Association for the Study of Animal Behaviour.     

Meadow Voles (Microtus pennsylvanicus) and Prairie Voles (M. ochrogaster) Differ in Their Responses to Over-Marks from Opposite- and Same-Sex Conspecifics

Over‐marking occurs when one individual deposits its scent mark on the scent mark of a conspecific. Previous studies have shown that meadow voles (Microtus pennsylvanicus) and prairie voles (M. ochrogaster) that were exposed to an over‐mark of two same‐sex conspecifics, later responded similarly to the top‐scent mark but differed in their response to the bottom‐scent mark. In the present study, we examined the responses of meadow voles and prairie voles to same‐sex and mixed‐sex over‐marks to ascertain whether their responses reflect the different tactics which males and females in promiscuous (meadow voles) and monogamous (prairie voles) species use to attract opposite‐sex conspecifics and to compete with same‐sex conspecifics. Males and females of both species spent more time investigating the mark of the top‐scent donor than that of the bottom‐scent donor of an over‐mark. Meadow voles exposed to a mixed‐sex over‐mark spent more time investigating the mark of the opposite‐sex conspecific independently of whether it was from the top‐ or bottom‐scent donor. In contrast, prairie voles spent more time investigating the mark of the opposite‐sex donor if it was from the top‐scent donor. These results suggest that: (i) over‐marking serves a competitive function; (ii) the scent marks of individuals attract multiple mates in promiscuous species such as the meadow vole; and (iii) the scent marks of individuals establish and maintain pair bonds between familiar opposite‐sex conspecifics in monogamous species such as the prairie vole.     

Meadow voles and prairie voles differ in the percentage of conspecific marks they over-mark

Many terrestrial mammals scent mark in areas containing the scent marks of conspecifics, and thus, may deposit their own scent marks on top of those that were deposited previously by conspecifics. This phenomenon, known as over-marking appears to play a role in same-sex competition or mate attraction. The present study determines whether meadow and prairie voles avoid over-marking the scent marks of conspecifics, target the scent marks of conspecifics and over-mark them, or randomly over-mark the scent marks of conspecifics. The data show that meadow and prairie voles adjust the number and location of scent marks that they deposit in areas marked previously by particular conspecifics. Male and female meadow and prairie voles target the scent marks of opposite-sex conspecifics and over-mark them. Female meadow and prairie voles also target the scent marks of female conspecifics. In contrast, male meadow and prairie voles over-mark the scent marks of male conspecifics in a random manner. By differentially over-marking the scent marks of conspecifics, voles may be able to communicate particular information to a variety of conspecifics.     

Male and Female Meadow Voles,Microtus pennsylvanicus,Differ in Their Responses to Heterospecific/Conspecific Over‐Marks

Voles use runways, paths, and trails that may also be used by rabbits and mink. These shared areas could contain the scent marks of conspecifics and heterospecifics. Thus, it is likely that the scent marks of heterospecifics may overlap or be overlapped by those of voles, forming over‐marks. Much is known about how voles respond to over‐marks of two different conspecifics. However, we do not know how they would respond to an opposite‐sex conspecific whose scent marks are in an over‐mark with the scent marks of predator or the scent marks of a non‐predator heterospecifics. We tested the hypothesis that meadow voles, Microtus pennsylvanicus, differ in their response to the scent mark of the opposite‐sex conspecific if the scent mark was overlapped by that of a mink, a vole predator, or rabbit, a vole non‐predator. We found that female but not male voles showed a preference for the scent marks of the opposite‐sex conspecifics that were part of the mink‐vole over‐mark when compared to those of opposite‐sex conspecifics that were not part of the over‐mark. This preference by female voles was independent of whether the male vole was the top‐scent donor or bottom‐scent donor of the over‐mark. Male and female voles showed no preference between the scent marks of the opposite‐sex conspecifics whose marks were part of or not part of the rabbit‐vole over‐mark. Sex differences in the manner that meadow voles respond to rabbit‐vole and mink‐vole over‐marks are discussed.     

