Linking quantity and acoustic properties of courtship vocalizations in male mice

The number of courtship ultrasonic vocalizations (USVs) in male mice is widely regarded as an indicator of sexual motivation. However, it remains unclear how variations in vocal quantity relate to the acoustic properties of these vocalizations. In this study, using ICR males subjected to castration, sham-operation, male co-housing, or female co-housing, we experimentally manipulated call rate and examined its relationship with multiple acoustic features. We found that call rate was positively correlated with syllable duration and negatively correlated with inter-syllable interval, and was also associated with bout-level temporal organization, indicating that males with higher vocal activity produce longer and more temporally dense vocal sequences. To explore strain effects, we re-analysed a previously published dataset of C57BL/6 J males and found positive correlations between call rate and both syllable duration and the degree of intra-syllable frequency modulation (cvfreq). Thus, while strain-specific differences exist, males with higher vocal activity in both strains tend to emit syllables with longer duration and/or greater frequency modulation. From a methodological perspective, our findings suggest that syllable duration offers a practical and biologically meaningful index for linking acoustic features with internal states. These results advance our understanding of how vocal quantity and quality interrelate in male courtship vocal communication.