Department of Biology, State University of New Yorh at Buffalo, a n d Auditory Research Laboratories, Princeton Uniuersity, Princeton, New Jersey ABSTRACT The burrowing squamates of the order Amphisbaenia lack external ears. The middle ear involves an extremely large stapes with the head of its shaft (columellar portion) projecting just posterior to and as far laterally as the quadrate. In most species a cartilaginous extracolumella articulates with the distal tip of the shaft and then stretches along the lower jaw and is attached to the skin in the region of the second infralabial. Records of the electrical potentials of the cochlea in some nine species show sensitivity curves whose maxima are approximately -20dB for a standard output of 0.1 pv. The zone of greatest sensitivity lies in the low frequencies, usually between 300 and 700 Hz. The sensitivity of amphisbaenians differs from that of snakes in being fairly good above levels of 700 Hz, where the latter drops off sharply. Inter- ruption of the extracolumella causes a 20 to 30 dB loss in sensitivity. Sound waves applied asymmetrically produce different electrical responses in the two ears, suggesting an explanation for behavioral observations on the utilization of the system. The form and function of the ear in amphisbaenians are hardly known. Since an external opening is lacking, the group has been characterized as ear-less, and only the structure of the middle ear has had brief mention in treatments of the anatomy of the skull. Some speculations have been offered about auditory function in general discussions of the life and hab- its of these reptiles (Gans, '60, '68). That hearing may play an important role in such activities as the finding of food and mates and predator avoidance seems a reasonable assumption for these curious animals whose life is mainly, sometimes exclusively, subterranean and whose vi- sion is apparently reduced to simple light perception (Gans and Bonin, '63). The present report presents evidence on the functional characteristics of the peripheral auditory mechanism in nine species of amphisbaenians. It includes ob- servations on the anatomy of the ear in two species as well as preliminary data on three others. For the functional studies we recorded the electrical potentials of the cochlea (Wever, '39, '66). These po- tentials represent the activity of the audi- tory hair cells and reflect the properties of the receptive and conductive systems J . EXP. ZOOL., 179 17-34 and of the mechanoelectric processes in the cochlea. The species studied (table 1) were: Am- phisbaena alba ( 3 specimens), Amphis- baena darwini trachura (1 specimen), Amphisbaena manni (7 specimens), Chir- india langi ( 2 specimens), Cynisca leu- cura ( 3 specimens), Zygaspis violacea ( 3 specimens), Monopeltis c. capensis ( 2 specimens), Trogonophis wi egmanni (2 specimens), and Bipes biporus (4 spe- cimens). METHOD The animals were anesthetized with 20% solution of ethyl carbamate in phys- iological saline, in dosages near 0.01 ml per gram of body weight. The sensitivity of the ear in terms of its cochlear poten- tials was determined by recording from an electrode in contact with the perilymph of the saccule, which is readily accessible through the dorsolateral surface of the posterior region of the head. The skull 1 The experi ments were carri ed out in the Auditory Research Laboratories of Princeton University wi th facilities provided by grants from the Nati onal I nsti- tute of Neurological Diseases and Stroke. The speci- mens were obtai ned wi th funds from NSF GB-6521X. 2 Present address: Department of Zoology, The Uni- versity of Michigan, Ann Arbor, Mi chi gan 48104. 17 18 CARL GANS AND ERNEST GLEN WEVER TABLE 1 The specimens used are deposited i n the Carl Gans (CG) collection. The species used, the catalogue numbers, the localities and the collectors are as follows: Amphisbnenn alba: (no No.), Brazil, (dealer); same species (CG 371 71, Venezuela, Edo. Carabobo, Valencia (S. M. McCown); same species (CG 3718), Brazil, S. P. J undiai, Sitio Bandeirante (W. C. A. Bokermann). Ampkisbaenn darwini trachurn: (CG 3752), Brazil, RGS, Tramandai, Emboaba (C. P. J aeger and S. Correa). Amphisbnena manni: (CG 3822, 3823, 3844, 3845, 3846, 3852, 3853, 3859, 3860, 3861, 3862, 3863) Hispaniola, Ile de l a Tortue, Palmistes (C. R. Warren). Bipes bi po~zt s: (CG 3804, 3805, 3806, 3808) Mexico, Baja California, Sul de l a Paz (L. W. Robbins). Chirindin langi: (CG 4033, 4035), Mozambique, 15 miles southeast