Change detection by thalamic reticular neurons

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1 Chnge detetion y thlmi retiulr neurons Xiong-Jie Yu 1,2, Xin-Xiu Xu 1, Shigng He 1 & Jufng He 1,2 The thlmi retiulr nuleus (TRN) is thought to funtion in the ttentionl serhlight. We nlyzed the detetion of devint ousti stimuli y TRN neurons nd the onsequenes of devine detetion on the TRN trget, the medil geniulte ody (MGB) of the rt. TRN neurons responded more strongly to pure-tone stimuli presented s devint stimuli (low pperne proility) thn those presented s stndrd stimuli (high proility) (devine-detetion index =.321). MGB neurons lso showed devine detetion in this proedure, leit to smller extent (devine-detetion index =.4). TRN neuron devine detetion either enhned (14 neurons) or suppressed (27 neurons) MGB neuronl responses to proe stimulus. Both effets were neutrlized y intivtion of the uditory TRN. Devine modultion effets were ross-modl. Devine detetion proly uses TRN neurons to trnsiently detivte surrounding TRN neurons in response to fresh stimulus, ltering uditory thlmus responses nd induing ttention shift. The dorsl thlmus relys sensory nd motor informtion to the ererl ortex nd reeives strong modultory input k from the ortex. Both thlmoortil nd ortiothlmi projetions send ollterls to the TRN of the ventrl thlmus 1,2. The TRN onsists of GABA-ontining (GABAergi) neurons, whih only projet k to the dorsl thlmus 2 4. These neurons exhiit urst firing tht, under different onditions, ours in synhrony with severl frequenies of rhythmi osilltions in the ortiothlmi networks 8. The lotion of the TRN, long with its firing ptterns nd unique onnetions, hs led to the proposl tht the TRN is importnt in the internl ttentionl serhlight 9 nd for oordinting multiple neuronl proesses y linking speifi nd nonspeifi pthwys. In the uditory setor of the TRN, neurons show vrious response ptterns to ousti stimuli, inluding the urst response, phsi ON response nd OFF response 11,12. TRN neurons re tuned to rod rnge of frequenies 12. Novel stimuli ttrt our ttention s result of devint stimulus preferene (DSP), phenomenon in whih neurons show inresed responsiveness to devint stimuli. Suh neurons hve een identified in severl rin regions, inluding the visul nd uditory ortex nd the inferior olliulus 13,14. Compred with thlmoortil neurons in the MGB, TRN neurons in the uditory setor show greter dpttion to repeted stimuli. Considering the potentil role of the TRN in the internl ttentionl serhlight or ttention shift, fst-dpting TRN neurons re likely to exhiit DSP. We used in vivo extrellulr reording nd n oddll proedure to nlyze DSP (tht is, devine detetion) in TRN neurons. We then investigted the funtionl implitions of TRN neuron DSP y exmining how the DSP ffets the responses of thlmoortil neurons ( TRN output trget) to uditory stimuli. Inter-modlity intertion ws investigted y exmining how preeding light stimulus ffets the uditory responses of the MGB neurons. RESULTS Devine detetion y TRN neurons To determine whether the fst dpttion of TRN neurons is stimulus speifi, we used n oddll proedure with omintion of two puretone stimuli (with stndrd frequeny f 1 of 14 khz nd devint frequeny f 2 of 8 khz). A lok of pure-tone stimuli onsisting of f 1,f 1,f 1,f 1,f 1,f 1 nd f 2 ws repeted 3 times. The f 1 stimuli with n interstimulus intervl (ISI) of ms eliited strong responses in oth the first nd seond trils of lok 1; however, dpttion ws pprent in susequent trils of this lok (trils 3 6; Fig. 1). Introdution of the f 2 stimulus t tril 7 evoked strong response (lok 1; Fig. 1). Reintrodution of the f 1 stimuli in loks 2 resulted in wek or sent responses. However, f 2 stimuli eliited strong responses in ll loks (Fig. 1). As shown y the rster displys of ll 3 loks (Fig. 1), the f 2 stimulus eliited muh stronger responses thn the stndrd frequeny. Next, the stndrd nd devint frequenies were swpped. The new stndrd frequeny stimulus (f 2, 8 khz) eliited strong responses in the first tril, wek responses in trils 2 4 nd no response in trils nd 6 (lok 1; Fig. 1). However, the new devint stimulus (f 1,14kHz) eliited strong response (lok 1; Fig. 1). The f 2 stimulus evoked strong response t tril 1 in lok 2, ut evoked wek or sent response in trils 2 6 (Fig. 1). Reintrodution of the oddll stimulus t lok 2 ws ssoited with only wek response. The neurons showed good responses to the first trils of the stndrd stimuli in the loks nd wek responses to the following trils of the stndrd stimuli (Fig. 1). Responses to devint f 1 stimuli (Fig. 1) were muh stronger thn those to stndrd f 1 stimuli of the sme frequeny (Fig. 1). The sme ws true of devint (Fig. 