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Superfast Muscles Set Maximum Call Rate in Echolocating Bats

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Science  30 Sep 2011:
Vol. 333, Issue 6051, pp. 1885-1888
DOI: 10.1126/science.1207309

Abstract

As an echolocating bat closes in on a flying insect, it increases call emission to rates beyond 160 calls per second. This high call rate phase, dubbed the terminal buzz, has proven enigmatic because it is unknown how bats are able to produce calls so quickly. We found that previously unknown and highly specialized superfast muscles power rapid call rates in the terminal buzz. Additionally, we show that laryngeal motor performance, not overlap between call production and the arrival of echoes at the bat’s ears, limits maximum call rate. Superfast muscles are rare in vertebrates and always associated with extraordinary motor demands on acoustic communication. We propose that the advantages of rapid auditory updates on prey movement selected for superfast laryngeal muscle in echolocating bats.

Laryngeal echolocation and insectivory characterize about 70% of present-day bat species (13). Over the course of an attack on a flying insect, bats increase their echolocation call emission rates as they progress from prey detection, through approach, to the terminal buzz (1, 2, 4) (Fig. 1A). Increasing call emission rates means more information updates per unit time from returning echoes on the relative position of the target. All aerial hawking bats studied to date produce the buzz, which is sometimes subdivided into “buzz I” and “buzz II” phase calls, the former occurring at rates of ~100 to 160 calls/s, and the latter ≥160 calls/s (1, 5) (Fig. 1B). Bats do not call at rates exceeding those reached during this final stage of aerial hawking attack (2, 4), and we hypothesize that call production, echo processing, or both limit maximum echolocation call rate.

Fig. 1

Echolocation and flight kinematics during aerial attack sequences in Myotis daubentonii. (A) Spectrogram and oscillogram of emitted echolocation calls as recorded on the center microphone of the array. (B) Instantaneous call repetition rates from sequence in (A). The start of buzz I is defined as call repetition rate >100 calls/s (cyan circles, light gray zone). In buzz II, the repetition rate is >160 calls/s (red circles, medium gray zone); here, peak fundamental frequency of the calls drops from 45 to 25 kHz. (C) Four reconstructed flight paths from a single bat using a 12-microphone array (10). Red arrows indicate flight direction.

Laryngeal nerve-cut experiments reveal that each call a bat emits is under active neuromuscular control (6, 7). Consequently, muscle performance might place an upper limit on the rate at which bats produce calls. Alternatively, if prey echoes overlap with or return after the next call is emitted, accuracy in target ranging may suffer as a result of ambiguity in matching echoes to calls (1, 8). While hunting, most species avoid potential ambiguity by not producing the next call until target echoes reach the bat’s ears (1), potentially limiting maximum call rates during the buzz. To investigate these hypotheses, we first measured sound production during aerial attack sequences in free-flying Daubenton’s bats (Myotis daubentonii, Vespertilionidae) using a 12-microphone array (Fig. 1C) (9) and determined when the start and the end of each prey echo (Fig. 2, A to C) would impinge upon the bat’s ears relative to both the source call and the next call emitted (10). Our data show that during a buzz, echoes from individual calls terminate before the start of the next call (Fig. 2C and fig. S1), suggesting no ambiguity in call-echo matching. In fact, for buzz II calls, the repetition rate could theoretically exceed 400 calls/s without any such ambiguity (Fig. 2C and fig. S1), a rate twice as high as the 190 calls/s observed in our study (Fig. 1). Our results also demonstrate that, because call duration decreases during the buzz, there is no overlap between a call and its echo until the bat is less than 5 cm from its target (Fig. 2B), corroborating previous estimates for this species and others (1, 5, 11). Assuming a processing time of ~20 ms for each received echo (12), any perceptual difficulties created by call-echo overlap at the end of a buzz may be negligible because by the time these echoes have been processed the insect will have already been taken or evaded the bat (9). In sum, our data and that of previous reports from the field (5) and lab (11) indicate that echo processing does not limit call repetition rate in the buzz.

