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1 doi:.38/nture277 d Time from sound onset (ms) Time from sound onset (ms) Firing rte (spikes/s) Firing rte (spikes/s) e f g h Frtion of neurons Frtion of neurons N = Neurometri slope Psyhometri slope N = Neurometri slope Psyhometri slope Supplementry Figure S. Responses of uditory ortil nd stritl neurons in the tsk., Rster from n exmple uditory ortex neurone shows monotoni response to frequeny ontent of the toneloud stimulus. Gry shding indites rt s position t the enter port., Tuning urve of neurone in pnel., Neurometri performne of neurone in pnel (red) is similr to ehviourl performne (gry). d, Cumultive histogrms of the rtio of neurometri nd psyhometri urve slopes ross the popultion of uditory ortil neurones. e, Rster from n exmple uditory stritum neurone with monotoni response to frequeny ontent of the toneloud stimulus. f, Tuning urve of neurone in pnel e. g, Neurometri performne of neurone in pnel e (red) ompred to ehviourl performne (gry). h, Cumultive histogrms of the rtio of neurometri nd psyhometri urve slopes ross the popultion of uditory stritl neurones. Supplementry Figure S2., Mirodrive rry for simultneous tetrode reording nd photostimultion., Close-up of tetrode/fier rrngement. The tetrodes re ut to terminte < µm of the optil fier tip.

2 I Depth from pi (normlized) II-IV V VI f Choie is 2 p = 9.3e Multiunit response khz N = 57 rts p =.56 d g Contrlterl is 5 p = 6.8e 6 Slope stim - Slope ontrol Slope stim + Slope ontrol N = 57 rts p = Preferred frequeny (khz) Frequeny preferene for ontrlterl hoie (otves) Frtion of neurons e Slope stim - Slope ontrol Slope stim + Slope ontrol h Contrlterl is 6 Asolute hoie is 2 ** ** *** * p = Supplementry Figure S3. Non-speifi ChR2 stimultion of uditory ortex ises hoies nd disrupts disrimintion performne., Expression of ChR2-Venus in the uditory ortex following injetion of AAV CAGS ChR2-Venus. Sle r - µm., Depth distriution of ChR2-Venus expressing neurones. Dshed lines mrk the pproximte lotion of lyer oundries. Error rs - 95% onfidene intervls., Exmple ehviourl session showing effets of non-speifi uditory ortex stimultion on ehviourl performne (lk - ontrol, lue - stimultion). d, Auditory ortex stimultion redued psyhometri performne mesured s the slope of the psyhometri urve. Bk rs indite sites with signifint effets (p <.5). e, Stimultion redued the slope of the psyhometri urve even for sites where stimultionevoked hoie is ws minor. Error rs - s.e.m., * - p<.5, ** - p<., *** - p<., signed-rnk test. f, Choie ised evoked y stimultion hd no signifint dependene on frequeny tuning of the stimultion site. g, Choie ises reltive to the stimulted hemisphere. Frequeny preferene ws relulted suh tht positive vlues reflet tuning for frequenies ssoited with the ontrlterl hoie port nd is the midpoint of the loud-of-tones stimulus (. khz). h, Stimultion tended to is responses towrd the hoie port ontrlterl to the stimulted ortex. Blk rs indite signifint ises (p <.5). 2

3 5 p =.95 5 p =.3 d 5 p =.69 Slope stim - Slope ontrol Slope stim + Slope ontrol Slope stim - Slope ontrol Slope stim + Slope ontrol Slope stim - Slope ontrol Slope stim + Slope ontrol 6 Asolute hoie is Slope stim - Slope ontrol Slope stim + Slope ontrol Supplementry Figure S. Effets of mnipultion of ortiostritl neurones on sujets ehviourl performne., While trgeted stimultion of ortiostritl neurones ised sujets hoies, it hd no effet on performne, quntified s the slope of the psyhometri urve., Axonl stimultion of ortiostritl neurones hd modest ut signifint effet on performne., During xonl stimultion, psyhometri slope ws redued only for sites with lrge hoie ises. Error rs - s.e.m. d, Intivtion of ortiostritl neurones hd no effet on performne. AAV CAGS ChR2 Stritum Stritum Stritum Auditory ortex Auditory ortex Auditory ortex Choie is N = 38 rts p =.65 Choie is 2 N = 52 6 rts p =.255 Choie is 6 N = 65 rts p = Preferred frequeny (khz) Preferred frequeny (khz) Preferred frequeny (khz) Supplementry Figure S5., ChR2 stimultion protool did not produe systemti hoie ises in uninjeted ontrol nimls., Choie ises evoked y stimultion of the xons of ortiostritl neurons depended on the frequeny preferene of the stimultion site., Arh stimultion protool did not produe systemti hoie ises in uninjeted ontrol nimls. 3

