Male Mouse Ultrasonic Vocalizations Research Articles

Investigating the acoustic directionality of mouse ultrasonic vocalizations

Across the animal kingdom, ultrasonic vocalizations (USVs) are used as a means of communication, navigation, and predation, and in some species (e.g., echolocating bats and whales), the directionality of the signal facilitates these processes. The utility and characteristics of mouse USVs, however, are less clear. Using an 8-channel microphone array, we recorded, localized, and assigned USVs to individual mice that were in isolation (n = 28) or socially interacting (male-female dyads, n = 13; 4-mouse, mixed-sex groups, n = 11). Males emitted a total of 3675 (100%), 22 268 (84.48%), and 73 488 (82.05%) of the assigned vocalizations when in isolation, interacting with a female, or socializing in a group. By examining the relationship between the USV sound intensity on the eight microphones and the head directions of the mice, we observed that male mice consistently produced directional USVs across each of the three behavioral contexts. Specifically, USV intensity was strongest in the direction that the mouse was facing and weakest in the opposite direction. Overall, these findings directly demonstrate that male mouse USVs are directional and may provide insight into the communicative function of mouse vocalizations.

Effect of Sociosexual Experience and Aging on Number of Courtship Ultrasonic Vocalizations in Male Mice.

Sexual behaviors are instinctually exhibited without prior training, but they are modulated by experience. One of the precopulatory behaviors in adult male mice, courtship ultrasonic vocalizations (USVs), has attracted considerable academic attention recently. Male mice emit ultrasounds as courtship behavior when encountering females. However, the modulatory effects of experience on USVs remain unclear. In the present study, we aimed to clarify the effects of sociosexual experience and aging on adult male vocalizations. First, we examined the effect of aging. The number of USVs decreased in an age-dependent manner. Following this, young adult male mice were co-housed for two weeks with normal female mice or ovariectomized (OVX) female mice, or housed without female mice, and the number of courtship USVs before and after co-housing were compared. In males housed with normal or OVX females, USVs increased significantly after co-housing. In contrast, males housed without females did not exhibit a significant increase of USVs. A facilitative effect of co-housing with female mice on vocalizations was also observed in aged males. In addition, females used as co-housing partners became pregnant, and the reproductive rate may be related to the vocal activity observed in the partnered males. These results indicate that sociosexual experience and aging affect vocalization activity, which may be related to courtship and/or reproductive function.

Ultrasonic Vocalizations of Male Mice Differ among Species and Females Show Assortative Preferences for Male Calls.

Male house mice (Mus musculus) emit ultrasonic vocalizations (USVs) during courtship, which attract females, and we aimed to test whether females use these vocalizations for species or subspecies recognition of potential mates. We recorded courtship USVs of males from different Mus species, Mus musculus subspecies, and populations (F1 offspring of wild-caught Mus musculus musculus, Mus musculus domesticus (and F1 hybrid crosses), and Mus spicilegus), and we conducted playback experiments to measure female preferences for male USVs. Male vocalizations contained at least seven distinct syllable types, whose frequency of occurrence varied among species, subspecies, and populations. Detailed analyses of multiple common syllable types indicated that Mus musculus and Mus spicilegus could be discriminated based on spectral and temporal characteristics of their vocalizations, and populations of Mus musculus were also distinctive regardless of the classification model used. Females were able to discriminate USVs from different species, and showed assortative preferences for conspecific males. We found no evidence that females discriminate USVs of males from a different subspecies or separate populations of the same species, even though our spectral analyses identified acoustic features that differ between species, subspecies, and populations of the same species. Our results provide the first comparison of USVs between Mus species or between Mus musculus subspecies, and the first evidence that male USVs potentially facilitate species recognition.