Fig. 2

Call-echo matching ambiguity does not limit echolocation call repetition rate. (A) High-frequency–time-resolution representation (28) and oscillogram of an approach and buzz II call showing smooth frequency sweeps. We calculated the time needed for the returning prey echo to reach the bat’s ears (10) and quantified call duration, the time from call offset to echo onset (ton), and the time from call onset to echo offset (toff) for each call. (B) Call duration versus distance to prey. Call duration decreases during the buzz down to ~200 μs. Exemplary traces of five bats are shown (cyan circles, buzz I; red circles, buzz II). Call-echo overlap occurs when call duration exceeds the time needed for sound to travel to and from the prey calculated (i) solely based on call duration (dashed black line) and (ii) after adjusting for bat flight speed (blue line). (C) Histograms of ton and toff (cyan bars, buzz I; yellow bars, buzz II) in relation to the next call (black bars, buzz I; red bars, buzz II) for a total of 432 calls from four approaches per bat from five bats. Prey echoes start (top) and end (bottom) at the bat’s ear well before the next call is produced. Vertical dotted lines indicate periods of higher repetition rates.

Next, we turned our attention to call production. Vespertilionids and other bat species generate calls in the larynx by flow-induced oscillation of vocal folds (6, 7, 13, 14), which terminate as very thin membranes (Fig. 3A). The tension in the folds and membranes determines their oscillation frequency, and thus the frequency of sounds produced. Tension is increased by rotation of the thyroid cartilage around the cricothyroid joint (Fig. 3B) and is mainly controlled by the bat’s massive cricothyroid muscle (Fig. 3C). The roles of several other laryngeal muscles controlling call duration are under debate (6, 7, 13). Because each echolocation call is under active neuromuscular control (6, 7), call repetition rates >100 calls/s require separate work-producing cycles of muscle contraction and relaxation at the same rate. Contraction cycles this fast lie outside the capability of typical vertebrate synchronous skeletal muscle (15), which suggests that bat laryngeal muscles may be part of a rare group of superfast muscles, defined here as muscles capable of producing work >100 Hz. Such muscles have only ever been identified in the sound-producing organs of toadfish (15, 16), rattlesnakes (15, 16), and birds (17, 18).

Fig. 3

Sound frequency modulation during echolocation calls. (A) Proposed frequency control mechanism of echolocation calls illustrated in schematic representation of a vespertilionid bat larynx [adapted from (7) with permission from R. A. Suthers]. Flow-induced self-sustained oscillations of the vocal folds (dark blue) and vocal membranes (light blue) generate ultrasonic pressure fluctuations. Their tension (σ) determines the fundamental frequency (f0) of self-sustained oscillations. (B) Rotation (blue arrows) of the thyroid cartilage around the cricothyroid joint (blue circle) causes the vocal folds and membranes to stretch and adduct while the arytenoid cartilage remains stationary (7). This rotation increases fold and membrane length and tension (red plus) and thereby increases the fundamental frequency of produced sound. (C) The anterior cricothyroid muscle (ACTM) controls cricothyroid rotation and consists of an anterior (ACTM-a) and posterior part (ACTM-p). Relaxation of the ACTM-a results in f0 decrease during phonation.

For the precise control of the terminal buzz, the main frequency modulating muscle must produce work and power at the repetition rates observed for the buzz (≤190 Hz) (Fig. 1). We conducted isometric twitch experiments on isolated muscle fibers of the main frequency modulating muscle (i.e., the anterior part of the anterior cricothyroid muscle) (Fig. 3C), which confirmed fast-twitch kinetics (Fig. 4A) and thus provided a necessary, but not sufficient, test of superfast muscle functionality (15). Twitch half-times measured 4.72 ± 0.38 ms at 39°C (N = 11 preparations from 7 individuals), faster than toadfish superfast swim-bladder muscle (5.8 ms at 25°C) (16) but slower than songbird superfast syringeal muscle (3.23 ms at 40°C) (17). The total contraction time was 8.27 ms during field stimulation of muscle fibers (10), almost twice as fast as the 12 to 16 ms reported for an anaesthetized big brown bat (Eptesicus fuscus, Vespertilionidae) in response to motor-nerve stimulation (13).

Fig. 4

Superfast performance of bat vocal muscle. (A) Isometric twitch recordings of ACTM-a preparations in vitro show extremely fast rise and decay times of 2.13 ± 0.33 ms (10 to 100% tension; N = 11 preparations from 7 individuals) and 6.14 ± 0.66 ms (100 to 10% tension), respectively. Downward arrow indicates stimulus. (B) Work-loop traces (five successive loops superimposed) of a preparation performing at cycle frequencies of 100, 140, 160, 180, and 200 Hz. At 180 and 200 Hz, increasing amounts of negative work appear (areas enclosed by clockwise rotation; asterisks) that counteract the production of positive work (areas enclosed by counter-clockwise rotation) in each work loop. (C) Both power and (D) work (mean + SD) demonstrate that the anterior cricothyroid muscle is capable of producing positive power and work during cyclic contractions up to 160 Hz, and in one case up to 200 Hz. Orange dots indicate performance of the preparation depicted in (B).