of Vila de Manica (D. G. Broadley, C. Gans and J . Visser). Cyni sca leucura: (CG 4126), Ghana, Legon (B. Hughes); same species: (CG 4151, 4152), Ghana, Legon, hill behind 2001. Dept. (B. Hughes). Monopeltis cnpensi s: (CG 4456, 4457), South Africa, O.F.S., Bothaville (J . Visser). Twgonopki s wi egmanni : (2 specimens, no No.), Maroc (Dealer). Zygnspi s violacea: CG 4546, 4558,4559), Zululand, Ndumu Game Reserve (A. C. Pooley). was exposed and a small hole was drilled of a size that was tightly filled by the ta- pered end of a fine steel needle that was insulated except at the tip. Two other electrodes, one of which was grounded, were located on inactive tissues of the region. The electrodes led to a differen- tial preamplifier that provided an 80-db rejection of stray potentials picked up between two of the electrodes and the grounded lead. The animal was stimulated with a series of tones that ordinarily covered the range from 100 to 10,000 Hz, but in some instances included lower frequencies. For aerial stimulation the tones were produced by a loudspeaker outside the shielded chamber in which the animal was kept and were conducted through the wall by a tube. This tube terminated in a sound cannula whose tip was located over the facial region close to the skin surface. To avoid any possibility of mechanical con- duction to the animal, this tube was held out of direct contact and the narrow gap between its rim and the skin was filled with petroleum jelly to make a tight acoustic seal. A probe tube leading to a condenser microphone ran concentrically within the sound tube and ended near its tip, and the system was calibrated to pro- vide readings of sound pressure. The physiological state of the animal was gen- erally monitored by display of an elec- trocardiogram. Vibratory stimulation for other experi- ments was produced by a crystal vibrator that actuated a steel needle, the blunted end of which could be applied to the body surface. The vibrator was fitted with an accelerometer and was calibrated by means of a "Fotonic sensor," a device utilizing fiber optics for the precise mea- surement of vibratory amplitudes. The stimuli were presented at the in- tensities necessary to produce some small but reliably observable level of potential, which, because of the physiological noise present, was set between 0.05 and 0.2 pv. I n the curves the results are reported for a standard output of 0.1 pv; when the measured values differed from this level the sound required was calculated, since our tests had shown that the response at low levels is a linear function of sound pressure. The earliest measurements were always repeated to check that the animal had not deteriorated during the test sequence. Most of the measurements were made at room temperature, which was about 23"C, since at higher temperatures the animals seemed to desiccate rapidly, mu- cous production increased, and we en- countered very significant mortality. Only in one instance ( Chi r i ndi a l angi ) were we able to obtain readings at higher temper- atures. These readings support the results of Werner ('68) that the general curve shape remains similar but shifts to a higher frequency range, while the sensi- tivity increases slightly. Unfortunately there are no published data for the pre- ferred body temperatures of any amphis- baenean. For this reason and because of the relatively minor changes in absolute values we deferred collection of temper- ature data until after behavioral observa- tions have been completed, when addi- tional specimens may be available. EAR AND HEARING I N AMPHISBAENIA (REPTILIA) 19 I n many of the experiments, special procedures were carried out to study the processes of sound perception and the transmission of vibrations to the cochlea. The effect of severing the extracolumella, singly or multiply, was studied in several animals. I n such operations the extra- columella was first exposed by dissecting away the overlying skin and muscle. The effect of this procedure on sensitivity was then checked prior to cutting the extra- columella itself. Systematic exploration of the lateral surface of the head with the mechanical vibrator was intended to locate the sensitive zones. The direction- ality of sound reception was tested by re- cording from both ears while stimulating with asymmetrically placed aerial sounds. After the electrical tests had been com- pleted, each anesthetized animal was per- fused through the circulatory system by means of a cannula inserted through the