Name: eliciting and analyzing male mouse ultrasonic vocalization (usv) songs
Uploaded: 2017-05-09T13:00:00
Duration: 8 min 44 s
Description: Watch this Scientific Journal Video about Eliciting and Analyzing Male Mouse Ultrasonic Vocalization USV Songs at JoVE.com

This work was supported by the Howard Hughes Medical Institute funds to EDJ. We thank Pr. Sylvie Granon (NeuroPSI - University Paris south XI - FRANCE) for lending us the speaker hardware. We also thank members of the Jarvis Lab for their support, discussions, corrections and comments on this work, especially Joshua Jones Macopson for help with figures and testing. We thank Dr. Gustavo Arriaga for help with the Mouse Song Analyzer software, upgrading it for us to V1.3, and other aspects of this protocol. v1.0 of the software was developed by Holy and Guo, and v1.1 and v1.3 by Arriaga.

<ol>
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The authors have nothing to disclose.

This protocol provides approaches to collect, quantify, and study male mice courtship vocalizations in the laboratory across a variety of mostly female-related stimuli. As presented previously in Chabout, et al. 23and in the representative results, the use of this method allowed us to discover context-dependent vocalizations and syntax that matter for the receiving females. The standardization of these stimuli will provide the collection of a reliable number of USVs and allow detailed analyses of the male&#39;s courtship songs and repertoires.
When a live female is present with the male, the protocol does not allow us to clearly identify the emitter of the vocalizations. However, previous studies showed that the majority of the vocalizations emitted in such context were by the male26,29. Most of the studies using a conspecific (male or female) as a stimulus for the males believe that the amount of female&#39;s vocalizations in these contexts is negligible4,5,22,30. However a recent paper used triangulation to localize the vocalization&#39;s of the emitter in group housed conditions31, and showed that within a dyad, the female contributes to ~10% of the USVs. In the present protocol the use of the anesthetized female allows the user to study the male vocalizations in the presence of a female without her vocalizing. In contrast to expectations of this recent study31, we found no difference in the number of syllables emitted between the FE and AF conditions23. It is possible that live females did not significantly contribute to the recordings or that the males vocalized less in the presence of live females versus anaesthetized females. Nevertheless, we believe that future experiments should consider the use of this triangulation method to assess the potential effect of the female contribution.
There are other software available that can do some the steps we have outlined, although we do not believe in a manner sufficient for the questions we asked using a combination of three programs: Software A, Mouse Song Analyzer software script C using software B, the syntax analysis software using a custom spreadsheet software D+E calculations, and syntax decorder using R. For example, a recent paper proposed a software named VoICE that allows the user to extract acoustic variables automatically from the sonograms or directly on units that had been manually selected by the user32. But, the automated or semi-automated sequence analyses are not as detailed as our approach. Some commercial software can automatically analyze the acoustic features, but do not provide an automatic classification of the syllables; the user has to sort the different syllables afterwards. Grimsley, Gadziola, et al. 33 developed a Table-based virtual mouse vocal organ program that clusters syllables based on shared acoustic features, but does not provide automatic detection of the syllables. Their program34 is unique in that it creates novel sequences from recorded songs using Markov models, and thus has more advanced features than simple editing.
Most prior communication studies on mice have focused on the emitter&#39;s side35,36. Few studies have explored the receiver&#39;s side30,37,38. Playback and discrimination protocols provide a simple test to study the receiver&#39;s side, such as the one also recently described by Asaba, Kato, et al. 39. In that study, the authors used a two-choice test box separated with acoustic foam instead of the Y-maze box described here. Both choice setups have advantages and disadvantages. First, the Y-maze doesn&#39;t isolate the sound from one arm to the other, but the two-choice box does. However, by using the Y-maze design, the animal can quickly evaluate the two songs that are played simultaneously, and move towards the preferred one. Nevertheless, playback experiments in general help experimenters determine the meaning and thus functions of the vocalizations generated for conspecific animals. In conclusion, after mastering the techniques of this protocol and analyses, readers should be able to address many questions that influence the context, genetics, and neurobiology of mouse USVs.
Using B6D2F1/J mice, the female associated stimuli almost always trigger USVs from the males we have tested in our lab. It is critical to collect enough syllables (&#62; 100 in 5 min) to be able to obtain strong statistically analysis. For troubleshooting, if no USVs are recorded (or not enough), check the configuration to make sure sounds are recorded. Do a live inspection of what is happening in the cage during recording by looking at the real-time sonogram on the computer screen after the introduction of the stimulus. Otherwise try to re-expose the male to a sexually mature/receptive female overnight and then house them alone for several days or up to a week before recording again. Based on anecdotal observations, we find that some males sing a lot on one day (for almost the full 5 min), and not much the next day, and then again another day. We do not know the reason why such within subject variability occurs, but we surmise it is probably a motivational or seasonal for the males, and the estrus state for female urine. If no USVs are recorded, try to record the animal on multiple days to pick up these variable effects. Unlike in songbirds, we have not noted overt differences in amount of singing based on time of day. We find that the males do not sing much (&#60; 100 syllables in 5 min) before they are 7 weeks old.
The detection methods presented here can extract thousands of syllables and all the acoustic parameters in a few minutes. But as any automatic detection method, it is very sensitive to background noise. Using Mouse Song Analyzer detection software with noisy recordings (for example from animals recorded with bedding) may require adjustment of the detection &#34;threshold&#34; to allow more flexibility. However, this will also increase the number of false positive syllables and the automatic detection might fail. Under such circumstances, manual coding can be used.
As stated earlier, the number, repertories, and latency of vocalizations are widely variable depending on the strain, thus one may have to change parameters (recording length, stimulus, automated syllable detection, etc.) for some strains to ensure optimal recordings for statistical analyses.

