Changes in CMDP and DPOAE during acute increased inner ear pressure in the guinea pig

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1 Eur Arh Otorhinolryngol (2008) 265: DOI /s OTOLOGY Chnges in CMDP nd DPOAE during ute inresed inner er pressure in the guine pig W. L. Vlk Æ H. P. Wit Æ F. W. J. Alers Reeived: 29 June 2006 / Aepted: 4 Septemer 2007 / Pulished online: 28 Septemer 2007 Ó Springer-Verlg 2007 Astrt During nd fter n inrese of inner er pressure, indued y injetion of rtifiil perilymph, the 2f 1 f 2 nd f 2 f 1 distortion produts (DPs) in ohler mirophonis (CM) nd otoousti emissions (OAE) were reorded in the guine pig. An inner pressure inrese of *600 P gve only smll hnges in CMDP nd DPOAE. Along with derese in f 1 mplitude, smll derese in mplitude of the 2f 1 f 2 nd smll inrese in the f 2 f 1 were mesured in CM. This mthes shift from symmetril position of the operting point for hir ell trnsdution, leding to n inrese in even-order distortion nd derese in odd-order distortion. Similr, derese in 2f 1 f 2 DPOAE ws expeted. This might e the se t the genertion sites ut this effet ws then more thn ompensted for y etter middle er trnsfer, ounting for the inrese of 0.4 db of the 2f 1 f 2 DPOAE mplitude. In onlusion, hnges of overll inner er fluid pressure hve minor effets on ohler funtion. This is relevnt finding for further understnding of diseses with hnged inner er fluid volumes, s Ménière s. Keywords Perilymph Injetion Inner er pressure 2f 1 f 2 f 2 f 1 CMDP DPOAE Guine pig Introdution The ohle is non-liner mehno-eletril trnsduer tht trnsforms sound pressure wves into eletril W. L. Vlk (&) H. P. Wit F. W. J. Alers Deprtment of Otorhinolryngology, University Hospitl Groningen, PO Box , 9700 RB Groningen, The Netherlnds e-mil: w.l.vlk@kno.umg.nl informtion, the so-lled mehnoeletri trnsdution (MET). The reverse eletromehnil trnsdution is performed y the outer hir ell (OHC) whih mehnilly influenes the virtion of the ohler prtition [7]. When the ohle is stimulted y two tones, distortion produts (DPs) n e mesured in ohler mirophoni potentils (CMDP) nd in the externl uditory metus s otoousti emissions (DPOAE). Both mirophoni nd ousti DPs re onsidered to e refletions of OHC eletromotility in the ohle. Oviously, it is of interest to find reltion etween these two funtionl modlities of the ohle nd the OHC in prtiulr. In this study the most prominent DPs, the 2f 1 f 2 nd f 2 f 1, hve een oserved while inresing inner er pressure. This ws done y rpid injetion of rtifiil perilymph into the sl tympni of the guine pig s ohle. Mesurement of speifi DPs my desrie hnges in gin nd operting point of the non-liner ohler mplifier. Furthermore, trnsfer hrteristis of forwrd nd kwrd trnsmission my e lrified y using CMDP nd DPOAE [17]. These dt ould help gining insight into pthophysiologil mehnisms involved in pthologilly inresed inner er pressures. When pressure vrition is in the norml physiologil rnge, this does not influene ohler funtion s mesured y ompound tion potentils in the guine pig [2]. In humns, during tivities like oughing or sneezing, perilymphti pressure my undergo sustntil flututions up to 10 mmhg (1.4 kp). An inresed pthologi inner er pressure my e the result of n inresed intrrnil pressure sine the perilymphti spe is linked to the surhnoid spe vi the ohler quedut. Humn OAE hve een proven to e very sensitive to modifitions of intrrnil pressure [10]. Another linil exmple of inresed inner er pressure or