ventricle into the ascending aortic ves- sels. Perfusion with physiological saline to flush out the blood was followed by a fixative containing mercuric chloride, potassium dichromate, sodium sulfate, and formaldehyde. The fixation was con- tinued for 40 to 60 minutes. The head was then removed and the tissues trimmed to a block containing both ears. This block was replaced in fixative, decalcified in 0.5% nitric acid, dehydrated in an as- cending series of alcohol from 10 to 100% by 10% steps, and embedded in celloidin. The hardened celloidin block was sec- tioned (horizontally, at an angle of about 15" toward the transverse) at 20 p inter- vals perpendicular to the long axis of the basilar membrane. Every section i n the region of the ear was mounted separately and stained with hematoxylin, azocarmine, and orange G, a combination giving good differentiation of the auditory hair cells. While the procedure yields excellent pres- ervation, it requires 10-12 months. At the time of this report the histolog- ical treatment is complete on 14 animals belonging to five species: three specimens of Amphisbaena alba, one of A. darwini trachura, five of A. manni , one of Trogo- nophis wiegmanni, and four of Bipes bi- porus. Several other specimens are still in preparation, and detailed morpholog- ical description is deferred until the his- tological study is completed. RESULTS Anatomical observations Earlier studies, especially those of Ver- sluys (1898) on the ear region and of Gans ('60) on the Trogonophidae, have included observations on the ear's struc- ture and some descriptions of the auditory mechanism. While none of the 140 species of the order (Gans, '67) has an external ear (fig. l), all have middle and inner ear structures, concealed below the uninter- rupted dermal segments covering the head. Fig. 1 Dorsal, lateral and ventral views of the head of Amphisbaenn mnnni (American Museum no. 49726), to show the pattern of dermal seg- mentation. The second left infralabial i s shaded. The line equals 1 mm to scale. 20 CARL GANS AND ERNEST GLEN WEVER Extracolurnel la' Fig. 2 Lateral view of the skull of Anzpkisbtre7zn c t ~ec n (Carnegie Museum no. 37636) showing the stapes and extracolumella. As in all other reptiles, the middle ear mechanism consists of columellar and ex- tracolumellar elements, but their form is quite distinct (Baird, '70). There is always an osseous coluniella, also called a stapes. This element has a broad footplate resting in the oval window and a relatively short shaft (columellar portion) the distal head of which connects with a generally cartilaginous extracol- umella that, except in Bipes, ultiniately attaches to the skin anterior to the an- gulus oris. In the genus Anzphisbaenn the extra- columella is a long, slender rod that ex- tends from the stapes anterolaterally and then anteriorly as shown in the drawing (fig. 2) and the photomicrograph (fig. 3) . Near its columellar articulation the extra- columella runs through dense connective and muscular tissues along the lateral side of the quadrate bone on the medial side of a muscle layer. More anteriorly beyond the articulation of the lower jaw it emerges as a structure lying freely in the subcutaneous space. Its anterior por- tion enters the dermal layer of the skin within which it shows broad connective tissue attachment; after a forward course as far as the two posterior teeth, it tapers Fig. 3 Photomicrograph of a section through to a slender thread and ends. the head of a specimen of A?np/ ?i sb~i e?ru mnnni, depicts further details of the showing the cochlear capsules and middle ear structures. The posterior portion of the extra- stapes and its connections in Amphis- columella on the left side had been interrupted bnenn mnnni. The footplate is broad, surgically; the system on the right is intact. equalling the diameter of the otic capsule, Figure Scale X 12. EAR AND HEARI NG I N AMPHISBAENIA (REPTI LI A) 21 Fig. 4 Photomicrograph at a higher power of the right i nner ear region of the same spe- cimen as in the previous figure. e, extracolumella; f, footplate of stapes; s, shaft of stapes. Scale X 60 of which it forms the anterolateral wall. The anterior portion of this ossicle makes a wide articulation with the prootic bone, while the smaller posterior portion is held to the lateral wall of the capsule by a wider ligament. The neck and headpiece extend farthest laterally. The articular surface is clearly indicated, while the con- nection to the expanded end of the extra- columella