Relative to humans, mice produce both low and high frequency vocalizations, the later known as ultrasonic vocalizations (USVs) above our hearing range. The USVs are produced in a variety of contexts, including from mother-pup retrieval, juvenile interactions, to opposite or same sex adult interactions4,12. These USVs are composed of a diverse multisyllabic repertoire which can be categorized manually9 or automatically10,11. The role of these USVs in communication has been under increasing investigation in recent years. These include using the USVs as readouts of mouse models of neuropsychiatric, developmental or behavioral disorders7,8, and internal motivational/emotional states13. The USVs are thought to convey reliable information on the emitter&#39;s state that is useful for the receiver14,15.
In 2005, Holy and Guo 11, advanced the idea that adult male mice USVs were organized as a succession of multisyllabic call elements or syllables similar to songbirds. In many species, a multisyllabic repertoire allows the emitter to combine and order syllables in different ways to increase the potential information carried by the song. Variation in this syntax is believed to have an ethological relevance for sexual behavior and mate preferences16,17. Subsequent studies showed that male mice were able to change the relative composition of syllable types they produce before, during, and after the presence of a female5,18. That is, the adult male mice use their USVs for courtship behavior, either to attract or maintain close contact with a female, or to facilitate mating19,20,21. They are also emitted in male-male interactions, probably to convey social information during interactions4. To capture these changes in repertoires, scientists usually measure the spectral features (acoustic parameters, such as amplitude, frequencies, etc.), number of USVs syllables or calls, and latency to the first USV. However, few actually look at the sequence dynamics of these USVs in detail22. Recently our group developed a novel method to measure dynamic changes in the USV syllable sequences23. We showed that syllable order within a song about (namely syntax) is not random, that it changes depending on social context, and that the listening animals detect these changes as ethologically relevant.
We note that many investigators studying animal communication do not attach to the term &#39;syntax&#39; the same exact meaning as syntax used in human speech. For animal communication studies, we simply mean an ordered, non-random, sequence of sounds with some rules. For humans, in addition, specific sequences are known to have specific meanings. We do not know if this is the case for mice.
In this paper and associated video, we aim to provide reliable protocols to record male mice&#39;s courtship USVs across various contexts, and perform playbacks. The use of three sequentially used software for: 1) automated recordings; 2) syllable detection and coding; and 3) in-depth analysis of the syllable features and syntax is demonstrated (Figure 1). This allows us to learn more about male mice USV structure and function. We believe that such methods ease data analyses and may open new horizons in characterizing normal and abnormal vocal communication in mouse models of communication and neuropsychiatric disorders, respectively.