Effects of amygdala lesions on male mouse ultrasonic vocalizations and copulatory behaviour

Mice produce ultrasonic vocalizations (USVs) in several behavioural contexts. In particular, male mice articulate a long series of various sounds to females during courtship behaviour. To determine the relationships between this kind of vocal behaviour and emotion, we examined the lesion effects of the amygdala, an important neural module in emotional behaviour, on USVs. We recorded USVs from mice in the lesion and the control (sham operation) groups upon presentation of females and compared USVs before and after surgery. We found that the mean syllable duration of the USVs shortened and the appearance rate of longer syllables decreased after the surgery. The main reasons for these alterations could be explained by the altered courtship behaviour. As reported previously, the mounting behaviour of the lesion group after surgery was markedly less than that of the control group. Therefore, the appearance rate of those longer syllables would decrease logically because longer syllables primarily appear during mounting and intromission. However, we can hypothesize another scenario for the alterations to vocal behaviour: effects on the direct amygdala-periaqueductal grey (PAG) projection might be involved in the increase in the appearance rate of shorter syllables owing to lesion-induced loss of emotions, such as vigilance. Overall, the results suggested two possible mechanisms of the amygdala lesions on the alteration of the vocal behaviour.

The Structure and Usage of Female and Male Mouse Ultrasonic Vocalizations Reveal only Minor Differences

Ultrasonic vocalizations (USV) of mice are increasingly recognized as informative dependent variables in studies using mouse models of human diseases. While pup vocalizations primarily serve to re-establish contact with the mother, adult male “songs” were considered to be courtship signals. Alternatively, mouse USVs may generally function as territorial signals. To distinguish between these two hypotheses, we compared the structure and usage of adult male and female USVs in staged resident-intruder encounters. If calls function primarily as courtship signals, males should respond stronger than females, specifically when presented with a female intruder. Refuting this hypothesis, we found that in response to female intruders, females called more than males (228±32 calls/min vs. 71±15 calls/min), and males called more to female than to male intruders (14±7.5 calls/min). There were no significant differences in the acoustic characteristics of the calls given by females and males. To control for the influence of the intruder's behavior on calling, we repeated the experiments using anaesthetized intruders. Again, females produced more calls to female than male intruders (173±17 calls/min vs. 71±15 calls/min), while males called more in response to female than male intruders (39±17 calls/min), and there were no acoustic differences in female and male calls. The vocal activity did not differ significantly with regard to intruder state (awake or anaesthetized), while the acoustic structure exhibited significant differences. Taken together, our findings support the view that calls do not mainly function as courtship signals, although they might serve both a territorial (sex-independent) and a courtship function. The comparison of responses to awake vs. anaesthetized intruders suggests that the latter are sufficient to elicit vocal activity. The subtle acoustic differences, however, indicate that the subject differentiates between intruder states.

Ultrasonic Vocalizations Induced by Sex and Amphetamine in M2, M4, M5 Muscarinic and D2 Dopamine Receptor Knockout Mice

Adult mice communicate by emitting ultrasonic vocalizations (USVs) during the appetitive phases of sexual behavior. However, little is known about the genes important in controlling call production. Here, we study the induction and regulation of USVs in muscarinic and dopaminergic receptor knockout (KO) mice as well as wild-type controls during sexual behavior. Female mouse urine, but not female rat or human urine, induced USVs in male mice, whereas male urine did not induce USVs in females. Direct contact of males with females is required for eliciting high level of USVs in males. USVs (25 to120 kHz) were emitted only by males, suggesting positive state; however human-audible squeaks were produced only by females, implying negative state during male-female pairing. USVs were divided into flat and frequency-modulated calls. Male USVs often changed from continuous to broken frequency-modulated calls after initiation of mounting. In M2 KO mice, USVs were lost in about 70–80% of the mice, correlating with a loss of sexual interaction. In M5 KO mice, mean USVs were reduced by almost 80% even though sexual interaction was vigorous. In D2 KOs, the duration of USVs was extended by 20%. In M4 KOs, no significant differences were observed. Amphetamine dose-dependently induced USVs in wild-type males (most at 0.5 mg/kg i.p.), but did not elicit USVs in M5 KO or female mice. These studies suggest that M2 and M5 muscarinic receptors are needed for male USV production during male-female interactions, likely via their roles in dopamine activation. These findings are important for the understanding of the neural substrates for positive affect.

