Female frog auditory sensitivity,male calls,and background noise: potential influences on the evolution of a peculiar matched filter

The matched filter hypothesis proposes that the tuning of females' auditory sensitivity matches the spectral energy distribution of males' signals. Such correspondence is expected to arise over evolutionary time, as it promotes conspecific information transfer and reduces interference from other sound sources. Our main objective was to determine the correspondence between the acoustic sensitivity of female frogs of Eupsophus roseus and the spectral characteristics of advertisement vocalizations produced by conspecific males. We also aimed to determine how auditory sensitivity is related to the characteristics of background noise. We analysed data on the auditory sensitivity of E. roseus females, and recordings of conspecific male vocalizations and of the acoustic environment during the breeding period of this species. Our results indicate a concordance between the auditory sensitivity of females and call spectra that would provide an appropriate detection of these signals. In addition, this matching is large relative to the correspondence between auditory sensitivity with the spectra of the abiotic and biotic background noise, with the last component being associated with calls of the related species Eupsophus vertebralis. This may be an adaptation of receivers confronting sound interference, which improves the capability of E. roseus to communicate sexually by means of acoustic signals. © 2013 The Linnean Society of London, Biological Journal of the Linnean Society, 2013, 110 , 814–827.     

An exception to the matched filter hypothesis: A mismatch of male call frequency and female best hearing frequency in a torrent frog

The matched filter hypothesis proposes that the tuning of auditory sensitivity and the spectral character of calls will match in order to maximize auditory processing efficiency during courtship. In this study, we analyzed the acoustic structure of male calls and both male and female hearing sensitivities in the little torrent frog (Amolops torrentis), an anuran species who transmits acoustic signals across streams. The results were in striking contradiction to the matched filter hypothesis. Auditory brainstem response results showed that the best hearing range was 1.6–2 kHz consistent with the best sensitive frequency of most terrestrial lentic taxa, yet completely mismatched with the dominant frequency of conspecific calls (4.3 kHz). Moreover, phonotaxis tests show that females strongly prefer high‐frequency (4.3 kHz) over low‐frequency calls (1.6 kHz) regardless of ambient noise levels, although peripheral auditory sensitivity is highest in the 1.6–2 kHz range. These results are consistent with the idea that A. torrentis evolved from nonstreamside species and that high‐frequency calls evolved under the pressure of stream noise. Our results also suggest that female preferences based on central auditory system characteristics may evolve independently of peripheral auditory system sensitivity in order to maximize communication effectiveness in noisy environments.     

The spectral structure of vocalizations match hearing sensitivity but imprecisely in Philautus odontotarsus

It is generally thought that for species using vocal communication the spectral properties of the sender’s calls should match the frequency sensitivity of the receiver’s auditory system. Nevertheless, few studies have investigated both sender and receiver characteristics in anuran species. In the present study, auditory brainstem responses (ABRs) were recorded in the serrate legged treefrog, Philautus odontotarsus, in order to determine if male call spectral structure and hearing sensitivity in males and females have co-evolved in this species. The results showed that the spectral structures of male vocalization match both male and female hearing sensitivity, even though the dominant frequencies of male calls (2.5 kHz) are mismatched with the regions of best frequency sensitivity (1.4 and 2.8 kHz). In addition, the results show that, in contrast with most previous ABR studies in non-human animals, but consistent with human studies, there are noticeable sex differences in peripheral auditory sensitivity in Philautus insofar as females exhibit lower auditory thresholds than males across the entire 1.8–18 kHz frequency range. The results also show that the dominant frequency of male calls is negatively correlated with body size, indicating that call characteristics reflect body size in this species which may be used by females during mate choice.     

