SEVERE HYPOXIA IMPAIRS LATERALIZATION IN A MARINE TELEOST FISH

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1 First post onlin on 30 Octobr 2014 as /jb J Exp Biol Avanc Accss Onlin th most Articls. rcnt vrsion First at post onlin on 30 Octobr 2014 as oi: /jb Accss th most rcnt vrsion at SEVERE HYPOXIA IMPAIRS LATERALIZATION IN A MARINE TELEOST FISH Tyron Lucon-Xiccato 1*, Juli J. H. Nati 2, Flip Rocco Blasco 3, Jacob L. Johansn 4, John F. Stffnsn 5, Paolo Domnici 6 1 Dipartimnto i Psicologia Gnral, Univrsità i Paova, Paova, Italy 2 Institut of Bioivrsity, Animal Halth an Comparativ Micin, Graham Krr Builing, Univrsity of Glasgow, Glasgow, Scotlan 3 Laboratório Zoofisiologia Bioquímica Comparativa, Dpartamnto Ciências Fisiológicas, Univrsia Fral São Carlos, São Paulo, Brasil 4 ARC Cntr of Excllnc for Coral Rf Stuis, School of Marin an Tropical Biology, Jams Cook Univrsity, Townsvill, Qunslan, Australia 5 Marin Biological Sction, Univrsity of Copnhagn, Hlsingør, Dnmark 6 CNR-IAMC, Istituto pr l Ambint Marino Costiro, Località Sa Marini, Torrgran, Oristano, Italy *Corrsponnc: T. Lucon-Xiccato, Dipartimnto i Psicologia Gnral, Via Vnzia 8, 35131, Paova, Italy. Phon: ; fax: ; -mail: tyron.luconxiccato@stunti.unip.it Publish by Th Company of Biologists Lt

2 Abstract In intrtial nvironmnts, th rcurring hypoxic conition at low ti is on of th main factors affcting fish bhaviour, causing broa ffcts on cological intractions. W assss th ffcts of hypoxia on latralization (.g. th tnncy to turn lft or right), a bhaviour rlat to brain functional asymmtry, which is thought to play a ky rol in svral lif history aspcts of fish. Using staghorn sculpin (Lptocottus armatus), a bnthic fish that typically inhabits th intrtial zon, w foun that hypoxia affcts bhavioural latralization at population-lvl. On avrag, staghorn sculpins show a istinct prfrnc for right turns unr normoxic conitions (> 90 % oxygn saturation), but an qual probability of turning right or lft aftr xposur to hypoxia for 2 hours (20 % oxygn saturation). Th spcific turning prfrnc obsrv in th staghorn sculpin control population is likly to hav an aaptiv valu, for xampl in prator-pry intractions by nhancing attack succss or survival from pratory attacks. Thrfor th altration of latralization xprss by staghorn sculpins unr hypoxic conitions may hav far-raching implications for spcis cology an trophic intractions. Morovr, our work raiss th n to stuy this ffct in othr spcis, in which a hypoxia-rivn isruption of latralization coul affct a wir rang of bhaviours, such as social intractions an schooling. Kywors: hypoxia, latralization, tour tst, staghorn sculpin