1) nd stndrd f 2 stimuli (Fig. 1). A omprison of verge responses to f 2 (Fig. 1) ndf 1 (Fig. 1d) s either stndrd or devint stimuli lerly showed tht the TRN neurons preferred devint stimuli. Anlysis of the TRN neuron exposed to pure 1 CAS-Hong Kong Joint Reserh Lortory for Visuo-Auditory Integrtion, Institute of Biophysis, Chinese Ademy of Sienes, Beijing, Chin. 2 Lortory of Applied Neurosiene, Deprtment of Rehilittion Sienes, The Hong Kong Polytehni University, Hung Hom, Kowloon, Hong Kong. Correspondene should e ddressed to J.H. (rsjufng@polyu.edu.hk) or S.H. (shignghe@moon.ip..n). Reeived Ferury; epted 19 June; pulished online 16 August 9; doi:.38/nn.2373 NATURE NEUROSCIENCE VOLUME 12 [ NUMBER 9 [ SEPTEMBER 9 116

2 Figure 1 Responses of TRN neuron to pure-tone stimuli of two frequenies presented in n oddll proedure. () Responses to 14-kHz frequeny tones (f 1 ) when presented s the stndrd tone nd to 8-kHz frequeny tones (f 2 ) when presented s the devint tone. Neuronl responses to the first of 3 loks re presented in the upper prt. Trings from the five loks re red from left to right nd top to ottom. The order of presenttion ws lok 1 (f 1,f 1, y, f 1,f 2 ), lok 2 (f 1,f 1, y, f 1,f 2 ), y lok 3 (f 1, y, f 1,f 2 ). Bloks were ontiguous in time. The lower prt of the figure shows the rster displys of 3 loks of seven sequentil tones. Only the first ms of the responses re shown in the tres nd rster disply, lthough the ISI ws ms. () Responses on reversl of the stndrd nd devint tones. (,d) Peristimulus time histogrms showing responses (tht is, firing rte) to stndrd nd devint stimuli. ** P o.1 (ANOVA). (e) Trk of reording eletrode, s shown y Nissl stin (left, the rrow indites the reording site) nd responses s funtion of stimulus frequeny t 7-dB SPL (right). Sle r represents mm. Pure tones of vrying frequenies (6 24 khz), seprted y 2-kHz intervl, were rndomly presented, with eh frequeny eing presented in trils. tones t 7-dB sound pressure level (SPL) reveled tht the responses rnged from 8 to khz (Fig. 1e). Inresing the ISI to 1 s nd ltering the pperne rtio to 9:1 of the stndrd to devint stimuli yielded similr results (see exmple of nother neuron in Fig. 2,, where f 1 ¼ 19.2 khz nd f 2 ¼ 21.2 khz, nd men dt from 19 neurons in Fig. 2). Tht is, the neuron hd stronger response to pure-tone stimuli when they were presented s the devint stimulus thn when they were presented s the stndrd stimulus. Devine detetion y MGB neurons Using the sme stimulus proedure nd prmeters, we found tht MGB neurons showed less differene in responses to pure-tone stimuli with different presenttion proilities (stndrd versus devint). In one MGB neuron (Fig. 3), we oserved omprle responses to f 1 (17.12 khz) stimuli s stndrd (9% frequeny of presenttion) nd s devint (%) s well s to f 2 (18.92 khz) stimuli s stndrd nd s devint. In ontrst, nother MGB neuron (Fig. 3) showed ovious differenes in responses to f 1 (11.42 khz) stimuli s stndrd nd s devint s well s to f 2 (12.61 khz) stimuli s stndrd nd s devint. Tril numer Tril numer 9% f 1 9% f 2 % f 1 % f 2 Firing rte (Hz) Blok numer Blok numer Firing rte (Hz) Order of the stimuli presenttion f 1 f 1 f 1 f 1 f 1 f 1 f 2 f 2 f 2 f 2 f 2 f 2 f 2 f 1 f 1 s devint f 1 s stndrd * ** Anlysis of the men responses of 41 MGB neurons using the sme proedure nd prmeters reveled trend towrd greter responses to devint stimuli thn stndrd stimuli (P ¼.8; Fig. 3). We dopted the stimulus-speifi dpttion index for eh frequeny s frequeny-speifi index SI(f i ), whih ws lulted dðf iþ sðf iþ using the eqution SIðf i Þ¼dðf ; where d(f iþ+sðf iþ i) nd s(f i ) were responses to the devint nd stndrd frequeny, respetively 16.In SI(f 2 ) versus SI(f 1 ) stter plots, the points of TRN neurons devited from the digonl more thn those of MGB neurons (Fig. 4). The greter the devition of the point from the digonl, the greter the stimulus-speifi dpttion/devine detetion tht the neuron showed. A normlized devine-detetion index SI, defined s dðf 1Þ+dðf 2Þ sðf 1Þ sðf 2Þ dðf 1Þ+sðf 1Þ+dðf 2Þ+sðf 2Þ Firing rte (Hz) e 4 3 TRN ** 8 khz s devint 8 khz s stndrd 14 khz s devint 14 khz s stndrd ; ws pproximtely 8% greter in TRN neurons (n ¼ 19) thn in MGB neurons (n ¼ 41) (.321 ±.1 versus.4 ±., men ± s.e.m., P o.1). The devine proedure lters MGB responses To investigte the funtionl implition of DSP in TRN neurons, we exmined neuronl responses in the MGB using devine proedure with n dded proe stimulus. The ojetive ws to f 2 s devint f 2 s stndrd * Tones s devint Tones s stndrd 1 n TRN = 19 Firing rte (Hz) 3 d ** 7-dB SPL Frequeny (khz) Figure 2 Differentil responses of TRN neurons to pure-tone stimuli of two frequenies presented in n oddll proedure. () Rster displys showing responses to tones of two frequenies (f 1, 19.2 khz; f 2, 21.2 khz) when presented s f 1 stndrd (9% ppering proility) nd f 2 devint (% ppering proility) or f 1 devint nd f 2 stndrd. The ISI ws 1 s. We smpled responses to the stndrd frequeny from tril numers 81 to ( trils), wheres we smpled ll of the responses to the devint frequeny ( trils,: numers 1 to ). () Peristimulus time histogrms (PSTHs) of responses shown in. () PSTHs showing the men responses of 19 TRN neurons. * P o. nd ** P o.1 (ANOVA). The ISI ws 1 s VOLUME 12 [ NUMBER 9 [ SEPTEMBER 9 NATURE NEUROSCIENCE