Traffic noise differentially impacts call types in a Japanese treefrog (Buergeria japonica)

Acoustic noise from automobile traffic impedes communication between signaling animals. To overcome the acoustic interference imposed by anthropogenic noise, species across taxa adjust their signaling behavior to increase signal saliency. As most of the spectral energy of anthropogenic noise is concentrated at low acoustic frequencies, species with lower frequency signals are expected to be more affected. Thus, species with low-frequency signals are under stronger pressure to adjust their signaling behaviors to avoid auditory masking than species with higher frequency signals. Similarly, for a species with multiple types of signals that differ in spectral characteristics, different signal types are expected to be differentially masked. We investigate how the different call types of a Japanese stream breeding treefrog (Buergeria japonica) are affected by automobile traffic noise. Male B. japonica produce two call types that differ in their spectral elements, a Type I call with lower dominant frequency and a Type II call with higher dominant frequency. In response to acoustic playbacks of traffic noise, B. japonica reduced the duration of their Type I calls, but not Type II calls. In addition, B. japonica increased the call effort of their Type I calls and decreased the call effort of their Type II calls. This result contrasts with prior studies in other taxa, which suggest that signalers may switch to higher frequency signal types in response to traffic noise. Furthermore, the increase in Type I call effort was only a short-term response to noise, while reduced Type II call effort persisted after the playbacks had ended. Overall, such differential effects on signal types suggest that some social functions will be disrupted more than others. By considering the effects of anthropogenic noise across multiple signal types, these results provide a more in-depth understanding of the behavioral impacts of anthropogenic noise within a species.     

The Acoustic Properties of Low Intensity Vocalizations Match Hearing Sensitivity in the Webbed-Toed Gecko,Gekko subpalmatus

The design of acoustic signals and hearing sensitivity in socially communicating species would normally be expected to closely match in order to minimize signal degradation and attenuation during signal propagation. Nevertheless, other factors such as sensory biases as well as morphological and physiological constraints may affect strict correspondence between signal features and hearing sensitivity. Thus study of the relationships between sender and receiver characteristics in species utilizing acoustic communication can provide information about how acoustic communication systems evolve. The genus Gekko includes species emitting high-amplitude vocalizations for long-range communication (loud callers) as well as species producing only low-amplitude vocalizations when in close contact with conspecifics (quiet callers) which have rarely been investigated. In order to investigate relationships between auditory physiology and the frequency characteristics of acoustic signals in a quiet caller, Gekko subpalmatus we measured the subjects’ vocal signal characteristics as well as auditory brainstem responses (ABRs) to assess auditory sensitivity. The results show that G. subpalmatus males emit low amplitude calls when encountering females, ranging in dominant frequency from 2.47 to 4.17 kHz with an average at 3.35 kHz. The auditory range with highest sensitivity closely matches the dominant frequency of the vocalizations. This correspondence is consistent with the notion that quiet and loud calling species are under similar selection pressures for matching auditory sensitivity with spectral characteristics of vocalizations.     

HEARING IN THE RED-BILLED FIREFINCH LAGON O S TIC TA SENEGALA AND THE SPANISH TIMBRADO CANARY SERINUS CANARIA: THE INFLUENCE OF NATURAL AND ARTIFICIAL SELECTION ON AUDITORY ABILITIES AND VOCAL STRUCTURE

ABSTRACT

We tested the auditory sensitivity of red-billed firefinches Lagonosticta senegal0061 and Spanish timbrado canaries Serinus canaria. Both these species produce songs and calls that are narrowband and relatively high in frequency, with spectral energy falling predominantly in the region of 3–6 kHz. Hearing thresholds were measured in these two species and compared to the auditory sensitivity of closely related species: the well studied zebra finch Taeniopygia guttata, and other strains of canary bred for song. Auditory thresholds were similar in both groups of birds, with firefinches having an audiogram typical for that of small birds. Timbrado canaries exhibited an audiogram with its greatest sensitivity in the relatively high region of 4–6 kHz, corresponding to the peak frequency of its calls. Critical ratios measured over a range of several octaves increased in a monotonie fashion at a rate of 2–3 dB per octave for both firefinches and timbrado canaries. Critical ratios in these two species are similar to what has been found in most other small passerine species, suggesting spectral resolving abilities similar to most small birds tested to date.     