3 Introuction Oxygn availability is on of th main factors affcting fish physiology, bhaviour, an cology. In marin intrtial nvironmnts, fish ar aily xpos to hypoxia vnts mainly u to tial rhythms. Morovr, th natural occurrnc of hypoxic vnts is xacrbat by human activitis, as wll as agricultur an ischarg of raw swag (Diaz an Rosnbrg, 2008; Richars t al., 2009). Hypoxia ffcts on fish biology hav bn intnsivly stui, showing numrous intrfrncs such as impair prformanc in rprouctiv output, growth, swimming an schooling (Domnici t al., 2007; Richars t al., 2009). Th brain oxygn rquirmnts ar among th highst of all th organs, thus brain snsitivity to hypoxia is likly to b non-trivial. Crtainly, nar lthal hypoxic conitions ar known to affct brain physiology (.g. Nilsson, 1990; Nilsson t al., 1993). Furthrmor, mormyri fish, charactriz by a hug brain oxygn uptak, ar known to show ruc brain siz in chronic hypoxic habitats at both intr- an intra-spcific lvl (Chapman an Huln, 2001). By contrast, th impact of hypoxia on bhaviours irctly rlat to brain function in fish rmains rlativly unxplor, though arguably knowlg of such ffcts coul b of grat cological rlvanc. In orr to assss altrations of brain functioning in hypoxic conitions in fish, w focus on bhavioural latralization, an xprssion of brain functional asymmtry that rflcts a iffrntial spcialization of th two hmisphrs in procssing information (Bisazza an Brown, 2011). In fish, vn though th xact volutionary procsss involv ar still not clar, th latralization pattrns obsrv in a population ar thought to rflct slctiv prssurs an to b important for th survival of iniviuals in many cological sttings such as fing, prator scaping, schooling, social an sxual intractions (.g. Bisazza t al., 1997; Bisazza t al., 2000; Daa t al., 2010; Bisazza an Brown, 2011). Although in som cass hmisphric spcialization coul rprsnt an a cost in information procssing (Bisazza an Brown, 2011), th vast of majority of vincs suggsts that it confrs svral avantags. A latraliz brain allows a sparation of task achivmnts: ach hmisphr can vlop highr spcialization in procssing spcific information, rsulting in gratr cognitiv prformancs. For xampl, mor latraliz iniviual fish (i.. thos with gratr iniviual-lvl latralization) outprform non-latraliz ons in spatial rorintation tasks or in simultanous activitis, as foraging an prator vigilanc (Sovrano t al., 2005; Daa an Bisazza, 2006). Morovr, latralization is also associat with nhanc ability to scap from a prator: strongly latraliz fish hav fastr raction tim, highr turning rats an longr istanc travll in scap rsponss (Daa t al., 2010). In som cass, a bias for a spcific irctionality of latralization is obsrv at population-lvl: a conspicuous proportion of

4 iniviuals in a population is latraliz in th sam irction (Bisazza an Brown, 2011). This bias appars to b rlvant in social spcis to archiv gratr coorination in movmnts (Bisazza t al., 2000), whras in non-grgarious spcis, it coul b involv in pry-prator intractions (Yasugi an Hori, 2012). In this xprimnt, w assss hypoxia ffcts on bhavioural latralization in th staghorn sculpins (Lptocottus armatus), a bnthic fish that typically inhabits th intrtial nvironmnt an may xprinc low oxygn availability (< 20 % oxygn saturation) uring straning in muflats an tipools (Palsson t al., 2008). W xpos two groups of staghorn sculpins to normoxia (> 90 % oxygn saturation) an non-lthal lvls of hypoxia that naturally occur in th sampling ara (20 % oxygn saturation) (Palsson t al., 2008; Sprs-Rosch t al., 2013), rspctivly, an w obsrv th subjcts in a turning prfrnc tst (tour tst) to valuat thir bhavioural latralization. W thn compar th population- an iniviual-lvl latralization of th two xprimntal groups using th Rlativ Latrality inx (L R ) an th Absolut Latrality inx (L A ), rspctivly. Th man of L R allows to assss th turning bias for a spcific irction at th population-lvl: positiv valus of L R inicat a right-turning bias, whras ngativ valus of L R inicat a lft-turning bias (Bisazza t al., 1997; Daa t al., 2010). Th man of L A rfrs to iniviual-lvl latralization an scribs th strngth of latralization irrspctiv to th irctionality (Bisazza t al., 1997; Daa t al., 2010). Bas on prvious work that tst th ffcts of nvironmntal strssors on latralization (.g. Nilsson t al., 2012), w hypothsiz that xposur to hypoxia may inuc a ruction of th iniviual-lvl latralization an/or th population-lvl latralization bias. Rsults Iniviuals wr subjct to hypoxia an control watr in small groups (s Matrials an Mthos sction). Grouping ha no significant ffct on latralization pattrn (L R : F(10,73)=1.278, p=0.259; L A : F(10,73)=0.725, p=0.698). Thr was a significant iffrnc in th population-lvl latralization (L R inx) of staghorn sculpins groups xpos to normoxia an hypoxia (t(82)=2.259, p=0.027) (Fig. 1). Unr normoxic conitions, th population of staghorn sculpins ha a man L R inx (31.91±5.87, man±s) significantly gratr than zro (t(41)=5.413, p<0.001), inicating a istinct turning prfrnc towars th right (Fig. 2). By contrast, staghorn sculpins xpos to hypoxic conitions ha a man L R inx (12.38±6.85) which was not iffrnt from zro (t(41)=1.767, p=0.085),