lacks an articular cleft and seems relatively rigid. I n Trogonophis the stapes has much the same form as in Amphisbaena, but the extracolumella is a short, plate-shaped cartilage expanding ventrally. Its forward extension passes only a short distance an- terior to the mandibular joint, and then connects to a broad band of dense connec- tive tissue that continues anterolaterally to fuse with the deep layer of the skin. In Bipes biporus the extracolumella takes still a different form. From the col- umellar articulation it extends laterally and somewhat posteriorly for a compara- tively short distance after which it ter- minates in a flat headpiece that lies just posterior to the quadrate bone. Its precise relations to the enveloping soft tissues need further study. The inner ear exhibits considerable uni- formity in the species examined so far. The basilar membrane has a distorted oval form, is nearly as wide as it is long, and bears an auditory papilla as shown in cross section of Amphisbaena alba in figure 5. In this section, which is from the midregion of the basilar membrane, there are 12 rows of hair cells whose cili- ary tufts are embedded in a thick tec- torial plate. These tufts extend into pits in the lower surface of the plate. The tec- torial membrane is attached to the plate near its middle as shown. This arrange- ment closely resembles that found in tur- tles (Wever, unpublished). Counts of the cochlear hair cells were made in the specimens for which serial sections were available and the preserva- tion and staining were suitable. The re- sults are given in table 2. The size of the 22 CARL GANS AND ERNEST GLEN WEVER Tectorial membrane Fig, 5 Drawing of the auditory papilla of Ampk i s bae na al ba, from a section transverse to the basilar membrane. TABLE 2 Nu mb e r of hai r cells Species Number of cells Animal number Left ear Right ear A m p hi s ba e na a l bn 1500 1667 1668 Amphi s bae na dtrrwini t i ncht i ra 1525 Amphi sbnent r mu n n i 1520 1524 1547 1548 Tr ogonophi s wi e g mann i 1736 Bi pes biporzrs 1522 1553 1554 1555 148 150 159 168 143 99 105 - 109 - 107 138 99 78 81 82 94 106 56 - 45 53 47 67 56 55 hair cell population varies with species. Amphisbaena alba has the largest num- ber, averaging 154 (N = 3) . For A. dar- wini trachura (N = 1), A. manni (N = 5) , and Trogonophis wi egmanni (N = 1) the numbers are hardly distinguishable, approximately 100. For Bipes biporus the number is smaller, mean 54 (N = 4). Cochlear potential studies 1. Amphisbaena alba. The solid-lined curve in figure 6 gives measurements of auditory sensitivity in terms of cochlear potentials for a specimen of Amphisbaena alba, under the usual conditions. This curve reaches its lowest level, represent- ing the greatest sensitivity, in the region of 200 to 600 Hz, and then, as the fre- quency increases, it rises, at first slowly and then more rapidly, up to 5000 Hz. Here an inversion appears, after which the upward rise continues. After these tests had been concluded, the extracolumella was cut anteriorly, leaving only its posterior two-thirds con- nected to the stapes. The results are shown by the broken curve of this figure. The sensitivity suffered a considerable loss, extending over the frequencies up EAR AND HEARI NG I N AMPHI SBAENI A (REPTI LI A) 1 2 3 4 5 6 7 8 9 1 2 3 4 5 6 7 8 9 1 23 Frequency Fig. 6 Auditory sensitivity to aerial sounds i n a specimen of Amphisbaena alba under normal conditions (solid line) and after interrupting the extracolumella (broken line). The curves represent for various frequencies the sound pressure, i n decibels relative to 1 dyne per sq cm, required to produce a cochlear potential of 0.1 pv. I n this and the following figures, frequency is in cycles per second (Hz). 100 1000 Frequency 10,000 Fig. 7 Sensitivity curves in a second specimen of Amphisbaena alba under normal condi- tions (solid line) and after two successive interruptions of the extracolumella, one in the anterior portion of this rod (long-dashed curve) and the other more posteriorly (short-dashed curve). to 2000 Hz and averaging 33 dB. For tones In a second specimen (solid-lined curve above this point at which observations of figure 7), the sensitivity was greatest could be made, there were no significant in the range from 100 to 290 Hz and fell changes. off fairly regularly for higher tones. When 24 CARL GANS AND ERNEST GLEN WEVER the extracolumella was first cut at an an- terior position (long-dashed curve), and then at a more posterior site (short- dashed curve) the sensitivity fell progres- sively. Apart from one inversion, the sec- ond of these impairment curves lies above the first, showing that sensitivity is re- lated to the intact length of the extra- columella. 