2 288 Eur Arh Otorhinolryngol (2008) 265: volume with n ompnying hnge of ohler funtion is the endolymphti hydrops in ptients with Ménière s disese. In previous study [11], we mesured solely the 2f 1 f 2 DPOAE s in response to miroinjetion of rtifiil perilymph into the ohle. Lrge hnges of *500 P in overll inner er pressure gve only smll men hnge of 0.7 db in 2f 1 f 2 DPOAE mplitude. The mgnitude of hnge ould (prtly) e explined y hnge in ovl window stiffness. In this study, the sme experimentl setup ws used, with the ddition of the simultneous reording of the 2f 1 f 2 nd f 2 f 1 CMDP. We ttempt to estlish etter understnding of inresed inner er pressure effets on DPs in oth CM potentils nd OAE s. Mterils nd methods Experiments were performed in eight guine pigs (Hrln Lortories, The Netherlnds; ody weight g) with positive Preyer reflex. Animl re nd use were in ordne with the priniples of the delrtion of Helsinki nd pproved y the niml experiment ommittee (protool numer 3047/3103). Generl nesthesi ws indued y intrmusulr dministrtion of ketmine/xylzine (60/3.5 mg/kg). Mintenne doses of the nestheti were dministered every hour. Musle relxtion ws otined with suinylholine (2.5 mg/kg). The nimls were rtifiilly ventilted through trheostom (Columus Instruments, model 7950). Body temperture ws mintined t 38 C with heting lnket. Hert rte ws monitored y skin eletrodes pled on oth sides of the thorx. The niml s hed ws kept in sttionry position y mens of steel olt fixed to the skull with dentl ement. Following retrouriulr inision, the ull nd externl uditory nl were exposed. Susequently, the ull ws opened equlizing middle er pressure to norml ir pressure [17]. By opening of the ull, the round window ws exposed. Through the round window memrne, the tip of doule-rreled miropipette ws inserted into sl tympni. The doule-rreled miropipettes were drwn from orosilite glss (1.5/ 0.84 mm dimeter per rrel) nd the tips were eveled (Nrishige EG-40). The totl tip dimeter ws round 60 lm, whih is ompromise etween low enough flow resistne for fluid injetion nd tip smllness. One rrel of the pipette ws used to mesure inner er pressure (WPI 900A miropressure system). Through the other rrel, rtifiil perilymph [9] ws injeted with onstnt flow rte y pplying ontrollle pneumti pressure to the rrel end. The injeted volume ws mesured s the displement of the fluid menisus in the pipette, for whih the inner dimeter is preisely known (0.84 mm). The fluid injetion rte ws lulted s the totl injeted volume divided y the totl injetion time. During the experiment the DPOAE s were ontinuously mesured using n Etymoti ER-10C DPOAE proe system. The two primry frequenies were set t 6 khz (f 1 ) nd 7.4 khz (f 2 ), mening f 2 /f 1 -rtio of The lowlevel intensities were set t, respetively, 65 db SPL (L1) nd 55 db SPL (L2). Full detils of the mesuring equipment re desried elsewhere [11, 12]. Cohler mirophoni potentils were reorded with two differentil Ni Cr eletrodes with dimeter of 50 lm. One eletrode ws pled in sl tympni through the ony wll of the sl turn of the ohle. The other ws pled in sl vestiuli next to the ovl window. The signls were routed vi n mplifier to spetrum nlyzer nd two lok-in mplifiers (SRS, model SR830 DSP). The lok-in mplifiers displyed the mplitude of the 2f 1 f 2 nd f 2 f 1 CMDP whih were referened to n eletroni 2f 1 f 2 nd f 2 f 1 -DP derived from the originl primries. A mesurement ontined sequene of ten miroinjetions of rtifiil perilymph of 10 s eh with puses of 40 s. The rte of miroinjetion ws 50 nl/s. The repetitive miroinjetion ws ontrolled with preision eletroni timer (Stnford DG535). During n experiment, Ntionl Instruments