Evolutionary diversification of the auditory organ sensilla in Neoconocephalus katydids (Orthoptera: Tettigoniidae) correlates with acoustic signal diversification over phylogenetic relatedness and life history

Neoconocephalus Tettigoniidae are a model for the evolution of acoustic signals as male calls have diversified in temporal structure during the radiation of the genus. The call divergence and phylogeny in Neoconocephalus are established, but in tettigoniids in general, accompanying evolutionary changes in hearing organs are not studied. We investigated anatomical changes of the tympanal hearing organs during the evolutionary radiation and divergence of intraspecific acoustic signals. We compared the neuroanatomy of auditory sensilla (crista acustica) from nine Neoconocephalus species for the number of auditory sensilla and the crista acustica length. These parameters were correlated with differences in temporal call features, body size, life histories and different phylogenetic positions. By this, adaptive responses to shifting frequencies of male calls and changes in their temporal patterns can be evaluated against phylogenetic constraints and allometry. All species showed well‐developed auditory sensilla, on average 32–35 between species. Crista acustica length and sensillum numbers correlated with body size, but not with phylogenetic position or life history. Statistically significant correlations existed also with specific call patterns: a higher number of auditory sensilla occurred in species with continuous calls or slow pulse rates, and a longer crista acustica occurred in species with double pulses or slow pulse rates. The auditory sensilla show significant differences between species despite their recent radiation, and morphological and ecological similarities. This indicates the responses to natural and sexual selection, including divergence of temporal and spectral signal properties. Phylogenetic constraints are unlikely to limit these changes of the auditory systems.     

Matching and Symmetry in the Frequency Recognition Curve of the Poison Frog Epipedobates trivittatus

In territorial species, males use signals to advertise territory ownership to other males. In species with acoustic communication, masking interference by heterospecific signals may impede male–male communication and affect the reproductive success of males. Frogs are thought to minimize masking interference by using species‐specific frequency channels for communication. For this strategy to work, a frequency match is expected between the advertisement call and the auditory sensitivity. A previous field study on the Amazonian frog Epipedobates femoralis supported this prediction, but also revealed an asymmetric decrease in the probability of male reaction towards synthetic calls. That males of E. femoralis reacted less towards low‐frequency (compared with high‐frequency) calls was interpreted as a mechanism that reduces masking interference by E. trivittatus, a species calling within a lower, partially overlapping, frequency range. If this hypothesis holds, then males of E. trivittatus should exhibit the opposite asymmetry pattern, i.e. react less towards high‐frequency (compared with low‐frequency) calls. We tested this prediction by conducting 25 playback experiments on 22 males of E. trivittatus. Male phonotactic reaction towards synthetic calls of various frequency values was evaluated as a binary variable (the male either approached or not the loudspeaker), by measuring the latency until first jump, and by calculating the linear approaching speed. As in E. femoralis, the maximum probability of positive reaction was matched to the call frequency. Against our expectations, the response curve was symmetric. We discuss whether these results reflect a lack of selective pressures, or a compromise between natural selection and physiological constraints on the shape of the frequency recognition curves.     

Hearing sensitivity and amplitude coding in bats are differentially shaped by echolocation calls and social calls

Differences in auditory perception between species are influenced by phylogenetic origin and the perceptual challenges imposed by the natural environment, such as detecting prey- or predator-generated sounds and communication signals. Bats are well suited for comparative studies on auditory perception since they predominantly rely on echolocation to perceive the world, while their social calls and most environmental sounds have low frequencies. We tested if hearing sensitivity and stimulus level coding in bats differ between high and low-frequency ranges by measuring auditory brainstem responses (ABRs) of 86 bats belonging to 11 species. In most species, auditory sensitivity was equally good at both high- and low-frequency ranges, while amplitude was more finely coded for higher frequency ranges. Additionally, we conducted a phylogenetic comparative analysis by combining our ABR data with published data on 27 species. Species-specific peaks in hearing sensitivity correlated with peak frequencies of echolocation calls and pup isolation calls, suggesting that changes in hearing sensitivity evolved in response to frequency changes of echolocation and social calls. Overall, our study provides the most comprehensive comparative assessment of bat hearing capacities to date and highlights the evolutionary pressures acting on their sensory perception.     

