Expression of Emotional Arousal in Two Different Piglet Call Types

13 Pages·2015·0.96 MB·English

RESEARCHARTICLE
Expression of Emotional Arousal in Two
Different Piglet Call Types
PavelLinhart1*,VictoriaF.Ratcliffe2,DavidReby2,MarekŠpinka1
1 DepartmentofEthology,InstituteofAnimalScience,Prague,Czechia,2 SchoolofPsychology,University
ofSussex,Brighton,UnitedKingdom
* [email protected]
Abstract
Humansaswellasmanyanimalspeciesrevealtheiremotionalstateintheirvoice.Vocal
featuresshowstrikinglysimilarcorrelationpatternswithemotionalstatesacrossmamma-
lianspecies,suggestingthatthevocalexpressionofemotionfollowshighlyconservedsig-
nallingrules.Tofullyunderstandtheprinciplesofemotionalsignallinginmammalsitis,
however,necessarytoalsoaccountforanyinconsistenciesinthewaythattheyareacousti-
callyencoded.Hereweinvestigatewhethertheexpressionofemotionsdiffersbetweencall
typesproducedbythesamespecies.Wecomparetheacousticstructureoftwocommon
OPENACCESS
pigletcalls—thescream(adistresscall)andthegrunt(acontactcall)—acrossthreelevels
Citation:LinhartP,RatcliffeVF,RebyD,ŠpinkaM ofarousalinanegativesituation.Wefindthatwhilethecentralfrequencyofcallsincreases
(2015)ExpressionofEmotionalArousalinTwo
witharousalinbothcalltypes,theamplitudeandtonalquality(harmonic-to-noiseratio)
DifferentPigletCallTypes.PLoSONE10(8):
e0135414.doi:10.1371/journal.pone.0135414 showcontrastingpatterns:asarousalincreased,theintensityalsoincreasedinscreams,
butnotingrunts,whiletheharmonicityincreasedinscreamsbutdecreasedingrunts.Our
Editor:BrentonG.Cooper,TexasChristian
University,UNITEDSTATES resultssuggestthattheexpressionofarousaldependsonthefunctionandacousticspeci-
ficityofthecalltype.Thefactthatmorevocalfeaturesvariedwitharousalinscreamcalls
Received:March19,2015
thaningruntsisconsistentwiththeideathatdistresscallshaveevolvedtoconveyinforma-
Accepted:July21,2015
tionaboutemotionalarousal.
Published:August14,2015
Copyright:©2015Linhartetal.Thisisanopen
accessarticledistributedunderthetermsofthe
CreativeCommonsAttributionLicense,whichpermits
unrestricteduse,distribution,andreproductioninany
medium,providedtheoriginalauthorandsourceare Introduction
credited.
Vocalizationshavelongbeenconsideredasawindowintoemotionalstatesinbothhumans
DataAvailabilityStatement:Allrelevantdataare
andotheranimals[1].Indeed,emotion-specificcallscanbetriggeredbythestimulationof
withinthepaperanditsSupportingInformationfiles.
basicemotionalbrainsystems[2],whilevocalizationsalonecaninduceappropriatebeha-
Funding:ThisstudywassupportedbytheCzech
viouralandphysiologicalresponses[3].Becauseoftheircloseassociationwithemotional
ScienceFoundation(GA14-27925S)andMinistryof
expression,vocalizationsandtheirspecificparametershavereceivedsignificantscientific
AgricultureofCzechRepublic(MZERO0714).The
attentionaspossibleindicatorsofemotionalstatesinanimals[4,5].Recently,adimensional
fundershadnoroleinstudydesign,datacollection
andanalysis,decisiontopublish,orpreparationof approachhasbeenadvocatedincharacterisinganimalemotionalstates[6].Themostcommon
themanuscript. modelcontainstwocoreaffectdimensions:arousal,whichquantifiestheintensityofthe
response,andvalence,whichquantifiestheaversivenessofthesituation—rangingfromavery
CompetingInterests:Theauthorshavedeclared
thatnocompetinginterestsexist. positive(pleasurable)toaverynegative(aversive)emotionalstate.Thedimensionalapproach
PLOSONE|DOI:10.1371/journal.pone.0135414 August14,2015 1/13
ExpressionofEmotionalArousalinTwoDifferentPigletCallTypes
providesaframeworkinwhichthesignallingofemotionalstatescanbeeasilycomparedacross
differentspecies.
Whenanimalsareexposedtodifferentemotionallyloadedsituations,theacousticfeatures
oftheircallsvarywiththeassumedemotionalvalenceand/oremotionalarousal[4].Particu-
larly,thepatternofchangesofcertainvocalfeaturesalongthearousalgradationissurprisingly
uniformamongspecies(forexamplebothpitchandamplitudeincreasewitharousal),suggest-
ingcommonunderlyingmechanisms[4,5].Thisremarkableconsistencyislikelyduetothe
factthattheprinciplesofvocalproduction[7,8]andperception[9,10]areconservativein
mammalianspecies.Interestinglyhowever,therearesomecallparameters,suchascalldura-
tionortonality,thatdonotshowsuchaconsistencyacrossanimalspecies[4].
