Functional topography of the secondary somatosensory cortex for nonpainful and painful stimuli: an fMRI study

NeuroImage 20 (2003) 1625–1638

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Functional topography of the secondary somatosensory cortex for nonpainful and painful stimuli: an fMRI study Antonio Ferretti,a,b,c,* Claudio Babiloni,d,g Cosimo Del Gratta,a,b,cMassimo Caulo,a,b Armando Tartaro,a,b Lorenzo Bonomo,a,b Paolo Maria Rossini,e,f,g and Gian Luca Romania,b,c a

b

Department of Clinical Sciences and Bio-imaging, University of Chieti, Italy ITAB–Institute for Advanced Biomedical Technologies, Fondazione Universita` Gabriele d’Annunzio, Chieti, Italy c INFM–National Institute for the Physics of Matter, Group of Chieti, Italy d Department of Human Physiology and Pharmacology, University of Rome La Sapienza, Italy e AFaR Department of Neurology, Fatebenefratelli Hospital, Isola Tiberina, Rome f Department of Clinical Neurosciences, University of Rome Campus Biomedico, Italy g IRCCS S. Giovanni di Dio, Via Pilastroni, Brescia, Italy Received 27 March 2003; revised 7 July 2003; accepted 14 July 2003

Abstract The regional activity of the contralateral primary (SI) and the bilateral secondary (SII) somatosensory areas during median nerve stimulations at five intensity levels (ranging from nonpainful motor threshold to moderate pain) was studied by means of functional magnetic resonance imaging (fMRI). The aim was to characterize the functional topography of SII compared to SI as a function of the stimulus intensity. Results showed that the galvanic stimulation of the median nerve activated the contralateral SI at all stimulus intensities. When considered as a single region, SII was more strongly activated in the contralateral than in the ipsilateral hemisphere. When a finer spatial analysis of the SII responses was performed, the activity for the painful stimulation was localized more posteriorly compared to that for the nonpainful stimulation. This is the first report on such a SII segregation for transient galvanic stimulations. The activity (relative signal intensity) of this posterior area increased with the increase of the stimulus intensity. These results suggest a spatial segregation of the neural populations that process signals conveyed by dorsal column-medial lemniscus (nonpainful signals) and neospinothalamic (painful signals) pathways. Further fMRI experiments should evaluate the functional properties of these two SII subregions during tasks involving sensorimotor integration, learning, and memory demands. © 2003 Elsevier Inc. All rights reserved. Keywords: Functional magnetic resonance imaging (fMRI); Primary somatosensory area (SI); Secondary somatosensory area (SII); Median-nerve electrical stimulation; Pain

Introduction Electrical stimulation of the median nerve at the wrist is a very popular approach for the study of somatosensory systems under physiological and clinical conditions, due to its low cost and the extreme feasibility of the procedure. Somatosensory-evoked neuromagnetic fields (SEFs) fol-

* Corresponding author. Institute for Advanced Biomedical Technologies, University G. D’Annunzio of Chieti, Via dei Vestini, 33, 66013 Chieti (CH), Italy. Fax: ⫹39-0871-3556930. E-mail address: [email protected] (A. Ferretti). 1053-8119/$ – see front matter © 2003 Elsevier Inc. All rights reserved. doi:10.1016/j.neuroimage.2003.07.004

lowing median nerve electrical stimulation have shown that contralateral primary (SI) and bilateral secondary (SII) somatosensory areas are deeply involved in the processing of nonpainful and painful stimuli (Peresson et al., 1992; Jousmaki et al., 1998; Tsutada et al., 1999; Kakigi et al., 2000). SI would process and encode the type and intensity of the sensory inputs, whereas SII would play a higher level sensory processing related to attention, learning, memory, and integration of nociceptive and nonnociceptive inputs (Ploner et al., 1999; Schnitzler and Ploner, 2000; Hamalainen et al., 2002). In a recent whole-head magnetoencephalographic (MEG) study from our institute (Torquati et