Do aggressive signals evolve towards higher reliability or lower costs of assessment?

It has been suggested that the evolution of signals must be a wasteful process for the signaller, aimed at the maximization of signal honesty. However, the reliability of communication depends not only on the costs paid by signallers but also on the costs paid by receivers during assessment, and less attention has been given to the interaction between these two types of costs during the evolution of signalling systems. A signaller and receiver may accept some level of signal dishonesty by choosing signals that are cheaper in terms of assessment but that are stabilized with less reliable mechanisms. I studied the potential trade‐off between signal reliability and the costs of signal assessment in the corncrake (Crex crex). I found that the birds prefer signals that are less costly regarding assessment rather than more reliable. Despite the fact that the fundamental frequency of calls was a strong predictor of male size, it was ignored by receivers unless they could directly compare signal variants. My data revealed a response advantage of costly signals when comparison between calls differing with fundamental frequencies is fast and straightforward, whereas cheap signalling is preferred in natural conditions. These data might improve our understanding of the influence of receivers on signal design because they support the hypothesis that fully honest signalling systems may be prone to dishonesty based on the effects of receiver costs and be replaced by signals that are cheaper in production and reception but more susceptible to cheating.     

Divergence in Female Duetting Signals in the Enchenopa binotata Species Complex of Treehoppers (Hemiptera: Membracidae)

Sexual communication often involves signal exchanges between the sexes, or duetting, in which mate choice is expressed through response signals. With both sexes acting as signalers and receivers, variation in the signals of males and females may be important for mate choice, reproductive isolation, and divergence. In the Enchenopa binotata species complex – a case study of sympatric speciation in which vibrational duetting may have an important role – male signals are species‐specific, females choose among males on the basis of signal traits that reflect species and individual differences, and female preferences have exerted divergent selection on male signals. Here, we describe variation in female signals in the E. binotata species complex. We report substantial species differences in the spectral and temporal features of female signals, and in their timing relative to male signals. These differences were similar in range to differences in male signals in the E. binotata complex. We consider processes that might contribute to divergence in female signals, and suggest that signal evolution in the E. binotata complex may be influenced by mate choice in both sexes.     

Auditory sensitivity and frequency selectivity in greater spear-nosed bats suggest specializations for acoustic communication

We investigated the relationship between auditory sensitivity, frequency selectivity, and the vocal repertoire of greater spear-nosed bats (Phyllostomus hastatus). P. hastatus commonly emit three types of vocalizations: group-specific foraging calls that range from 6 to 11 kHz, low amplitude echolocation calls that sweep from 80 to 40 kHz, and infant isolation calls from 15 to 100 kHz. To determine if hearing in P. hastatus is differentially sensitive or selective to frequencies in these calls, we determined absolute thresholds and masked thresholds using an operant conditioning procedure. Both absolute and masked thresholds were lowest at 15 kHz, which corresponds with the peak energy of isolation calls. Auditory and masked thresholds were higher at sound frequencies used for group-specific foraging calls and echolocation calls. Isolation calls meet the requirements of individual signatures and facilitate parent-offspring recognition. Many bat species produce isolation calls with peak energy between 10 and 25 kHz, which corresponds with the frequency region of highest sensitivity in those species for which audiogram data are available. These findings suggest that selection for accurate offspring recognition exerts a strong influence on the sensory system of P. hastatus and likely on other species of group-living bats.     