Oneaspectofemotionalsignallingwhichhasnotyetbeenconsideredisthatemotional
statesmaybeencodeddifferentlyacrosscalltypeswithintherepertoireofthesamespecies.
Whilecalltypeswithqualitativelydifferentacousticstructuresorfunctions,suchasalarmcalls
(e.g.[11]),contactcalls(e.g.[12]),ornon-situation-specificcallslikedogbarks[13]orpig
grunts(e.g.[14])haveallbeeninvestigated(reviewedby[4,5]),distresscallshavereceiveddis-
proportionatelygreaterscrutiny(e.g.[15,16,17]),perhapsbecauseanimalsvocalizemoreread-
ilyandconspicuouslywhentheyexperiencestress.However,becausecalltypesaredefinedby
overalldifferencesintheiracousticstructure,assumedtoreflectspecificfunctions[18],varia-
tioninemotionalstatemaybeencodedindifferentwaysacrosscalltypes.
Emotionsinducechangesintheautonomicnervoussystem.Changesinemotionalarousal
triggermodificationsinphysiologicalprocessessuchasrespiration,muscletensionandsaliva-
tion[19,20].Becausetheseprocessesaredirectlyinvolvedinvocalproduction,acousticparam-
etersthatarecontrolledbytheseprocessesshouldvaryconsistentlywiththeemotionalstateof
thesignallerindependentlyfromtheparticularcalltypeproduced.Forinstance,emotionally
drivenvariationinrespiratoryairflowmaysimilarlymodulatetheamplitudeofvocalisations
acrosscalltypes.Thus,anycalltypemightreflecttheemotionalstateoftheanimalindepen-
dentlyofitsfunctionorspecificstructure[14].However,fewstudieshavereportedhowdiffer-
entcalltypesmayencodethesameemotionalstateinmammals[14,21].
Wetestedthehypothesisthatacousticparameterswithintwofunctionallyandstructurally
differentcalltypeswouldchangeinasimilarwaywithincreasingemotionalarousalindomestic
pigletvocalisations.Pigletsemittwobasiccalltypes(Fig1).Highfrequencycalls(screamsand
squeals)aremainlygiveninsituationsofurgentthreat.Incontrastlowfrequencycalls(grunts)
arenotsituation-specificastheyareproducedacrossawiderangeofnon-emergencysocialsitu-
ationsaswellasurgentdistresssituations[22].Thesignallingfunctionofgruntsindistresscon-
textsremainsunclear,butassocialcallsitispossiblethattheycouldfacilitateindividual
recognition.Althoughtransitionalvocalisationsaresometimesproduced,screamsandgrunts
representwellseparatedcalltypes[22,23].Inadditiontotheirfunctionaldifferences,theproduc-
tionofbothcallsalsodiffers:gruntsareemittednasallywhilescreamsareemittedorally,though
bothareassumedtooriginatefromthevocalfolds[24].Thestructureofscreamcallshasalready
beenshowntoreflectarousalinpiglets(e.g.[16]),whilepotentialassociationsbetweengruntcall
structureandtheemotionalstateofthesignallerremainlargelyunexplored(butsee[14,16]).
Inthecurrentstudy,wedirectlycomparetheacousticstructureofpigletscreamsandgrunts
emittedinthesameemotionalandphysiologicalcontexts.Tocontrolforpotentialdifferences
inemotionalstatebetweenindividuals,wecomparescreamandgruntcallsthatwereemitted
inimmediatesuccessionacrossthreebroadlevelsofarousal,ensuringthatthetwovocalisa-
tionswereproducedwhenthepigletwasinanequivalentphysiologicalstate.Asthepurposeof
thisstudyistodemonstrategeneralprinciples,ratherthantodeterminespecificindicatorsof
particularemotionalstates,wefocusonasmallnumberofcommonlyinvestigatedacoustic
parameters:duration,amplitude,centralfrequencyandharmonic-to-noiseratio.Inaccordance
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ExpressionofEmotionalArousalinTwoDifferentPigletCallTypes
Fig1.Typicalscreamandgruntcallsandthecourseofthebacktestsession,depictingthecallingactivity(spectrogramandoscillogram),
strugglingpresence,andstrugglingforceover60softhebacktestonasecondbysecondbasis.
doi:10.1371/journal.pone.0135414.g001
withpreviousresearch,weexpectthatbothgruntsandscreamsshouldbelonger,moreintense,
harsher(lowerharmonic-to-noiseratio)andhavemoreenergyinhigherfrequencieswith
increasingarousal.