LVIEW 1 ws used for reording of the following output signls: mplitude nd phse of the 2f 1 f 2 DPOAE, 2f 1 f 2 CMDP, f 2 f 1 CMDP, f 1 CM, nd inner er pressure. Clultions nd fits to relevnt portions of the otined reordings were mde off-line with n pproprite softwre pkge. Results Suessful repeted sequenes of injetion with rte of 50 nl/s (n = 10) were performed in ll guine pigs (n = 8). The men stedy stte inner er pressure mesured prior to injetion of rtifiil perilymph ws 205 P. To determine the rise nd fll times of the urves from the mesured signls, proper fits were otined with simple exponentil funtion: expð t=sþþ: In some ses (Figs. 1, 2, 4) the urves needed n extr term in the formul for proper fit: expð t=sþ=½1 þ f1 expð t=sþgš þ : Aording to Wit et l. [13] time onstnts s were lulted from the fits. In Fig. 1, the verged reorded inner er pressure during nd fter injetion of 0.5 ll of rtifiil perilymph is shown. At the strt of injetion the men pressure inresed

3 Eur Arh Otorhinolryngol (2008) 265: Fig. 1 Averged hnges (n = 8) in inner er pressure (P) during nd fter injetion of 0.5 ll of rtifiil perilymph into sl tympni. The gry re depits the injetion period. Stndrd error rs re shown t t = 5, 15, nd 40 s. The shpe of the men inner er pressure urve t the strt of injetion (time xis orresponds with Fig. 1). The reording (filled irles) is fitted (roken line) with n exponentil funtion, yielding time onstnt s. The sme is done for inner er pressure reovery diretly fter injetion pressure (P) pressure (P) = 1.16 s 400 pressure (P) = 1.25 s to n *600 P higher level within seonds. At the end of injetion, the pressure immeditely deresed, returning to its initil vlue within seonds. This pttern ws onsistently oserved in ll individul experiments. The time onstnt for inrese nd susequent derese of inner er pressure were 1.25 nd 1.16 s, respetively (Fig. 1, ). The verged f 1 signl in the CM (Fig. 2) showed mirrored ehvior in respet to the inner er pressure. Diretly fter the strt of injetion, the f 1 CM deresed y 30 lv. The orresponding time onstnt ws 1.17 s (Fig. 2), whih is very lose to the time onstnt for pressure inrese. Conversely, t the end of injetion the f 1 mplitude returned to its initil vlue with s = 3.49 s (Fig. 2), whih is slow with respet to the orresponding s for pressure. The verged DPOAE 2f 1 f 2 -mplitude showed similr typil ourse s the inner er pressure (Fig. 3). Almost diretly fter the strt of injetion the 2f 1 f 2 - mplitude inresed to reh mximum of 0.4 db ove its originl vlue t the end of the injetion period. When the injetion ws instntneously terminted, the mplitude dropped nd returned to slightly ove its onset level. There ws smll oservle time lg with respet to the pressure urve. The time onstnt for inrese nd Fig. 2 Averged hnges (n =8)inf 1 CM mplitude (lv) during nd fter injetion of 0.5 ll of rtifiil perilymph into sl tympni. The gry re depits the injetion period. Stndrd error rs re shown t t = 5, 15, nd 40 s. The shpe of the men f 1 CM mplitude urve t the strt of injetion (time xis orresponds with Fig. 2). The reording (filled irles) is fitted (roken line) with n exponentil funtion, yielding time onstnt s. The sme is done for f 1 CM mplitude reovery diretly fter injetion f1 CM (µv) f1 CM (µv) = 3.49 s f1 CM (µv) = 1.17 s