2. Amphi sbaena darwi ni trachura. The sensitivity of a specimen of Amphi sbaena darwi ni trachura with the sound source placed forward on the lower jaw is rea- sonably uniform up to 600 Hz, after which the curve rises sharply showing a loss of sensitivity, and then falls rapidly to the point of best response at 1000 Hz (fig. 8). The sensitivity decreases for the upper frequencies. Even in the low-tone region the sensitivity is slight. A second sound probe position near the center of the sec- ond infralabial gave somewhat greater sensitivity (fig. 9). The form of the curve is similar except that the sharp rise at 100 1000 Frequency 10, 000 Fig. 8 A normal sensitivity function i n a specimen of Anzphisbflena dwwi ni t mchzi m (left ear). + 60 - 20 Frequency Fig. 9 Sensitivity functions for the right ear of the same specimen as i n the preceding figure, under normal conditions (solid line) and after cutting the extracolumella (broken line). EAR AND HEARI NG I N AMPHI SBAENI A (REPTI LI A) 25 700 Hz is absent. The extracolumella was then severed just anterior to its connec- tion with the stapes, so that nearly all of this element was disconnected. The result was a large loss of sensitivity, which for the tones up to 1000 Hz averaged 40 dB and for higher tones up to 7000 Hz aver- aged 23.5 dB. 3. Amphisbaena manni. The best sen- sitivity of Amphisbaena manni is between 600 and 1500 Hz (fig. 10); i t declines fairly rapidly for lower and higher tones. I n a second specimen (fig. ll), the form of the curve is similar, but the most sen- sitive region extends from 500 to 1000 Hz, and the degree of sensitivity is a little Frequency Fig. 10 Sensitivity function for a specimen of Amphisbtrena manni , under normal conditions. 1 2 3 4 5 6 7 8 9 1 2 3 4 5 6 7 8 9 1 Frequency Fig. 11 Sensitivity functions for another specimen of Amphisbnena manni , under normal conditions (solid line) and after interrupting the extracolumella (broken line). 26 CARL GANS AND ERNEST GLEN WEVER less. When the extracolumella was severed fig. 12), maximum sensitivity occurred at just anterior to its connection to the 500 Hz, which was lower than for aerial stapes, the loss of sensitivity (broken stimulation. The most sensitive zone was curve) averaged 41 dB for tones up to also more restricted, but the level of sen- 2500 Hz and 20.3 dB for higher tones. sitivity was otherwise similar to that with I n response to a vibrating needle ap- aerial stimulation. The extracolumella plied to the side of the face (solid line, was then severed just anterior to its con- 1 2 3 4 5 6 7 8 9 1 2 3 4 5 6 7 8 9 1 Frequency Fig. 12 Sensitivity functions for a specimen of Amphisbaena manni under stimulation with mechanical vibrations applied through a needle to the skin surface at the side of the head, for normal conditions (solid line) and after severing the extracolumella (broken line). + 40 - 40 Fig. 13 Normal sensitivity curves for aerial sounds i n a specimen of Chirindia langi at two body temperatures as indicated. EAR AND HEARING IN AMPHISBAENIA (REPTILIA) 27 nection to the stapes, and the measure- ments repeated (dashed line) with only slight effect on sensitivity. 4. Chirindia langi. Two curves for a specimen of Chirindia langi, taken respec- tively at 24.4"C and 29.4"C, are shown in figure 13. At the lower temperature the curve reaches its lowest point (greatest sensitivity) at 500 Hz, whereas at the higher temperature this point is in the higher frequencies, around 3000 Hz. There is also a shift i n the two ends of the curve; at the higher temperature the low end is raised and the high end is lowered. This type of modification of audi- tory sensitivity was shown to be a regular occurrence in extensive experiments on lizards by Werner ('68). 5. Cynisca leucura. The curve from Cynisca leucura indicates that the great- est responsiveness is at 300 and at 500 Hz, with a rapid decline to 800 Hz, then a leveling and a further decline for the upper frequencies (fig. 14). Fig. 14 conditions. An aerial sensitivity curve for a specimen of Cynisca leucura under normal Frequency Fig. 15 conditions. An aerial sensitivity curve for a specimen of Zygaspis violacea under normal 28 CARL GANS AND ERNEST GLEN WEVER 6. Zygaspis uiolacea. The greatest sensitivity of a specimen of Zygaspis vi- olacea occurred at 300 and at 500 Hz with a sharp loss in the intermediate zone as in Cynisca (fig. 15). The sensitivity decline at higher frequencies is typical of Phese animals. 