3 tivte the TRN neurons tht exerted inhiition/disinhiition on the reording MGB neuron nd to investigte whether the devine proedure modulted MGB neuronl responses to the proe stimulus. The devine proedure onsisted of sequene of pure-tone f 1 stimuli (stndrd), pure-tone f 2 stimuli (devint), nd proe stimulus (p), f 1, f 1, f 1, f 1, f 1, f 1, f 1, f 1, f 1, f 1, f 2 nd p (proe stimulus: noise-urst or pure tone of the est frequeny, f 3 ). In the ontrol proedure, the f 2 in the devine proedure ws repled y f 1. When n MGB neuron ws seleted, preliminry sreening ws performed to ensure tht the MGB neuron responded to the proe stimulus of either noise-urst or pure-tone of f 3 nd filed to respond to pure-tone f 1 or f 2 stimulus. Two of the MGB neurons tht we exmined (Fig. ) responded to the proe stimuli under oth ontrol nd devine proedures, ut they filed to respond to pure tones of f 1 (3.44 khz) nd f 2 (33.64 khz) (nother two exmples of MGB neurons re shown in Supplementry Fig. 1). One neuron (Fig. ) showed weker responses to the proe stimulus (p: f 3 ¼ 8-kHz tones) under the devine proedure thn under the ontrol proedure. We nmed this MGB neuron devinesuppressed neuron. Conversely, nother neuron (Fig. ) hd stronger response to the proe stimulus (p: noise) in the devine proedure nd ws referred to s devine-enhned neuron. We used n index to mesure the modultory effet of the devine proedure on MGB neuronl response (IDC), defined s RD RC R D+R C ; where R D nd R C re the responses to the proe stimulus in the devine nd ontrol proedures, respetively). A totl of 77 MGB neurons were exmined using the devine proedure. Of these, 27 were devinesuppressed neurons nd 14 were devine-enhned neurons (Fig. ). Aordingly, the verge mplitude of the suppression (IDC ¼.263 ±.47, n ¼ 27) showed trend of eing greter thn the mplitude of enhnement (IDC ¼.144 ±.37, n ¼ 14) (P ¼.98). SI (f 2 ) 1.. TRN MGB Tril numer Tril numer Firing rte (Hz) 3 MGB neuron 1 MGB neuron 2 9% f 1 9% f 2 9% f 1 9% f 2 % f 1 f 1 s devint f 1 s stndrd % f 2 % f 1 % f f 2 s devint f 2 s stndrd Figure 3 Differentil responses of MGB neurons to pure-tone stimuli presented in n oddll proedure. () Rster displys (upper two rows) nd Peristimulus time histogrms (PSTHs) (lower row) showing responses to tones of two frequenies (f 1, khz; f 2, khz) when presented s f 1 stndrd/f 2 devint or f 2 stndrd/f 1 devint. () Rster displys (upper two rows) nd PSTHs (lower row) showing responses to tones of two frequenies (f 1, khz; f 2, khz) when presented s f 1 stndrd/f 2 devint or f 2 stndrd/f 1 devint. () PSTHs showing the men responses of 41 MGB neurons. The ISI ws 1 s. ** P o.1;, not signifint (P ¼.88). f 1 s devint f 1 s stndrd ** ** f 2 s devint f 2 s stndrd Tones s devint Tones s stndrd 1 n MGB = 41 TRN intivtion suppresses devine modultion of MGB To determine whether the hnges in MBG neuron responses to the proe stimulus of the devine proedure were used y the DSP of TRN neurons, we investigted the effet of lidoineindued TRN intivtion on MGB neuronl responses to the proe stimulus (Supplementry Fig. 2). Before TRN intivtion, one MGB neuron (Fig. 6,) exhiited responses to the proe stimulus (noise urst), ut not to the pure tone of f 1 (3.44 khz) or f 2 (33.64 khz) (Fig. 6). Responses to the proe stimulus were signifintly weker (P o.1) under the devine proedure thn under the ontrol proedure (Fig. 6). After TRN intivtion, the neuron responded to the pure-tone stimulus f 2, ut not f 1. In ddition, responses to the proe stimuli were signifintly inresed under oth ontrol (182%, P o.; Fig. 6 ) nd devine proedures (3%, P o.1; Fig. 6 ), implying tht the TRN strongly inhiited the MGB. However, responses to the proe stimuli did not differ etween the ontrol nd devine proedures fter TRN intivtion (P ¼.27; Fig. 6,). This result indites tht the devine modultion of the MGB ws primrily ttriutle to the TRN. Firing rte (Hz) SI (f 1 ) Figure 4 Devine preferenes of TRN nd MGB neurons. Stter plot of SI(f 2 ) versus SI(f 1 ) in ll TRN (red) nd MGB neurons (lk). MGB Responses re modulted y preeding light stimulus The ove experiments were designed to investigte devine modultion in the sme modlity. We next investigted whether MGB neuronl responses ould e modulted in ross-modl mnner. To do this, we introdued light stimulus efore the proe sound stimulus (light-sound, light-sound proedure). One of the MGB neurons tht we exmined (Fig. 7,) hd signifintly stronger NATURE NEUROSCIENCE VOLUME 12 [ NUMBER 9 [ SEPTEMBER

