elastic modulus of cetacean auditory ossicles
TRANSCRIPT
Elastic Modulus of Cetacean AuditoryOssicles
ANDREW A. TUBELLI,1* ALEKS ZOSULS,1 DARLENE R. KETTEN,2,3
AND DAVID C. MOUNTAIN1
1Department of Biomedical Engineering, Hearing Research Center, Boston University,Boston, Massachusetts
2Department of Biology, Woods Hole Oceanographic Institution, Marine Research Facility,Woods Hole, Massachusetts
3Department of Otology and Laryngology, Harvard Medical School, Massachusetts Eye andEar Infirmary, Boston, Massachusetts
ABSTRACTIn order to model the hearing capabilities of marine mammals (ceta-
ceans), it is necessary to understand the mechanical properties, such aselastic modulus, of the middle ear bones in these species. Biologically realis-tic models can be used to investigate the biomechanics of hearing in ceta-ceans, much of which is currently unknown. In the present study, theelastic moduli of the auditory ossicles (malleus, incus, and stapes) of eightspecies of cetacean, two baleen whales (mysticete) and six toothed whales(odontocete), were measured using nanoindentation. The two groups ofmysticete ossicles overall had lower average elastic moduli (35.2 6 13.3GPa and 31.6 6 6.5 GPa) than the groups of odontocete ossicles (53.3 6 7.2GPa to 62.3 6 4.7 GPa). Interior bone generally had a higher modulus thancortical bone by up to 36%. The effects of freezing and formalin-fixation onelastic modulus were also investigated, although samples were few and noclear trend could be discerned. The high elastic modulus of the ossicles andthe differences in the elastic moduli between mysticetes and odontocetes arelikely specializations in the bone for underwater hearing. Anat Rec, 297:892–900, 2014. VC 2014 Wiley Periodicals, Inc.
Key words: cetacean; ossicles; middle ear; hearing; elasticmodulus
INTRODUCTION
Information on the mechanical properties of bone inthe marine mammal (cetacean) auditory system is lim-ited. Cetaceans possess an auditory system that is ana-tomically similar to that of terrestrial mammals but ismorphologically different, being highly specialized forunderwater use (Ketten, 2000). Cetacean ossicles aremore massive than their terrestrial counterparts; addi-tionally, the anterior process of the malleus forms a syn-ostosis with the tympanic bone, a characteristic alsofound in few terrestrial mammals such as bats, shrews,and mice (Fleischer, 1978). In vivo measurement of hear-ing capability in cetaceans is restricted by marine mammalprotection laws, and behavioral and electrophysiologicalmethods of obtaining hearing information tell us littleabout the mechanics of the auditory system. A way around
this issue is to simulate sound reception with biologicallyrealistic models (e.g. Gan et al., 2004; Elkhouri et al.,2006; Cranford et al., 2008; Homma et al., 2009; Tubelliet al., 2012).
Grant sponsor: Joint Industry Program and the LivingMarine Resources program; Grant number: N00244-100-0053.
*Corresponding to: Andrew A. Tubelli, Hearing ResearchCenter and Department of Biomedical Engineering, Boston Uni-versity, 44 Cummington St., Boston, MA 02215. Fax: 617-353-6766. E-mail: [email protected]
Received 12 January 2013; Revised 9 September 2013;Accepted 15 January 2014.
DOI 10.1002/ar.22896Published online 13 February 2014 in Wiley Online Library(wileyonlinelibrary.com).
THE ANATOMICAL RECORD 297:892–900 (2014)
VVC 2014 WILEY PERIODICALS, INC.
Transmission of sound through the cetacean middleear is not well understood. Biophysical models providethe best evidence for how sound can travel through themiddle ear. These models are highly dependent on themechanical property values of tissues that are used asparameters. Elastic modulus, a key mechanical propertythat is used to describe the stiffness of a material, is onesuch mechanical property. Elastic modulus is defined asthe ratio of stress per unit area that a material under-goes divided by the resultant strain (deformation). Inorder to calculate the frequency response of cetaceans asaccurately as possible (Tubelli et al., 2012), the mostrealistic values of elastic modulus must be used.