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			<td height="21" style="height:21px;width:297px;">Sound proof beach cooler</td>
			<td style="width:429px;">See Gus paper has more info on specific kind</td>
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			<td style="width:307px;">Inside dimensions (L 27 x W 23 x H 47 cm):</td>
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			<td height="42" style="height:42px;width:297px;">Condenser ultrasound microphone CM16/CMPA</td>
			<td style="width:429px;">Avisoft Bioacoustics, Berlin, Germany</td>
			<td style="width:129px;">#40011</td>
			<td>Includes extension cable&#160;</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:297px;">Ultrasound Gate 1216H sound card</td>
			<td style="width:429px;">Avisoft Bioacoustics, Berlin, Germany</td>
			<td style="width:129px;">#34175</td>
			<td>12 channel sound card</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:297px;">Ultrasound Gate Player 216H</td>
			<td style="width:429px;">Avisoft Bioacoustics, Berlin, Germany</td>
			<td style="width:129px;">#70117</td>
			<td>2 channels playback player</td>
		</tr>
		<tr height="22">
			<td height="22" style="height:23px;width:297px;">Ultrasonic Electrostatic Speaker ESS polaroid</td>
			<td style="width:429px;">Avisoft Bioacoustics, Berlin, Germany</td>
			<td style="width:129px;">#60103</td>
			<td>2 playback speakers</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:297px;">Test cage&#160;</td>
			<td style="width:429px;">Ace</td>
			<td style="width:129px;">#PC75J</td>
			<td>30 x 8 x 13 cm height; plexiglas</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:297px;">plexiglas separation</td>
			<td style="width:429px;">home made</td>
			<td style="width:129px;">-</td>
			<td>4 x 13 cm plexiglas with 1cm holes</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:297px;">Video camera</td>
			<td style="width:429px;">Logitech</td>
			<td style="width:129px;">C920</td>
			<td>logitech HD Pro webcam C920</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:297px;">Heat pad&#160;</td>
			<td style="width:429px;">Sunbeam</td>
			<td style="width:129px;">722-810-000</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:297px;">Y-maze&#160;</td>
			<td style="width:429px;">Home made</td>
			<td style="width:129px;">-</td>
			<td>Inside dimensions (L 30 x W 11 x H 29 cm):</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:297px;">Tweezers</td>
			<td style="width:429px;"></td>
			<td style="width:129px;"></td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:297px;"></td>
			<td style="width:429px;"></td>
			<td style="width:129px;"></td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:297px;">Software</td>
			<td style="width:429px;"></td>
			<td style="width:129px;"></td>
			<td></td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:297px;">&#160;Avisoft Recorder (Software A)</td>
			<td style="width:429px;">Avisoft Bioacoustics, Berlin, Germany</td>
			<td style="width:129px;">#10101, #10111, #10102, #10112</td>
			<td>http://www.avisoft.com</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:297px;">&#160;MATLAB R2013a (Software B)</td>
			<td style="width:429px;">MathWorks</td>
			<td style="width:129px;">-</td>
			<td>MATLAB R2013a (8.1.0.604)</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:297px;">&#160;Mouse Song Analyzer v1.3 (Software C)</td>
			<td style="width:429px;">Custom designed by Holy, Guo, Arriaga, &#38; Jarvis; Runs with software B</td>
			<td style="width:129px;"></td>
			<td>http://jarvislab.net/wp-content/uploads/2014/12/Mouse_Song_Analyzer_v1.3-2015-03-23.zip</td>
		</tr>
		<tr height="22">
			<td height="22" style="height:22px;width:297px;">&#160;Microsoft Office Excel 2013 (Software D)</td>
			<td style="width:429px;">Microsoft</td>
			<td style="width:129px;">-</td>
			<td>Microsoft Office Excel</td>
		</tr>
		<tr height="36">
			<td height="36" style="height:36px;width:297px;">Song Analysis Guide v1.1 (Software E)</td>
			<td style="width:429px;">Custom designed by Chabout &#38; Jarvis. Excel calculator sheets, runs with software D</td>
			<td style="width:129px;"></td>
			<td>http://jarvislab.net/wp-content/uploads/2014/12/Song-analysis_Guided.xlsx</td>
		</tr>
		<tr height="22">
			<td height="22" style="height:22px;width:297px;">Syntax decorder v1.1 (Software F)</td>
			<td style="width:429px;">Custom designed by Sakar, Chabout, Dunson, Jarvis - in R studio</td>
			<td style="width:129px;"></td>
			<td>https://www.rstudio.com/products/rstudio/download/</td>
		</tr>
		<tr height="22">
			<td height="22" style="height:22px;width:297px;">&#160;Graphiz (Software G)</td>
			<td style="width:429px;">AT&#38;T Research and others&#160;</td>
			<td></td>
			<td>http:// www.graphviz.org;&#160;</td>
		</tr>
		<tr height="22">
			<td height="22" style="height:22px;width:297px;">&#160;Avisoft SASLab (Software H)</td>
			<td style="width:429px;">Avisoft Bioacoustics, Berlin, Germany</td>
			<td style="width:129px;">#10101, #10111, #10102, #10112</td>
			<td>http://www.avisoft.com</td>
		</tr>
		<tr height="22">
			<td height="22" style="height:22px;width:297px;"></td>
			<td style="width:429px;"></td>
			<td style="width:129px;"></td>
			<td></td>
		</tr>
		<tr height="22">
			<td height="22" style="height:22px;width:297px;"></td>
			<td style="width:429px;"></td>
			<td style="width:129px;"></td>
			<td></td>
		</tr>
		<tr height="22">
			<td height="22" style="height:22px;width:297px;">Reagents</td>
			<td style="width:429px;"></td>
			<td style="width:129px;"></td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:297px;">Xylazine (20mg/ml)</td>
			<td style="width:429px;">Anased</td>
			<td style="width:129px;">-</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:297px;">Ketamine HCL (100mg/ml)</td>
			<td style="width:429px;">Henry Schein</td>
			<td style="width:129px;">#045822</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:297px;">distilled water</td>
			<td style="width:429px;"></td>
			<td style="width:129px;"></td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:297px;">Eye ointment&#160;</td>
			<td>Puralube Vet Ointment&#160;</td>
			<td>NDC 17033-211-38</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:297px;">Cotton tips</td>
			<td style="width:429px;"></td>
			<td style="width:129px;"></td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:297px;">Petri dish</td>
			<td style="width:429px;"></td>
			<td style="width:129px;"></td>
			<td></td>
		</tr>
	</tbody>
</table>