To demonstrate that superfast muscles are responsible for bats’ ability to produce the buzz, we measured the mechanical performance of isolated bundles of muscle fibers by subjecting them to various strain cycles and stimulation regimes, that is, the work-loop technique (10). We found that bat laryngeal muscle produces both positive power and work at cycle frequencies up to 180 Hz and, in one case, up to 200 Hz (Fig. 4, B to D). At cycle frequencies of >180 Hz, the work loops deform to a figure-of-eight configuration (Fig. 4B). Beyond this point, negative work (Fig. 4B) starts to counteract the positive work in each loop. Above 200 Hz, negative work outweighs positive work, resulting in a negative net amount of work and power for all preparations. Thus, despite their extreme performance, Daubenton’s bats’ superfast laryngeal muscles perform up to, but not beyond, the cycle frequencies observed at the highest call rates in the buzz. Superfast muscles therefore allow, but limit, the maximum rate of bat echolocation call production.

Superfast muscles have been previously reported in a small number of species of reptiles, birds, and ray-finned fishes (1520). We can now add mammals to that list. Studies of superfast synchronous muscle in nonmammalian species, most notably the oyster toadfish (Opsanus tau), indicate several hallmark adaptations to the basic contractile architecture (15, 19, 20) that allow for extremely rapid kinetics at every step in excitation-contraction coupling and relaxation. Histological studies of bat laryngeal muscle have revealed some similar modifications, including increased sarcoplasmic reticulum (21) and mitochondrial density (22). Intriguingly, the laryngeal and extraocular muscles of rats and rabbits express the myosin heavy-chain isoform MYH13 (23, 24), which exhibits detachment rates from actin faster than is typical of skeletal muscles (25) and belongs to an ancient cluster of myosin genes (26). Whether similar isoforms are expressed in laryngeal echolocating bats remains to be seen.

We propose that the development of superfast muscles was crucial to the success of bats as aerial predators. Two innovations—powered flight and echolocation—are thought to have allowed bats to exploit the previously unrealized foraging niche of night-flying insects (14). Although a number of other vertebrates use echolocation for orientation (e.g., oilbirds, cave swiftlets, and a few tongue-clicking Rousettus spp. from the otherwise nonecholocating Old World fruit bats), only toothed whales and laryngeal echolocating bats use echolocation to detect prey, and only these animals produce buzzes (27). The ubiquity of buzzes in today’s aerial hawking bats when taking prey (1, 2, 9) suggests that the capacity to emit short echolocation calls at very high rates evolved to enhance bats’ success in capturing night-flying insects. We suggest that the demands of an active sensory system specialized for target acquisition, rather than simply orientation, selected for functional superfast vocal muscles needed to power the terminal buzz.

Supporting Online Material

www.sciencemag.org/cgi/content/full/333/6051/1885/DC1

Materials and Methods

Fig. S1

References

References and Notes

  1. Materials and methods are available as supporting material on Science Online.
  2. Acknowledgments: We thank R. Suthers for permission to reprint Fig. 3 from (7); R. Langill, P. Martensen, and F. Mortensen for technical assistance; G. Lutz for providing carbon microspheres; T. Gardner for sparse time-frequency Matlab scripts; and F. Jensen, D. Canfield, A. Surlykke, and A. West for use of equipment. F. Andrade, S. Brinkløv, P. Faure, B. Fenton, B. Galef, J. van Leeuwen, P. Madsen, M. Nydam, A. Surlykke, H. ter Hofstede, and S. Zawadzki commented on the manuscript. This study was funded by grants from the Danish Research Council (FNU), Carlsberg Foundation, and Grass Foundation to C.P.H.E.; Company of Biologists to A.F.M.; Oticon Foundation to L.J.; and FNU to J.M.R. Animal capture and experimentation was approved by Skov- og Naturstyrelsen (Denmark). C.P.H.E. and J.M.R. conceived of the study. C.P.H.E. and A.F.M. conducted physiological experiments; C.P.H.E., L.J., and J.M.R. conducted behavioral experiments. All authors contributed to data analysis. C.P.H.E. and J.M.R. wrote the manuscript. The data reported in this paper are available from C.P.H.E and J.M.R.
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