There are few publications that describe the mechani-cal properties of the ear bones, especially of cetacean earbones. The research groups that have measured the elas-tic modulus of cetacean ear bones are summarized inTable 1. Each study used a different method to measureelastic modulus and each obtained significantly differentresults. There are only a couple known studies that mea-sure the material properties of terrestrial mammalianossicles, in human (Speirs et al., 1999) and rabbit (Soonset al., 2010). These values are also listed in Table 1. Thevalues obtained for these two terrestrial species are con-siderably less than those of cetaceans. This difference inossicular stiffness between terrestrial and aquatic mam-mals, when combined with the anatomical similaritiesand morphological differences, suggests a similar yetmodified mechanism for sound reception in cetaceans.
The ossicles of the middle ear are the three smallestbones found in all mammals. The ossicles are also irreg-ularly shaped. Figure 1 shows anatomical reconstruc-tions of the intact ossicular chain in a mysticete (baleenwhale) and an odontocete (toothed whale) species. As aresult, conventional mechanical tests such as bendingand compression tests, which require regularly shapedsamples of a certain size, are not suitable to measurematerial properties, such as elastic modulus, of theossicles. The tympanic bones are much larger than theossicles, thereby allowing bending tests to be used inCurrey (1979) and Zioupos et al. (1997). Speirs et al.
(1999) used compression testing on the ossicles but notedthe artifact associated with using such small samples.Lees et al. (1996) used sonic velocity and density meas-urements to calculate elastic moduli with the caveatthat the results were to be regarded as representative ofelasticity rather than regarded as precise values.Instead, nanoindentation is a more suitable method ofelastic modulus measurement. Nanoindentation uses aspecialized tip that indents a material of interest andcan determine material properties at the level of themicrostructure. Nanoindentation has been used to mea-sure the elastic modulus in a number of studies of otherbones (e.g. Zysset et al., 1999; Fan and Rho, 2003; Don-nelly et al., 2005). Additionally, since bone is heterogene-ous with a complex structure, nanoindentation canmeasure the modulus of different regions of the bonerather than treating the modulus as a bulk parameter.
The goal of this study was to measure the elastic mod-uli of the ossicles via nanoindentation for eight cetaceanspecies: minke whale (Balaenoptera acutorostrata), finwhale (Balaenoptera physalus), common dolphin (Delphi-nus delphis), short-finned pilot whale (Globicephalamacrorhynchus), Atlantic white-sided dolphin (Lageno-rhynchus acutus), harbor porpoise (Phocoena phocoena),striped dolphin (Stenella coeruleoalba), and bottlenosedolphin (Tursiops truncatus). The sensitivity of theseproperties to dehydration and to fixation by freezing orformalin was also examined.
MATERIALS AND METHODS
Sample Preparation
A total of 45 ossicles were obtained from 17 cetaceanspecimens under a permit for scientific research on ceta-cean tissues issued to D. Ketten. Table 2 lists all speci-mens used. Due to the nature of some of the deaths, notall information could be collected. Specimens rangedfrom calf to adult. Full maturation of the cetacean tym-panic bulla occurs 1 year after birth (de Buffr�enil et al.,2004), so most animals were presumed to have fullydeveloped ossicles, with the exception of the bottlenose
TABLE 1. Average value of elastic modulus (E) measured for auditory bones
Source Bone MethodAverage E 6 standard
deviation (GPa)
Cetacean BullaCurrey (1979) B. physalus tympanic bulla Three-point bending 31.3 6 0.97Lees et al. (1996) B. physalus tympanic bulla Estimation from sonic velocity
and density51.9
B. physalus malleus Estimation from sonic velocityand density
56.5
B. physalus periotic Estimation from sonic velocityand density
52
T. truncatus periotic Estimation from sonic velocityand density
73.6
Zioupos et al. (2000) B. physalus tympanic bulla Three-point bending 33.5Zioupos (2005) B. physalus tympanic bulla Nanoindentation 40.2 6 5.1
Terrestrial ossiclesSpeirs et al. (1999) Human malleus/incus Compression 2–3Soons et al. (2010) Rabbit malleus/incus Two-needle indentation 16.4 6 2.8
Caput malleus 16.3 6 2.9Collum malleus 15.6 6 1.8Corpus incudis 16.8 6 3.1Crus longum incudis 17.1 6 3.8
ELASTIC MODULUS CETACEAN AUDITORY OSSICLES 893
dolphin calf. This animal may or may not have had fullydeveloped ossicles. There were no known pathologies asso-ciated with the auditory bones in any of the specimens.