Methods
Ethicsstatement
TheexperimentwasapprovedbytheInstitutionalAnimalCareandUseCommitteeofthe
InstituteofAnimalScienceandtheCzechCentralCommitteeforProtectionofAnimals,Min-
istryofAgriculture(decisionMZe1244).Thisstudywasapartoflargerprojectinvestigating
theconsistencyofarousalsignallingacrossdifferentcontextsandsituations.Forthisreason,
werecordedpigletsinanumberofpositiveandnegativenaturalandexperimentalsituations
withdifferentlevelsofarousal.Thebacktestsituation(commonlyusedinpigstostudyindivid-
ualcopingstyles[25,26,27])correspondstoamildlystressfulsituationcausedbytheseparation
ofthepigletfromitslitterandbeingplacedinanunnaturalposition.Thestressinducedbythe
backtestisconsideredmildandisnotprolonged(thewholeprocedure,includinghandling,
lastsabout3minutes).Allofthepigletswerehealthywhentheywerereturnedtotheirhome
penimmediatelyafterthetest.Pigletswerecarefullyandgentlyhandledandtransportedina
straw-beddedcratetoreducestress.Nopainrelieverswerenecessary.
Animals
Piglets(LargeWhite×Landrace)werebornandkeptattheexperimentalfarmoftheInstitute
ofAnimalScience,Prague,Czechia.Sowsandpigletswerehousedin2.3mx2mstraw-bedded,
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ExpressionofEmotionalArousalinTwoDifferentPigletCallTypes
concretefloorpensequippedwitha‘walkaround’ellipsoidfarrowingcrate(2.3mx1.4m).
Sowswerefedastandardlactationdiettwiceaday.Waterwasavailableadlibitumseparately
forthesowandforthepiglets.Pigletswereweighedandindividuallymarkedatday1postpar-
tum(bodymarker).Weaimedtotesteightpigletsperlitter.Overall,12littersweresubjected
tobacktest.In3litters,wecouldonlytest6,6,and4pigletsperlitterduetosmalllittersizeor
verylowweight(<1.00kg)leadingto88pigletswithintermediateweights(mean+-sd=1.84
kg+-0.44kg;range:1.04kg–2.80kg)beingsubjectedtobacktest.Thenumberofmaleand
femalepigletswasbalanced(44males/44females).
Eachlitterwastestedwithinasinglepausebetweennursingepisodesstarting5minutes
afternursingended.Pigletsweretestedinarandomizedorder.Eachpigletwasgentlyremoved
fromthelitterandtransported(c.a.30s)inastraw-beddedbaskettoaseparatetestingroom
wherethebacktestwasperformed.Ingeneral,pigletsremainedcalmandsilentduringpen
removalandtransportation.Thewholeproceduretookabout3minutes.
Backtest
Weusedthebacktestasamodelsituation,whichiscommonlyusedinpigstostudyindividual
copingstyles.Forthepurposeofthisstudy,wewerenotinterestedincopingstylesofpiglets
butinsteadfocusedonbehaviourofthepigletsduringthebacktest.Weadaptedtheprocedure
fromHessingetal.[27]andRuisetal.[26].Thepigletwasgentlyputonitsbackonadigital
scaleandkeptinthispositionfor60sbyplacingonehandoverthepiglet’sthoraxandholding
itsrightforelegwiththeotherhand.Thehindlegsremainedfreetomove.Thesmallestforce
possiblewasusedtokeepthepigletinthesupinepositiontoensurethattheweightreadingson
thescalematchedthestrugglingforceofthepigletascloselyaspossible.Oneperson(PL)per-
formedallofthetesting.Althoughthescalereadingsareonlyapproximationsofthestruggling
force,theyallowedustounambiguouslydifferentiateperiodsofmoreversuslessintensestrug-
gling.Werecordedthenumber,latencyanddurationofstrugglingperiods.Onestrugglewas
definedasacontinuousseriesofwriggles.Subsequentstruggleshadtobeseparatedatleastby
1sofresttobetreatedastwoseparatestruggles.Thewholeprocedurewasvideotaped.We
notedwhetherthepigletwasstrugglingornotonasecond-by-secondbasis.Wealsoidentified
themomentwhenthepigletwasstrugglingwiththemaximumintensity(astheabsolutemaxi-
mumweightonthescale).