Scent wars: the chemobiology of competitive signalling in mice

Many mammals use scent marks to advertise territory ownership, but only recently have we started to understand the complexity of these scent signals and the types of information that they convey. Whilst attention has generally focused on volatile odorants as the main information molecules in scents, studies of the house mouse have now defined a role for a family of proteins termed major urinary proteins (MUPs) which are, of course, involatile. MUPs bind male signalling volatiles and control their release from scent marks. These proteins are also highly polymorphic and the pattern of polymorphic variants provides a stable ownership signal that communicates genome-derived information on the individual identity of the scent owner. Here we review the interaction between the chemical basis of mouse scents and the dynamics of their competitive scent marking behaviour, demonstrating how it is possible to provide reliable signals of the competitive ability and identity of individual males.     

The role of scent marking in the social communication of wild golden lion tamarins, Leontopithecus rosalia

The role of scent marking in the social communication of mammals is widely variable. One reason for this variation is that the function of scent marking may vary with different ecological and social conditions. The purpose of this study was to test four nonexclusive hypotheses explaining the role of scent-marking frequency in different ecological and social contexts for wild golden lion tamarins. Relative to ecological contexts, we compared scent-marking frequency during seasons of abundant and scarce food resources. Relative to social contexts, we compared scent-marking frequency when groups were isolated and when groups were in the presence of neighbouring groups. We found that the tamarins used scent marking to mark the location of food resources. Additionally, males used scent marking to communicate intrasexual dominance within their groups, while females did not. Our results also indicate that alpha females increased their scent-marking frequency to communicate to members of other groups, while the presence of members of other groups did not elicit a similar response by alpha males. We did not find evidence for a territorial function of scent marking in golden lion tamarins. Copyright 2003 Published by Elsevier Science Ltd on behalf of The Association for the Study of Animal Behaviour      

Food Deprivation Suppresses a Preference for the Top-Scent Mark of an Over-Mark in Meadow Voles (Microtus pennsylvanicus)

Food availability affects whether mammals communicate their interest in interacting with opposite‐sex conspecifics. This study examined the responses of voles to over‐marks, and factors that influence the formation and maintenance of a preference for the top‐scent in an over‐mark. Specifically, we investigated how food deprivation affected the amount of time male and female voles exposed to an over‐mark, later responded to the marks of the top‐ and bottom‐scent donors when subsequently presented with the two scents side by side. Males and females that were not food deprived and males that were food deprived 6 h before exposure to an over‐mark later maintained a preference for the donor of the top‐scent mark compared with the donor of the bottom‐scent mark of the over‐mark. Females that were food deprived for 6 h before or after exposure of the over‐mark and males food deprived 6 h after the exposure to the over‐mark showed no preference for the top‐scent mark donor. Re‐feeding females that were food deprived for 6 h before exposure to an over‐mark was sufficient to restore their preference for the mark of the top‐scent male over that of the bottom‐scent male. The different responses of food‐deprived male and female voles to over‐marks of opposite‐sex conspecifics may be associated with differences in their tactics for interacting with potential mates and the higher energetic costs of reproduction in female voles than in male voles.     

Scent Marking in Female Prairie Voles: a Test of Alternative Hypotheses

We conducted three experiments with females in different stages of reproductive condition to test alternative hypotheses for the function of scent marking in female prairie voles, Microtus ochrogaster . The three reproductive categories were isolated females prior to sexual stimulation (anoestrous), sexually stimulated (oestrous) and lactating. Females in different reproductive condition were given the opportunity to scent mark clean unmarked substrate or areas that had previously been marked by adult females or adult males. The numbers of scent marks deposited by females did not differ statistically for females in different reproductive condition. However, there was a trend for anoestrous females to mark the most, oestrous females less, and lactating females the least. The lack of scent marking by lactating females might be to reduce conspicuousness to conspecifics or predators. Oestrous females tended to mark the most in the area marked previously by males, although the difference was not statistically significant.
Our results provide some support for a mate-attraction hypothesis and a territorial-defense hypothesis, but were most consistent with a self-advertisement hypothesis. Over marking was uncommon and did not differ by experiment or sex of previous donor. Our results suggest that the number and placement of scent marks by females are highly variable and function primarily to convey individual identity.     