Four ears were preserved via formalin fixation andone ear was preserved via freezing. The approximateduration of time that each ear was preserved is indi-cated in Table 1. Within the specified duration, the earsmay have been thawed and refrozen or taken out of andplaced back into formalin. This would have been withina 48-h timeframe for either observation or nondestruc-tive experimentation. The remaining non-preserved earswere chilled at 4�C without preservation and disarticu-lated within several days after animal death.
The bones were cleaned of soft tissue with dental toolsand then dehydrated in air. The bones were then embed-ded in underwater epoxy (Sea Goin’ Poxy Putty, Perma-lite Plastics Inc., Costa Mesa, CA) onto a 15 mmmagnetic specimen disc, cured in an oven at 38�C for 1 hand then left at room temperature to cure for at least 24h more. Heating is not expected to have any effect on thebone samples. Collagen in bone does not start to denatureuntil 60�C; furthermore, collagen denaturation has littleeffect on elastic modulus up to 200�C (Wang et al., 2001).
Once the epoxy cured, the samples were ground flatusing a miniature milling machine (Micro-Mark, BerkeleyHeights, NJ). Samples were kept hydrated with distilledwater during milling. The resulting surface was polishedusing a sequence of silicon carbide abrasive paper withprogressively finer grit sizes (400, 600, 1,500) and fin-ished with 0.5 and 0.1 micron diamond paste on low nappolishing cloth. The surface was washed off with distilledwater and blotted dry. Atomic force microscopy was usedto verify that the polishing technique produced a smoothenough surface. The resulting RMS roughness for a pol-ished specimen was on the order of 2 to 23 nm.
Nanoindentation
The indentation was done using a Triboindenter(Hysitron Inc., Minneapolis, MN). The system uses theOliver-Pharr method (Oliver and Pharr, 1992) to deter-mine the reduced modulus, the effective modulus of boththe indenter and the specimen together, via the unload-ing portion of a force-displacement curve. The reducedmodulus is calculated using Eq. (1), where S is the con-tact stiffness, calculated as the slope of a power-law fitover a region of 95% to 20% of the unloading force-displacement curve, and Ac is the contact area.
Er5
ffiffiffi
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The elastic modulus of the sample can then be calcu-lated from the reduced modulus using Eq. (2), where thePoisson’s ratio and the elastic modulus of the tip (mtip
and Etip, respectively) are known. A diamond Berkovichtip was used (approximately 150 nm radius) for indenta-tion, with mtip equal to 0.07 and Etip equal to 1,140 GPa.
1
Er5ð12v2
sampleÞEsample
1ð12v2
tipÞEtip
(2)
The Poisson’s ratio of the sample (msample) wasassumed to be 0.3, a value commonly used for bone, for
Fig. 1. Anatomical reconstructions of (a) a minke whale and (b) abottlenose dolphin ossicular chain, medial view. Orange: segment oftympanic bone, red: malleus, green: incus, blue: stapes. ap: anteriorprocess, mb: manubrial region of the body of the malleus, cb: centralregion of the body of the malleus, mh: head/articular region of themalleus, bo: body of the incus, lp: long process of the incus, sp: shortprocess of the incus, he: head of the stapes, ba: base of the stapes,ac: anterior crus, pc: posterior crus. The short process of the minkewhale incus is occluded. The black scale bar in each panel corre-sponds to 5 mm in length.