Threelevelsofarousal
Previousstudieshaveuseddifferentapproachestoinduceandassessemotionalarousalinani-
mals,including,forinstance,presentingmoreandlessdangerouspredators,avoidance/pref-
erenceforthesituation,administrationofpsychoactivedrugs,aswellasusingtheintensityof
thebehaviouralandphysiologicalresponsetosituationorstimulus[4,5].Inthepresentstudy,
weusethestrugglingofpigletsasabehaviouralindicatoroftheiremotionalarousal.Thebackt-
estrepresentsanegatively-valenced,stressfulsituationleadingtoanincreaseincortisollevels
inpiglets[28].Piglet'sstrugglingtoescapecanbeviewedasabehaviouractivatedbythepun-
ishmentavoidancesystem[6].Theresponsesofpigletstothebacktestvaryfromtonicimmo-
bility,lyingstillbutalerted,tofiercestruggling.Theselevelscorrespondwellwiththe
descriptionofarousalbyRussel[29]:“Theverticaldimension,arousal,rangesfromsleep,then
drowsiness,throughvariousstagesofalertnesstofreneticexcitement.Thefeelingisone’ssense
ofmobilizationandenergy.”Althoughweuseonlybehaviouralindicatorsofarousal,itisvery
likelythatthefrequentlyusedphysiologicalindicatorsofarousalsuchasheartrateandrespira-
tionrate(seee.g.[30])wouldreflectstrugglingintensityaswell.Wethereforeconsiderstrug-
glingasagoodindicatorofarousalinthebacktestsituation.
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ExpressionofEmotionalArousalinTwoDifferentPigletCallTypes
Foreachpiglet,weidentifiedthreebroadarousallevelsduringthebacktestbasedonqualita-
tiveandquantitativebehaviouralindicators.Thebacktestsituationisassumedtobeexperi-
encedasnegativeandunpleasant,yetnotpainful,bypiglets.Thebehaviourofpigletsduring
thetestvariedmarkedlyfromlyingstill(=noorlittlemuscleactivation)tointensestruggling,
indicatingaprogressiveactivationofthepunishmentavoidancesystem[6].Weidentified
threedistinctarousallevels:1)Lowarousallevel(LOW):thepigletliesstillonthescalewithout
struggling,thepigletresiststhesituationpassively;2)mediumarousallevel(MEDIUM):the
pigletshowsactiveresistanceandstrugglestoescapewiththelowtoveryhighresistanceforce
(wetooktheweightnecessarytokeeppigletinasupinepositionasaproxyfortheresistance
intensity);3)maximumsituationarousallevel(MAXIMUM):thepigletshowsthemostinten-
siveactiveresistancewhilststruggling(maximumweightnecessarytokeeppigletinasupine
positionduringthebacktest).
Audiorecordingandanalyses
Adirectionalmicrophone(SennheiserME66,flatresponse40–20000Hz,maximumsound
pressurelevel=128dBSPL)equippedwithafoamwindshieldwasplaced1.5mabovethepig-
letduringthetrial.VocalizationswererecordedonadigitalsolidstaterecorderMarantz
PMD671intoa24-bit,48000Hz,mono,PCM(wav)format.Therecordingvolumegainwas
keptconstantovertherecordings.Tomeasurethesoundpressurelevel,therecordingswere
calibratedwithareferencebeepsoundwhosesoundlevelwasmeasuredatthemicrophone
withalso-TechSLM-1352ASoundlevelmeter(fast,A-weighting).Pigletswereabletomove
theirheadsfreely,thereforetheheadwasnotalwaysdirectlypointinginthedirectionofthe
microphone,whichcouldhaveaffectedthemeasurementsofthesoundpressurelevel.Never-
theless,headmovementsoccurredinallstagesoftestandhencethisvariationwasunlikelyto
causeanysystematicbiasinourresults.RecordingswereanalysedinAvisoftSASLabPro[31].
Allcallswereidentifiedautomaticallyinthefirststage(-15to-20dBthresholdsforcallonset
andoffsetdependingontherecording).Annotatedrecordingswerethenreviewedmanually
andanymisclassificationswerecorrected(c.a.1–5%;non-detectedcalls,partlydetectedcalls–
incorrectonsetsoroffsets,mergedcalls,etc.).
Weselected6callsperpiglet:3screamcalls(highfrequencycalls)and3gruntcalls(lowfre-
quencycalls),withonecallperarousallevelforeachcalltype(Fig1).Weusedthepeakfre-
quencyasacriteriontoclassifythecallasagruntorascreaminaccordancewithprevious
studies[23].Becausethedistributionofpeakfrequencyvalueswasclearlybi-modal,callswith
apeakfrequencyabove1000Hzwereclassifiedasscreams,whilsttheremainingcallswere
classifiedasgrunts.Screamcallswerealwaysselectedfirstforeacharousallevel.TheMAXI-
MUMscreamcallwasthecallemittedinthetimeofthemostintensivestrugglingresistance
+/-onecall.TheMAXIMUMgruntcallwasthecallimmediatelyneighbouringtheMAXI-
MUMscreamoragruntcallproducedamaximumof1sfromonsetoroffsetofthescream.