4 6 MGB neuron 3 Control proedure Devine proedure 6 MGB neuron 4 Control proedure Devine proedure Tril numer 3 1, 2, 1, 2, 3 1, 2, 1, 2, 18 n MGB = 77 Tril numer f 1 f 1 p f 1 f 2 p f 1 f 1 p f f 2 p Count 12 6 Firing rte (Hz) 1, , 1,6 1,6 1, p p p p ** Control proedure * Control proedure Devine proedure Devine proedure 1,66 1,6 1, responses to the proe stimulus (noise urst) in the light-sound proedure thn in the sound proedure without the light stimulus (sound proedure) (Fig. 7,). This neuron showed no responses to light stimuli (Supplementry Fig. 3). On TRN intivtion, the neuronl response to the proe stimulus inresed signifintly in oth the sound proedure nd the light-sound proedure (P o.1; Fig. 7 ), gin implying tht the TRN strongly inhiits the dorsl thlmus. However, fter TRN intivtion, the neuronl responses to the proe stimuli in the sound nd light-sound proedures did not differ (P ¼.26; Fig. 7,). Notly, 1,6 1,6 1, 1,6 1,6 1, 1,66 1,6 1,6 1,6 1.. IDC. 1. Figure Effet of the devint-stimulus proedure on MGB neuronl responses to proe stimulus. (,) Rster displys showing responses of two MGB neurons to sequentil uditory stimulus proedures. In the ontrol stimulus proedure (left olumns), pure tones of f 1 (lue rs) were presented 11 times efore proe stimulus (lk r). In the devine proedure (right olumns), pure tones of f 1 (lue rs) were presented ten times efore devint stimulus, pure tone of f 2 (red r) nd proe stimulus (lk r). The tones nd proe stimuli hd -ms durtion with -ms rise-fll time. In, f 1 ¼ 3.44 khz, f 2 ¼ khz nd p ¼ 8kHz.In, f 1 ¼ 3.44 khz, f 2 ¼ khz nd p ¼ noise urst. Upper rster displys show neuronl responses during the whole time (2 s), wheres the lower ones show neuronl responses t the first ms fter the proe stimulus (mgnified 1: from the upper displys). Peristimulus time histogrms of MGB neuronl responses re shown t the ottom. * P o. nd ** P o.1 (ANOVA). () Distriution of the modultory effet (IDC) of TRN devine detetion on MGB neurons. Blk rs showed modultion effets ove the threshold nd gry rs showed modultion effets elow the threshold. Figure 6 Effet of TRN intivtion on modultion of MGB neuronl responses y devint proedures. (,) MGB neuronl responses to sequentil stimulus proedures in the presene of n intt () orintivtedtrn(). In the ontrol stimulus proedure, pure tones of f 1 were presented 11 times efore proe stimulus (left olumns). In the devine proedure, pure tones of f 1 were presented ten times efore devint stimulus, pure tone of f 2 nd the proe stimulus (right olumns). Neuronl responses for the whole 2 s (upper row) nd the first ms fter the proe stimulus (lower row, mgnifition 1:) re shown. f 1 ¼ 3.44 khz (lue rs), f 2 ¼ khz (red rs) nd p ¼ noise urst (lk rs). () Peristimulus time histogrms of MGB neuronl responses re shown t the ottom. ** P o.1;, not signifint, P ¼.27 (ANOVA), ontrol versus devine proedures under the sme onditions. # P o. nd ## P o.1, intt versus intive TRN. Tril numer Tril numer Firing rte (Hz) , 2 ** 1, Control proedure 1, 1,6 TRN intivtion olished the effet of the preeding light stimulus on the uditory response of the MGB neuron. We exmined the effet of light stimulus on the uditory responses of 118 MGB neurons. Of these neurons, 23 hd enhned uditory responses following the light stimulus nd hd suppressed responses. The mgnitude of the enhnement effet ws greter thn tht of the suppressive effet (.176 ±.26 versus.73 ±., P o.1). TRN intt 6 3 2, 1, 2, 6 3 1,6 1, p p p p 1,66 Devine proedure Control proedure Devine proedure 6 6 1,6 # ## 1,6 1, , 1, 2, 1,6 TRN intivted ,6 1, 1, 2, 1,6 1,6 1, 1,66 1, VOLUME 12 [ NUMBER 9 [ SEPTEMBER 9 NATURE NEUROSCIENCE