Temporal and Spatial Distribution of Male Scent Marks in the Polygynous Greater Sac-Winged Bat

Scent marks are relatively long-lived signals that can be perceived by conspecifics when the producer is absent. Therefore, it is often not obvious to whom the signal is directed. In daytime roosts of the polygynous greater sac-winged bat, males scent mark territories with facial gland secretions. Territories are a valuable resource for males, as they offer exclusive courtship opportunities, which results in increased male reproductive success and, consequently, increased male–male competition over territories. The information encoded in male scent marks could, therefore, be either directed at females as part of an olfactory courtship display or at male competitors as part of territorial behaviour. We expected territorial males to scent mark in the morning, shortly before females return to the territory and close to female roosting sites, if scent marks are directed at females as part of the courtship display. And we expected harem males to scent mark at the territory boundaries, where male–male encounters are most likely to occur, if scent marks are directed at male competitors. We found that males marked more frequently in the afternoon, at a time when all females have already left the territory, and harem males marked at the territory boundaries and not inside their territory in the area where females roost. At boundaries males fan volatiles from specialised wing sacs towards competitors outside the territory. Scent marking of male Saccopteryx bilineata might therefore be congruent with the assessment-hypothesis, which states that scent marks offer intruders the possibility to make an olfactory assessment of the territory owner without direct physical interaction. Thus, scent marks of male S. bilineata are most likely influenced by male–male competition and not by female choice.     

Female marmoset monkeys (Callithrix jacchus) can be identified from the chemical composition of their scent marks.

The present study analyzed 42 organic solvent extracts of scent mark pools from five dominant female common marmosets by gas chromatography (GC) and combined GC and mass spectrometry. We determined whether there were qualitative or quantitative differences between the chemical composition of scent marks from individual females. Gas chromatography and mass spectral analysis detected the same 162 chemicals in 86% (36/42) of scent mark pools from five dominant females. This near identical chemical composition of scent marks suggested there were few, if any, qualitative differences between the chemical composition of scent marks from individual females. Instead, quantitative differences in scent may provide the key factor distinguishing individual females. Using the relative concentration of highly volatile chemicals detected by GC in scent marks, linear discriminant analysis classified scent mark pools to their correct donor approximately 91% of the time. Such highly reliable statistical matching of scent to donor suggested that each individual female common marmoset has a unique ratio of highly volatile chemicals in their scent marks which may permit individual identification of females from odors in their scent alone.     

Olfactory Experience Affects the Response of Meadow Voles to the Opposite‐Sex Scent Donor of Mixed‐Sex Over‐Marks

Scent marking and over‐marking are important forms of communication between the sexes for many terrestrial mammals. Over the course of three experiments, we determined whether the amount of time individuals investigate the scent marks of opposite‐sex conspecifics is affected by 4 d of olfactory experience with those conspecifics. In Experiment 1, female meadow voles, Microtus pennsylvanicus, spent more time investigating the scent mark of the novel male conspecific than that of the familiar male donor, whereas male voles spent similar amounts of time investigating the scent mark of the familiar female and a novel female conspecific. In Experiment 2, voles were exposed to a mixed‐sex over‐mark in which subjects did not have 4 d of olfactory experience with either the top‐scent donor or the bottom‐scent donor. During the test phase, male and female voles spent more time investigating the scent mark of the opposite‐sex conspecific that provided the top‐scent mark than that of a novel, opposite‐sex conspecific. Male and female voles spent similar amounts of time investigating the scent mark of the bottom‐scent donor and that of a novel opposite‐sex conspecific. In Experiment 3, voles were exposed to a mixed‐sex over‐mark that contained the scent mark of an opposite‐sex conspecific with which they had 4 d of olfactory experience. During the test phase, male voles spent more time investigating the mark of the familiar, top‐scent female than the scent mark of a novel female donor but spent similar amounts of time investigating the mark of the familiar, bottom‐scent female and that of a novel female donor. In contrast, female voles spent more time investigating the mark of a novel male donor than that of either the familiar, top‐scent male or that of the familiar, bottom‐scent male. The sex differences in the responses of voles to scent marks and mixed‐sex over‐marks are discussed in relation to the natural history and non‐monogamous mating system of meadow voles.     