894 TUBELLI ET AL.
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ELASTIC MODULUS CETACEAN AUDITORY OSSICLES 895
this analysis. Varying the value of Poisson’s ratiobetween 0.2 and 0.4 has little effect on the elastic modu-lus (Rho et al., 1997).
Nanoindentation was performed at various regions onthe flat polished surface of the bone samples at roomtemperature. All indents were load-controlled and useda load/unload rate of 600 mN/s. A maximum load of 6,000mN was used to provide sufficient depth penetration andthereby prevent surface roughness from affecting thedata (Donnelly et al., 2005); indentation depth rangedfrom approximately 300 to 500 nm. A hold time of 30 swas used to account for the viscoelastic properties ofbone (Fan and Rho, 2003). Calibration of the data wasdone using a quartz sample (reduced modulus of 69.9GPa).
Regions were chosen based on anatomical location.These locations are shown in Fig. 2 with the exception ofthe short process of the incus. More detailed informationon the macrostructure of cetacean ossicles can be foundin sources such as Fraser and Purves (1960), Fleischer(1978), Solntseva (2007), and Mead and Fordyce (2009).Since the ossicles were oriented randomly within theepoxy, indents were made only in regions that could beseen on the final polished surface. Two categorizations ofbone were made for ossicular measurements: an outer(“cortical”) layer and an inner, less uniformly calcifiedlacunate core (“interior”). In humans, the ossicles arecomposed of a mixture of enchondral and intrachondrialbone with a dense outer cortex and a less dense interiorcharacterized by scattered lacunae (Gulya, 2007). Thus,the distribution of these bone types in cetacean ossicleswas found to be morphologically similar to that observedin human ossicles (Kirikae, 1960; Galioto and Marley,1965). These distinct bone regions (here referred to as"cortex" and "interior") were clearly distinguishable inthe histological sections and are labeled in Fig. 2. It isimportant to note that the predominant bony elementsof the ossicles, which include particularly intrachondrialbone, are quite unlike those of long bones, such as thefemur, which are typically used for bone mechanicalproperty measurements. Indentation of the corticalregion was not always possible because the cortex can bevery thin in some portions. Indentation near the bone-epoxy boundary was avoided. Indentation regions weretypically defined as a square area of 0.64 mm2. A total of16 indents were performed in each indentation region.Spacing between individual indents was not uniform,but all indents were spaced greater than 5 lm apart.
In the majority of indentations performed, there was asingle indentation region per anatomical region;
Fig. 2. Histological slices of (a) a striped dolphin malleus, (b) a com-mon dolphin incus, and (c) a harbor porpoise stapes. The slices arerepresentative of the surface of a nanoindentation sample. Histologywas completed by first decalcifying the sample in EDTA and thenembedding the sample in celloidin. The sample was sectioned at 25lm, and every 10th section was stained with hematoxylin and eosinand mounted on glass slides. Bone types and anatomical regions dis-tinguished in experiments are labeled, with the exception the shortprocess of the incus (not shown). ap: anterior process, mb: manubrialregion of the body of the malleus, cb: central region of the body of themalleus, mh: head/articular region of the malleus, bo: body of theincus, lp: long process of the incus, he: head of the stapes, ba: baseof the stapes, ac: anterior crus, pc: posterior crus. The black scale barin each panel corresponds to 1 mm in length.
896 TUBELLI ET AL.
however, there were four cases in which there were twoindentation regions per anatomical region. The interiormanubrial region of a bottlenose dolphin malleus (T-tru53, left ear) was indented twice to test variability withinan anatomical region. The cortical region of the long pro-cess of the incus was indented twice in three species (B-acu 22, right ear; B-acu 23, right ear; G-mac 64, leftear). On the cut surface of the bone of these three sam-ples, the thick cortex could be seen flanking the interiorbone. One indentation region was assigned to the cortexon each side of the interior bone, again to test variabilitywithin an anatomical region.