5 Tril numer 3 TRN intt 3 TRN intivted d 4 n MGB = Firing rte (Hz) 3 S L S S L S With light With light Without light Without light ## ** ## Count 1.. IDC. 1. Figure 7 Effet of preeding light stimulus on MGB neuronl responses to sound stimuli. (,) Responses to light nd sound stimulus proedures in the presene of n intt () or intive TRN (). MGB neurons were tested in ontrol proedure (left pnels) in whih only proe stimulus (S) ws presented or ross-modlity proedure (right pnels) in whih light stimulus (L, green r) ws presented efore the proe stimulus (S, lk r). The proe stimulus ws -ms noise urst with -ms rise-fll time. () Peristimulus time histogrms of responses to the proe stimulus in the presene of n intt (left) or intive TRN (right). ** P o.1 (ANOVA) for sound versus light-sound proedures under the sme onditions nd ## P o.1 (ANOVA) for intt versus intive TRN. (d) Distriution of the ross-modlity modultory effet on MGB neurons. The modultory effet (IDC) ws defined s RLS RS ;where R LS nd R RLS+RS S re the responses to the proe in light-sound nd sound proedures, respetively. Negtive vlues indite n inhiitory effet of the light stimulus nd positive vlues indite n enhned effet. Blk rs show modultion effets ove the threshold nd gry rs show modultion effets elow the threshold. DISCUSSION Using the oddll proedure, we found tht TRN neurons hve sustntilly stronger responses to pure-tone stimulus when it ppers s the devint stimulus thn when it ppers s the stndrd stimulus. Although some MGB neurons lso hd devine preferene, the devine-detetion index ws sustntilly greter in TRN neurons thn in MGB neurons. The im of the seond prt of our study ws to investigte the funtionl implition of DSP y TRN neurons. Of the 77 MGB neurons tht we exmined with the devine proedure, 41 were modulted y the devine proedure, with 27 eing devinesuppressed nd 14 eing devine-enhned neurons. The mplitude of the suppression ws greter thn tht of the enhnement. The effet of devine detetion on MGB neuronl responses to the proe stimulus vried with the frequeny used in the testing proedure (Supplementry Fig. 4). After the TRN ws intivted, the MGB neurons showed sustntil inreses in their responses to the proe stimuli under oth ontrol nd devine proedures. However, responses to the proe stimuli did not differ etween the ontrol nd devine proedures, inditing tht the modultory effet of devine detetion on the MGB neurons were mostly ttriutle to TRN neurons. Of the 118 MGB neurons tht we exmined for the ross-modl modultory effet, 23 hd enhned uditory responses following introdution of the light stimulus nd neurons hd wekened responses. The TRN neurons send strong inhiitory projetions to the dorsl thlmus 1 4. Our finding tht MGB neurons showed sustntilly inresed responses to the sme uditory stimuli fter the uditory TRN ws intivted (Fig. 6) onfirms previous findings tht TRN inhiition of the dorsl thlmus is strong nd my lst for few seonds 17. Injetion of lidoine in the TRN not only intivtes TRN neurons, ut would lso e expeted to lok the ommunition of the xons pssing through the TRN (tht is, the ortiothlmi nd thlmoortil xons). Although the xons of the thlmoortil neurons were ffeted, the effet of the lidoine ws lolized nd did not lter the genertion of tion potentils in the MGB (Figs. 6 nd 7). With the use of ontrol onditions efore nd fter the drug pplition, the modultion effet of the diret ortiothlmi projetion to the MGB ws shown to e miniml s this pthwy is primrily exittory 17,18 nd MGB neurons showed inresed responses (Figs. 6 nd 7). Using n oddll proedure 16, we hve shown for the first time, to the est of our knowledge, tht fst-dptive TRN neurons in the uditory setor hve DSP. In the uditory system, DSP hs previously een reported in inferior olliulus neurons 13,14 nd ortil neurons 16. Using the sme proedure nd prmeters, we found tht MGB neurons hd smller DSP thn TRN neurons. Although MGB neurons were lso sensitive to devint stimuli, TRN neurons were more sensitive with ISIs on the order of hundreds of milliseonds, phenomenon tht is linked to novelty detetion 14,16. Using proedure tht ould exmine the resulting effet of devine detetion y the TRN neurons, we found tht some MGB neurons showed response enhnement, wheres others showed response suppression. The suppressive effet of DSP y TRN neurons on MGB neurons is understood to e used diretly y the inhiitory input to the MGB neurons. However, the enhnement of MGB neuronl responses is proly used y disinhiition of the TRN neurons or thlmi interneurons. The DSP of one TRN neuron would inhiit other TRN neurons through their ollterls either synptilly or eletrilly 3, They my lso inhiit thlmi interneurons. In either se, this would result in the disinhiition in the dorsl thlmus (Supplementry Fig. ). Shifts in the ttentionl serhlight during novelty detetion my involve projetions from the prefrontl ortex nd from the limi system to the TRN, whih re referred to s the nonspeifi pthwys 1,,22. The distriuted limi-ortil network is n essentil omponent for memory retrievl nd novelty detetion 23 2.Deresed tivity in the nuleus umens, whih provides other limi inputs NATURE NEUROSCIENCE VOLUME 12 [ NUMBER 9 [ SEPTEMBER