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Fig. 1. The five intensity levels used for electrical stimulation of the median nerve. Fig. 2. (A) Results of the group analysis showing the activated areas in the somatosensory cortex at the stimulation level I5 (painful) superimposed onto axial and coronal sections of an individual brain (images are displayed using the neurological convention, i.e., right is right, left is left). Top, axial view. Left, activation occurring in contralateral SI; right, activation in SII. Bottom, coronal view. Left, anterior SII areas and contralateral SI; right, posterior SII areas and contralateral SI.

al., 2002), SI and SII responses have been found to be differently modulated by the intensity of electrical stimulation ranging from the sensory to the weak painful level. The SI response progressively increased with the intensity of the stimulation, whereas the SII response showed a dip for stimulus intensities between motor and pain thresholds. One of the possible interpretations of this effect was the exis-

tence of two neuronal populations in SII diversely sensitive to nonpainful and painful stimuli. However, the spatial resolution of MEG does not allow the spatial discrimination of two closely located neural populations such as those hypothesized for the processing of painful and nonpainful stimuli in SII areas. A powerful approach for the study of the somatotopy of

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Fig. 2. (B) Results of the group analysis showing the activated areas in the somatosensory cortex at the stimulation level I1 (nonpainful) superimposed onto axial and coronal sections of an individual brain. Top, axial view. Left, activation in contralateral SI; right, activation in SII. Bottom, coronal view. Left, activation in anterior SII areas; right, activation in conralateral SI.

SI and SII is functional magnetic resonance imaging (fMRI). In recent fMRI studies using nonpainful electrical stimulation of the median and tibial nerves (Del Gratta et al., 2000, 2002), a rough but clear somatotopic organization of the SII areas was demonstrated. The median nerve area was more anterior and more inferior than the tibial nerve area. Median and tibial nerves induced a bilateral SII activation, stronger in the contralateral than ipsilateral hemisphere. Similar results have been obtained in another fMRI study (Ruben et al., 2001) in which the second finger, the fifth finger, and the hallux were stimulated. On the other hand, fMRI studies have shown a marked activation of bilateral SII areas to painful stimulations in healthy and diseased human subjects (Bornho¨ vd et al.,

2002; Brooks et al., 2002; Lin and Forss, 2002). Finally, a recent event-related fMRI experiment (3.0 T) has disclosed separate SII representations for painful and nonpainful intramuscular electrical stimulations applied to the abductor pollicis brevis (Niddam et al., 2002). In this experiment activation elicited by two levels of stimulus intensity was compared. In the present study, we investigated the responses of the contralateral SI and bilateral SII by means of 1.5 T fMRI during galvanic median nerve stimulations at five stimulus intensities, ranging from nonpainful motor threshold to moderate pain. A detailed study of the functional topography of SII as a function of the stimulus intensity was performed.

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Methods Subjects and stimulation procedures Eight healthy volunteers ranging in age from 19 to 22 years (4 males, 4 females) were enrolled in this study. All of them were right handed according to the Edinburgh Inventory (Oldfield, 1971). All subjects gave their written informed consent according to the Declaration of Helsinki (World Medical Association Declaration of Helsinki, 1997) and could request an interruption of the investigation at any time. The general procedures were approved by the local Institutional Ethics Committee. The electric stimulus was a rectangular pulse with 1.9-Hz frequency and 400-␮s duration and was delivered to the right median nerve at the wrist via nonmagnetic AgCl electrodes. The stimulation levels were determined by a series of increasing and decreasing stimulus intensities at the beginning of each recording block. The experimental design included five levels of galvanic stimulation intensity ranging from nonpainful to moderately painful levels (I1 to I5), according to a subjective scale (Fig. 1). This kind of stimulation mainly excites Aalpha and Abeta fibers at nonpainful levels and mainly excites Adelta fibers at painful levels (York, 1985; Babiloni et al., 2001). The first level (I1) was the motor threshold stimulus, the fourth level (I4) was the slight pain stimulus, and the fifth level (I5) was above the slight pain stimulus. With reference to a subjective scale for pain ranging from 0 (no sensation) to 10 (pain tolerance threshold), first level (I1) corresponded to 2 (painless thumb muscle twitch), second level (I2) corresponded to 3, third level (I3) corresponded to 4, fourth level (I4) corresponded to 5 (slight pain), and the fifth level (I5) corresponded to 6 (moderate pain). The stimulation current for the first level (I1) reaching the motor threshold varied across subjects in the range of 3–13 mA (mean value 7.0 ⫾ 0.3 mA), whereas the maximum stimulation current for the fifth level (I5) never exceed 50 mA. Individual stimulation levels were assessed outside the scanner just before the fMRI session. To maintain a stable level of vigilance and to minimize the effects of habituation, the fMRI runs at different stimulation levels were randomized across subjects. The stimulation level in each run was kept constant. Each participant was given a brief training session in which he learned to stare at a visual fixation point and to minimize blinking and eye movements during the recording block. Subjects were not required to score the perceived stimulus during the experiment in order to avoid any effect due to the movement of the fingers used for the scoring. fMRI recordings BOLD contrast functional imaging was performed with a Siemens Magnetom Vision scanner at 1.5 T by means of T2*-weighted echo planar imaging (EPI) free induction