All samples were kept dry until indented. The sam-ples were rehydrated in 0.9% saline a half hour beforeindentation. Rehydration of bone after drying was foundto have an insignificant effect on elasticity (Currey,1988).
RESULTS
There were a few samples where some data pointswere excluded from analysis. For the Atlantic white-sided dolphin, the long process of the incus exhibitedseveral large cracks from sample preparation thataffected the integrity of the region. For the bottlenosedolphin, suspected machine error occurred on the sameday as indentation of four regions, three incudal and onemallear. All indents from each of these regions wereignored during analysis.
There were four specimens that were formalin-fixedbefore indentation, two minke whale ears and two bot-tlenose dolphin ears. Both formalin fixed minke earshad ossicles that had a higher elastic modulus than theirfresh counterparts: malleus, 49% higher; incus, 32%higher; stapes, 12% higher. For the bottlenose dolphin,however, the malleus and incus moduli were 6% and33% lower, respectively, and the stapes modulus was11% higher. One specimen, a minke whale ear, was fro-
zen and thawed before indentation. The thawed incuswas 14% higher than its fresh counterpart and thethawed stapes was 11% lower than its fresh counterpart.Since the sample size was so low and variability amongmembers of the same species is uncharacterized, no cleartrend can be discerned for preserved bone.
Both interior and cortical regions were indented in 21ossicles, five of which were fixed. Formalin and frozenossicles did not show any difference in trend from thefresh ossicles. In 15 of the 21 ossicles, interior bone over-all had a higher elastic modulus than cortical bone(Fig. 3). In most of these ossicles, the average modulusof interior bone was 1% to 36% higher than the averagemodulus of cortical bone. The fin whale was a specialcase, where interior bone was twice as stiff as corticalbone. Looking at cortical vs. interior within specificregions, there are 15 cases of a statistically significantdifference between cortical and interior bone (unpairedt-test, P <0.05), 11 of which had a higher interior modu-lus, and 13 cases of no statistically significant difference.Statistically significant results were not correlated withany particular regions.
In the 12 ears in which all three ossicles wereindented, there were no consistent differences in averagestiffesses of the three ossicles within an ear.
In six out of eight cases in which the anterior processof the malleus was indented, it had the lowest modulusof any mallear region (Fig. 4). In five of those six cases,the anterior process was significantly lower than thenext highest modulus within the same bone (unpaired t-test, P <0.05). Only two mysticete mallei were indented,both minke whale specimens. The average anterior pro-cess elastic moduli for these two ears were 60% and 62%of the maximum elastic moduli for their respective mal-lei. For the odontocetes, the average anterior processelastic moduli were between 73% and 94% of the maxi-mum modulus for their respective mallei. No other trendwas observed for the mallear regions.
Fig. 3. Average elastic moduli and standard deviations for cortical and interior bone in each ossicle inwhich both were measured. (a) Mysticete and (b) odontocete bones. Results from bones that were fixedby formalin or freezing are marked. M: malleus, I: incus, S: stapes. Circles represent the mean, lines rep-resent the median, boxes correspond to the lower and upper quartiles, and whiskers correspond to thehigh and low values.
ELASTIC MODULUS CETACEAN AUDITORY OSSICLES 897
There were no consistent differences observed in theaverage elastic moduli for stapedial or incudal regionsacross specimens.
There were four ossicles in which there were twoindentation regions specified for an anatomical region(T-tru 53, manubrial region of the malleus, interior; B-acu 22, long process of the incus, cortical; B-acu 23, longprocess of the incus, cortical; G-mac 64, long process ofthe incus, cortical). The difference between each indenta-tion region within a single bone was significant for allexcept for the short-finned pilot whale incus (unpaired t-test, P< 0.05).