7. Monopeltis c . capensis. Best sen- sitivity for a specimen of Monopeltis c. capensis occurred at two points in the low-frequency region, 200 and 500 Hz, with lesser responses between, and with +40 the usual decline in the high frequencies (fig. 16). In another specimen, the normal curve (solid line) also shows two points of great sensitivity, at 200 and 600 Hz (fig. 17). Severing of the extracolumella again led to a more than 20 dB loss of sensitivity (broken curve in fig. 17). Another experiment documented the directional characteristics of the amphis- baenid ear. The animal was stimulated via sound tube and response recorded from - 40 100 1000 Frequency 10,000 Fig. 16 An aerial sensitivity curve for a specimen of Monopel t i s c. capensi s, under nor- mal conditions. 100 1000 Frequency 10,000 Fig. 17 Sensitivity curves for a second specimen of Monopel t i s c. capensi s under normal conditions (solid line) and after interrupting the extracolumella (broken line). EAR AND HEARING IN AMPHISBAENIA (REPTILIA) 29 both saccules. The results obtained from the right ear are shown by the solid line of figure 18 and those from the left ear by the broken line. In general (all except 2 points), there is a clear difference in favor of the ipsilateral ear. A greater dif- ferential response occurred when stimula- tion was carried out by placing the sound tube at a distance of 5 cm from the right side of the face. 100 8. Trogonophis wiegmanni. A speci- men of Trogonophis wiegmanni showed excellent sensitivity with the maximum falling between 300 and 500 Hz (fig. 19), in an experiment extended down to 40 Hz. Figure 20 compares results for aerial and vibratory stimulation in a second speci- men. The aerial curve is much like the previous one, though the region of good sensitivity is broader. The curve for vibra- 2 3 4 5 6 7 8 9 1 9 1 1000 10,000 Frequency Fig. 18 Aerial sensitivity curves obtained i n a specimen of Monopeltis c. capensis with an electrode applied to the left saccule (solid line) and an electrode applied to the right sac- Cole (broken line). For both sets of observations the sound tube was close to the surface of the face on the left side. + 40 3 +20 2- 2 0 P u m T2 m a 0. -u S 3 & 5; - 20 - 40 100 1000 Frequency Fig. 19 A sensitivity curve for a specimen of Trogonophis wiegmnnni. 100 30 CARL GANS AND ERNEST GLEN WEVER 2 3 4 5 6 7 8 9 1 2 3 4 5 6 7 8 9 1 4 5 6 7 8 9 1 Frequency 1000 10,000 Fig. 20 Sensitivity curves obtained i n another specimen of Trogonophis wiegmnnni with aerial stimulation (solid line) and with vibratory stimulation (broken line). The ordinate scale represents decibels relative to a sound pressure of 1 dyne per sq cm for the aerial stimuli and decibels relative to an amplitude of 1 mp for the mechanical stimuli. Fig. 21 A sketch of the left side of the head of a specimen of Trogonophis wiegmcinni showing the scalation and the locations used for stimulating with a vibrating needle. As figure 2 has shown, the extracolumella runs from a deep connection with the stapes (around positions 18-19) forward along the infralabials (to about position 15). Its fibrous extension continues to about position 14. tory stimulation is irregular, and shows two regions of good sensitivity, one at 500 Hz and another at 1500-2500 Hz. Further tests explored the reception of vibratory stimuli by the lateral surface of the head. The blunted end of the vibrat- ing needle was brought into firm contact with the supra- and infralabilas and with those segments posteriorly in line well be- yond the position of the stapes (see num- bered circles in fig. 21). At each place the contact pressure of the needle was ad- justed to give a maximum response at a frequency of 400 Hz, and the stimulus amplitude varied to produce a constant Figure 22 shows the amplitudes, in de- cibels relative to an arbitrary zero level output of 0.2 pv. EAR AND HEARING I N AMPHISBAENIA (REPTILIA) 31 - 10 5 P 2 11 0 -L > 0. required for the standard response, as a function of distance from the tip of the jaw for both the lower jaw (solid line) and the upper jaw (broken line). In this graph, unlike the others, the lower the curve the less the sensitivity. It is seen that along both upper and lower jaws there is no significant variation of sensi- tivity with position until the vibrator reaches the zone where the extracolum- ella lies deep to the jaw muscles; sensi- tivity then sinks sharply to a new and much lower level. 