decay (FID) sequences with the following parameters: TR 3 s, TE 60 ms, matrix size 64 ⫻ 64, FOV 256 mm, in-plane voxel size 4 ⫻ 4 mm, flip angle 90°, slice thickness 3 mm, and no gap. A standard head coil was used and the subject’s head was fixed by foam pads to reduce involuntary movement. Functional volumes consisted of 22 transaxial slices parallel to the AC-PC line including the cortical regions of interest (SI, SII). The experimental paradigm was a block design alternating a state of stimulation of 30 s with a control state having the same duration. For each stimulus intensity a run of 60 vol was acquired starting with a control period. A high-resolution structural volume was acquired at the end of the session via a 3D MPRAGE sequence with the following features: axial, matrix 256 ⫻ 256, FOV 256 mm, slice thickness 1 mm, no gap, in-plane voxel size 1 ⫻ 1 mm, flip angle 12°, TR ⫽ 9.7 ms, TE ⫽ 4 ms. Data analysis Raw data were analyzed by means of the Brain Voyager 4.6 software (Brain Innovation, The Netherlands). Due to T1 saturation effects, the first two scans of each run were discarded from the analysis. Preprocessing of functional scans included motion correction and removal of linear trends from voxel time series. A three-dimensional motion correction was performed by means of a rigid body transformation to match each functional volume to the reference volume (the third volume, since the first two were discarded to avoid the T1 saturation effect) estimating three translation and three rotation parameters. These parameters were stored in log files and inspected to check that estimated movement was not larger than 3 mm or 3° (approximately one voxel) for each functional run. Preprocessed functional volumes of a subject were coregistered with the corresponding structural data set. Since the 2D functional and 3D structural measurements were acquired in the same session, the coregistration transformation was determined using the Siemens slice position parameters of the functional images and the position parameters of the structural volume. In some cases, after accurate visual inspection, this transformation was adjusted to account for subject movement between functional and anatomical scans. Structural and functional volumes were transformed into the Talairach space (Talairach and Tourmoux, 1988) using a piecewise affine and continuous transformation. Functional volumes were resampled at a voxel size of 3 ⫻ 3 ⫻ 3mm. Statistical analysis was performed for individual subjects and stimulus intensities using the general linear model (GLM) (Friston et al., 1995) with correction for temporal autocorrelation (Bullmore et al., 1996; Woolrich et al., 2001). To account for the hemodynamic delay, the boxcar waveform representing the rest and task conditions was convolved with an empirically founded hemodynamic response function (Boynton et al., 1996). No spatial or temporal smoothing was performed in this analysis.