6 to the TRN, ws found in lrge group of individuls with tinnitus 26. Deresed limi innervtion of the TRN neurons would e expeted to result in disinhiition of the MGB. This might ount for inresed tivity in the uditory pthwy of people suffering from tinnitus 26. The nonspeifi pthwys my supplement speifi pthwys to the TRN,22,27. Indeed, the integrtion of speifi nd nonspeifi pthwys through the TRN is involved in ttention nd ognition. Some neurons in the TRN re multi-modl nd hve projetions tht ross nulei, suggesting tht the TRN hs n intermediry role 11,12,28. The ft tht MGB neuronl responses to sound stimuli were modulted y preeding light stimulus indites tht the intermodl intertion ould our t the TRN (Supplementry Fig. ). Devine detetion y TRN neurons would enle these ells to reognize new stimulus, while inhiiting other TRN neurons through their ollterls. Our finding tht the MGB neuron ws influened y preeding pure tones of frequeny to whih it normlly did not respond (Fig. nd Supplementry Fig. 1) indites tht the modultory effet spreds eyond its response frequeny rnge. The devine detetion effet would indue shift in the serhlight to diret ttention to the stimulus 9 nd the shift of ttention through the TRN ould e ross-modl. The strong dpttion or hitution tht TRN neurons undergo in response to repetitive stimuli might redue the redundny of the environment nd shrpen ontrsts etween differing stimuli. Finlly, in the devine proedure tht we used, some MGB neurons shifted from toni mode to urst mode (Supplementry Fig. 1). The step funtion of the input-output reltionship in urst mode (tht is, the mode in whih thlmi neurons re hyperpolrized y TRN input) would enle thlmi neurons to respond more strongly thn if they were in the toni mode This is nother potentil mehnism underlying devine detetion-indued enhnement of the MGB neuronl response nd my supplement those tht we propose here (Supplementry Fig. ). METHODS Methods nd ny ssoited referenes re ville in the online version of the pper t Note: Supplementry informtion is ville on the Nture Neurosiene wesite. ACKNOWLEDGMENTS The uthors thnk A. Plmer nd M. Wlle for their ritil redings nd omments. This work ws supported y the Nturl Siene Foundtion of Chin (Overses Coopertion Fund) nd the Hong Kong Grnts Counil (PolyU 412/6M). AUTHOR CONTRIBUTIONS X.-J.Y., S.H. nd J.H. designed the experiments. X.-J.Y. nd X.-X.X. performed the experiments. X.-J.Y. nd J.H. nlyzed the results nd wrote the mnusript. Pulished online t Reprints nd permissions informtion is ville online t reprintsndpermissions/. 1. Jones, E.G. Some spets of the orgniztion of the thlmi retiulr omplex. J. Comp. Neurol. 162, (197). 2. Steride, M., Jones, E.G. & MCormik, D.A. Thlmus: Orgniztion nd Funtion (Elsevier Siene, Oxford, 1997). 3. Yen, C.T., Conley, M., Hendry, S.H. & Jones, E.G. The morphology of physiologilly identified GABAergi neurons in the somti sensory prt of the thlmi retiulr nuleus in the t. J. Neurosi., (198). 4. Houser, C.R., Vughn, J.E., Brer, R.P. & Roerts, E. GABA neurons re the mjor ell type of the nuleus retiulris thlmi. Brin Res., (198).. Deshênes, M., Mdrig-Domih, A. & Steride, M. Dendrodendriti synpses in the t retiulris thlmi nuleus: struturl sis for thlmi spindle synhroniztion. Brin Res. 334, (198). 6. Steride, M. & Deshênes, M. The thlmus s neuronl osilltor. Brin Res. 3, 1 63 (1984). 7. Bl, T. & MCormik, D.A. Mehnisms of osilltory tivity in guine-pig nuleus retiulris thlmi in vitro: mmmlin pemker. J. Physiol. (Lond.) 468, (1993). 8. MCormik, D.A. & Prine, D.A. Aetylholine indues urst firing in thlmi retiulr neurones y tivting potssium ondutne. Nture 319, 42 4 (1986). 9. Crik, F. Funtion of the thlmi retiulr omplex: the serhlight hypothesis. Pro. Ntl. Ad. Si. USA 81, (1984)..Llinás, R. & Pré, D. in Thlmus (eds. Steride M., Jones E.G. & MCormik D.A.) 1 16 (Plenum, New York, 1997). 11. Shosku, A. & Sumitomo, I. Auditory neurons in the rt thlmi retiulr nuleus. Exp. Brin Res. 49, (1983). 12. Simm, G.M., de Riupierre, F., de Riupierre, Y. & Rouiller, E.M. Dishrge properties of single units in uditory prt of retiulr nuleus of thlmus in t. J. Neurophysiol. 63, 21 (199). 13. Mlone, B.J. & Semple, M.N. Effets of uditory stimulus ontext on the representtion of frequeny in the geril inferior olliulus. J. Neurophysiol. 86, (1). 14.Pérez-González, D., Mlmier, M.S. & Covey, E. Novelty detetor neurons in the mmmlin uditory midrin. Eur. J. Neurosi. 22, ().. Yu, X.-J., Xu, X.-X., Chen, X., He, S.-G. & He, J. Slow reovery from exittion of thlmi retiulr nuleus neurons. J. Neurophysiol. 1, (9). 16. Ulnovsky, N., Ls, L. & Nelken, I. Proessing of low-proility sounds y ortil neurons. Nt. Neurosi. 6, (3). 17. Zhng, Z. et l. Cortiofugl projetion inhiits the uditory thlmus through the thlmi retiulr nuleus. J. Neurophysiol. 