5 inicating a ruction of th avrag turning prfrnc towars th right an an incras in th frquncy of iniviuals with lft-turning bias (Fig. 2). Th L A inx of hypoxia trat group was not significantly iffrnt from th control group (hypoxia: 37.14±4.12; control: 41.43±4.17; t(82)=0.876, p=0.384) showing thr was no ffct of hypoxia on th iniviual-lvl latralization (Fig. 1). Discussion This stuy shows that hypoxia xposur influncs population-lvl latralization but not iniviual-lvl latralization in staghorn sculpins. On avrag, control subjcts xhibit a right- bias latralization at population-lvl, rval by a istinct prfrnc for right turns whn facing th barrir in th tour tst. In contrast, in th group xpos to hypoxia a smallr proportion of subjcts show this right-turning bias an thrfor this group show no latralization bias at population-lvl. Sinc bhavioural latralization is an xprssion of brain asymmtry, th loss of th turning prfrnc towars th right unr hypoxic conitions is likly to b caus by changs in brain hmisphr functioning or nurotransmittr functions, an coul hav irct implications for th cology an fitnss of iniviuals in th wil. A shift in information procssing btwn brain hmisphrs coul xplain th ffct of hypoxia xposur on population-lvl latralization. In svral spcis, lft hmisphr controls bhaviours in non-strssful situations, whras right hmisphr controls mrgncy rsponss (Rogrs, 2010). Som vincs an thortical work suggst that strssful situations coul caus a shift of information procssing btwn th two hmisphrs (i.. from lft to right control; Rogrs, 2010). In lin with Rogrs (2010), it is possibl that th strss u to hypoxia caus trat subjcts to incras right-hmisphr control an hnc turn mor frquntly to th lft (Bisazza t al., 1997), rsulting in a loss of th original population right-turning bias. In fish, a larg within- spcis variation in bhavioural coping to strss situation has bn rport. For xampl, iffrnt iniviuals of juvnil sa bass iffrntially altr th risk-taking bhaviour in hypoxic conition, likly bcaus of iniviual variation in snsitivity to hypoxia (Killn t al., 2012). Thrfor, it may b rlvant to trmin whthr hypoxia ffcts on bhavioural latralization ar mor intns in iniviuals with low tolranc to hypoxia. Th ffct on th population-lvl latralization may also b caus by a physiological chang in nural transmittr functions within th brain following hypoxia xposur. Such changs hav prviously bn rport for CO 2 xposur: bhavioural latralization of coral rf fish larva