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In addition, to search for activated areas that were consistent for the entire group of subjects, a statistical group analysis was performed as well. In this analysis the time series from each run and subject were z-normalized and concatenated prior to the GLM computation. Individual and group statistical maps were thresholded at P ⬍ 0.0004 at the voxel level and a cluster size of at least four voxels was required. These thresholds and an estimate of the spatial correlation of voxels (Forman et al., 1995; 3dFWHM routine of AFNI package, Cox, 1996) were used as input in a Monte-Carlo simulation (AlphaSim routine of AFNI package, Cox, 1996; Forman et al., 1995) in order to assess the overall significance level (the probability of a false detection for the entire functional volume). In this way we obtained P ⬍ 0.05 as the significance level corrected for multiple comparisons. Individual thresholded statistical maps were then superimposed on the respective structural scans for the localization of significantly activated areas, while thresholded group activation maps were superimposed on the (Talairach transformed) structural scan of one of the subjects. Due to the expected close position of the different activation sites in SII and to the well-known problem of intersubject anatomical variability, the relationship between the stimulus intensity and the BOLD signal in each area was studied using data from individuals and is discussed in the next section. Regions of interest (ROIs) Cortical areas corresponding to contralateral SI and bilateral SII were traced independently by two expert experimenters along the lines outlined in previous literature (Baraldi et al., 1999; Picard and Strick, 1996). Anatomical landmarks included the posterior bank of the central sulcus and the postcentral gyrus at the omega zone for the contralateral SI, and the upper bank of the lateral sulcus near the posterior pole of the insula for the bilateral SII. The mediolateral extension of SI was based on the omega zone landmark, while the medio-lateral extension of SII ranged from the lip of the upper bank of the lateral sulcus to its fundus. Note that we used the omega fold to delimitate the “hand” area in SI, following the lead of several fMRI studies showing marked “hand” sensorimotor responses in the omega zone (Puce, 1995; Rumeau et al., 1994; Toyokura et al., 1999; Wood et al., 1988; Pizzella et al., 1999). Regions of interest in the cortical areas outlined above were determined by considering the whole mask obtained from voxels activated at any stimulus level. The mean time course of the fMRI signal of voxels belonging to a given ROI was analysed to inspect the effect of different stimulus conditions. Attention was devoted to distinguish activated areas in the posterior insula from activated areas in SII, the latter being located in the deep surface of the parietal lobe corresponding to the parietal operculum (so that the SII

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locations were more lateral with respect to the insular cortex). The subject’s responses to different stimulation intensities were characterized by evaluating the BOLD contrast signal intensity variation in each ROI. In order to avoid the transient hemodynamic response the first three scans of each epoch were discarded, so that the baseline signal level was calculated as the mean of the last seven scans of the rest epochs and the activation signal as the mean of the last seven scans of the task epochs. Then the relative signal variation between baseline and activation was calculated. The regional comparison of activation was undertaken by means of the analysis of variance (ANOVA) for repeated measures. Mauchley’s test was used to evaluate the sphericity assumption. The number of degrees of freedom was corrected by means of the Greenhouse-Geisser procedure, and the Duncan test was used for post hoc comparisons. The dependent variable of the ANOVA analysis was the BOLD signal relative variation between the stimulation and the rest conditions and the factors were the Stimulus Intensity (I1, I2, I3, I4, I5) and ROI (contralateral SI, contralateral SII, and ipsilateral SII).

Results Group analysis A consistent activation across subjects was observed at the maximal stimulus intensity (I5) in contralateral SI, bilateral SII, and bilateral insular cortex. A noteworthy finding was that two distinct regions in the anterior-posterior axis of SII were active during the painful stimulation (level I5) in both the contralateral (cSII) and the ipsilateral (iSII) hemispheres. We termed these areas anterior (iSIIa and cSIIa) and posterior (iSIIp and cSIIp) subregions of SII. At the motor threshold stimulation level, only SI, iSIIa, and cSIIa were significantly activated. The group statistical maps for the I5 (moderate pain) and the I1 (motor threshold) intensity levels are shown in Figs. 2a and b, respectively, superimposed on an individual (Talairach transformed) structural image. The location of SI, iSIIa, and cSIIa activation did not change when considering different stimulation intensities. Talairach coordinates of activated areas in SI, SII, and insula were derived from the centroids of clusters of activation during the stimulation at the I1 and I5 levels. These coordinates are listed in Table 1. The ANOVA analysis performed on the coordinates of the SII areas across subjects yelded a high significance level (P ⬍ 0.0003) when the y coordinates (anterior-posterior direction) of SIIa and SIIp were compared, with no significant interaction between the hemispheres. When the x coordinates of SIIa and SIIp were compared only the contralateral pair showed a significant difference (P ⬍ 0.01). The z coordinates did not show any significant difference.