4 e 2 f z sore z sore z sore z sore FRlight - FRpre FR FRprepre light -+FR FRlight FRlight + FRpre g 2 2 g 2 2 h h Spikes s f Spikes s 2 z sore e z sore d Spikes s # neurons # neurons z sore d Spikes s z sore 2 2 Supplementry of ortiostritl ortiostritlneurones neuronesresults results suppression SupplementryFigure Figure S6. S6. ChR2 ChR2 stimultion stimultion of in in suppression of of firing of most uditory ortil ells., Ativity of n uditory ortex neurone inhiited y photostimultion firing of most uditory ortil ells., Ativity of n uditory ortex neurone inhiited y photostimultion of of ortiostritl nm) ws ws presented presentedfor for ms mstt5 5 mw (lue r)., Most ortil ortiostritlneurones. neurones. Blue Blue light light (73 nm) mw (lue r)., Most ortil neurones ells.light Lightmodultion modultion index ws lulted ompring neuronesre reinhiited inhiitedy ystimultion stimultion of ortiostritl ortiostritl ells. index ws lulted ompring firingrtes rtesduring during2 2ms msfollowing following nd nd preeding preeding the tringle - medin lightlight firing the onset onsetofofthe thelight lightpulse. pulse.open Open tringle - medin modultion;filled filledtringle tringle-- ell ell in in pnel pnel., of of thethe neurone in pnel modultion;, Peristimulus Peristimulustime timehistogrm histogrm(psth) (PSTH) neurone in pnel PSTHofofputtive puttive nrrow-spiking nrrow-spiking interneurone ortiostritl neurones...d,d,psth interneurone tivted tivtedyystimultion stimultionof of ortiostritl neurones. e-h, Menstndrdized stndrdizedpsths PSTHs for for neurones neurones grouped ss highlighted in pnel.. e-h, Men groupedy ylight lightmodultion modultionindex, index, highlighted in pnel -3. mm -. mm -5.6 mm -6. mm -3. mm -. mm -5.6 mm -6. mm 2.5 Normlized ell density 3 Distne from Bregm (mm) 6.5 Supplementry Figure S7. -, Retrogrde leling of ortiostritl neurones following injetions of HSV- mcherry into dorsl (top) nd ventrl (ottom) uditory stritum., In eh setion, we quntified the numer of leled ortiostritl neurones in primry uditory ortex per mm3 of tissue. We then normlized these vlues y the men density ross ll setion in given rin. Density of ortiostritl neurones in primry uditory ortex followed udl grdient for injetion in pnel (lk, open irles) nd rostrl grdient for injetion in pnel (gry, filled irles). W W W. N A T U R E. C O M / N A T U R E

5 preferred hoies preferred hoies preferred hoies preferred hoies preferred hoies preferred hoies preferred hoies Pre 2% lidoine Post p =. p =.7e p =.8 Pre 2% lidoine Post p =. p =. Pre 25 μm TTX Post p = 9.5e 5 p = 2.2e 5 Pre 25 μm TTX Post p = 5.2e 6 p =.7e 6 Pre 25 μm TTX Post p =.2 p =.6 Pre 25 μm TTX Post p = 2.e 3 Tones pref - Tones nonpref p =.6 Pre 25 μm TTX Post p =.3 Tones pref - Tones nonpref p =.5 p =. p =.83 p =.6e 3 p = 3.9e p =.3 p =.2 Tones pref - Tones nonpref Supplementry Figure S8. Choie ises evoked y stimultion of ortiostritl neurones do not depend on reurrent ortil exittion. Psyhometri urves from ontrol (lk) nd stimultion (lue) trils for the ehviorl session efore intivtion (left olumn), during the intivtion session (enter olumn) nd following reovery from intivtion (right olumn). The p-vlues quntify signifine of the stimultionevoked hoie is. 5