6 maintain in high CO 2 concntration is impair (Domnici t al., 2011) an th ffct is thought to b miat by changs in brain physiology that altr nuronal transmittrs functions (Nilsson t al., 2012). Extrm hypoxia xposur triggrs svral physiological changs on fish brain: most fish spcis rapily los ion homostasis in thir brain rsulting in a brakown of brain functioning (Nilsson t al., 1993), whras othrs show brain protctions against hypoxia as an incras or a ristribution of inhibitory nurotransmittrs that supprss brain functioning an limit O 2 rquirmnts (Nilsson, 1990). Similar physiological changs in th brain coul xplain th loss of th population-lvl right-turning bias obsrv in staghorn sculpin xpos to hypoxia. As spcis aapt to iffrnt nvironmnts coul show iffrnt snsitivity to hypoxic strss, a prspctiv fil of invstigation is th btwn-spcis variation of th rport ffct. Fish communitis of our sampling ara may aily xprinc hypoxic conitions similar to th on rcrat in our xprimnt or vn mor svr (< 20 % oxygn saturation) (Palsson t al., 2008), with an incrasing occurrnc/intnsity bcaus of human-rivn pollution (Diaz an Rosnbrg, 2008; Richars t al., 2009). As a consqunc, th loss of population-lvl right- turning bias obsrv in hypoxia trat staghorn sculpins might hav critical cological rprcussions for th spcis itslf as wll as othr spcis. Littl is known rgaring th cological maning of population-lvl bias in latralization, howvr sinc th iniviuals obsrv in our stuy wr wil caught fish, thir population-lvl latralization pattrns ar likly to hav an aaptiv valu in thir natural nvironmnt. For instanc, in som fish spcis, scaping prator an attacking pry show latraliz bhaviours an th outcom of pry-prator intractions is rlat to th spcific irctionality of latralization (.g. th prfrnc for approach th pry from right or lft) of th two iniviuals involv (Yasugi an Hori, 2012). In a similar fashion, th latralization pattrn of th staghorn sculpins population us in this stuy (i.. most of th iniviuals ha a right turning prfrnc in th tour tst) may b aapt to match thir pry latralization pattrns in orr to nhanc attack succss, or altrnativly to surviv from pratory attacks (Vallortigara an Rogrs, 2005). Th iffrnc btwn control an hypoxia trat subjcts in population-lvl latralization suggsts that som of th iniviuals xpos to hypoxia shifts thir turning prfrnc from right-bias to lft-bias. Thrfor, in hypoxic conition staghorn sculpins coul suffr th svr fitnss costs of a cras in pration succss or in survival from prator attacks. Aitionally, in grgarious fish spcis population-lvl bias in latralization givs avantags for coorination uring schooling (Bisazza an Brown, 2011). Staghorn sculpins ar solitary an thus o not school. Howvr, similar ffcts of hypoxia on population-lvl latralization may subsist in othr fish spcis as wll, incluing thos involv in social

7 bhaviours, possibly affcting thir schooling activity. Thrfor it may b worthwhil to tst th ffct of hypoxia on latralization in grgarious spcis. In our xprimnt, iniviual-lvl latralization was not affct by hypoxia xposur. Inspction of th frquncy istribution plots (Fig. 2) offrs a possibl intrprtation for th fining that hypoxia has an ffct on latralization at th population-lvl but not at th iniviual-lvl. In normoxia, iniviual sculpins ar mainly ithr non-latraliz or right latraliz, whil hypoxia xposur rsults in som iniviuals to b lft-bias. Hnc, xposur to hypoxia might caus right-latralis iniviuals to bcom lft-latraliz (s pak at LR=-60), rsulting in th loss of th original si bias in th whol population, whil maintaining a similar lvl of iniviual latralization as in normoxia. A similar ffct was foun in a coral rf fish, whr xposur to lvat CO 2 lvl affct population-lvl but not iniviual-lvl latralization (Domnici t al., 2014). In contrast, stuis on othr spcis rport an ffct of xposur to lvat CO 2 lvl in iniviual-lvl but not in population-lvl latralization (.g. Nilsson t al., 2012). At th currnt stag of rsarch, it is not clar what trmins this incongrunc of th rsults. On possibility is that th iffrnt strssors affct latralization only at iniviual- or population-lvl in ach spcis. Howvr, on shoul also consir th possibility that th nvironmntal strssors affct simultanously both iniviual- an population-lvl latralization but that iffrncs in th ffct sizs limit th statistical infrnc. Our rsults show that hypoxia can isrupt latralization in fish an suggst broa implications for wil populations, sinc hypoxia is foun across most of th worl s aquatic cosystms, an hypoxic aras ar spraing worlwi u to incras utrophication, simnt loaing an pollution from human activitis (Richars t al., 2009). Morovr, in coastal aras vry oftn fish ar simultanously xpos to othr strssors in aition to hypoxia, such as incras tmpratur, lvat CO 2 or ammonia. Som of ths strssors ar known to affct latralization (tmpratur an lvat CO 2 ; Domnici t al., 2014), hnc thy coul potntially intract with hypoxia in affcting bhavioural latralization. Thrfor, broar invstigations on th synrgistic ffct of iffrnt nvironmntal factors on a wi rang of spcis woul b of high valu to unrstan th unrlying physiological procsss an cological implications of incrasing hypoxic zons worlwi. Matrials an Mthos Exprimntal subjcts