Figure 5 shows the average bulk elastic moduli andstandard deviations calculated for each species for freshears. These values assume homogeneous and isotropicbone within a species. Odontocete species had higherelastic moduli, ranging from 53.3 6 7.2 GPa to 62.3 6 4.7GPa. The short-finned pilot whale had the greatest elas-tic modulus value. The two mysticete species, fin whaleand minke whale, had elastic moduli of 35.2 6 13.3 GPaand 31.6 6 6.5 GPa, respectively. When fixed ears areincluded in the data set, the elastic modulus for theminke whale is raised to 34.8 6 7.2 GPa and the elasticmodulus for the bottlenose dolphin is lowered from58.0 6 5.3 GPa to 56.0 6 6.7 GPa.
DISCUSSION
Given the small amount of data for formalin-fixed andfrozen-thawed bone in this study, no specific conclusioncan be made regarding how preservation affects bone.Formalin-fixed bones exhibited higher elastic modulithan their fresh and frozen counterparts for the minkewhale yet showed lower elastic moduli for the bottlenosedolphin malleus and incus. Overall, other studies havefound that formalin fixation causes either no change ora slight insignificant increase in elastic modulus (Sedlinand Hirsch, 1966; Currey et al., 1995; Nazarian et al.,2009; Unger et al., 2010). As is the case with the one
frozen-thawed specimen in this study, freezing has beenshown to have no effect on the mechanical properties ofbone (Sedlin and Hirsch, 1966; Nazarian et al., 2009;Unger et al., 2010).
In general, interior bone had a somewhat higher elas-tic modulus than cortical bone, although the differenceswere often small. While there are studies that havemeasured a higher macroscopic trabecular (comparableto interior bone here) modulus (Roy et al., 1999; Turneret al., 1999; Hengsberger et al., 2002), the results are incontrast with the typical result that cortical bone has ahigher macroscopic elastic modulus than trabecular bone(Rho et al., 1993, 1999; Zysset et al., 1999; Bayraktar
Fig. 4. Average elastic moduli and standard deviations for mallear regions. Results from bones thatwere fixed by formalin are marked. G-mac: short-finned pilot whale, S-coe: striped dolphin, P-pho: harborporpoise, D-del: common dolphin, T-tru: bottlenose dolphin, L-acu: Atlantic white-sided dolphin, B-phy:fin whale, B-acu: minke whale.
Fig. 5. Average elastic moduli and standard deviations for eachcetacean species, fresh ears only.
898 TUBELLI ET AL.
et al., 2004). In some cases, the cortical layer was thin,making accurate measurements difficult. The differencemay also be attributed to the difference in structure ofthe bone compared with typically measured long bones.
No attention was given to indentations in different ori-entations. Lees et al. (1983) observed that differences inproperties were caused by inhomogeneity, not anisotropy.Currey (1979) likewise observed that the structure of thetympanic bulla was randomly arranged. The ossicles areassumed to have a similar random arrangement.
It is common to consider a bulk modulus for theossicles when modeling the middle ear (Homma et al.,2009; Cranford et al., 2010; Tubelli et al., 2012). Thisassumption implies that the ossicles behave as rigidbodies. Based on the study by Soons et al. (2010) on rab-bit ossicles, measured elastic moduli were not found tobe significantly different among regions, making bulkossicular moduli justifiable. However, the results of thecurrent study contradict these findings by showing alarge variability in elastic moduli of different anatomicalregions within a bone, even within the same anatomicalregion. These results suggest that ossicles should notnecessarily be treated as bulk masses having isotropicmaterial properties. This may be especially true forhigh-frequency animals, such as many dolphins are. Atvery high frequencies, the ossicles do not necessarilymove as rigid bodies. The tympanic bulla has many com-plex movements at audible frequencies for these species(Cranford et al., 2010).
The elastic moduli of cetacean ossicles are of particu-lar interest when considering their function. The middleear of cetaceans is stiffness-dominated (i.e. the combinedstiffnesses of the components have a greater effect onthe frequency response of the ear than mass) (Fleischer,1978; Miller et al., 2006). The anterior process of themalleus forms a synostosis with the tympanic bone. Thisconnection, along with the annular ligament of the sta-pes, is the major source of stiffness within the middleear of cetaceans (Fleischer, 1978; Miller et al., 2006). Incetaceans, the proposed axis of rotation of the ossicularchain runs directly through the anterior process(Fleischer, 1978; Hemil€a et al., 1999). The other side ofthe axis is the connection of the short process of theincus to the periotic bone via the posterior incudal liga-ment. The torsional stiffness is dominated by the ante-rior process. The anterior process is the portion of thebone that twists, with the bulk of the malleus awayfrom the axis (Fleischer, 1978). There is a greater needfor the anterior process to be comparatively less stiff sothat the malleus can rotate. The results from this studycan be explained by this concept. There were a total ofeight mallei that had their anterior processes indented.Six of those eight mallei showed their lowest elasticmodulus within the anterior process. No explanation canbe given for why the anterior process elastic modulus forthe remaining two mallei were larger than otherregions.
Overall, the elastic moduli measured for the ossiclesare comparatively similar to results from other studiesof cetacean ear bones, summarized in Table 1. The finwhale ossicles from this study were quantitatively simi-lar to the three-point bending results for the fin whaletympanic bone measured by Currey (1979) and Ziouposet al. (2000). Some of the odontocetes, particularly theshort-finned pilot whale, had unusually high moduli
which were quantitatively similar to that of the Mesoplo-don densirostris (Blainville’s beaked whale) rostrum(65.3 6 6.1 GPa), the most highly mineralized bone cur-rently known (Zioupos et al., 2005). The rostrum playsan important role in directing sound to the tympanicbulla in odontocetes (Norris, 1964; Cranford et al., 2008).Odontocete ossicles also have higher densities than mys-ticete ossicles, both of which have higher densities thanterrestrial mammalian ossicles (Nummela et al., 1999).The high mineralization, density, and stiffness of theossicles, tympanic bulla, and rostrum, show the presenceof a specialized pathway for sound reception. Whilethere is no proposed theory that mysticetes possess thesame mechanism of sound reception, the tympanic bullaregion could function similarly to odontocetes (Yamatoet al., 2012).
The differences in elastic moduli observed betweenmysticetes and odontocetes provide further context forthe proposed hearing differences between the twogroups. A stiffer ear typically corresponds to higher fre-quency hearing (Fleischer, 1978; Miller et al., 2006).While the actual hearing ranges for mysticetes areunknown, the range is likely to at least contain the fre-quencies of vocalizations. Mysticetes generally havelower-frequency vocalizations than odontocetes, whichemploy high-frequency echolocation. Myticetes are alsogenerally much larger in body size, which is correlatedwith a more massive auditory bulla, and subsequentlymore massive ossicles, which scale isometrically (Num-mela et al., 1999). Larger mass usually correlates withlower frequencies (Ketten, 2000). In any case, higherelastic moduli in the ossicles, combined with higher den-sities and mineralization, seem to be an evolutionaryspecialization in cetaceans functionally related to under-water auditory perception.
ACKNOWLEDGEMENTS
The authors thank the Living Marine Resources (LMR)program and the Joint Industry Program (JIP) for fund-ing this study. Specimens were generously provided bythe International Fund for Animal Welfare’s MarineMammal Rescue and Research Group and by Dr. Joy Rei-denberg of the Mount Sinai School of Medicine. Theauthors also thank Maya Yamato, Julie Arruda and ScottCramer at the Woods Hole Oceanographic InstitutionComputerized Imaging Facility for help in scanning andpreparing specimens and JiaPeng Xu of the Materials Sci-ence and Engineering department at Boston Universityfor his assistance in using the experimental equipment.
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