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Fig. 3. Activated areas in contralateral SI and bilateral SII in two subjects during stimulation at level I5. In the second subject activation was superimposed on the inflated cortex. Note the anterior and posterior subregions of SII.

Single subject analysis The aforementioned BOLD activation in SI, cSIIa, iSIIa, cSIIp, and iSIIp was observed in all subjects after the

stimulation at maximal stimulus intensity (I5, moderate pain). Fig. 3 shows the BOLD cortical activation after the moderately painful stimulation (I5) in two representative subjects.

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Fig. 4. BOLD signal time course (average across subjects) in activated areas in SI and SII (anterior and posterior collapsed) during electrical median nerve stimulation at motor threshold (I1, blue) and at maximal stimulus intensity (I5, red). Error bars are standard errors.

The fMRI signal time course during the rest and task conditions was derived for each subject and cortical area. This signal time course, averaged across epochs and subjects, is shown in Fig. 4 for the stimulations at I1 and I5 with SIIa and SIIp collapsed. In contrast the signal time course of the separate SIIa and SIIp areas is illustrated in Fig. 5. In all cases the signal intensity increased reaching an initial peak about 6 s (i.e., two scans) after the stimulus onset (placed at the end of scan 6), and then decreased to a plateau level in the following part of the stimulation. A Student’s t test was performed in order to detect statistical differences between the signal time courses during stimulation at I1 and I5. The

comparison was carried out separately for the first half (scans 7–11) and the second half (scans 12–16) of the stimulation period. In the first half of stimulation a significant difference was observed for SI (P ⬍ 0.01) and iSII (P ⬍ 0.02) but not for cSII (P ⬎ 0.05). In the second half of stimulation the difference was significant for SI (P ⬍ 0.001), iSII (P ⬍ 0.004), and cSII (P ⬍ 0.002). When anterior and posterior SII subregions were considered, only posterior areas showed significant differences when comparing the signal during stimulations at I1 and I5. In the first half of stimulation a significant difference was observed for iSIIp (P ⬍ 0.005) and cSIIp (P ⬍ 0.04). In the second half

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Table 1 Group analysis of the BOLD cortical activation after the galvanic median nerve stimulations: Talairach coordinates (centroids of activated clusters) of the peak activity within SI, SII subregions, and insula during stimulation at motor threshold (I1) and moderate painful (I5) stimulus intensity Area

x

y

z

Nonpainful stimulation (I1) ⫺36 ⫺35 ⫺43 ⫺23 48 ⫺22 Painful stimulation (I5) ⫺36 ⫺34 ⫺41 ⫺22 48 ⫺23 ⫺49 ⫺31 45 ⫺35 ⫺34 ⫺2 31 13

Contralateral SI Contralateral SIIa Ipsilateral SIIa Contralateral SI Contralateral SIIa Ipsilateral SIIa Contralateral SIIp Ipsilateral SIIp Contralateral Insula Ipsilateral Insula

48 17 19 48 18 20 19 21 15 15

of stimulation a significant difference was observed for iSIIp (P ⬍ 0.003) and cSIIp (P ⬍ 0.001) as well. As a general behavior, the observed differentiation between the BOLD response in a comparison of the two stimulus intensities was larger in the second half of stimulation. A similar result was found also in Krause et al. (2001) for SI when stimulating fingers at two nonpainful intensity levels. Statistical analysis First, activation in SI and SII was compared considering SII as a single activated region (SIIa and SIIp collapsed). An ANOVA analysis was used to evaluate the influence of the factors Stimulus Intensity (I1, I2, I3, I4, I5) and ROI (SI, cSII, iSII) on the fMRI BOLD activation (relative variation). There was a statistical main effect for the factor Stimulus Intensity (F(4,28)⫽5.4; P ⬍ 0.002), indicating that BOLD activation increased with the increment of the stimulus intensity from nonpainful (I1) to painful (I4, I5). Furthermore, there was a statistical main effect of the factor ROI (F(2,14) ⫽ 7.6; P ⬍ 0.006), indicating that BOLD activation was stronger for the SI and cSII with respect to the iSII. The BOLD response as a function of stimulus intensity is reported in Fig. 6 for each ROI (mean values across subjects).