An ephemeral sex pheromone in the urine of female house mice ( Mus domesticus)

From previous research, the ultrasonic vocalizations of male mice (Mus domesticus) to female mouse urine were hypothesized to be learned as a result of classical conditioning during adult heterosexual encounters. According to this interpretation, a previously neutral conditioned stimulus in female urine comes to elicit vocalizations as a result of its association with some other unknown unconditioned stimulus associated with adult females. However, the research from which this hypothesis was derived utilized urine collected from females housed in metabolic cages. Three experiments further examined the classical conditioning hypothesis using two types of female urine: (i) metabolic-cage-collected urine and (ii) freshly voided urine. Experiment 1 demonstrated that, in contrast to vocalizations to metabolic-cage-collected urine, adult heterosexual experience was not necessary for males to vocalize to freshly voided female urine. In addition, unlike metabolic-cage-collected urine (Experiment 3), freshly voided urine remained a potent stimulus for eliciting vocalizations during repeated testing (Experiments 2 and 3). Finally, freshly voided urine appeared to cause a previously neutral stimulus (cotton swab) to acquire ultrasound eliciting properties (Experiment 2). We suggest from these findings that two chemosignals that elicit vocalizations from males may exist in female mouse urine: (i) a potent, but volatile or easily degraded, unconditioned stimulus to which males vocalize without sexual experience and (ii) a nonvolatile, chemically stable conditioned stimulus.

Courtship ultrasonic vocalizations in male Swiss-Webster mice: Effects of hormones and sexual experience

The effects of testosterone and its metabolites (estradiol and 5α-dihydrotestosterone) on male mouse ultrasonic vocalizations were investigated using Swiss-Webster mice. In the first experiment daily treatment with 50 μg or 200 μg of testosterone propionate (TP) restored ultrasonic calling in castrated sexually naive males. Treatment with 200 μg of dihydrotestosterone propionate (DHTP) also activated ultrasound production, but treatment with 50 μg of DHTP or estradiol benzoate (EB 0.5 μg and 2 μg/day) failed to have a significant effect on this measure. In experiment two, the animals used had shown male sexual behavior before castration. These sexually experienced males showed a marked reduction in ultrasound production after castration but significantly increased ultrasound production after daily treatment with either TP (50 μg or 200 μg), DHTP (50 μg or 200 μg) or 2 μg of EB. Hormones that facilitated ultrasound calling also activated other components of male sexual behavior in this and previous studies with Swiss-Webster mice. The results indicate that hormone dose and precastration sexual experience interact to determine the effects of postcastration treatment on male vocalizations.

Female behavior is affected by male ultrasonic vocalizations in house mice

The effects of male-mouse ultrasonic vocalizations on female preference behavior were investigated using Swiss-Webster mice. Intact females spent more time with an intact tethered male than with a devocalized tethered male during 3-min preference tests. Female preference behavior was similar during estrus and diestrus; but, a preference for the vocalizing male was absent after ovariectomy, and returned following replacement treatment with ovarian hormones. Similar effects on female behavior were obtained using synthetic 70 kHz ultrasounds presented behind one of two devocalized tethered males during preference tests. The results indicate that ultrasounds maintain the female close to the vocalizing male and may facilitate copulation.

Olfactory and vomeronasal mediation of ultrasonic vocalizations in male mice

The effects of deafferentation of the vomeronasal, olfactory or both systems of male mice on ultrasonic vocalizations in response to a male or female stimulus were analyzed. Regardless of the technique used, disruption of the vomeronasal system led to consistent reductions in ultrasonic vocalizations. Each animal receiving vomeronasal tract cuts, bulbectomies or a combined treatment of vomeronasal tract transection and intranasal flush with ZnSO 4 markedly suppressed responding to the stimulus animals. Animals treated with intranasal ZnSO 4 alone or those receiving control procedures continued to respond to the female stimulus animals at rates similar to pretreatment. These results indicate that the vomeronasal system is directly involved in the perception of female mouse chemosignals and, hence, mediates the male's responses to these signals.

Effects of medial preoptic lesions on male mouse ultrasonic vocalizations and copulatory behavior

In male mice, lesions in the medial preoptic area (MPA) reduced the proportion of animals showing mounting behavior but failed to abolish ano-genital exploration of the female. The lesions did not effect the amount of 70 kHz ultrasound elicited by intact females or by soiled bedding obtained from female-occupied cages. These results suggest components of male sexual behavior are mediated by neural systems outside the MPA.

Social status and ultrasonic vocalizations of male mice

Dominant male mice (Mus musculus) emit much more 70-kHz ultrasound in response to females than do subordinates. Neither dominant nor subordinate males emit much ultrasound in response to male stimuli. Inhibition of initial courtship by social subordination may be a central factor contributing to the greater reproductive fitness of dominant males.