4 290 Eur Arh Otorhinolryngol (2008) 265: Fig. 3 Averged hnges (n =8)in2f 1 f 2 DPOAE mplitude (db SPL) during nd fter injetion of 0.5 ll of rtifiil perilymph into sl tympni. The gry re depits the injetion period. Stndrd error rs re shown t t =5, 15, nd 40 s. The shpe of the men 2f 1 f 2 DPOAE mplitude (lp) urve t the strt of injetion (time xis orresponds with Fig. 3). The reording (filled irles) is fitted (roken line) with n exponentil funtion, yielding time onstnt s. The sme is done for 2f 1 f 2 DPOAE mplitude reovery diretly fter injetion 2f1-f2 DPOAE (db SPL) 2f1-f2 DPOAE (µp) = 3.43 s f1-f2 DPOAE (µp) = 1.52 s susequent derese of the 2f 1 f 2 -mplitude were 1.52 nd 3.43 s, respetively (Fig. 3,). In Fig. 4, the typil temporry derese for the verged 2f 1 f 2 CMDP mplitude reltive to the f 1 CM is shown. The time onstnt for derese in mplitude t the onset of injetion ws 1.12 s (Fig. 4), whih orresponds with s for pressure. The reovery is somewht slower (2.28 s, Fig. 4), ut still fster thn the time onstnt for 2f 1 f 2 DPOAE reovery. Finlly, in Fig. 5, the verged results of the f 2 f 1 CMDP mplitude reltive to f 1 CM depit n inrese during injetion of rtifiil perilymph. When the injetion ws terminted, the f 2 f 1 CMDP mplitude returned to the preinjetion level. Unfortuntely, due to low signl-to-noise rtio for the f 2 f 1 CMDP, the otined urve ould not e fitted properly nd no time onstnts ould e derived. Disussion The most striking result of this study is the miniml hnge in mplitude of the DPs in oth OAE nd CM during n ute inrese of inner er pressure. Besides, mplitudes hnged to norml when inner er pressure returned to its Fig. 4 Averged hnges (n =8)in2f 1 f 2 CMDP reltive to f 1 CM mplitude during nd fter injetion of 0.5 ll of rtifiil perilymph into sl tympni. The gry re depits the injetion period. Stndrd error rs re shown t t = 5, 15, nd 40 s. The shpe of the men 2f 1 f 2 CMDP (reltive to f 1 CM) mplitude urve t the strt of injetion (time xis orresponds with Fig. 4). The reording (filled irles) is fitted (roken line) with n exponentil funtion, yielding time onstnt s. The sme is done for 2f 1 f 2 CMDP (reltive to f 1 CM) mplitude reovery diretly fter injetion 2f1-f2 CMDP / f1 CM 2f1-f2 CMDP (µv) = 2.28 s f1-f2 CMDP (µv) = 1.12 s

5 Eur Arh Otorhinolryngol (2008) 265: f2-f1 CMDP / f1 CM Fig. 5 Averged hnges (n =8)inf 2 f 1 CMDP reltive to f 1 CM mplitude during nd fter injetion of 0.5 ll of rtifiil perilymph into sl tympni. The gry re depits the injetion period initil level. These results indite dynmilly stle inner er whih is le to ope with pressure inrese of 600 P with only miniml nd reversile ohler funtion loss. Aording to Böhmer [2] this level of pressure inrese would still e in the physiologil rnge of the guine pig er vrying etween 100 nd +700 P. The inner er is n elsti struture ompletely enompssed y one, in whih pressure hnges depend on the ompline of the memrnous strutures [14]. In norml ers, hydrostti pressure in the perilymph equls pressure in the endolymph, nd pressure hnges pplied to one omprtment re immeditely trnsmitted to the other through the elsti wlls. The perilymph is in dynmi equilirium with the ererospinl fluid vi the ohler quedut. An exess of perilymph will espe through the ohler quedut restoring inner er fluid level. When inresing inner er pressure, ohler disturne with funtionl loss is expeted. The oserved derese in CM f 1 mplitude thus seems ovious. An inresed ererospinl fluid pressure lso suppresses the CM [16]. The inrese in 2f 1 f 2 DPOAE is ounterintuitive, ut in ordne with previous pulition in whih we demonstrted the sme hnge in ohler funtion s mesured y 2f 1 f 2 DPOAE s [11]. Aording to Büki et l. [3] hnge in stiffness of the ovl window might slightly improve middle er trnsfer nd thus explin the smll inrese in DPOAE mplitude. In