BecausetheMEDIUMarousallevellastedaconsiderableamountoftimeandthereweretypi-
callymanyMEDIUMscreamcallswithinatrial,weselectedonescreamatrandomandpaired
itwithaMEDIUMgruntwithin2sfromtheMEDIUMscreamcall.Thesameselection
approachwasusedinthecaseoftheLOWscreamandgruntcalls.Asaresultofthisselection
processweobtainedthreepairsofscreamandgruntcallsperpiglet.Wedidnotanalysemore
thanonecallfromeacharousallevelasthebehaviourofthepigsvariedrapidlyduringthetest,
forexample,theintensityofstrugglingchangedveryquicklyintheMEDIUMarousallevel,
whilstcallsinLOWarousallevelwereinfluencedbytheirtemporalproximitytostruggling
bouts.Therefore,comparisonsbasedonaverageswouldnotsuitthepurposeofourstudy.
Withineachscream/gruntpairwecouldbesurethatthetwocallswereemittedinavery
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ExpressionofEmotionalArousalinTwoDifferentPigletCallTypes
similarphysiological(heartrate,respirationrate,muscletension,etc.)andemotional/motiva-
tionalstatewithouttheneedtomeasurethephysiologicalandemotionalstateofthepiglets
directly.Finally,wehadafullsetof6callsfor30piglets,providing180analyzedcallsintotal
forthemainanalysis.Fortherestofthepigletswewerenotabletogetacompletesetofgrunts
andscreamsfromeacharousallevel.
Wedecidedtofocusouranalyseson4acousticparametersofthecalls:duration(s);root-
(cid:1)
mean-squaresoundintensity(dBSPL);central frequency(Hz;frequencydividingtheaverage
spectrumofthecallintwopartsofequalenergy;50%quartilefrequencyinAvisoftSASLab—
usedasthespectrumdistributioncharacteristic),andtonality(dB,harmonic-to-noiseratio).
Thesefourparameterscomprisesomeofthemoststudiedvocalparametersandhaveprevi-
ouslybeenlinkedtospecificphysiologicalchanges[4].Wedidnotevaluatethefundamental
frequency(F0)becauseitwasproblematictoobtaintheF0foralargeamountofthecalls
(becausetheyweretoonoisyand/orshort)andwewantedtofocusonlyonthoseparameters
thatwouldbeeasilycomparablebetweenthescreamandgruntcalltypes.
Theduration,root-mean-squareamplitudeandcentralfrequencyweremeasuredautomati-
callyinthedetectedcallswiththe‘Automaticparametermeasurement’toolinAvisoftSASLab.
Thespectrogramsettingswere:FFTlength=1024,windowlength=0.046s,window
type=Hamming.Thesoundintensitywascalibratedwithabeepofknownintensity(see
above)atthebeginningoftherecordingbyusingthefunction‘Calibration/SPLwithreference
sound’inAvisoftSASLab.Theharmonic-to-noiseratiowasmeasuredinPRAAT[32]using
the‘ToHarmonicity(cc)’functionwiththestandardsettings(Timestep:0.01s;Minimum
pitch:75Hz;Silencethreshold:0.1;Periodsperwindow:1.0).Thisfunctioncomputesacoustic
periodicityonthebasisofacross-correlationmethodindependentlyofwhetherapitchcanbe
determined[33].
Besidestheanalysesofacousticcallstructure,wewerealsointerestedinhowpigletsvocalize
whentheywerestrugglingversuswhentheywerestill.Becausethedurationofeachbeha-
viouralresponsedifferedperpiglet,wecomputedthecallingrates(grunts/s;screams/s)ineach
stage(still/struggling)separately.