Neural Processing of Short-Term Recurrence in Songbird Vocal Communication

Background

Many situations involving animal communication are dominated by recurring, stereotyped signals. How do receivers optimally distinguish between frequently recurring signals and novel ones? Cortical auditory systems are known to be pre-attentively sensitive to short-term delivery statistics of artificial stimuli, but it is unknown if this phenomenon extends to the level of behaviorally relevant delivery patterns, such as those used during communication.

Methodology/Principal Findings

We recorded and analyzed complete auditory scenes of spontaneously communicating zebra finch (Taeniopygia guttata) pairs over a week-long period, and show that they can produce tens of thousands of short-range contact calls per day. Individual calls recur at time scales (median interval 1.5 s) matching those at which mammalian sensory systems are sensitive to recent stimulus history. Next, we presented to anesthetized birds sequences of frequently recurring calls interspersed with rare ones, and recorded, in parallel, action and local field potential responses in the medio-caudal auditory forebrain at 32 unique sites. Variation in call recurrence rate over natural ranges leads to widespread and significant modulation in strength of neural responses. Such modulation is highly call-specific in secondary auditory areas, but not in the main thalamo-recipient, primary auditory area.

Conclusions/Significance

Our results support the hypothesis that pre-attentive neural sensitivity to short-term stimulus recurrence is involved in the analysis of auditory scenes at the level of delivery patterns of meaningful sounds. This may enable birds to efficiently and automatically distinguish frequently recurring vocalizations from other events in their auditory scene.     

Female reproductive state influences the auditory midbrain response

Female behavioral responses to sensory stimuli can be highly variable across the reproductive cycle. Female green treefrogs (Hyla cinerea) use the male vocal signal to locate and choose a mate. Gravid females approach a vocalizing male to mate but do not approach if they have recently mated. Such differences in behavioral response may be due in part to shifts in the neural representation of auditory information in the brain. In this study, we investigated the influence of female reproductive state on neural responses in the auditory midbrain to both communication signals (advertisement calls) and non-communication sounds (band limited noise bursts). Recently mated females exhibited significantly reduced response strengths compared to females not recently mated. Reduced response strengths in post-mated females were in response to both noise bursts and male advertisement calls but were limited to the lower frequency range corresponding to the amphibian papilla of the peripheral auditory system. Our results therefore show that the ability of social signals to stimulate the auditory system differs in females depending on their reproductive state, and that the differential effect on low versus high spectral sensitivities may influence the way the two spectral peaks of male advertisement calls are represented.     

A comparison of auditory brainstem responses and behavioral estimates of hearing sensitivity in Lemur catta and Nycticebus coucang

Primates depend on acoustic signals and cues to avoid predators, locate food, and share information. Accordingly, the structure and function of acoustic stimuli have long been emphasized in studies of primate behavioral and cognitive ecology. Yet, few studies have addressed how well primates hear such stimuli; indeed, the auditory thresholds of most primate species are unknown. This empirical void is due in part to the logistic and economic challenges attendant on traditional behavioral testing methods. Technological advances have produced a safe and cost‐effective alternative—the auditory brainstem response (ABR) method, which can be utilized in field conditions, on virtually any animal species, and without subject training. Here we used the ABR and four methods of threshold determination to construct audiograms for two strepsirrhine primates: the ring‐tailed lemur (Lemur catta) and slow loris (Nycticebus coucang). Next, to verify the general efficacy of the ABR method, we compared our results to published behaviorally‐derived audiograms. We found that the four ABR threshold detection methods produced similar results, including relatively elevated thresholds but similarly shaped audiograms compared to those derived behaviorally. The ABR and behavioral absolute thresholds were significantly correlated, and the frequencies of best sensitivity and high‐frequency limits were comparable. However, at frequencies ≤2 kHz, ABR thresholds were especially elevated, resulting in decreased agreement with behavioral thresholds and, in Lemur, the ABR 10‐dB range starting points were more than 2 octaves higher than the behavioral points. Finally, a comparison of ABR‐ and behaviorally‐derived audiograms from various animal taxa demonstrates the widespread efficacy of the ABR for estimating frequency of best sensitivity, but otherwise suggests caution; factors such as stimulus properties and threshold definition affect results. We conclude that the ABR method is a promising technique for estimating primate hearing sensitivity, but that additional data are required to explore its efficacy for estimating low‐frequency thresholds. Am. J. Primatol. 72:217–233, 2010. © 2009 Wiley‐Liss, Inc.     