ontrst to the 2f 1 f 2 DPOAE, the 2f 1 f 2 CMDP showed derese in mplitude during injetion. Unfortuntely, in literture very little n e found out the reltion etween DPOAE nd CMDP. Still, it were Kemp nd Brown [6] who found lose prllels in the ehvior of the 2f 1 f 2 DPOAE nd CMDP during nd fter noxi in the guine pig. Group ltenies nd third-tone suppression were found to e very similr, linking oth signls to hve ommon origin. The DPOAE nd CMDP re generted in response to the sme ousti stimuli. The ousti primries re trnsferred through the externl nd middle er to eventully reh the sensory epitheli in the ohle. The ohle s non-liner mehno-eletril trnsduer trnsforms sound pressure wves into eletril informtion. This forwrd trnsdution proess is lled the MET. The CM potentil is diret result of this proess nd is thought to e minly produed y OHCs. On the other hnd, there is lso reverse eletromehnil trnsdution in whih the OHC mehnilly influenes the virtion of the ohler prtition [7]. Sound is trnsmitted k to the externl er where the ousti DPOAE n e mesured; the kwrd trnsmission. The rise nd fll times of the DPs in response to hnges in inner er pressure were derived from the fitted urves (Figs. 2, 3, 4). As oth CMDP nd DPOAE were mesured, this yielded informtion on forwrd nd kwrd trnsmission. As ntiipted, the time onstnts for the CMDP were smller thn the time onstnts for the DPOAE. Indeed, the ltter would trvel k nd forth the middle er whih understndly osts more time. However, the differene hs not the form of simple longer dely. The explntion for this oservtion remins tenttive. The verged differenes were 0.4 nd 1.15 s for the 2f 1 f 2 DP during inrese nd derese of inner er pressure, respetively. At pressure inrese ll mesured signls lmost diretly followed the hnge in inner er pressure. Time onstnts vried from 1.17 to 1.52 s with respet to 1.25 s for the inner er pressure urve. However, with pressure derese there ws dely of 1 2 s for ll mesured signls. Mesured differenes nnot e ounted for y dely in mesuring equipment, for this ws \0.18 s. Assumed tht the overll stiffness of the stpes-ovl window-system governs the ehvior of the DPOAE 2f 1 f 2 mplitude, it is expeted tht it would follow stiffness hnges diretly nd not 2 s lter, s oserved. It is oneivle tht delyed hnge in the endoohler potentil (EP) ould e the use of differene in s vlues. However, in experiments y Slt nd DeMott [9] the EP followed temporry pressure hnge with dely of \0.1 s. So, the mehnisms for the oserved delys in inner er pressure reovery remin to e eluidted. Two-tone distortions re generted during non-liner mehnil mplifition in the ohle. Genertion of the 2f 1 f 2 depends on symmetri omponent of non-liner trnsfer funtion wheres the f 2 f 1 depends on symmetri omponents. A hnge of the operting point is refleted in the level of the f 2 f 1 distortion. In the geril low-frequeny tones of 5 Hz were used to is the position of the ohler prtition y Frnk nd Kossl [4, 5]. By low-frequeny modultion of the 2f 1 f 2 DPOAE, Bin et l. [1] desried the forwrd MET y sigmoid-