Statisticalanalyses
StatisticalanalyseswerecarriedoutinR.Wilcoxonsigned-ranktestswereusedtodetermine
whethercallingratesincreasedbetweenthestrugglingandstillstagesofthebacktest.Forthe
mainanalysisweusedcallsfrom30pigletsforwhomwehadcompletesampleofcalls(grunts
andscreamsfromallthreelevelsofarousal,seealso‘Audiorecordingandanalyses’).Weused
linearmixed-effectmodels(‘lme’functionfrom‘nlme’package)toassesstheeffectofthe
arousalleveloneachofthevocalparametersseparately(arousallevelasfixedfactorwiththree
levels:LOW,MEDIUM,andMAXIMUM).Thecentralfrequencyanddurationwerefirstlog
transformedduetonon-normality.Wealsoincludedaninteractiontermbetweencalltypeand
arousalleveltoestablishwhetheranyoftheacousticparametersresponddifferentlytochanges
inarousaldependingonthecalltype.Wealsoaddedthefixedeffectsoftestingorder,piglet
sex,andpigletweighttocontrolfortheirpotentialinfluenceonthecallstructure.Thetest
orderwasincludedasacovariatebecauseitprovidedanindicationoftimeaftermilkintake
(westartedtestingeachlitter5minutesafternursingandthetestingofeachpiglettookabout3
minutes)whichcouldaffecttheemotionalstateofpiglets[15].Litterandpigletidentitywere
includedasrandomfactors(pigletnestedwithinlitter).Weusedabackwardsstepwisemethod
toremovenon-significantfixedvariablesfromthemodelsuntilonlysignificanteffects
remained.Testingorder,pigletsex,andpigletweightwereneversignificant,thereforewe
reportonlythemodelsincludingthefixedeffectsofarousallevelandcalltype.Wechecked
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ExpressionofEmotionalArousalinTwoDifferentPigletCallTypes
thatthedistributionofresidualswasapproximatelynormalanddidnotsystematicallyvary
acrossthefittedvalues(withconstantvarianceandnotrends).Therewereminorissueswith
theresidualvarianceinthecaseofcentralfrequency,suggestinglargervarianceinthescream
callsthaningruntcalls.Wethereforeusedthe‘weights’argumentofthe‘lme’functionthat
enablestheheteroscedasticityofthewithin-errorgrouptobemodelled(i.e.allowsdifferent
variancesforeachfactorlevel)[34].Weusedthe‘ghlt’functionfrom‘multcomp’toperform
post-hoctestswithinthefinallinearmixedmodels.
Results
Behaviourofthepigletsduringthebacktest
ThebehaviourofthepigletsissummarizedinTable1.Pigletsstartedtostrugglefromthestart
ofthebacktest(latencytostrugglewas4.3sonaverage),performedaround3boutsofstrug-
glingduringthetest(2.7boutsofstrugglingonaverage),andstruggledforalmosthalfofthe
time(25.0sonaverage).Pigletsalsovocalizedintensively.Onaverage,pigletsemitted
mean±SD=50.3±21.6callsduringthebacktest;takingeachcalltypeseparately,therewere
slightlymoregrunts(29.8onaverage)thanscreams,butthenumberofscreamswasalsohigh
(20.5onaverage).
Thecallingratewashigherwhenthepigletswerestrugglingforbothscreams(Wilcoxon
signedranktest:N=87,Z=-8.03,r=-0.61,p<0.001)andgrunts(Wilcoxonsignedranktest:
N=88,Z=-4.69,r=-0.35,p<0.001).
Arousalandcallparameters
Screamsdifferedfromgruntsinalloftheinvestigatedacousticparameters(allp<0.001,see
theeffectofcallstypeinTable2fordetails).Thearousallevelsignificantlyaffectedtheinten-
sityandcentralfrequencyofthecalls.Withincreasingarousal,callsbecamelouderandhigher.
However,alloftheacousticparameterswerealsoinfluencedbytheinteractionbetweencall
typeandarousallevel,suggestingthatthetwocalltypesrespondeddifferentlytochanging
arousallevels(seeFig2),whichwasconfirmedbypost-hoctests(S1Table).
Thecalldurationdidnotsignificantlydifferbetweenthearousallevelsforeitherofthetwo
calltypes(S1Table,allp>0.19).Thesignificantarousal(cid:1)calltypeinteractionthusprobably
Table1. Descriptivestatisticsforpigletbehaviourduringthebacktest.
allpiglets(n=88) pigletsinanalysis(n=30)
mean sd mean sd
struggles(n) 2.71 1.15 2.9 1.12
strugglingduration(s) 24.88 12.58 23.07 11.7
latencytostruggle(s) 4.3 9.2 3.1 4.3
pigletweight(g) 1830 450 1870 430
maximumresistanceforce(maximumweight/pigletweight) 1.46 0.16 1.47 0.18
vocalizations(n) 50.3 21.6 56.7 20.1
screams(n) 20.5 18.6 21.9 16.8
grunts(n) 29.8 15.1 34.8 12.9
screams/s—relaxed 0.10 0.17 0.09 0.12
screams/s–struggling 0.67 0.43 0.76 0.32
grunts/s–relaxed 0.35 0.29 0.51 0.32
grunts/sstruggling 0.81 0.61 0.78 0.45
doi:10.1371/journal.pone.0135414.t001
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Table2. Effectsofcalltype(scream/grunt),arousallevel(LOW/MEDIUM/MAXIMUM)andtheirinteractiononthefouracousticparameters.
Variable effect df F p
Duration(log) calltype 1,145 99.18 <0.001
arousal 2,145 0.95 0.389
calltype*arousal 2,145 3.29 0.040
Root-mean-squareamplitude calltype 1,145 100.00 <0.001
arousal 2,145 28.18 <0.001
calltype*arousal 2,145 8.82 <0.001
Centralfrequency(log) calltype 1,145 772.67 <0.001
arousal 2,145 13.27 <0.001
calltype*arousal 2,145 5.29 0.006
Harmonic-to-noiseratio calltype 1,145 13.29 <0.001
arousal 2,145 2.04 0.134
calltype*arousal 2,145 15.48 <0.001
doi:10.1371/journal.pone.0135414.t002
arosefromtheslight,non-significantextensionofthescreamsandshorteningofthegrunts
withincreasingarousal.