Table 3 Pearson correlation factors (r) and significance of correlation (P) between the stimulus intensity and the BOLD response in cSI, cSIIa, cSIIp, iSIIa, iSIIp, i insula, and c insula Area

r⫽

P⬍

cSI cSIIa cSIIp iSIIa iSIIp i insula c insula

0.94 0.16 0.98 0.19 0.95 0.91 0.96

0.02 0.79 0.003 0.76 0.01 0.03 0.01

Second, SII activation was evaluated considering the anterior and posterior subregions of the SII as separate functional subregions. A two-way ANOVA design was used to evaluate the influence of the factors Stimulus Intensity (I1, I2, I3, I4, I5) and ROI (cSIIa, iSIIa, cSIIp, iSIIp) on the BOLD activation (relative variation). A statistical main effect was observed for the factor ROI (F(3,21) ⫽ 5.8; P ⬍ 0.004), indicating that both cSIIa and cSIIp showed a higher response with respect to iSIIa and iSIIp. A statistical main effect was observed also for the factor Stimulus Intensity but at a lower significance level (F(4,28) ⫽ 2.9; P ⬍ 0.04). Furthermore, a significant interaction (F(12,84) ⫽ 2.4; P ⬍ 0.01) between the two factors revealed that the posterior SII showed an increasing BOLD activation with the stimulus intensity, while a practically constant BOLD activation was observed in the anterior SII areas as a function of the stimulus intensity. Post hoc comparisons of BOLD response at different stimulation intensities were performed by means of the Duncan test for each posterior SII area (Table 2). Finally, the Pearson correlation between the stimulus intensity and the BOLD response was calculated for each area. The correlation coefficients and the related significance values are reported in Table 3. The BOLD response as a function of stimulus intensity is shown in Fig. 7 for the subregions in SII (mean values across subjects). Activation in insular cortex Compared to the activation in SII, the BOLD cortical activation at contralateral and ipsilateral insula was much

Table 2 Post hoc comparisons (Duncan test, P values when ⬍0.05) of BOLD response at different stimulation intensities for cSIIp and iSIIp cSIIp I1 I1 I2 I3 I4 I5

0.01 0.001 0.00002

iSIIa I2

0.01

I3

I4

I5

0.01

0.001

0.00002 0.01 0.05

0.05

I1

0.01 0.01 0.0004

I2

0.03 0.03 0.002

I3

I4

I5

0.01 0.03

0.01 0.03

0.0004 0.002

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Fig. 5. BOLD signal time course (average across subjects) in subregions of SII during electrical median nerve stimulation at I1 (blue) and I5 (red). Error bars are standard errors.

less evident. The group statistical map is shown in Fig. 8. In single subject analysis, a statistically significant BOLD activation was recognized only in 5 out of 8 subjects and was clearly evident only for painful stimuli I4, I5. In insular areas, the average BOLD cortical activation slightly increased as a function of the stimulus intensity (Fig. 8). Note that this function seems to be different from that of the posterior subregions of SII. In particular the ipsilateral insula showed a pain versus nonpain effect (i.e., it was activated only at the painful levels I4, I5). Nevertheless these results should be confirmed on an extended number of subjects.

Discussion The above described results indicate that the contralateral SI and bilateral SII increase their activation with the increase of the stimulus intensity from nonpainful to painful. Note that this is true only when the SII is considered as a single region. Furthermore, the contralateral SI and SII are characterized by a stronger activation with respect to the ipsilateral SII, as an interesting hemispherical asymmetry given the well-known bilateral SII involvement in the processing of nonpainful and painful stimuli (Schnitzler and Ploner, 2000; Timmermann et al, 2001; Lin and Forss 2002; Torquati et al., 2002; Frot and Mauguiere 2003). This hemi-

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Fig. 6. Mean BOLD response across subjects as a function of the stimulus intensity in activated areas in SI and in SII (anterior and posterior collapsed). Error bars are standard errors.