eliciting and analyzing male mouse ultrasonic vocalization (usv) songs

Mice produce ultrasonic vocalizations (USVs) in a variety of social contexts throughout development and adulthood. These USVs are used for mother-pup retrieval1, juvenile interactions2, opposite and same sex interactions3,4,5, and territorial interactions6. For decades, the USVs have been used by investigators as proxies to study neuropsychiatric and developmental or behavioral disorders7,8,9, and more recently to understand mechanisms and evolution of vocal communication among vertebrates10. Within the sexual interactions, adult male mice produce USV songs, which have some features similar to courtship songs of songbirds11. The use of such multisyllabic repertoires can increase potential flexibility and information they carry, as they can be varied in how elements are organized and recombined, namely syntax. In this protocol a reliable method to elicit USV songs from male mice in various social contexts, such as exposure to fresh female urine, anesthetized animals, and estrus females is described. This includes conditions to induce a large amount of syllables from the mice. We reduce recording of ambient noises with inexpensive sound chambers, and present a quantification method to automatically detect, classify and analyze the USVs. The latter includes evaluation of call-rate, vocal repertoire, acoustic parameters, and syntax. Various approaches and insight on using playbacks to study an animal&#39;s preference for specific song types are described. These methods were used to describe acoustic and syntax changes across different contexts in male mice, and song preferences in female mice.