Dissimilarities in auditory tuning in midwife toads of the genus Alytes (Amphibia: Anura)

The auditory sensitivity in three species of the anuran genus Alytes (Alytidae) was examined to determine patterns of intra‐ and interspecific variation, relating these measurements to behavioural preferences measured in previous studies and to the adaptive and evolutionary significance of this sensory function. The audiograms obtained with multi‐unit recordings in the torus semicircularis of 13 Alytes cisternasii, 10 Alytes obstetricans, and eight Alytes dickhilleni show two regions of enhanced sensitivity, between approximately 100–500 and 1200–2400 Hz, with minimum thresholds at approximately 40 and 45 dB SPL, respectively. The mean and range of the high‐frequency region differed among species, although the sensitivity, measured as minimum thresholds, was similar. The region of high‐frequency sensitivity was centred at approximately the frequency of the advertisement call in A. cisternasii but, in A. obstetricans and A. dickhilleni, was centred at frequencies higher than the conspecific calls. These results contrast with preferences for lower frequencies exhibited by Alytes in female phonotactic and in male evoked vocal responses. Such loose relationships between signals and receivers suggest that the divergence of the sound communication system in Alytes has implied environmental and phylogenetic factors in addition to sexual selection processes.     

Correlation between auditory sensitivity and vocalization in anabantoid fishes

Several anabantoid species produce broad-band sounds with high-pitched dominant frequencies (0.8–2.5 kHz), which contrast with generally low-frequency hearing abilities in (perciform) fishes. Utilizing a recently developed auditory brainstem response recording-technique, auditory sensitivities of the gouramis Trichopsis vittata, T. pumila, Colisa lalia, Macropodus opercularis and Trichogaster trichopterus were investigated and compared with the sound characteristics of the respective species. All five species exhibited enhanced sound-detecting abilities and perceived tone bursts up to 5 kHz, which qualifies this group as hearing specialists. All fishes possessed a high-frequency sensitivity maximum between 800 Hz and 1500 Hz. Lowest hearing thresholds were found in T. trichopterus (76 dB re 1 μPa at 800 Hz). Dominant frequencies of sounds correspond with the best hearing bandwidth in T. vittata (1–2 kHz) and C. lalia (0.8–1 kHz). In the smallest species, T. pumila, dominant frequencies of acoustic signals (1.5–2.5 kHz) do not match lowest thresholds, which were below 1.5 kHz. However, of all species studied, T. pumila had best hearing sensitivity at frequencies above 2 kHz. The association between high-pitched sounds and hearing may be caused by the suprabranchial air-breathing chamber, which, lying close to the hearing and sonic organs, enhances both sound perception and emission at its resonant frequency. Accepted: 26 November 1997     

Pre-existing sensory biases in the spectral domain in frogs: empirical results and methodological considerations

In many species of anurans, advertisement calls excite only one of the two inner-ear organs. One prediction of the pre-existing bias hypothesis is that signal innovations that additionally excite the “untapped” organ will be more behaviorally effective than normal calls. However, recent studies have shown that females of three species with single-peaked calls that stimulate only the basilar papilla (BP) preferred single-peaked synthetic calls with a frequency typical of conspecific calls to two-peaked calls that also stimulated the amphibian papilla (AP). We report that in spring peepers (Pseudacris crucifer) that also produce single-peaked calls, females did not show a preference in choices between single-peaked and two-peaked synthetic calls. Thus, the addition of energy exciting the AP had a neutral effect on signal attractiveness. Together, these results are unsupportive of the pre-existing bias hypothesis. An alternative hypothesis is that positive fitness consequences of responding to sounds providing extraordinary spectral stimulation are required for a novel call to become established as a mate-attracting signal. Testing these ideas requires a taxonomically broader examination of responses to sounds with novel spectral complexity, and attention to some methodological details will improve the comparability of such studies.     