9. Bipes biporus. Our results for Bipes bi porus are to be regarded as preliminary, - - I I I I because at the time of these tests we were unaware that the extracolumella termi- nated posterior to the quadrate. Presum- ably, the sound tube for aerial stimula- tion should not have been located adjacent to the anterior portion of the head. Four animals tested with aerial sounds showed a range of sensitivity represented by the two curves of figure 23. The solid-lined curve indicates a level of best sensitivity that is about the same as observed in other amphisbaenians, but differs in that this falls in the high tones, around 3000 Hz. The dashed curve shows rather little sensitivity, with irregularities, and does + 40 +20 P V al D 2 $ 0 m al a U S 3 L 64 -20 - 40 100 1000 10,000 Frequency Fig. 23 Aerial sensitivity curves i n two specimens of Bipes biporus, chosen to show the range obtained. 32 CARL GANS AND ERNEST GLEN WEVER v) - 3+20 V a, 6 1 ' 0 3 Q E, : -20 7, 0 L +- .- > 1+40E - 40 100 2 3 4 5 6 7 8 9 1 2 3 4 5 6 7 1000 Frequency 10,000 Fig. 24 Sensitivity curves obtained for vibratory stimulation in a specimen of Bipes biporiis with the needle applied to the side of the face near the stapes ((solid line) and to the front region of the lower j aw (broken line). not agree in form with the other curve. Whether these differences are due to the manner of presentation of the sounds or to other conditions of the tests, or repre- sent actual individual variations, is dif- ficult to say. Figure 24 shows two curves obtained by vibratory stimulation, one (solid line) by placing the vibrating needle on the side of the face in the region of the stapes and the other (broken line) by placing it on the front edge of the lower jaw. The two functions have much the same form, but, in general, stimulation near the stapes shows the greater sensitivity. The level of sensitivity is similar to that found by this method in other species. DI SCUSSI ON The observations clearly show that am- phisbaenians are sensitive to aerial sounds, and that these sounds are con- ducted to the inner ear by way of the skin along the lower jaw, the extracolumella, and the stapes. In amphisbaenians gener- ally the extracolumella projects anteriorly to provide a dermal connection lateral to the mandible. Bi pes bi por us represents an exception since its extracolumella is short and attaches to the skin in the immediate vicinity of the quadrate. In the members of the family Amphis- baenidae (genera Amphi s bae na, Chi r i ndi a, Cyni s ca, Monopel t i s, and Zygas pi s ) , the extracolumellar rod extends far forward into the skin layer, lateral to the lower jaw. I n Tr ogonophi s , the extracolumella does not itself run as far forward as a cartilaginous structure but is extended by a strong band of connective tissue that evidently serves the same purpose. Gans ('60) showed that this connective tissue apparently serves as the starting point for the ossification of the extracolumella seen in the trogonophid genera Di pl omet opon and Agamodon. Here the tissue has be- come ossified, and many of the connec- tions run to the upper rather than the lower lip. Most amphisbaenian ears have a region of greatest sensitivity in the low frequen- cies, usually between 300 and 700 Hz. This zone often contains two points of high sensitivity separated by a less sensi- tive zone. I n Amphi s bae na manni the best region seems to be slightly higher (be- tween 700 and 1000 Hz), and it may be still higher in Bi pes bi por us . Most of the amphisbaenian species yielded sensitivity curves whose maximum was approximate- ly -20 dB though there was considerable variation and Tr ogonophi s repeatedly reached - 40 dB. The levels of cochlear potentials may be compared with similar results on other squamates, which also showed large vari- EAR AND HEARING IN AMPHISBAENIA (REPTILIA) 33 ation; a rough mean for lizards is around -40 dB (for a response of 0.1 ~v ) , but many species are far more sensitive, up to -60 dB or better. Various species of the Sceloporus group give lower maxima around -20dB although their regions of good response are broader than those found in amphisbaenians (Wever and Peterson, '63, and Wever unpublished). The chameleons so far studied vary great- ly according to the special mechanisms that have been developed as substitutes for the absent tympanic membrane and round window; usually