In present protocol, changes in vocal behavior and syntax of male B6D2F1/J mice were characterized. Generally, using this protocol we were able to record, on average per male per 5 min session, 675 &#177; 98.5 classified syllables in response to female UR, 615.6 &#177; 72 in FE, 450 &#177; 134 in AF, 75.6 &#177; 38.9 in AM and 0.2 &#177; 0.1 in male UR (n = 12 males). The rates were ~ 130 syllables/min for female UR, ~ 120 syllables/min for FE, or ~ 100 syllables/min for AF contexts (Figure 5A). Males produce a much larger amount of syllables in response to freshly collected urine relative to overnight collected urine10,23. Males also sing considerably less in presence of an anesthetized male or fresh male urine Figure 5A). Males also change their repertoire across context23. For example, B6D2F1/J males significantly increase production of multiple pitch jump &#34;m&#34; category syllables in the female urine condition (Figure 5B). They also change acoustic features of individual syllables across context. For example, B6D2F1/J males sing syllables at a higher amplitude and bandwidth in the female urine context, and higher spectral purity in the awake female context compared to the others (Figure 6)23.
This protocol also provides a means for measuring dynamic features of sequence and thus syntax changes. Using an adapted method from Ey, et al. 22, we use the ISI to define the gaps between sequences (Figure 7A)23, and then use the gaps to distinguish and analyze temporal patterns of syllable sequences. We showed longer sequence lengths are produced in the awake female context (Figure 7B)23. This technique allows us to calculate the ratio of complex sequences (composed by at least 2 occurrences of the &#34;m&#34; syllable type) versus simple sequences (composed of one or no &#34;m&#34; type, and thus mostly by &#34;s&#34; type). We found that with B6D2F1/J males the female urine context triggered a higher ratio compared to the others (Figure 7C)23, indicating they produced more complex syllables in the female urine condition, but also that such syllables are distributed over more sequences.
We are also able to calculate the conditional transition probabilities from one syllable type to another (24 transition types total including the transitions from and to the &#34;silence&#34; state)23. We found that in different contexts, the mice&#39;s choice of transition types for given starting syllables differs, and that there is more syntax diversity in the female urine condition (Figure 8)23. These observations are consistent with previous reports that show that males can change the acoustic features or repertoire composition of their vocalizations in response to different stimuli and experiences4,5,24.
Finally, the present protocol provides guidance to test female preferences with playbacks. We found that B6D2F1/J females prefer more complex songs (containing 2 or more &#34;m&#34; syllables) relative to simple songs23. Most females chose to stay more often on the side of the Y-maze that played the complex songs (Figure 4B).
<img alt="Figure 1" src="/files/ftp_upload/54137/54137fig1.jpg" /> 
Figure 1: Flow Chart of Software Use and Analyses. Each program and associated code is given a letter name to help explain their identity and use in the main text. In ( ) are the specific programs we use in our protocol. <a href="http://cloudflare.jove.com/files/ftp_upload/54137/54137fig1large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>
<img alt="Figure 2" src="/files/ftp_upload/54137/54137fig2.jpg" /> 
Figure 2: Set up for Recording Male Mice Songs. (A) Picture of a sound attenuation recording box and set up to record USV vocalizations. (B) Example sonogram of a recording made with Software A (Table 1), including detailed spectral features calculated by &#34;Mouse Song Analyzer v1.3&#34;: duration, inter-syllable interval (ISI), peak frequency min (Pf min), peak frequency max (Pf max), peak frequency start (Pf start), peak frequency end (Pf end), and bandwidth. (C) Sonograms of another male singing to a live female, inside the sound attenuation box and outside of the box on the lab bench in the same room. Our anecdotal observations indicate that the recordings in the box of the same animal show larger volume (stronger intensity) and less harmonics, but no evidence of echoes in the box without sound foam. <a href="http://cloudflare.jove.com/files/ftp_upload/54137/54137fig2large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>
<img alt="Figure 3" src="/files/ftp_upload/54137/54137fig3.jpg" /> 