Seasonal plasticity in auditory processing of the envelope and temporal fine structure of sounds in three songbirds

Songs mediate mate attraction and territorial defence in songbirds during the breeding season. Outside of the breeding season, the avian vocal repertoire often includes calls that function in foraging, antipredator and social behaviours. Songs and calls can differ substantially in their spectral and temporal content. Given seasonal variation in the vocal signals, the sender–receiver matching hypothesis predicts seasonal changes in auditory processing that match the physical properties of songs during the breeding season and calls outside of it. We tested this hypothesis in white-breasted nuthatches, Sitta carolinensis, tufted titmice, Baeolophus bicolor, and Carolina chickadees, Poecile carolinensis. We measured the envelope-following response (EFR), which quantifies phase locking to the amplitude envelope, and the frequency-following response (FFR), which quantifies phase locking to the temporal fine structure of sounds. Because songs and calls of nuthatches are amplitude modulated at different rates, we predicted seasonal changes in EFRs that match the rates of amplitude fluctuation in songs and calls. In chickadees and titmice, we predicted stronger FFRs during the spring and stronger EFRs during the winter because songs are tonal and calls include amplitude-modulated elements. In all three species, we found seasonal changes in EFRs and FFRs. EFRs varied across seasons and matched the amplitude modulations of songs and calls in nuthatches. In addition, female chickadees had stronger EFRs in the winter than in the spring. In all three species, FFRs during the spring tended to be stronger in females than in males. We also found species differences in EFRs and FFRs in both seasons; EFRs and FFRs tended to be higher in nuthatches than in chickadees and titmice. We discuss the potential mechanisms underlying seasonality in EFRs and FFRs and the implications of our results for communication during the breeding season and outside of it, when these three species form mixed-species flocks.     

Phenotypic integration and the evolution of signal repertoires: A case study of treefrog acoustic communication

Animal signals are inherently complex phenotypes with many interacting parts combining to elicit responses from receivers. The pattern of interrelationships between signal components reflects the extent to which each component is expressed, and responds to selection, either in concert with or independently of others. Furthermore, many species have complex repertoires consisting of multiple signal types used in different contexts, and common morphological and physiological constraints may result in interrelationships extending across the multiple signals in species’ repertoires. The evolutionary significance of interrelationships between signal traits can be explored within the framework of phenotypic integration, which offers a suite of quantitative techniques to characterize complex phenotypes. In particular, these techniques allow for the assessment of modularity and integration, which describe, respectively, the extent to which sets of traits covary either independently or jointly. Although signal and repertoire complexity are thought to be major drivers of diversification and social evolution, few studies have explicitly measured the phenotypic integration of signals to investigate the evolution of diverse communication systems. We applied methods from phenotypic integration studies to quantify integration in the two primary vocalization types (advertisement and aggressive calls) in the treefrogs Hyla versicolor, Hyla cinerea, and Dendropsophus ebraccatus. We recorded male calls and calculated standardized phenotypic variance–covariance ( P ) matrices for characteristics within and across call types. We found significant integration across call types, but the strength of integration varied by species and corresponded with the acoustic similarity of the call types within each species. H. versicolor had the most modular advertisement and aggressive calls and the least acoustically similar call types. Additionally, P was robust to changing social competition levels in H. versicolor. Our findings suggest new directions in animal communication research in which the complex relationships among the traits of multiple signals are a key consideration for understanding signal evolution.