8 No physical invasiv manipulation was prform on th fish. All animal car an xprimntal protocols follow th guilins of Institutional Animal Car an Us Committ at th Univrsity of Washington (IACUC prmit numbr ). Eighty-four staghorn sculpins wr us ovrall (stanar lngth ±2.91 mm, man±s). Forty-two subjcts wr xpos to hypoxia, an forty-two wr us as control. Each subjct was tst only onc. Staghorn sculpins wr sampl by bach sining at th Jackson bach ( N, W) on San Juan Islan (Washington, US) in August Bfor th xprimnts, fish wr maintain at Friay Harbor Laboratoris (Univrsity of Washington) for 24 hours. Maintnanc tanks (130 x 60 x 26 cm) wr kpt at a constant sawatr flow, through unr natural light conitions, an at tmpratur of 11.64±0.52 C. Exprimntal tratmnt Subjcts wr transfrr in small groups (7-8 iniviuals) to a tratmnt tank (45x50 cm) fill with 25 cm of sawatr (s supplmntary matrials Fig. S1). Watr tmpratur was maintain nar constant uring th tratmnt (11.48±0.06 C ) in orr to avoi any confouning ffct of tmpratur on th oxygn consumption of th subjcts. To avoi arial rspiration of th subjcts, th watr surfac was covr with a panl ma of transparnt synthtic glass. Th tratmnt tank was connct to an xtrnal sump (40 cm in iamtr, 60 cm high) fill with 50 cm of sawatr which allow constant rcirculation of th watr into th systm. In th bottom of th sump, 5 stanar air stons wr install, 1 connct to an air pump an 4 connct to a nitrogn tank which was proviing nitrogn to th sump. A galvanic oxygn prob (Oxyguar Hany, Oxyguar Intrnational, Birkro, Dnmark) connct to an oxygn rgulator (PR 5714, PR Elctronics, Ron, Dnmark) an a solnoi valv controll nitrogn flow through th air stons, allowing rgulation of oxygn saturation in th sump, an, inirctly in th tratmnt tank. For th hypoxia group, w cras oxygn saturation in th tratmnt tank from normoxia (> 90 % oxygn saturation) to 20 % oxygn saturation ovr a 20 minuts prio an kpt oxygn saturation constant for 120 minuts (20.29±0.74 % oxygn saturation). For th control group, oxygn saturation was kpt at normoxia (98.34±1.02 % oxygn saturation) for 140 minuts an air insta of nitrogn was injct in th sump via th air stons. Latralization assssmnt Immiatly aftr th n of th tratmnt (normoxia or hypoxia), subjcts wr tst for thir bhavioural latralization using a tour tst (Bisazza t al., 1997), a procur prviously aopt to stuy latralization in bnthic fishs (Bisazza t al., 2000). Th apparatus consist of a