Figure 3: Screenshot of the &#34;Mouse Song Analyzer v1.3&#34; whis_gui Window Showing the Different Options Available for Analyses. The parameters shown are ones used to record male USVs in the figures and data analyses presented (except min note duration was 3 ms). <a href="http://cloudflare.jove.com/files/ftp_upload/54137/54137fig3large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>
<img alt="Figure 4" src="/files/ftp_upload/54137/54137fig4.jpg" /> 
Figure 4: Female Choices Between Playbacks of Complex and Simple Songs. (A) Picture of the Y-maze apparatus used and dimension measurements. (B) Time spent by the females in each arm, playing either from a complex (female urine elicited) or more simple (awake female elicited) song from the same male. Data are presented for n = 10 B6D2F1J female mice as mean &#177; SE, with individual values also shown; 9 of the 10 females showed a preference for the more complex syllable/sequence song. * p &#60; 0.05 paired student t-test. Figure modified from Chabout, et al. 23 with permission. <a href="http://cloudflare.jove.com/files/ftp_upload/54137/54137fig4large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>
<img alt="Figure 5" src="/files/ftp_upload/54137/54137fig5.jpg" /> 
Figure 5: Number of Syllables Emitted and Repertoire Across Conditions. (A) Syllable production rate of males in different contexts. (B) Repertoire compositions of males when in the presence of female urine (UR), anesthetized female (AF), awake female (FE), and anesthetized male (AM) contexts. Data are presented as mean &#177; SEM. * p&#60;0.03; ** p&#60;0.005; *** p&#60;0.0001 for post-hoc paired student t-test after Benjamini and Hochberg correction (n = 12 males). Figure from Chabout, et al. 23 with permission. <a href="http://cloudflare.jove.com/files/ftp_upload/54137/54137fig5large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>
<img alt="Figure 6" src="/files/ftp_upload/54137/54137fig6.jpg" /> 
Figure 6: Examples of Spectral Features in Different Context. (A) Amplitude. * p&#60; 0.025 for post-hoc paired student t-test after correction. (B) Frequency range or Bandwidth. *: p &#60; 0.041; **: p &#60; 0.005; ***: p &#60; 0.0001 after correction. (C) Spectral purity of the syllables. * p: &#60; 0.025; **: p &#60; 0.005; ***: p &#60; 0.0001 after correction. Abbreviations: female urine (UR), anesthetized female (AF), awake female (FE), and anesthetized male (AM). Figures modified from Chabout, et al. 23 with permission. <a href="http://cloudflare.jove.com/files/ftp_upload/54137/54137fig6large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>
<img alt="Figure 7" src="/files/ftp_upload/54137/54137fig7.jpg" /> 
Figure 7: Sequence Measurements. (A) Usage of the ISI to separate the sequence. Short ISI (SI) and medium ISI (MI) are used to separate syllables within a sequence, and long ISI over 250 ms (LI) separate two sequences. (B) Length of the sequences, measured as number of syllables per sequence, produced by males in different contexts. *: p &#60; 0.025; ** p &#60; 0.005; *** p &#60; 0.0001 after correction. (C) Ratio of complex songs over simple songs produced by males in different contexts. * p &#60; 0.041; ** p &#60; 0.005; *** p &#60; 0.0001 after correction. Data are presented as mean &#177; SEM (n = 12 males). Figure from Chabout, et al. 23 with permission. <a href="http://cloudflare.jove.com/files/ftp_upload/54137/54137fig7large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>
<img alt="Figure 8" src="/files/ftp_upload/54137/54137fig8.jpg" /> 
Figure 8: Syllable Syntax Diagrams of Sequences Based on Conditional Probabilities for Each Contexts. Arrow thickness is proportional to the conditional probability occurrence of a transition type in each context averaged from n = 12 males: P (occurrence of a transition given the starting syllable). For clarity, rare transitions below a probability of 0.05 are not shown. Figure from Chabout, et al. 23 with permission. <a href="http://cloudflare.jove.com/files/ftp_upload/54137/54137fig8large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>

Watch this Scientific Journal Video about Eliciting and Analyzing Male Mouse Ultrasonic Vocalization USV Songs at JoVE.com

Eliciting and Analyzing Male Mouse Ultrasonic Vocalization USV Songs

Mice produce a complex multisyllabic repertoire of ultrasonic vocalizations (USVs). These USVs are widely used as readouts for neuropsychiatric disorders. This protocol describes some of the practices we learned and developed to consistently induce, collect, and analyze the acoustic features and syntax of mouse songs.

Eliciting and Analyzing Male Mouse Ultrasonic Vocalization (USV) Songs

Mice produce ultrasonic vocalizations (USVs) in a variety of social contexts throughout development and adulthood. These USVs are used for mother-pup ...

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