99, (8). 18. He, J., Yu, Y.Q., Xiong, Y., Hshikw, T. & Chn, Y.S. Modultory effet of ortil tivtion on the lemnisl uditory thlmus of the Guine pig. J. Neurophysiol. 88, 4 (2). 19. Lndismn, C.E. et l. Eletril synpses in the thlmi retiulr nuleus. J. Neurosi. 22, 2 9 (2).. Long, M.A., Lndismn, C.E. & Connors, B.W. Smll lusters of eletrilly oupled neurons generte synhronous rhythms in the thlmi retiulr nuleus. J. Neurosi. 24, (4). 21. Wrren, R.A., Agmon, A. & Jones, E.G. Osilltory synpti intertions etween ventroposterior nd retiulr neurons in mouse thlmus in vitro. J. Neurophysiol. 72, (1994). 22. Zikopoulos, B. & Brs, H. Prefrontl projetions to the thlmi retiulr nuleus form unique iruit for ttentionl mehnisms. J. Neurosi. 26, (6). 23. Knight, R. Contriution of humn hippompl region to novelty detetion. Nture 383, (1996). 24. Tiitinen, H., My, P., Reinikinen, K. & Náátánen, R. Attentive novelty detetion in humns is governed y pre-ttentive sensory memory. Nture 372, 9 92 (1994). 2. Crpenter, G.A. & Grosserg, S. Norml nd mnesi lerning, reognition nd memory y neurl model of ortio-hippompl intertions. Trends Neurosi. 16, (1993). 26. Mühlu, M. et l. Struturl rin hnges in tinnitus. Cere. Cortex 16, (6). 27. Llinás, R.R. & Pré, D. Of dreming nd wkefulness. Neurosiene 44, 21 3 (1991). 28. Crtree, J.W. & Is, J.T. New intrthlmi pthwys llowing modlity-relted nd ross-modlity swithing in the dorsl thlmus. J. Neurosi. 22, (2). 29. Jhnsen, H. & Llinás, R. Eletrophysiologil properties of guine-pig thlmi neurones: n in vitro study. J. Physiol. (Lond.) 349, 226 (1984). 3.Jhnsen, H. & Llinás, R. Ioni sis for the eletro-responsiveness nd osilltory properties of guine-pig thlmi neurones in vitro. J. Physiol. (Lond.) 349, (1984). 31. Shermn, S.M. A wke-up ll from the thlmus. Nt. Neurosi. 4, (1). 32. Guillery, R.W. & Shermn, S.M. Thlmi rely funtions nd their role in ortioortil ommunition: generliztions from the visul system. Neuron 33, (2). 117 VOLUME 12 [ NUMBER 9 [ SEPTEMBER 9 NATURE NEUROSCIENCE

7 ONLINE METHODS Animls. We used mle nd femle Wistr rts (28 36 g) with len externl ers. Anesthesi ws indued with 1. g per kg of ody weight urethne (% (wt/wt) solution, intrperitonel, Sinophrm Chemil Regent) nd mintined throughout surgery nd reording with. g per kg per h urethne. Atropine sulfte (. mg per kg, suutneous) ws dministered min efore indution of nesthesi to inhiit trhel seretion. A lol nestheti (2% (wt/vol) xyloine) ws lierlly pplied to the wound. Rts were surgilly prepred s desried previously Briefly, rts were mounted in stereotxi devie nd midline inision ws mde in the slp. A rniotomy ws performed to vertilly ess the MGB nd the uditory setor of the TRN nd the dur mter ws then removed. Throughout the experiment, the rt ws kept on heting lnket nd ody temperture ws mintined t C. All niml proedures were pproved y the Animl Sujets Ethis Suommittees of the Institute of Biophysis, Chinese Ademy of Sienes nd The Hong Kong Polytehni University. Reording. Tungsten miroeletrodes with impednes of 2 7 MO (Frederik Her) were stereotxilly implnted into the TRN nd MGB from the top of the rin, ording to rt rin tls 36. The vertil oordinte of the eletrode rry ws mesured from point slightly ove the ortil surfe. For reording, eletrodes were positioned with stepping-motor mirodrive, whih ws ontrolled from outside the soundproof room. The signl reorded y the miroeletrode, together with the ousti stimulus signl, ws mplified nd stored using OpenEX (Tuker-Dvis Tehnologies) nd Axosope softwre (Axon Instruments). The time of spike ourrene reltive to stimulus delivery ws lulted with Mtl softwre (Mthworks). Antomil onfirmtion. After the reording session, the rts were deeply nesthetized with sodium pentoritl. They were killed y trnsrdil perfusion of with.9% (wt/vol) sline followed y 4% (wt/vol) prformldehyde in.1 M phosphte uffer (ph 7.3). The rins were removed nd stored overnight in.1 M phosphte uffer ontining 3% (wt/vol) surose. Trnsverse thlmi setions of -mm thikness were stined using the Nissl method. Imges of Nissl-stined setions were overlid onto physiologil mp using the eletrode penetrtion trks nd lesions for guidne. We used eight rts to onfirm the orret positioning of the eletrodes. Aousti stimuli. Aousti stimuli were digitlly generted using omputerontrolled Auditory Worksttion (Tuker-Dvis Tehnologies) nd delivered to the er vi oupled eletrostti speker (EC1, Tuker-Dvis Tehnologies) mounted in proe. The SPL of the output of the proe ws lirted with ondenser mirophone (Center Tehnology). Oddll proedure. Stimulus frequenies eliiting optiml responses in TRN nd MGB neurons were determined. During