The Structure,Stability and Pheromone Binding of the Male Mouse Protein Sex Pheromone Darcin

Mouse urine contains highly polymorphic major urinary proteins that have multiple functions in scent communication through their abilities to bind, transport and release hydrophobic volatile pheromones. The mouse genome encodes for about 20 of these proteins and are classified, based on amino acid sequence similarity and tissue expression patterns, as either central or peripheral major urinary proteins. Darcin is a male specific peripheral major urinary protein and is distinctive in its role in inherent female attraction. A comparison of the structure and biophysical properties of darcin with MUP11, which belongs to the central class, highlights similarity in the overall structure between the two proteins. The thermodynamic stability, however, differs between the two proteins, with darcin being much more stable. Furthermore, the affinity of a small pheromone mimetic is higher for darcin, although darcin is more discriminatory, being unable to bind bulkier ligands. These attributes are due to the hydrophobic ligand binding cavity of darcin being smaller, caused by the presence of larger amino acid side chains. Thus, the physical and chemical characteristics of the binding cavity, together with its extreme stability, are consistent with darcin being able to exert its function after release into the environment.     

Relative Contributions of Urine and Anal-Sac Secretions in Scent Marks of Large Felids

SYNOPSIS. A hypothesis generated from field observations oflions and tigers was tested in a zoo setting. The presence ofa whitish material in urine marks, detected by visual inspection,led to the surmise that analsac secretions were expelled alongwith urine. This hypothesis was evaluated by labelling anal-sacsecretions of various felids with an inert dye. The animalswere monitored thereafter for signs of dye in urine-marks, onfeces or in their enclosures. The failure to find anal-sac secretionsin urine or on feces led to a search for an alternate explanationfor the whitish material. A likely answer was found in the confirmationof considerable lipid in the bladder urine of lions and tigers,which separates to form a visible, whitish layer. Relative levelsof urinary lipid reflect general condition of an animal as theycorrelate with kidney fat reserves (Hewer et al., 1948), sothe amount of lipid in a scent mark could serve as an indicatorof condition. Furthermore, because lipid retards the releaseof volatile compounds (Regnier and Goodwin, 1976), lipid ina scent mark will extend release time of a pheromonal message.     

Scent-marking behaviour by male prairie voles,Microtus ochrogaster,in response to the scent of opposite- and same-sex conspecifics

We conducted an experiment using the prairie vole (Microtus ochrogaster) to test predictions associated with the proposed functions of scent marking as a sexual attractant, in reproductive competition, and as a self-advertisement. We allowed an oestrous female, an anoestrous female, and an adult male to scent mark three portions of a clean substrate and then exposed a second male to this substrate for secondary marking. We did not support a sexual attraction hypothesis in that males did not place more scent marks in response to oestrous than anoestrous females. Similarly, we did not support a reproductive competition hypothesis in that males did not place more scent marks in response to marks of males than to those of females or bare substrate. Males did not overmark the scent of males or females and thus we did not support a scent-masking or scent-blending hypothesis. In that males deposited scent similarly in response to males, females, and on bare substrate, our results suggest that the frequency and placement of scent marks by males function primarily to advertise individual identity in an area.     