the sensitivity is relatively poor but in some species may approach -20 dB (Wever, '68, '69a,b). The general shapes of the curves are nevertheless remarkably similar in lizards and amphisbaenians. The sensitivity of some species of snakes agrees with those determined for amphis- baenians in giving maxima around -20 dB in the low frequencies from 200 to 700 Hz. The sensitivities of snakes do, however, drop off most rapidly above 700 Hz (Wever and Vernon, '60). Among non-squamate reptiles, turtles always exhibit the greatest sensitivity in the low range. Their curves are generally quite sharply peaked here with a particu- larly rapid decline in sensitivity for high tones. The maximum sensitivity level of turtles varies widely, with some species having the same magnitude as amphis- baenials while others reach sensitivities up to -60 dB. Crocodilians are generally more sensitive showing both a broader range of good sensitivity and maxima around -60 dB (Wever and Vernon, '57). Experiments well document the differ- ential function of the middle ear appara- tus. The sensitivity, particularly in the low range (below 1000 Hz) drops drastic- ally (by 20 to 30 dB) whenever the extra- columella is severed (figs. 6, 7, 9, 11, 17). Supplementary and sequential transec- tions of the extracolumella show (cf. figs. 3, 7) that the length remaining intact re- lates to the level of absolute sensitivity, which in some cases dropped by as much as 40 dB. The experiments with the mechanical vibrator show that the response to signals applied anywhere along the head (1) is fairly constant regardless of the site of application and (2) does not diminish when the extracolumella is transected. These experiments (on Amphisbaena alba and A. manni ) suggest that the extracol- umella functions in transmitting aerial sounds or other low-displacement vibra- tions from the anterior labial surface to the inner ear. From these observations, the signals under usual life conditions could be aerial sounds or vibrations of the substratum. Probably both are utilized under suitable conditions. The structure of the labial system should permit re- sponse to subterranean vibration when the face is held in contact with the tun- nel wall. The experiments on Monopeltis capen- si s document that at least this species has a small but significant differential sensitivity to asymmetrical sounds, a sen- sitivity which is probably enhanced by central bilateral inhibition. This then suggests an explanation for the earlier observation (Gans, '60) that these animals appear to discover their prey by sound. It also opens the way to a variety of be- havioral analyses. LITERATURE CITED Baird, I. L. 1970 The anatomy of the reptilian ear. I n: The Biology of the Reptilia. Vol. 2b. C. Gans and T. S. Parsons, eds. Academic Press, London, pp. 192-275. Gans, C. 1960 Studies on amphisbaenids (Am- phisbaenia: Reptilia). 1. A taxonomic revision of the Trogonophinae and a functional inter- pretation of the amphisbaenid adaptive pattern. Bull. Amer. Mus. Nat. Hist., 119: 129-204. A checklist of the recent amphis- baenians (Amphisbaenia: Reptilia). Bull. Amer. Mus. Nat. Hist., 135: 61-106. 1968 Relative success of divergent pathways i n amphisbaenian specialization. Amer. Nat., 102: 345-362. Gans, C., and J . J . Bonin 1963 Acoustic activ- ity recorder for burrowing animals. Science, 140: 398. Versluys, J . 1898 Die mittlere und aussere Ohrsphare der Lacertilia und Rhynchocephalia. Zool. J b', Abt. Anat., 12: 161406. Werner, Y. L. ,1968 The function of the i nner ear i n lizards: temperature effects i n Gekko- noidea and I guanidae. Proc. 11th Ann. Meet- ing, SOC. Study of Amphib. Rept., J . Herpetol., . 2: 178. Wever, E. G. 1939 The electrical respogses of the ear. Psychol. Bull., 36: 143-187. 1966 Electrical potentials of the coch- lea. Physiol. Revs., 46: 102-127. 1968 The ear of the chameleon: Chn- miicleo s enegal emi s and Chiimcleleo qziilensis. J . Exp. Zool., 168: 423436. 1969a The ear of the chameleon: the round window problem. J . Exp. Zool., 171: 1-6. 1967 34 CARL GANS AND ERNEST GLEN WEVER 1969b The ear of the chameleon: Clzn- Wever, E. G., and J . A. Vernon 1957 Auditory mneleo koknelii and Ckamcteleo jnchsoni. J . responses i n the spectacled cai man. J . Cell. and Exp. Zool., 171: 305312. Wever, E. G., and E. A. Peterson 1963 Audi- ___ 1960 'The problem of hearing i n snakes. tory sensitivity i n three iguanid lizards. J . Audi- tory Res., 3: 205-212. Comp. Physiol., 50: 33-340. J . Auditory Res., 1 ; 77-83.