spherical asymmetry could reasonably be explained by considering the convergent somatosensory inputs to the contralateral SI and SII. Contralateral SII receives both nociceptive and tactile inputs from periphery via ventroposterior inferior and/or ventro-dorso-medial thalamic nuclei (Apkarian and Shi, 1994; Schnitzler and Ploner, 2000). Futhermore, the contralateral SII receives central inputs from primary somatosensory cortex of the same hemisphere (Barbaresi et al., 1994; Barba et al., 2001; Forss et al., 2001). In the other hemisphere, the ipsilateral SII receives nociceptive, transcallosal inputs not only from opposite SII (Schnitzler and Ploner, 2000; Frot and Mauguiere 2003) but also from ipsilateral thalamic nuclei (Olausson et al., 2001; Fabri et al., 2002). Regarding the tactile system, the existence of ipsilateral inputs from periphery to SII is more controversial; i.e., there is pro evidence in hemispherectomized and unilateral stroke patients (Forss et al., 2001; Olausson et al., 2001) but con evidence in patients with resection of corpus callosum (Fabri et al., 2001, 2002). As a major result, our data showed that the SII was

characterized, in both hemispheres, by two distinct areas of BOLD activation in the anterior-posterior direction. These areas exhibited different responses as a function of the stimulus intensity. The posterior but not anterior SII increased its activation as a function of the increasing intensity of the galvanic stimulation. A reasonable explanation is that SII is characterized by at least two populations diversely sensitive to pain. This explanation is also in agreement with that suggested by the MEG results of Torquati et al. (2002), as discussed in the Introduction. Our results extend previous evidence, showing separate SII representations for the painful and nonpainful intramuscular electrostimulation, obtained with two levels of stimulus intensity and 3.0 T event-related fMRI (Niddam et al., 2002). In that study, nonpainful stimulation activated mainly the lateral segment of SII, while the painful stimulation activated both the lateral and the medial part of SII. In our study, the nonpainful stimulation activated roughly the same lateral SII subregion, while the painful stimulation activated an additional SII subregion located more posteri-

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Fig. 7. Mean BOLD response across subjects as a function of the stimulus intensity in activated areas in SII subregions discussed in the main text. Error bars are standard errors.

orly rather than more medially. Indeed, the shift toward the posterior direction was found in all subjects with respect to the SII subregion responding to the nonpainful stimuli. Finally, we found an involvement of the bilateral anterior insula during painful stimulation, in substantial agreement with Niddam et al. (2002). Keeping in mind these data, it can be speculated that at least two SII representations may account for different kinds of galvanic stimulations, namely a more medial representation for muscle nociceptive receptors (Niddam et al., 2002) and a more posterior representation for skin nociceptive receptors as those involved by a median nerve stimulation at the wrist. These different representations raise the issue of a parallel processing of distinct pain features in SII with respect to SI. A comparison of our results with those of intracerebral electroencephalo-

graphic recordings (Frot et al., 2001; Frot and Mauguiere, 2003) is difficult because the regions covered by the recording contacts do not completely overalp the regions activated in our study. Indeed, the position of the sampling electrode array was too anterior with respect to the posterior SII subregions. Further investigations with intracerebral electroencephalographic recordings might unveil the temporal dynamics of the SII activation during nonpainful compared to painful stimulations. The different sensitivity to the nonpainful and painful stimuli of the SII subregions is consistent with the different features of the nociceptive and tactile systems. Tactile stimulation triggers a sequential engagement of contralateral SI, contralateral SII, and ipsilateral SII, thus suggesting a strong hierarchical control of information processing in ipsilateral

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Fig. 8. Results of group analysis showing activated areas during stimulation at level I5 in insular regions. BOLD response as a function of stimulus intensity in the contralateral and ipsilateral insula: mean values across subjects. Error bars are standard errors.