6 3 n.s. * 3 Choie is 2 Normlized hoie is 2 Pre Intivtion Post Stimultion tril numer Supplementry Figure S9., Intivtion of ortil reurrent exittion does not olish light-evoked hoies ises. The dt summrized re the sessions in Supplementry Figure S8. Bises evoked y stimultion were redued on intivtion sessions ut the differene ws not sttistilly signifint (p =.3, Wiloxon rnksum test). This redution in hoie is persisted following reovery from intivtion (p <.5, Wiloxon rnksum test, ompred to pre-intivtion sessions) nd likely reflets sujets dpttion to repeted stimultion of the sme site., Consistent with this, we oserved tht the mgnitude of stimultionevoked hoie ises deyed during single stimultion session. For sites where we olleted t lest 75 stimultion trils nd stimultion produed hoie is of t lest 5 tones/s, we inned the stimultion trils in loks of 25 trils nd independently mesured the stimultion-evoked hoie is in eh lok. We normlized these mesurements y the men hoie is during tht session. Error rs re s.e.m. nd the line is n exponentil fit to the dt. Effets of stimultion were gretest t the strt of the session nd deyed with time onstnt of 68 stimultion trils (95% onfidene intervl, trils). Similr dpttion ws oserved during stimultion of re MT in mques. Slzmn, C. D., Mursugi, C. M., Britten, K. H. & Newsome, W. T. Mirostimultion in visul re MT: effets on diretion disrimintion performne. J. Neurosi. 2, (992). 6

7 d.3.3 rt PZ57 rt PZ57 rt PZ3 rt PZ3 Response time (s) Response time (s) # of rts Tones low - Tones low - Tones low - Tones low e 5 f 2.5 r =.66 g h p = 2.8e 5 RT tones/s RT tones/s (ms) Psyhometri urve slope RT tones/s RT tones/s (ms) - Tones low i j k l p =.3 p =.9 p =.9 Chronometri is Chronometri is Preferred frequeny (khz) Choie is Preferred frequeny (khz) Chronometri is Chronometri is Response time (s) Tones low 2 p =.8 2 Choie is Supplementry Figure S. Effets of stimulus strength nd mnipultion of ortiostritl neurones on response times. -, Psyhometri nd hronometri funtions for rt tht shows slower response times on diffiult trils. -d, Psyhometri nd hronometri funtions for rt tht shows no effet of stimulus strength on response times. e, Distriution of effets of stimulus strength on rts response times (see methods). Blk - rts showing signifint (p<.5) effets of stimulus strength on response time. f, Rts tht respond slower on diffiult trils show etter psyhometri performne. Green nd lue dots indite the rts in pnels - nd -d respetively. Error rs - 95% onfidene intervls. g-h, Psyhometri nd hronometri funtions from single session during stimultion of ortiostritl xons. Blk - ontrol trils; lue - stimultion trils; error rs - s.e.m. i, Shifts of the rts hronometri urves evoked y stimultion of ortiostritl neurones depended on the frequeny tuning of the stimultion site. Filled irles - HSV-medited stimultion; open irles - xonl stimultion. j, Stimultion-evoked hronometri shifts were orrelted with stimultion-evoked hoie ises. k, Arh-intivtion of ortiostritl neurones tended to hve the opposite effet of rts hronometri urve, ut effets did not reh signifine. l, Intivtion-evoked hronometri shifts tended to orrespond to intivtioned-evoked hoie ises. 7

8 d µm µm 2 µm 7 9 µm Spikes s Spikes s Spikes s Spikes s Supplementry Figure S. Sptil drop-off of Arh-intivtion effiieny. -d, PSTH of n exmple puttive ortiostritl neurone, illuminted with green light (5 mw) t fiers,, 2 nd 9 µm from the reording eletrode in the wke niml without uditory stimultion. Green r shows timing of light delivery. This neurone shows reound firing fter relese from inhiition. 8