6 292 Eur Arh Otorhinolryngol (2008) 265: shped funtion, whih relted the displement of the silr memrne to the response of the hir ells. By injeting rtifiil perilymph into sl tympni of the ohle, the silr memrne is temporrily displed. Thus, y this displement towrd sl medi, the operting stte of the ohler mplifier is lso influened. We mesured smll derese in the mplitude of the 2f 1 f 2 distortion in CM (with respet to the mplitude of f 1 in CM) nd smll inrese in the f 2 f 1 in CM. Using the model y Bin et l. [1] this leds to the onlusion tht the operting point for hir ell trnsdution is somewht shifted from symmetri position. This shift leds to n inrese in even-order distortion nd derese in oddorder distortion. Along these lines of resoning we would lso expet the 2f 1 f 2 DPOAE to derese. This might e the se t the genertion sites [15], ut this effet is then more thn ompensted for y n improved middle er trnsfer. Anyhow, the oserved hnges re smll. Chnges of overll inner er fluid pressure hve minor effets on ohler funtion. It my e tht reting n rtifiil endolymphti hydrops, y injeting rtifiil endolymph into sl medi insted of sl tympni, hs lrger effet. This is urrently tested in further reserh, whih will hopefully give insight into diseses with pthologilly hnged inner er fluid volumes, s in Ménière s disese. Aknowledgments This study ws supported y the Heinsius Houolt Foundtion nd is prt of the reserh progrm of our deprtment: Communition through Hering nd Speeh. The progrm is inorported in the Sensory Systems Group of the Groningen Grdute Shool for Behviourl nd Cognitive Neurosienes (BCN). We thnk Hns Segenhout nd Wilm Hiemstr for their invlule ontriution in performing the experimentl work in this study. Referenes 2. Böhmer A (1994) Hydrostti pressure in the inner er fluid omprtments nd its effets on inner er funtion. At Otolryngol Suppl 507: Büki B, Avn P, Lemire JJ, Dordin M, Chzl J, Riri O (1996) Otoousti emissions: new tool for monitoring intrrnil pressure hnges through stpes displements. Her Res 94: Frnk G, Kossl M (1996) The ousti two-tone distortions 2f 1 f 2 nd f 2 f 1 nd their possile reltion to hnges in the operting point of the ohler mplifier. Her Res 98: Frnk G, Kossl M (1997) Aoustil nd eletril ising of the ohler prtition. Effets on the ousti two tone distortions f 2 f 1 nd 2f 1 f 2. Her Res 113: Kemp DT, Brown AM (1984) Er nl ousti nd round window eletril orreltes of 2f 1 f 2 distortion generted in the ohle. Her Res 13: Kim DO (1986) Ative nd nonliner ohler iomehnis nd the role of outer-hir-ell susystem in the mmmlin uditory system. Her Res 22: de Kleine E, Wit HP, vn Dijk P, Avn P (2000) The ehvior of spontneous otoousti emissions during nd fter posturl hnges. J Aoust So Am 107: Slt AN, DeMott JE (1998) Longitudinl endolymph movements indued y perilymphti injetions. Her Res : Slt AN, DeMott JE (1999) Longitudinl endolymph movements nd endoohler potentil hnges indued y stimultion t infrsoni frequenies. J Aoust So Am 106: Vlk WL, Wit HP, Alers FW (2004) Effet of ute inner er pressure hnges on low-level distortion produt otoousti emissions in the guine pig. At Otolryngol 124: Vlk WL, Wit HP, Alers FW (2004) Evlution of ohler funtion in n ute endolymphti hydrops model in the guine pig y mesuring low-level DPOAEs. Her Res 192: Wit HP, Thlen EO, Alers FW (1999) Dynmis of inner er pressure relese, mesured with doule-rreled miropipette in the guine pig. Her Res 132: Wit HP, Wrmerdm TJ, Alers FW (2000) Mesurement of the mehnil ompline of the endolymphti omprtments in the guine pig. Her Res 145: Withnell RH, Shffer LA, Tlmdge CL (2003) Genertion of DPOAEs in the guine pig. Her Res 178: Yoshid M, Uemur T (1991) Trnsmission of ererospinl fluid pressure hnges to the inner er nd its effet on ohler mirophonis. Eur Arh Otorhinolryngol 248: Zhng M, As PJ (1997) Effets of middle er pressure on otoousti emission mesures. J Aoust So Am 102: Bin L, Chertoff ME, Miller E (2002) Deriving ohler trnsduer funtion from low-frequeny modultion of distortion produt otoousti emissions. J Aoust So Am 112: