Abstract Phosphenes represent a perceptual eVect oftranscranial magnetic stimulation (TMS) or electric stimu-lation of visual cortical areas. One likely neural basis forthe generation of static phosphenes is the primary visualcortex (V1) although evidence is controversial. A peculiarfeature of V1 is that it has sparse callosal connections withthe exception of a central portion of visual Weld representa-tion. In contrast, visually responsive cortical areas in theparietal lobe have widespread callosal connections. Thus,interhemispheric transfer (IT) time of oV-centre phosph-enes should be slower when generated by V1 than by visualparietal areas. To verify this possibility, in Exp. 1 we mea-sured IT of phosphenes generated by TMS applied to V1and in Exp. 2 we measured IT of phosphenes obtained byTMS applied to posterior parietal cortex. In both experi-ments, we obtained static bright circular phosphenesappearing in the contralateral hemiWeld. We measured ITtime behaviorally by comparing unimanual simple reactiontime to the onset of a phosphene under crossed or uncrossedhemiWeld-hand condition (PoVenberger paradigm). In keep-ing with our prediction, we found a substantially longer ITtime for V1 than for parietal phosphenes. Additionally, anIT similar to that obtained with V1 stimulation was foundwhen participants were asked to imagine the phosphenespreviously experienced during TMS. In conclusion, thepresent results suggest that IT of phosphenes either generated
by V1 TMS or imagined is subserved by slower callosalchannels than those of real visual stimuli or parietal phos-phenes.
Keywords TMS · Primary visual cortex · Posterior parietal cortex · Corpus callosum · Interhemispheric transfer
Introduction
There is ample evidence that transcranial magnetic stimula-tion (TMS) applied to the occipital lobe produces phosph-enes (for reviews see Amassian et al. 1998; Cowey andWalsh 2001; Pascual-Leone et al. 2000). The neural siteand the neurophysiological mechanisms are still unsettledbut several studies suggest V1 possibly including V2/V3 asresponsible sites (Amassian et al. 1994; Cowey and Walsh2000; Kammer 1999, 2007; Kammer et al. 2001, 2005;Kastner et al. 1998; Meyer et al. 1991; Pascual-Leone andWalsh 2001; Sparing et al. 2002). Phosphenes can beobtained by stimulation over large parts of the occipitallobe yielding visual Weld locations usually in the contralat-eral lower hemiWeld. They appear often to be circular andbright white or gray and move with the eyes (Kammer2007; Kammer et al. 2005). All these properties suggestthat they represent the activation of visuotopic areas and areconsistent with a primary visual cortex (V1) locus of gener-ation, although the contribution of adjacent V2 and V3 can-not be excluded.
To try and cast further light on the cortical sites generat-ing phosphenes, in the present study we exploited the wellknown neuroanatomical and neurophysiological evidence(Aboitiz et al. 1992; Berlucchi 1972; Clarke and Miklossy1990; Pandya and Seltzer 1986) that in cats and primates
C. A. Marzi · F. Mancini · S. Savazzi (&)Department of Neurological and Vision Sciences, Section of Physiology, University of Verona, Strada le Grazie 8, 37134 Verona, Italye-mail: [email protected]
C. A. Marzi · F. Mancini · S. SavazziNational Institute of Neuroscience, Verona, Italy
the callosal connections have a diVerent density in diVerentcortical areas. In V1 they run in the splenium and are lim-ited to a central narrow strip of representation of the verti-cal meridian at the cytoarchitectonic boundary between V1and V2. As a consequence, the topographical representationof the visual Weld in each hemisphere is basically restrictedto the contralateral half (Lavidor and Walsh 2004). This isnot the case in visually responsive areas in the parietal lobewhere there is a callosally mediated large representation ofthe ipsilateral visual hemiWeld (Ffytche et al. 2000; Grosset al. 1977; Marzi et al. 1982). Thus, on the basis of thisanatomo-physiological arrangement a straightforward pre-diction is that an oV-centre stimulus processed by V1 andadjacent visual areas should take longer to cross to the otherhemisphere in comparison to a stimulus processed by parie-tal areas with widespread callosal connections. To studyinterhemispheric transfer (IT) time we employed thePoVenberger paradigm, that is, a simple unimanual reactiontime (RT) task with lateralized visual stimuli that has beendevised by PoVenberger in 1912 (Berlucchi et al. 1971;Marzi 1999; Marzi et al. 1991; Zaidel and Iacoboni 2003).With this method IT time is measured by subtracting RT ofaveraged uncrossed hand-hemiWeld conditions from that ofaveraged crossed hand-hemiWeld combinations. The latterrequire an IT and usually yield a longer RT than the former.The obtained value, the so-called crossed-uncrossed diVer-ence (CUD), is consistent across studies and is consideredas a reliable measure of commissural IT time. On average itamounts to about 4 ms in healthy participants but is consid-erably slower in patients with either an agenesis or a surgi-cal section of the corpus callosum (Marzi et al. 1991;Zaidel and Iacoboni 2003).
In the present study we carried out two experiments: inExp. 1, we tested the CUD for RT to onset of phospheneselicited by TMS over the occipital lobe while in Exp. 2 weused a similar procedure but phosphenes were generated byapplying TMS to the parietal lobe in a visually responsivearea in the intraparietal sulcus.
Experiment 1
The general strategy used in this and the subsequent experi-ment was to ask participants to respond as quickly as possi-ble to the onset of a TMS-generated phosphene. In addition,in Exp. 1, we tested RT for visually presented and for imag-ined stimuli. In the latter condition, we asked participants toimagine the phosphenes experienced with TMS in the pre-vious session and to react to them as quickly as possible.Visual imagery of simple stimuli is likely to be anotherfunction subserved by V1 (Kosslyn and Thompson 2003;Kosslyn et al. 2001; Marzi et al. 2006; Savazzi et al. 2008)and, together with phosphenes, represents a form of
“artiWcial phenomenal vision” as deWned by Stoerig (2001).If imagined phosphenes are generated in V1 and transferredto the other hemisphere through V1 callosal connectionsthey should take as long as TMS-phosphenes to transferfrom one hemisphere to the other. Finally, for a compari-son, we presented reproductions of the phosphenes drawnby each participant following TMS and, in diVerent trialblocks, simple visual stimuli (circles). Our prediction wasthat stimuli that use callosal connections at the level of V1,such as TMS-generated or imagined phosphenes shouldyield a substantially slower IT than visually presented stim-uli that, as shown by various studies (Iacoboni 2006; Iaco-boni and Zaidel 2004; Marzi 1999; Tettamanti et al. 2002;Weber et al. 2005) are likely to undergo IT at the level ofparietal or prefrontal cortical areas.
Methods
Participants
Seven right-handed (4 females) naive participants took partin the experiment. Their age ranged between 26 and 33years (mean 27.4) and they had normal or corrected-to-nor-mal visual acuity. All gave informed consent and the exper-iment was carried out according to the principles laid downin the 1964 Declaration of Helsinki.
Apparatus, stimuli and procedure
There were three types of sessions that for the sake of brev-ity will be referred to as: TMS, perception and imagination.In all sessions, participants were seated in front of a LCDmonitor (AOC LM520) with the eyes at 57 cm from thecenter of the screen.
The TMS session was subdivided in two parts:In a preliminary part we assessed the individual intensity
threshold for phosphene perception. The coil used was aWgure-of-eight Magstim (Magstim 220, The Magstim Com-pany Ltd, UK) with a diameter of 70 mm. The orientation ofthe coil was upside-down, with bends oriented downward,in order to avoid unspeciWc activation of neck and shouldermuscles. We did not attempt a systematic investigation ofthe eVect of diVerent coil orientation because it was beyondthe aim of the study and in the light of recent evidence thatcoil orientation does not signiWcantly inXuence phospheneperception with single pulses at an intensity of 100% of sin-gle phosphene threshold (Sparing et al. 2005). We localizedthe scalp area in which single-pulse TMS elicited circum-scribed and lateralized phosphenes using the minimumintensity capable of inducing a phosphene on 95% ofstimulations (mean 76.4% of maximum stimulator outputintensity; SD 7.1%). The mean number of single-pulse stim-ulations for each participant was about 20 for each side. The
coordinates of the stimulation sites over the scalp overlyingthe two hemispheres were established for each subject. Thesuccessful stimulation site was considered as that yielding avivid, lateralized, and circumscribed phosphene in the con-tralateral hemiWeld. The position of the phosphene wasreconstructed over the scalp on the basis of the International10-20 EEG System with participants wearing an electrodecap. This is a simple method that yields satisfactory resultscomparable to more elaborate stereotaxic methods (Herwiget al. 2003; Sparing et al. 2008). Unless a high resolutionlocalization is required (see e.g. Schönfeldt-Lecuona et al.2005), this method can be used without the expensive aid ofbrain imaging. In all participants, the most eVective site foreliciting phosphenes was close to O1 and O2, see Fig. 1.Only one participant did not meet the above criterion andwas discarded. The other six either experienced a phos-phene on the Wrst TMS or soon thereafter, in keeping withwhat has been found elsewhere (Fernandez et al. 2002;Kammer et al. 2005). At the end of the preliminary sessionall participants were able to experience a phosphene imme-diately following TMS.
In the experimental part participants were to maintainthe gaze on the Wxation point (a small circle in the center ofthe screen) and to press as quickly as possible followingappearance of a phosphene the space-bar of a PC keyboardwith the index-Wnger of either the right or the left hand.Trials were self initiated by pressing the space-bar of thePC keyboard and the participant was informed that couldstop the experiment at any moment. The temporal sequence
of occurrence of TMS stimuli was randomized with theconstraint that the inter-stimulus interval should not beshorter than 3 s, as recommended by safety instructions(Anand and Hotson 2002; Wassermann 1998). In order tokeep the gaze on the Wxation point, participants kept theireyes open during TMS stimulation (see Deblieck et al.2008). The intensity of stimulation was the same as thatused in the preliminary phase and the number of TMS stim-uli per participant was 40 for each of the four hemisphere/hand conditions. The order of responding hand and hemi-sphere of stimulation was counterbalanced across partici-pants: half of the participants followed an ABBA sequencewhile the other half followed a BAAB sequence of alternat-ing hands. As to side of hemisphere stimulation, half of theparticipants received TMS over the right hemisphere Wrstand the other half over the left hemisphere Wrst. In caseTMS did not elicit a vivid phosphene, the trial was re-administered to the subject.
In the perception session, the basic apparatus and proce-dure were the same as in the TMS session. The visual stim-uli were of two kinds presented in separate sub-sessions:(1) 1° diameter circles presented 6° to the right or left of theWxation point with an exposure duration of 96 ms and witha luminance of 2.7 cd/m2 over a background luminance of0.001 cd/m2; (2) stimuli similar to the phosphenes per-ceived by each participant in the preliminary phase anddrawn on a transparent sheet of paper placed on the PC-monitor so as to reproduce the exact visual Weld positionand shape. The phosphene-like stimuli were presented onthe monitor with an exposure duration and an intensity sim-ilar to that of the circles. An acoustic warning stimulus (200ms duration) prompted the participants to maintain Wxationsteady. The interval between acoustic warning and visualstimulus was randomized within the temporal window of300–600 ms and 20 catch trials were presented to discour-age the participants from responding to the tone rather thanto the visual stimulus.
In the imagination session apparatus and general proce-dure were similar to the previous sessions; participantswere asked to keep their gaze on the central Wxation pointand, immediately after hearing the acoustic warning stimu-lus, to imagine a phosphene similar to that experienced inthe previous TMS session and to react to the image bypressing the response key as quickly as possible. No visualstimulus was presented on the screen and RTs were mea-sured from the onset of the tone. Control trials in whichparticipants were asked to react to the acoustic tone withoutimaging visual stimuli enabled us to check the occurrenceof unwanted responses to the tone rather than to the imag-ined stimuli. Clearly the former were 4–5 times quickerthan the latter.
At the beginning of each block participants wereinformed on the hemiWeld and the position of the stimuli to
Fig. 1 Coil position. Black circles indicate individual position ofTMS coil with reference to O1 and O2 (gray circles) in Exp. 1 and P3and P4 in Exp. 2
be imagined. The latter corresponded to that in which theyperceived the phosphenes in the TMS session. The hand tobe used for response was varied from block to block in abalanced sequence. It has been previously shown that in asimple RT paradigm like that employed in the present studyspatial compatibility eVects do not play a signiWcant role(Anzola et al. 1977; Berlucchi et al. 1977). However, giventhat visual imagery may yield spatial compatibility eVectsin a choice paradigm (Tlauka and McKenna 1998), wethought worthwhile to check their possible contribution tothe CUD with our imagined stimuli. We did that by testingparticipants with arms either in an anatomical or in acrossed position. This procedure allows to disentangle cal-losal IT from spatial compatibility eVects (Anzola et al.1977; Berlucchi et al. 1977; Tlauka and McKenna 1998)because if spatial rather than interhemispheric eVects arecrucial, in the crossed arm position one should obtain areversed CUD.
All participants were tested Wrst in the TMS sessionwhile the order of the perception and imagination sessionswas counterbalanced across participants. The same was truefor the circles and the phosphene-like stimuli in the percep-tion session and for the anatomical versus crossed armsposition in the imagination session. The sequence of thevarious testing conditions is shown in Fig. 2.
The range of accepted RTs was 140–650 ms in the TMSand perception sessions, while in the imagination sessionwe decided to accept all RTs longer than 140 ms given thatthere were diVerences in the imagery speed of participants(outliers were detected by means of the Grubbs’ test,extreme studentized deviate method, Grubbs 1969, and dis-carded). Excluded trials were a minuscule minority. Also,the proportion of omission errors was negligible. Eyemovements were controlled by an infrared camera placed infront of the participants; it turned out that all of them had avery stable Wxation. Participants were adapted to the roomambient light for a few minutes prior to testing.
Results
Phosphenes
As mentioned above, all our participants, with the excep-tion of one who was excluded from the study, found it
relatively easy to perceive well-deWned phosphenes imme-diately after each single-pulse TMS. This was not the casein other studies (e.g. Sparing et al. 2005) although manyhave had a success rate comparable to ours (Boroojerdiet al. 2000; Fernandez et al. 2002; Kammer 1999; Marg andRudiak 1994; Meyer et al. 1991; Ray et al. 1998). Figure 3ashows visual Weld position and shape of the phosphenes asdrawn from memory by the participants. Four out of sixparticipants perceived the phosphenes in the lower contra-lateral visual Weld, while the remaining two perceived thephosphenes in the upper contralateral hemiWeld. In all ourparticipants phosphenes were circular and bright gray andhad a mean width of 2.2° (SD 0.98). Phosphenes were alllocated within the most central 5° with respect to verticaland horizontal meridian (inner edge, mean 1.26°; SD 0.55)and had a rather sharp contour that could be drawn withgood accuracy on a screen.
Reaction time
Participants found no diYculty in reacting quickly to theappearance of a phosphene, as shown by overall RT whichwas reasonably fast although clearly slower than that forvisually presented stimuli, see Table 1 for mean RT foreach testing condition. The same holds for reacting toimagined phosphenes that yielded overall slower RTs sub-stantially similar to those obtained in previous experiments(Marzi et al. 2006; Savazzi et al. 2008).
The RT scores were statistically analyzed by means of athree-way ANOVA with stimulus (TMS-phosphenes, cir-cle stimuli, phosphene-like stimuli, imagined phospheneswith responding hand in anatomical or in crossed position),visual hemiWeld (left and right) and hand (left and right) asfactors. The main eVect of stimulus was signiWcant[F(4,20) = 25.774, P < 0.001]: a series of post hoc t testswith Bonferroni correction showed that RT for TMS-phos-phenes and RT for imagined phosphenes (with respondinghand either in anatomical position or in crossed position)were reliably slower than RT to circle or phosphene-likestimuli presented on the screen. Importantly, there was noreliable diVerence between RT to imagined stimuli withcrossed or uncrossed hands, thus ruling out spatial compat-ibility eVects. The interaction visual hemiWeld £ hand wassigniWcant [F(1,5) = 28.438, P < 0.005] indicating that RTs
Fig. 2 Temporal order of sessions. Light gray indicates perception sessions; dark gray indicates imagination sessions
in the uncrossed conditions (582.80 ms) were faster thanthose in the crossed conditions (603.10 ms). This diVerencereXects IT time averaged across conditions of stimulusgeneration.
The crucial Wnding was that the three-way interactionstimulus £ visual hemiWeld £ hand was highly signiWcant[F(4,20) = 3.938, P < 0.01] indicating that IT time varied
with the various conditions of stimulus generation. A seriesof post-hoc t tests conducted on CUD scores with Bonfer-roni correction showed a larger CUD for TMS-phosphenes(40.35 ms) and imagined phosphenes (37.40 ms) than withvisually presented stimuli (circles = 3.24 ms; phosphene-like = 3.86 ms), see Fig. 4a. Notably, the CUD for TMS-induced and imagined phosphenes did not diVer from each
Fig. 3 Position and shape of phosphenes in each participant in (a) Exp. 1 and (b) Exp. 2
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436 Exp Brain Res (2009) 192:431–441
other and the same was true for the two types of visuallypresented stimuli.
Discussion
In Exp. 1, we compared IT of one kind of “artiWcial phe-nomenal vision”, that is, TMS-induced phosphenes, withanother kind of “artiWcial vision” that is, imagined phosph-enes, and two kinds of real vision, that is, visually pre-sented phosphene-like and circle visual stimuli. The aimwas to establish whether TMS-generated phosphenes aretransferred from one hemisphere to the other through thecallosal connections of V1/V2. We found that IT of bothTMS-generated and imagined phosphenes was about ten
times slower than that of visually presented stimuli. Impor-tantly, this diVerence was independent of overall RT: aver-aged together, the two kinds of artiWcial phenomenal visionyielded RTs about three times longer than real vision whilethe CUD was about ten times longer. Moreover, there wasno statistical correlation between RT and CUD in the vari-ous conditions of stimulus generation. By the same token,overall RT was about the double for imagined versus TMS-induced phosphenes but the CUD was almost identical, seeTable 1.
This pattern of results suggests two considerations: Wrst,the locus of IT of TMS-generated and imagined phosph-enes may be similar; second, this locus may be diVerentfrom that subserving IT of visually presented stimuli. It isworth mentioning that the present results with imaginedphosphenes conWrm a recent study on interhemispherictransmission and summation of imagined visual stimuliwhich yielded an IT time roughly similar to that found inthe present study (Savazzi et al. 2008). In that study, weused simple light patches as stimuli and this shows that theimagination results of the present experiment are not pecu-liar to phosphenes. What could be the locus of IT of phos-phenes? As mentioned in the Introduction, it is very likelythat both TMS-induced and imagined phosphenes are gen-erated in V1. The idea that phosphene and visual imageryshare a V1 locus has been convincingly conWrmed by aTMS study in which phosphene threshold was lowered byvisual imagery (Sparing et al. 2002). It is well establishedthat in monkeys and humans V1 and V2 are devoid of cal-losal connections except that at the V1/V2 border in a zonerepresenting a narrow strip of central vision (Clarke andMiklossy 1990; Lavidor and Walsh 2004; Pandya and Selt-zer 1986). The existence of a sharp division at the foveabetween the representation of the two hemiWelds has beenrecently conWrmed by mapping the visual Weld loss in aquadrantanopic patient, or scotomata temporarily inducedby TMS in healthy participants (Chiang et al. 2004). Proba-bly then, TMS- or imagery-generated phosphenes, that inour experiment are perceived as located between 1° and 5°of eccentricity from the vertical meridian, have used sparse
Table 1 Mean RTs in the various crossing conditions and sessions
callosal connections. Hence, their IT is slower than that ofvisually presented stimuli which are likely to be transferredthrough widespread callosal connections in parietal andprefrontal cortex (Iacoboni 2006; Iacoboni and Zaidel2004; Marzi 1999; Omura et al. 2004; Tettamanti et al.2002; Weber et al. 2005). Before concluding that phosph-enes transfer at the level of V1/V2 one should consider thata TMS pulse applied over V1/V2 stimulates not only localinterneurons but also excitatory projection neurons. Thus, itmight induce neural activity in extrastriate visual areas suchas V5 which have extensive callosal connections and there-fore this activation might spread to contralateral hemi-sphere. Furthermore, the neural conduction time of V1–V5projection Wbers is fast and presumably beyond the tempo-ral resolution of the behavioral paradigm used in this study.This in principle is a possibility to keep in mind; however,it has been shown that phosphenes’ IT time as estimatedwith our behavioral method is broadly in keeping with thatfound by Ilmoniemi et al. (1997) by using TMS appliedunilaterally over the occipital lobe. They found EEG activ-ity related to TMS in the opposite occipital cortex within 20ms. This value is compatible with a slow callosal channeland is in broad agreement with IT time measured electro-physiologically with reference to the two early visual com-ponents (P1 and N1) of the ERP (Brown et al. 1999;Hoptman et al. 1996; Ipata et al. 1997; Rugg et al. 1984;Saron et al. 2003). Clearly, IT measures obtained eitherwith coupled TMS-EEG experiments or with ERP arecloser to those found in the present study for TMS- andmentally-generated phosphenes than those one would haveexpected by a transfer via the widespread V5 callosal con-nections. Moreover, it is well known that, with some excep-tions, see below, electrophysiological estimates of IT timegive slower values than those obtained behaviorally withvisually presented stimuli. This is keeping with the possi-bility that TMS- and imagery-generated phosphenes trans-fer at the level of the callosal connections of V1 whereas ITwith visually presented stimuli does not depend upon thevisual callosal connections but rather on anterior callosalportions such as the genu as shown by fMRI studies(Omura et al. 2004; Tettamanti et al. 2002; Weber et al.2005), that is, the anterior portion of the corpus callosuminterconnecting prefrontal areas (Pandya and Seltzer 1986).This possibility is reinforced by the results of Rugg et al.1984 (see also Lines et al. 1984) who found a short (3–4ms) CUD for visual ERPs recorded over more anteriorsites, presumably reXecting the premotor route that is mea-sured by behavioral CUDs.
In addition, other fMRI studies found a critical role ofthe posterior parietal cortex (Iacoboni 2006; Iacoboni andZaidel 2004) in visuomotor interhemispheric integration,thus again reinforcing the idea that simple visuomotorinformation cross at anterior portions of the corpus callo-
sum presumably under the form of a “go” signal rather aspurely visual information. In keeping with this possibility isthe well established Wnding that in healthy participants theCUD is not signiWcantly aVected by visual parameters (Ber-lucchi et al. 1971; Fendrich et al. 2004; Milner and Lines1982; Zaidel and Iacoboni 2003).
One possible alternative explanation of at least part ofour results is that the slow IT for TMS-generated phosph-enes might be related to an inhibition of callosal transmis-sion concomitant with phosphene generation. In fact,Marzi et al. (1998) found that single pulse TMS stimula-tion over extrastriate cortex delayed IT time while leavingintrahemispheric RT unimpaired. That study, however,diVered from the present substantially since TMS parame-ters and location were chosen so as to avoid the productionof phosphenes or scotomata. More relevant to the presentWnding is a recent experiment by Chiang and Lavidor(2005) who conWrmed and extended Marzi et al. (1998)results of a higher susceptibility to TMS of the callosalpathway with respect to the intrahemispheric pathway. Atvariance with Marzi et al. (1998), they used repetitiverather than single pulse TMS and stimulated V1 ratherthan extrastriate areas. They found a lengthened IT timefor the crossed conditions with stimulus presentation tothe hemiWeld contralateral to the hemisphere over whichTMS was applied. Their interpretation was that as a conse-quence of TMS the visual stimulus on the stimulated sidewas too weak to access the callosal pathway. It is worthpointing out that in both the above studies the eVect ofTMS on IT time was relatively small: the CUD was 13 msin Marzi et al. (1998) and 22 ms in Chiang and Lavidor(2005) while the CUD for phosphenes in the present studyamounted to 40 ms. Thus, it seems unlikely that our resultsare related to a callosal inhibition. Furthermore, the CUDfor phosphenes was very similar to that in the imagerycondition, a situation phenomenologically akin to phos-phene perception but not induced by TMS. A Wnal possi-bility as to the pathways for IT of TMS-generated andimagined phosphenes is the superior colliculus and itscommissure. This possibility is suggested by the broadanalogy between the CUD of TMS and imagined phosph-enes and that of split-brain participants (see Corballis2002; Savazzi et al. 2007).
Experiment 2
An interesting question is whether TMS applied to visuallyresponsive cortical areas other than V1 might generatephosphenes. So far, although visually responsive areas havebeen described in both human and non-human primates inparietal, temporal and frontal cortex, to our knowledge,there is no evidence of TMS generated phosphenes from
these areas with the exception of moving phosphenesobtained for stimulation of V5/MT (Amassian et al. 1998;Antal et al. 2004; Cowey and Walsh 2000; Silvanto et al.2005, 2007).
A crucial conWrmation of our hypothesis that the slowIT of phosphenes is related to a callosal transfer throughthe sparse callosal connections of the visual cortex(Clarke and Miklossy 1990) would be to Wnd a faster ITfor phosphenes generated in areas with more widespreadcallosal connections than V1. Such a result would alsoconWrm that the lengthened CUD found for TMS-gener-ated phosphenes in Exp. 1 was not related to callosal inhi-bition. Widespread callosal connections have beendescribed both in monkeys (Pandya and Seltzer 1986) andin humans (Aboitiz et al. 1992; Zarei et al. 2006) in theparietal lobe. In contrast to the occipital lobe which isrichly endowed with well-deWned retinotopic visual Weldrepresentations in diVerent cytoarchitectonic areas, theorganization of visually responsive areas in the parietallobe is less well understood although several maps havebeen found in both anterior and posterior parietal cortex(Grefkes and Fink 2005; Orban et al. 2006; Schluppecket al. 2005; Sereno et al. 2001; Silver et al. 2005). Veryrecently, more deWned retinotopic maps of the contralat-eral hemiWeld have been identiWed along the intraparietalsulcus (Schluppeck et al. 2005; Swisher et al. 2007) thusreinforcing the general concept that topographic maps tilethe cortex continuously from primary visual cortex to pos-terior parietal cortex. In particular, Swisher et al. 2007have described two newly identiWed areas (IPS3 andIPS4) with a contralateral hemiWeld representation thatlay in a rostral position potentially amenable to TMSsince this position is in correspondence of the electrodelocations P3 and P4 of the 10–20 International System(Herwig et al. 2003). In view of this opportunity wedecided to apply TMS to this area using as landmarks theabove electrode locations.
Method
Participants
Six right-handed (4 females) participants took part in theexperiment. All of them were the same as those tested inExp. 1. Their age ranged between 26 and 33 years (mean27.4). All of them gave again informed consent and theexperiment was carried out according to the principles laiddown in the 1964 Declaration of Helsinki.
Apparatus, stimuli and procedure
They were identical to those of Exp. 1 with the diVerencethat the coil was positioned over the P3–P4 locations of the
International 10–20 System (Fig. 1b). Interestingly, thesame TMS parameters as those used for stimulation of O1/O2 turned out to be optimal for P3/P4 as well. As describedby Herwig et al. (2003) these locations correspond to Brod-mann areas 7 and 40 and in reference to the maps providedby Swisher et al. (2007) are probably close to the newlydescribed visual areas in the medial bank of the intrapa-rietal sulcus, namely, IPS3 and IPS4 (Swisher, personalcommunication, October 2007) containing a map of contra-lateral visual hemiWeld. Another diVerence with Exp.1 wasthat we skipped testing with imagery stimuli and visuallypresented circle stimuli. As in Exp. 1, participants were Wrsttested for RT to phosphenes and then for RT to visuallypresented drawings of the same phosphenes experiencedwith TMS. The experiment took place on a diVerent daywith respect to Exp. 1.
Results
Phosphenes
Participants experienced phosphenes broadly similar tothose following occipital stimulation, see Fig. 3b. Theywere bright gray, circular and located in the contralateralhemiWeld. Their width was of 2.6° (SD 0.59). They were alllocated within the most central 5° with respect to verticaland horizontal meridian (inner edge, mean 1.32°, SD 0.46).Thus, in contrast to stimulation of V5/MT which yieldsmoving phosphenes, the parietal area we stimulated (P3/P4)yielded static phosphenes very similar to those obtained byTMS applied to O1/O2. A more thorough characterizationof the phosphenes obtained from the two locations requiresa further dedicated study.
Reaction time
The RT scores were statistically analyzed by means of athree-way ANOVA with stimulus (TMS-phosphenes,phosphene-like stimuli), visual hemiWeld (left and right)and hand (left and right) as factors. The main eVect ofstimulus was signiWcant [F(1,4) = 6.883, P < 0.05] indicat-ing that RT for TMS-phosphenes (390.93 ms) was reliablyslower than that to phosphene-like (256.92) stimuli pre-sented on the screen. The interaction visual hemiWeld £hand was signiWcant [F(1,4) = 9.171, P < 0.05] indicatingthat, overall, RTs in the uncrossed conditions (325.84 ms)were faster than those in the crossed conditions (331.08ms). The crucial result was that the three-way interactionstimulus £ visual hemiWeld £ hand was not signiWcant[F(1,4) = 0.054, P = 0.825] indicating that IT time did notvary with the two conditions of stimulus presentation(TMS-phosphenes = 4.95 ms; phosphene-like = 5.53 ms,Fig. 4b).
This result shows that phosphenes similar to those obtainedby occipital TMS yield a much shorter IT when obtained byparietal stimulation. Importantly, IT time with parietalphosphenes is similar to that with visually presented stim-uli. This reinforces the possibility that parietal phosphenestransfer, as is the case with visually presented stimuli, viathe widespread callosal connections interconnecting theparietal lobes in the two sides. An important question iswhether the phosphenes observed following TMS over P3
and P4 are generated in the posterior parietal cortex or inV1. In principle, one possibility is that TMS activated V1antidromically through cortico-cortical connections fromV1 to parietal cortex. This would explain the similaritybetween phosphenes elicited by stimulation of parietal andoccipital sites. However, clear antidromic eVects of TMS inthe cortex have not been described and this possibility,although theoretically possible, is unlikely. Another expla-nation for parietal phosphenes might be related to an ortho-dromic activation of parietal–occipital connections. This isa more reasonable possibility although it would not explainthe diVerence in IT time between parietal and occipitalstimulation.
Finally, another possibility would be stimulation of theoptic radiation, see Marg and Rudiak (1994). Two consid-erations go against this possibility: Wrst, to excite the opticradiation one needs deep stimulation, second one obtainsphosphenes located throughout the visual Weld while weobtained only paracentral phosphenes.
General conclusions
The main thrust of this study is twofold: First, by using ITtime as a marker of the callosal site of transfer we foundthat both phosphenes generated by V1 TMS and imaginedsimilar stimuli are likely to transfer at the level of V1/V2for the reasons detailed above in the “Discussion” of Exp.1. In contrast, visually presented stimuli use more anteriorcallosal connections, as shown by several neuropsychologi-cal and brain imaging studies using the PoVenberger para-digm (Iacoboni 2006; Iacoboni and Zaidel 2004; Marziet al. 1999; Omura et al. 2004; Tettamanti et al. 2002;Weber et al. 2005). Second, we obtained novel evidence ofphosphenes generated by TMS applied to an area close tothe intraparietal sulcus in posterior parietal cortex. Thesephosphenes show an IT time that is much faster than that ofoccipital phosphenes and similar to that obtained with visu-ally presented stimuli. This dissociation between IT timefollowing stimulation of occipital and parietal areas doesnot necessarily imply that the sites for phosphene percep-tion following TMS correspond to the site of IT. The
subjective experience of phosphenes is likely to be sub-served by V1: as shown by several studies (Cowey andWalsh 2000; Pascual-Leone and Walsh 2001; Silvantoet al. 2005, 2007), without V1 there is no awareness ofTMS induced phosphenes. Therefore, it is possible thatTMS over parietal cortex generated activity that reachedconsciousness through activation of V1. This would explainwhy the location, size and shape of phosphenes are similarfor parietal and occipital phosphenes. However, it is rea-sonable to hypothesize that the site of callosal transmissionis independent from that mediating phosphene awareness.We believe that callosal transmission of phosphenes occurshomotopically at the location of TMS application (seeIlmoniemi et al. 1997). This holds for both parietal andoccipital stimulation with the diVerence that in the lattercase site of IT and phosphene perception coincides whilethis is not the case for the former. It is well known from thepioneering studies of Libet (for a recent review see Libet2006) on electrical stimulation of somatosensory cortex andthat of Pollen (2004, 2006) on stimulation of the visual cor-tex that electrical current directly applied over the corticalsurface yields perceptual eVects only after 0.5–1 s. Interest-ingly, Pollen used RT to the onset of phosphenes, as in thepresent study, and found values in broad keeping with oursobtained with TMS. The interpretation given by Libet andPollen on the reasons for this delay is diVerent but the veryexistence of the phenomenon suggests that awareness ofphosphene perception might occur later than the callosaltransmission necessary for manual response. Therefore, wespeculate that following TMS to posterior parietal cortextwo parallel neural events occur: a callosal transmission tothe other hemisphere when a motor response with the ipsi-lateral hand is required and an orthodromic transmission toV1 for conscious experience. This sequence of eventsmight mimic what happens during natural visual stimuluspresentations: the activation of V1 that gives rise toconscious perception might normally depend on prior pari-etal stimulation and orthodromic stimulation of V1. Theparietal contribution is by-passed when V1 is activated byTMS and IT may then depend on the slower processes wesee in the split brain. If this is correct, one would also haveto assume that an imagined stimulus also by-passes parietalactivation.
Acknowledgments The study has been Wnanced by the ItalianMIUR. We thank Paola Cesari for the use of the TMS facility at theFaculty of Motor Science of the University of Verona.
Addendum
This article is dedicated to Giovanni Berlucchi who hastaught the senior author how to do science honestly, crea-tively and enthusiastically. Giovanni, and his colleagues
and friends Carlo Umiltà and Giacomo Rizzolatti have thehistorical merit of having revitalized the PoVenberger para-digm with their seminal 1971 paper, which is an example ofgreat behavioral science, neat and clever. Since then thePoVenberger paradigm has been employed many times inboth normal and brain damaged people always providingimportant information for understanding how the cerebralhemispheres interact. In fact, the present study has beencarried out using stimuli that do not really exist in the exter-nal world but are in the mind of the beholder. We trustGiovanni will appreciate this bizarre use of the PoVenber-ger paradigm.
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