Theintensityincreasedwitharousalonlyinthescreamcalls(S1Table,forgruntcallsall
p>0.18);LOWscreamswerelessintensethaneitherMEDIUM(z=-6.43,p<0.001)or
MAXIMUMscreams(z=-7.73,p<0.001).
Thecentralfrequencyincreasedinboththescreamandgruntcalls(S1Table).Inscreams,
theLOWarousalleveldifferedonlyfromtheMAXIMUMarousallevel(z=-3.64,p=0.002),
whileingruntstheLOWarousalleveldifferedfromboththeMEDIUM(z=-3.85,p<0.001)
andMAXIMUMarousallevels(z=-4.53,p<0.001).Thecentralfrequencyincreasedmuch
moreinthegruntcallsthaninthescreamcalls.
Theharmonic-to-noiseratioincreasedwitharousalinthescreamcalls(S1Table).LOW
arousalscreamswerelessharmonicthanMAXIMUMarousalscreams(z=-3.02,p=0.020).
Incontrast,theharmonicityofthegruntsdecreasedwitharousal,asbothMEDIUM(z=4.64,
p<0.001)andMAXIMUM(z=4.10,p<0.001)gruntswerelessharmonicthantheLOW
arousalgrunts.
Discussion
Inthisstudy,wetestedwhethertheacousticstructureofpigletdistresscallsandcontactcalls
encodedcuestoemotionalarousal,andhowspecificacousticfeaturessignalthelevelofarousal
acrossthetwocalltypes.Wefoundthatpigletsincreasethegruntandscreamratewhen
aroused(i.e.strugglingduringbacktest).Increasedcallingratehasbeenoftenfoundtobeasso-
ciatedwithincreasedarousal[4].Further,wefoundthattheacousticstructureofbothscreams
(distresscalls)andgrunts(contactcalls)doesvarywitharousal.Thisisinagreementwithpre-
viousstudiesofpigvocalizations[14,16]and,moregenerally,withtheoreticalandexperimen-
talstudiesshowinghowemotionalstateaffectsthephysiologyofanorganism,whichisinturn
reflectedinitsvocalizations[1,7,20].Suchphysiologicalchangesareassumedtoaffectthepro-
ductionofalltypesofvocalizations,asdocumentedbythewiderangeofcontextsinwhich
callshavebeenfoundtoreflectemotionalstate[4,5].Indeed,signallingsituationurgencypre-
cedesthecontextualspecificityofcalls[35].
Althoughtheemotionalstatewassignalledinbothcalltypes,wefoundthatarousalwas
encodeddifferentlyinscreamsandgrunts.Theeffectofsituationalarousalontheacoustic
structureoftwocalltypesormorehavepreviouslybeenstudied,butcallparameterswere
PLOSONE|DOI:10.1371/journal.pone.0135414 August14,2015 8/13
ExpressionofEmotionalArousalinTwoDifferentPigletCallTypes
Fig2.Scream(emptycircles)andgrunt(fullcircles)parametersinthethreearousallevels.Meansand95%confidenceintervalsareshown.Duration
andcentralfrequencyaredisplayedinlogarithmicscale.N=180(6callsfrom30individuals).
doi:10.1371/journal.pone.0135414.g002
averagedeitherfromasubsetofcalls[36]orfromallofthecalls[14]withinthesameexperi-
mentwithoutconsideringtheactualphysiologicalstateinwhichthecallswereemitted.Our
studyisthefirsttodirectlycomparehowemotionalstateisencodedintwodifferentcalltypes
underthesameconditions.Thephysiologicalconditionsonthevocalfolds(e.g.tension,sub-
glottalpressure,etc.)andinthevocaltract(e.g.salivation)arelikelytheproximatefactors
affectingexpressionofemotionsinvocalizations[4,8,20].However,theyaredifficulttocontrol
andmeasureinliveanimals.Theadvantageofourstudywasthatweanalysedscreamsand
gruntsemittedinclosesuccession(within2seconds)ineachsituation.Thefacteachscream
andgruntpairwasemittedinclosesuccessionallowsustoconfidentlyassumethattheemo-
tionalandphysiologicalstatewascomparableforbothcalltypes.