9 tank (120 x 50 cm) with a cntral runaway (75 x 10 cm) (s supplmntary matrials Fig. S2). A prpnicular barrir was install 6 cm aftr ach n of th runaway. At th start of ach trial, a singl fish chosn at ranom was introuc into th xprimntal arna an lft for 1 minut to bcom accustom to th nvironmnt. Subsquntly, by using a pair of small fish nts, th subjct was gntly conuct up to half of th runway, in orr to ncourag it to swim towar th barrir. For ach subjct th procur was rpat tn tims altrnating btwn th two symmtrical ns of th runway (Domnici t al., 2014). Th irction of turns in front of th barrir was rcor. For ach subjct, w comput th Rlativ Latrality inx (L R ) an th Absolut Latrality inx (L A ) (Bisazza t al., 1997). L R was comput using th formula: L R = [(turns to th right - turns to th lft) / (turns to th right + turns to th lft)]*100, with valus ranging from -100 to +100, inicating complt prfrnc for lft an right turning, rspctivly. L A, ranging from 0 to 100, was comput accoring th formula: L A = L R (Bisazza t al., 1997). Statistical analysis Bcaus L R an L A inics ar prcntag ata, w prform arcsin squar root transformation bfor th analysis (Sokal an Rohlf, 1995). To al with th ngativ L R valus, w transform th absolut valu of this variabl an subsquntly w a th corrsponing sign to ach obsrvation. In th main txt, untransform ata ar rport for consistncy with prvious litratur. Sinc th subjcts wr xpos to hypoxia or control watr in small groups, w conuct a prliminary analysis to tst whthr th group ha a significant ffct on th L R an L A inics using a on-way ANOVA. A man L R nar zro inicats that a givn sampl of th population is nithr lft- nor right-bias in its turning tnncy (Bisazza t al., 1997). Thrfor, th man L R of ach group was compar to a thortical zro using a on sampl t-tst (Bisazza t al., 1997). W us inpnnt sampl t-tst to compar man of L R an L A inxs btwn th control group an hypoxia trat group. All analyss wr carri out in R statistical softwar (vrsion 3.0.2, R Founation for Statistical Computing). Acknowlgmnts Th authors woul lik to thanks th Friay Harbor Laboratoris staff for logistical support, th 2013 Fish Swimming class for fish collction an hlpful iscussions, an Anglo Bisazza for commnts on th arlir vrsion of th manuscript.

10 Author Contributions T.L.X, J.J.H.N, F.R.B, J.L.J, J.F.S an P.D. conciv an sign th xprimnts; T.L.X., J.J.H.N an F.R.B prform th xprimnts. T.L.X, J.J.H.N an F.R.B. analys th ata; T.L.X, J.J.H.N, F.R.B., J.L.J, J.F.S an P.D. wrot th papr. Compting intrsts Th authors clar no compting financial intrsts. Rfrncs Bisazza, A., Pignatti, R. an Vallortigara, G. (1997). Latrality in tour bhaviour: intrspcific variation in pocilii fish. Anim. Bhav. 54, Bisazza, A., Cantalupo, C., Capocchiano, M. an Vallortigara, G. (2000). Population latralisation an social bhaviour: a stuy with 16 spcis of fish. Latrality 5, Bisazza, A. an Brown, C. (2011). Latralization of cognitiv functions in fish. In Fish cognition an bhaviour (. C. Brown, K. Lalan an J. Kraus), pp Cambrig: Blackwll Publishing. Chapman, L. J. an Huln, K. G. (2001). Implications of hypoxia for th brain siz an gill morphomtry of mormyri fishs. J. Zool. 254, Daa, M. an Bisazza, A. (2006). Dos brain asymmtry allow fficint prformanc of simultanous tasks? Anim. Bhav. 72, Daa, M., Koolhaas, W. H. an Domnici, P. (2010). Bhavioural asymmtry affcts scap prformanc in a tlost fish. Biol. Ltt. 6, Diaz, R. J. an Rosnbrg, R. (2008). Spraing a zons an consquncs for marin cosystms. Scinc 321, Domnici, P., Lfrancois, C. an Shingls, A. (2007). Hypoxia an th antiprator bhaviours of fishs. Philos. Trans. R. Soc. B-Biol. Sci. 362, Domnici, P., Allan, B., McCormick, M. I. an Munay, P. L. (2011). Elvat carbon ioxi affcts bhavioural latralization in a coral rf fish. Biol. Ltt. 8, Domnici, P., Allan, B. J., Watson, S. A., McCormick, M. I. an Munay, P. L. (2014). Shifting from right to lft: th combin ffct of lvat CO2 an tmpratur on bhavioural latralization in a coral rf fish. PLoS ONE 9:87969.