SII. Conversely, painful stimulation induces a mainly parallel activation of this distributed network (Ploner et al., 1999; Treede et al., 2000). This parallel organization bypasses several cortico-cortical and transcallosal connections, reduces the distance from periphery to ipsilateral SII, and shortens the processing time of the painful stimulus. The existence of two SII subregions is also consistent with the complexity of thalamic projections from several relays (Apkarian and Shi, 1994; Schnitzler and Ploner, 2000) within a multifunctional network involved in (1) noxious stimulus recognition, learning, and memory, (2) autonomic reactions to noxious stimuli, and (3) affective aspects of pain-related learning and memory (Ploner et al., 1999, 2000; Schnitzler and Ploner, 2000; Treede et al., 2000; Hamalainen et al., 2002). The existence of a separate SII representation for painful stimuli is not surprising. Animal studies using microelectrode recordings have shown that several areas located in the parietal operculum and functionally connected to SII take part in the processing of painful and nonpainful inputs,

namely retroinsular, granular insula (Burton and Fabri, 1995), and 7b (Robinson and Burton, 1980a, 1980b) areas. In humans, intracranial evoked potentials have demonstrated that the parasylvian cortex (particularly SII) includes separate cortical relay stations in the central processing of pain (Ploner et al., 1999; Treede et al., 2000; Frot et al., 2001, 2003). Along this line, the present study confirmed the extreme sensitivity of the SII subregion to painful stimuli as opposite to the scarce activation of the insula. A reasonable explanation is that the SII activation may mainly reflect sensory stimulus recognition, whereas insular activation may mainly reflect a further processing stage related to the emotional and attentional coloring of the stimulus (Treede et al., 2000; Brooks et al., 2002; Frot and Mauguiere 2003). Here the emotional and attentional coloring of the stimulus was probably modest, due to the features of the painful stimulation (namely, repetitive and brief). A word of note is in order when comparing group results on activation loci in SII and in the posterior insular cortex from different studies. Indeed, given the close position of

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the two areas in the brain, the classification of the activated areas based only on the Talairach coordinates should be considered cautiously. In fact, the anatomy of a single brain may deviate significantly from the Talairach brain making normalization difficult. This might determine between-subject coregistration errors, leading to a blurring of activated areas. In this regard, our location of contralateral anterior SII (⫺43, ⫺23, 17) is close to the location reported in the PET studies with tonic and thermal stimulation by Craig et al. (2000) (⫺36, ⫺22, 24) and Derbyshire and Jones (1998) (⫺40, ⫺20, 16) and in the fMRI study by Brooks et al. (2002) (⫺39, ⫺20, 20) which were classified as posterior insula. However, in our study, we classified the activation for each individual subject by projecting statistical maps onto the structural brain image: this allowed a safe localization of the cortical activity in the surface of the parietal operculum (SII) rather than in the Insula.

Conclusions We evaluated the SI and SII activity by means of fMRI during the galvanic median nerve stimulation at intensity levels ranging from motor threshold to pain. SI showed an increasing response function with increasing stimulus intensities. When considered as a single region, SII also showed a similar response function. When the detailed activation pattern in SII was considered, the activity for the painful stimulation was localized more posteriorly compared to that for the nonpainful stimulation, suggesting a spatial segregation of the neural populations that process painful and nonpainful signals. This was the first report on such a SII segregation for transient galvanic stimulations. The use of the galvanic stimulation allowed a direct comparison of the results with those of previous experiments using fMRI (Niddam et al., 2002) and whole-head magnetoencephalographic techniques (Torquati et al., 2002). However, further investigations should try to replicate the present results with stimulations exhibiting higher nociceptive specificity such as CO2 laser beam (Bromm et al., 1998; Frot and Mauguiere 2003) and inducing no overt muscular twitches. Moreover, future fMRI experiments should evaluate the functional properties of these two SII subregions during tasks involving sensorimotor integration, learning, and memory demands. In this framework, the use of more powerful fMRI scanners (3 to 7 T) and proper experimental design might address the relationship between possible multiple painful representations in SII and distinct painful sensations referred to as first and second pain sensation (Ploner et al., 1999). Finally, the use of intracranial EEG recordings could reveal fine grains of the spatial and temporal properties of the SII subregions in the processing of nonpainful and painful phasic stimuli and confirm the results suggested by our study.

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Acknowledgments This work was partially supported by a grant from the Italian Ministry of Research to the Center of Excellence on Aging of the University.

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