Wefoundsignificantinteractionsbetweenarousallevelandcalltypeforallofthefour
acousticparameters.Theseresultsindicatethatthefunctionand/orstructureofcalltypes
affectstheacousticalencodingofemotions.Sucheffectscouldexplainwhystudiesinvestigat-
ingvocalcorrelatesofemotionsinanimalsvocalisations[4,5]aswellasinhumanspeechor
vocalisations[37]havefounddifferentandsometimescontradictingresults.Pigletgruntsare
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ExpressionofEmotionalArousalinTwoDifferentPigletCallTypes
contactcallswithanacousticstructuresuitableforshort-rangecommunication(e.g.short
durationandlowintensity),whilescreamsaredistresscalls'designed'tobeheard(withe.g.a
longdurationandhighintensity).Wesuggestthatconstraintslinkedtotheproductionof
thesetwocalltypes[24]causetheobserveddifferencesintheencodingofemotions.
Neitherthedurationofthescreamsnorthedurationofthegruntswassignificantlyaffected
byarousal.Despitethefactthattherelationshipbetweencalldurationandarousalhasreceived
muchscrutiny,theresultshavebeencontradictory:increasingarousalhasbeenfoundtobe
associatedwitheitheranincrease,adecrease,ornochangeincallduration[4,5].Increasedcall
durationhelpsdetectionandlocalization[38],andconflictingresultsareparticularlyreported
inalarmandisolationcontexts[5]whenfacilitatinglocalizationmaynotalwaysbebeneficial
andmight,forinstance,attractunwantedattentionfrompredators.Hence,thecontextofcall
emissionandwhetherthecallisdesignedtobeeasilydetectedornotcouldconstraintheway
thatemotionalarousalisexpressedincallduration.
Bothscreamsandgruntsincreasedincentralfrequencywithincreasingarousal.Thisisin
accordancewiththegeneraltrendobservedacrossmammalianspeciesthatcallpitchincreases
witharousal[4,5].Twoaspectsofvocalproductionarelikelytoaffectthepitchofscreamsand
grunts.First,heightenedactivitywhenthepigletswerestrugglingtoescape(MEDIUMand
MAXIMUMarousallevels)islikelytoresultinincreasedmuscletension,breathingrateand
breathingamplitude,leadingtoanincreaseinthesubglottalpressureandhigherF0,which
can,inturn,influencetheenergyspectrumincludingthecentralfrequency.Second,theneed
foranincreasedairsupplyduringstrugglingmayresultinmorefrequentandgreatermouth
opening,therebyraisingtheformantfrequencies[7,20].Wesuggestthatdifferencesinthepro-
ductionofscreamsandgruntscouldexplainwhytherateofincreaseishigherforgrunts.Loud
callssuchasscreamsaretypicallyemittedorallywithanopenmouth[24].Althoughgruntsare
typicallyemittednasallywithaclosedmouthinstatesoflowarousal[24],ourpigletsfre-
quentlyopenedthemouthwhilstproducinggruntsinhighlevelsofarousal,thusconsiderably
shorteningthelengthofthevocaltractandmodifyingitsresonanceproperties[24].
Highactivityconnectedwithintensebreathingduringstruggling(MEDIUMandHIGH
arousallevels),causeshighersubglottalpressure[7,20],andshouldleadtoanincreaseinthe
amplitudeofbothcalltypes.Increasingcallamplitudewitharousalhasbeenfoundacrossa
widerangeofspeciesandcontexts[4].However,theamplitudeincreasedsignificantlyonlyfor
screams,whiletheamplitudeofthegruntswasnotaffectedbyarousal.Thismaybeaconse-
quenceofthefactthatgruntsarerelativelyfaintvocalizationsusedincloserangecommunica-
tion,andthatbecausegruntsaretypicallyproducednasally,thisdoesnotallowfora
substantialincreaseinamplitude.Anothercontributingfactorcouldbethatnasalcavities
absorbmoreacousticenergythanoralcavities[24].Moreworkonthemechanismsofvocal
productioninpigsisclearlyneededtoconfirmorrefutethesespeculations.
Wealsofoundthattheharmonic-to-noiseratioshowedoppositechangesforscreamsand
forgrunts–itincreasedwitharousalinscreamswhileitdecreasedingrunts.Previouslabora-
torystudieshavedescribedtransitionstochaoticvibrationregimeswithincreasingsubglottal
pressure[39,40].However,thisdoesnotseemtohappenwithpigletscreams.Inourstudy,the
harmonicityofthescreamcallsactuallyincreased(i.e.chaoticvibrationsdecreased)with
arousal,aspreviouslyobservedinscreamsemittedintheextremelypainfulsituationofuna-
nesthetisedcastration[16].Increasedtonalityincallsgiveninmoreurgentsituationshasalso
beendocumentedinothermammalspecies[11].Itispossiblethatcertaincalltypes,including
pigletscreams,needsubstantialairflowbeforetheyattainoptimumphonation.Ithasindeed
beensuggestedthat,inhumans,increasedjitter(=lowervoicetonality)couldbeassociated
withbothhypotensionandhypertensionofthevocalfolds[20].
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