the mjority of the experiments, we seleted two frequenies, f 1 nd f 2, tht ordered entrl frequeny (f 1 f 2 ) 1/2 equling its est frequeny nd tht hd rtio of.141 otves. Stimulus intensity ws set to 7 db. In most ses, the stndrd nd devint stimuli ourred t rtio of 9 to 1. Otherwise, this rtio ws 6 to 1, s speified. The stimuli were presented in repetitive sequene of nine stndrd tones followed y single devint tone. The ISI ws either ms or 1, ms, with sttistil omprisons eing performed on dt otined with n ISI of 1, ms. PSTHs were lulted from over 18 trils for the stndrd tone nd trils for the devint tone. Anlysis of MGB neuronl responses using the devine proedure. The devine proedure ws designed to unover possile modultory effets of the TRN devine-preferene on the responses of MGB neurons to uditory stimuli. A few prmeters of the devine proedure were modified to eliit mximl effet. The devine proedure onsisted of the following sequene of stimuli: f 1,f 1,f 1,f 1,f 1,f 1,f 1,f 1,f 1,f 1,f 2 nd p. In the ontrol proedure, f 2 ws repled with f 1 to generte the following sequene: f 1,f 1,f 1,f 1,f 1,f 1,f 1,f 1,f 1, f 1,f 1 nd p. The rtionle for this ws tht the TRN neurons would e tivted y f 2 in the devine proedure, ut not y the finl f 1 in the ontrol proedure. MGB neuronl responses to the proe stimulus were ompred under the devine nd ontrol proedures. Differenes would show the modultory effet of the TRN devine preferene. To exlude the lst dpttion effet of the responses to the preeding sequene of tones (f 1, y, f 1 or f 1,y, f 2 )onthe responses to the proe stimulus, we hose frequenies of f 1 nd f 2 tht evoked no response from the MGB neuron eing reorded. The ISI ws typilly set t ms etween onseutive f 1 stimuli nd etween f 1 nd f 2.Thef 2 stimulus ws seprted from the proe stimulus y ms, or s otherwise speified. The inter-lok stimulus ws set t 3 s. When the ISI etween f 2 nd p ws set t ms, the proe stimulus followed f 2 without ny dely, s the durtion of ll stimuli (f 1,f 2 nd p) ws set to ms. Bloks of the devine nd ontrol proedures were rndomly presented y the omputer. An index tht mesured the modultory effet on the uditory response of the MGB neurons y the preeding devine proedure, IDC, ws defined s R D R C R D+R C ; where R D nd R C re the responses to the proe stimulus in the devine nd ontrol proedures, respetively. A negtive vlue indites suppressive effet nd positive vlue indites n enhned effet. The threshold for determining modultory effet, IDC th, ws set on neurony-neuron sis. The IDC th for eh individul neuron ws lulted using the ove eqution with responses from the odd (3 trils) nd even numer (3 trils) of trils in the ontrol proedure. The definitions for IDC nd IDC th lso pply to the ross-modl proedure. When the solute vlue of IDC exeeded the solute vlue of IDC th, the neuron ws onsidered to hve undergone modultion nd ws inluded in the sttistis. Comintion of light nd sound stimuli. A light stimulus ( ms) ws presented nd ws followed y noise urst with -ms intervl (light-sound proedure). Only the sound stimulus ws presented in the ontrol proedure (sound proedure). White light ws delivered through n rry of light diodes, whih were pled inside the soundproof hmer nd ontrolled eletronilly outside the hmer. No sound ws deteted while swithing the light diodes. MGB neuronl responses to the sound stimulus were ompred under the lightsound nd sound proedures. An initil exmintion of the neuronl response to the visul stimulus lone ws performed to exlude the dpttion effet. The light-sound nd sound proedures were rndomly presented every 3 s. Neuronl response dt were sorted for eh proedure using homemde progrm. Drug pplition. Lidoine ws purhsed from Sigm. Lidoine injetions (.3 ml, mg ml 1 ) were dministered in the TRN of seven rts using miroinjetion system (Hmilton). A tungsten miroeletrode ws glued to the injetion glss pipette to monitor the tivity of the TRN. The distne etween the two tips ws pproximtely mm. Dt nlysis. Spike detetion ws rried out with OpenEX softwre (Tuker- Dvis Tehnologies). Differenes etween vrying onditions were nlyzed with ANOVA. In ll nlyses, P o. ws onsidered to e signifint. 33. He, J. On nd off pthwys segregted t the uditory thlmus of the guine pig. J. Neurosi. 21, (1). 34. Guo, Y.P. et l. Cortiothlmi synhroniztion leds to -fos expression in the uditory thlmus. Pro. Ntl. Ad. Si. USA 4, (7). 3. Xiong, Y., Yu, Y.Q., Chn, Y.S. & He, J. Effets of ortil stimultion on uditoryresponsive thlmi neurones in nesthetized guine pigs. J. Physiol. (Lond.) 6, (4). 36. Pxinos, G. & Wtson, C. The Rt Brin in Stereotxi Coordintes (Elsevier Ademi Press, Amsterdm, ). doi:.38/nn.2373 NATURE NEUROSCIENCE