11 Killn, S. S., Marras, S., Ryan, M. R., Domnici, P. an McKnzi, D. J. (2012). A rlationship btwn mtabolic rat an risk taking bhaviour is rval uring hypoxia in juvnil Europan sa bass. Funct. Ecol. 26, Nilsson, G. E., Prz-Pinzon, M., Dimbrg, K. an Winbrg, S. (1993). Brain snsitivity to anoxia in fish as rflct by changs in xtracllular K+ activity. Am. J. Physiol.-Rgul. Intgr. Comp. Physiol. 264, R250-R253. Nilsson, G. E. (1990). Long-trm anoxia in crucian carp: changs in th lvls of amino aci an monoamin nurotransmittrs in th brain, catcholamins in chromaffin tissu, an livr glycogn. J. Exp. Biol. 150, Nilsson, G. E., Dixson, D. L., Domnici, P., McCormick, M. I., Sørnsn, C., Watson, S. A. an Munay, P. L. (2012). Nar-futur carbon ioxi lvls altr fish bhaviour by intrfring with nurotransmittr function. Nat. Clim. Chang. 2, Palsson, W. A., Pacunski, R. E., Parra, T. R. an Bam, J. (2008). Th ffcts of hypoxia on marin fish populations in southrn Hoo Canal, Washington. Amrican Fishris Socity Symposium Sris 64, Richars, J. G., Farrll, A. P. an Braunr, C. J. (2009). Hypoxia. In Fish Physiology, Vol. 27 (. A. P. Farrll an C. J. Braunr), pp San Digo: Acamic Prss. Rogrs, L. J. (2010). Rlvanc of brain an bhavioural latralization to animal wlfar. Appl. Anim. Bhav. Sci. 127, Sokal, R. R. an Rohlf, F. J. (1995). Biomtry: th principls an practic of statistics in biological rsarch. Nw York: WH Frman an Company. Sovrano, V. A., Daa, M. an Bisazza, A. (2005). Latraliz fish prform bttr than nonlatraliz fish in spatial rorintation tasks. Bhav. Brain Rs. 163, Sprs-Rosch, B., Manic, M., Groom, D. J. an Richars, J. G. (2013). Critical oxygn tnsions as prictors of hypoxia tolranc an tissu mtabolic rsponss uring hypoxia xposur in fishs. J. Exp. Mar. Biol. Ecol. 449, Vallortigara, G. an Rogrs, L. J. (2005). Survival with an asymmtrical brain: avantags an isavantags of crbral latralization. Bhav. Brain Sci., 28, Yasugi, M. an Hori, M. (2012). Latraliz bhavior in th attacks of largmouth bass on Rhinogobius gobis corrsponing to thir morphological antisymmtry. J. Exp. Biol. 215,

12 Fig. 1. Rlativ Latrality Inx (L R ) an Absolut Latrality Inx (L A ) of subjcts xpos to control conition (ark bars) or hypoxia conition (gry bars). Fig. 2. Rlativ frquncy istribution of Rlativ Latrality Inx (L R ) in subjcts xpos to control conition (ark bars) or hypoxia conition (gry bars).

13 LatralityInx(man±s) h Journal of Exprimntal Biology ACCEPTED AUTHOR MANUSCRIPT * LR NS LA

14 Rlativfrquncy(%) RlativLatralityInx(LR)