Scent marking by voles in response to predation risk: a field-laboratory validation

Predators use scent to locate their prey, and prey animals oftenalter their behavior in response to predation risk. I testedthe hypothesis that voles would decrease their frequency ofscent marking in response to predation risk. I conducted trialsin which prairie voles, Microtus ochrogaster, and woodland voles,M. pinetorum, were allowed to scent mark ceramic tiles placedin their runways in the field. The tiles were subjected to oneof three treatments: scented with odor from mink, Mustela vison(a rodent predator); rabbit, Oryctolagus cuniculus (a nonpredatormammal control); and no odor (control). No significant differenceswere found in the frequency of scent marking in response tothe three treatments for either species. To validate that volesdid not decrease their scent marking in response to predationrisk, I brought male prairie voles from the field site intothe laboratory and allowed them to scent mark white paper substratetreated with mink odor, rabbit odor, or no odor. No significantdifferences were found in the frequency of scent marks in responseto the three treatments. These results differ from what waspredicted based on laboratory studies with other species ofrodents that show avoidance, reproductive suppression, decreasedactivity, and reduced scent marking in response to odors ofpredators. Voles appear to scent mark different substrates andunder a wide variety of social and environmental situations,and this is not influenced by the presence of odor from a predator.     

Genetic clues from olfactory cues: brown hyaena scent marks provide a non-invasive source of DNA for genetic profiling

Brown hyaena scent marks were tested for their potential to provide DNA suitable for PCR. Up to 100% of scent marks were successfully amplified. The approach can potentially be adapted to other species with similar pasting behaviour and scent mark morphology, or to samples that are environmentally exposed/degraded/inhibitor-rich or where there are very limited amounts of sample.     

Scent-marking displays provide honest signals of health and infection

Males of many species produce scent marks and other olfactorysignals that function to intimidate rivals and attract females.It has been suggested that scent marks provide an honest, cheat-proofdisplay of an individual's health and condition. Here we reportseveral findings that address this hypothesis in wild-derivedhouse mice (Mus musculus domesticus). (1) We exposed males tofemale odor, which induces an increase in testosterone, andfound that sexual stimulation significantly increased the males'scent-marking and the attractiveness of their scent marks tofemales. (2) We challenged sexually stimulated males with anonreplicating strain of bacteria (Salmonella enterica C5TS)to activate immunity and found that this significantly decreasedthe males' scent-marking and the attractiveness of their scentmarks to females. (3) We collected scent marks from infectedand sham-infected males when they were sexually stimulated ornot, and we found that females could significantly discriminatethe scent marks of infected versus control males, but only whenthe males were sexually stimulated. Taken together, our resultsindicate that male mice modulate their scent-marking displaydepending on their health and perceived mating opportunities.Increased scent marking enhances males' attractiveness to females,scent marks provide an honest indicator of bacterial infection(and perhaps immune activation), and females are able to assessthe health of males more easily when males mark at a high rate.     

Meadow Voles (Microtus pennsylvanicus, Arvicolidae) Over-mark and Adjacent-mark the Scent Marks of Same-sex Conspecifics

Scent over-marking occurs when an animal deposits its scent mark on top of the scent mark of a conspecific; adjacent-marking occurs when an animal deposits its scent mark next to the scent mark of a conspecific. Given that male rodents usually scent mark more than females and that animals spend more time investigating the odor of the top-scent donor of an over-mark, I tested the following three hypotheses. First, male meadow voles deposit more scent marks than female meadow voles. Second, male meadow voles will deposit more over-marks and adjacent-marks in response to the scent marks of a same-sex conspecific than females would. Third, meadow voles spend more time investigating the odor of the second vole placed in the arena than that of the first vole placed in the arena. To test these hypotheses, two age-matched, like-sex conspecifics (first vole and second vole) were placed successively into an arena in which they were allowed to freely explore and scent mark for 15 min. The first hypothesis was not supported. The first and second vole, independently of sex, deposited a similar number of scent marks. The second hypothesis was also not supported by the data: more conspecific scent marks were over-marked by the second female than by the second male. The third hypothesis was supported by the data. After investigating a scented arena, males and females spent more time investigating the odor of the second vole than that of the first vole. Sex differences in scent-marking behaviors of meadow voles are unlike those reported for other species of rodents.