www.elsevier.com/locate/ynimg NeuroImage 31 (2006) 710 – 720

Rapid Communication

Two different reorganization patterns after rehabilitative therapy: An exploratory study with fMRI and TMS Farsin Hamzei,a,b,* Joachim Liepert, a Christian Dettmers, c Cornelius Weiller, b and Michel Rijntjes b a

Department of Neurology, University Medical Center Hamburg Eppendorf, Germany Department of Neurology, University Clinic Freiburg, Germany c Schmieder-Kliniken Konstanz, Germany b

Received 12 September 2005; revised 13 December 2005; accepted 15 December 2005 Available online 3 March 2006 We used two complementary methods to investigate cortical reorganization in chronic stroke patients during treatment with a defined motor rehabilitation program. BOLD (‘‘blood oxygenation level dependent’’) sensitive functional magnetic resonance imaging (fMRI) and intracortical inhibition (ICI) and facilitation (ICF) measured with transcranial magnetic stimulation (TMS) via paired pulse stimulation were used to investigate cortical reorganization before and after ‘‘constraint-induced movement therapy’’ (CI). The motor hand function improved in all subjects after CI. BOLD signal intensity changes within affected primary sensorimotor cortex (SMC) before and after CI showed a close correlation with ICI (r = 0.93) and ICF (r = 0.76) difference before and after therapy. Difference in number of voxels and ICI difference before and after CI also showed a close correlation (r = 0.92) in the affected SMC over the time period of training. A single subject analysis revealed that patients with intact hand area of M1 (‘‘the hand knob’’) and its descending motor fibers (these patients revealed normal motor evoked potentials [MEP] from the affected hand) showed decreasing ipsilesional SMC activation which was paralleled by an increase in intracortical excitability. This pattern putatively reflects increasing synaptic efficiency. When M1 or its descending pyramidal tract was lesioned (MEP from the affected hand was pathologic) ipsilesional SMC activation increased, accompanied by decreased intracortical excitability. We suggest that an increase in synaptic efficiency is not possible here, which leads to reorganization with extension, shift and recruitment of additional cortical areas of the sensorimotor network. The inverse dynamic process between both complementary methods (activation in fMRI and intracortical excitability determined by TMS) over the

* Corresponding author. University Clinic of Freiburg, Department of Neurology, Breisacherstrasse 64, 79106 Freiburg im Breisgau, Germany. Fax: +49 761 270 5310. E-mail address: [email protected] (F. Hamzei). Available online on ScienceDirect (www.sciencedirect.com). 1053-8119/$ - see front matter D 2006 Elsevier Inc. All rights reserved. doi:10.1016/j.neuroimage.2005.12.035

time period of CI illustrates the value of combining methods for understanding brain reorganization. D 2006 Elsevier Inc. All rights reserved. Keywords: Stroke Rehabilitation

reorganization;

BOLD;

Cortical

excitability;

Different patterns of reorganization in human and non-human primates after CNS lesion have been described: perilesional extension, shifts from primary to secondary motor areas and to the homologous areas of the non-affected hemisphere, and an increase and decrease in ipsilesional primary sensorimotor (SMC) activation during acute and subacute phases of stroke. This reflects a dynamic process in cortical reorganization during recovery (Chollet et al., 1992; Weiller et al., 1992, 1993; Nudo et al., 1996; Cramer et al., 1997; Dettmers et al., 1997; Cao et al., 1998; Rouiller et al., 1998; Rossini et al., 1998; Seitz et al., 1998; Seitz and Azari, 1999; Darian-Smith et al., 1999; Nelles et al., 1999, 2001; Marshall et al., 2000; Calautti et al., 2001; Feydy et al., 2002; Ward et al., 2003). Especially, whether an increase or a decrease of fMRI (functional magnetic resonance imaging) or PET (positron emission tomography) activation within ipsilesional primary motor cortex is related to improved hand function is discussed (Weiller et al., 1992; Ward et al., 2003). The meaning of fMRI or PET activation changes in the affected primary motor cortex is still unclear in electrophysiological terms. The aim of this exploratory study was to better understand the mechanisms of brain reorganization during treatment-induced motor recovery in chronic stroke patients with no spontaneous motor improvement in the last 3 months who were treated with a specific motor rehabilitation program. We used complementary methods which give the opportunity to interpret cortical reorganization from different perspectives. fMRI and paired pulse stimulation with transcranial magnetic stimulation (TMS) were applied to the same individual subjects during the process of motor improvement. Since reorganizational changes in the acute phase can be

F. Hamzei et al. / NeuroImage 31 (2006) 710 – 720

influenced by spontaneous recovery, we focussed on chronic stroke patients with no further spontaneous motor improvement. Patients were treated with ‘‘constraint-induced movement therapy’’ (CI). CI is a rehabilitation approach developed through the combined efforts of behavioral psychologists, neuroscientists and rehabilitation teams (Taub et al., 2002; Morris and Taub, 2001). Its idea originated in non-human primate studies. Monkeys did not use the upper extremity after unilateral sensory loss through dorsal rhizotomy (Twitchell, 1954; Knapp et al., 1958). After immobilization of the non-affected arm they learned to reuse the deafferented arm (Knapp et al., 1963; Taub and Berman, 1968; Taub, 1977, 1980). This initial observation was systematically reevaluated and later transferred to rehabilitation in chronic stroke patients (Wolf et al., 1989; Taub and Wolf, 1997; Taub et al., 1993, 1998; Ostendorf and Wolf, 1981). CI involves training of the affected arm, restraining of the non-affected arm (immobilization), shaping techniques and conditional responses. Previous reports have documented CI’s clinical efficacy, in despite of the long delay between brain lesion and onset of rehabilitation (Taub et al., 1993, 1998; Kopp et al., 1999; Kunkel et al., 1999; Van der Lee et al., 1999; Miltner et al., 1999; Dromerick et al., 2000; Dettmers et al., 2005; Rijntjes et al., 2005; Ko¨no¨nen et al., 2005). With the use of TMS, Liepert et al. (1998) showed an enlargement of the ipsilesional cortical hand representation, an extension and shift of the center of the output map in the affected hemisphere after massed practice, e.g., after CI (Liepert et al., 1998, 2000, 2001; Wittenberg et al., 2003). Reduced excitability of the related cortical area has been demonstrated after immobilization of one extremity (Liepert et al., 1995; Wittenberg et al., 2003). Thus, TMS demonstrates use-dependent reorganization in the affected hemisphere. But in a well-defined group with chronic stroke patients treated with this same therapy, divergent results are obtained in imaging studies (Johansen-Berg et al., 2002; Schaechter et al., 2002; Wittenberg et al., 2003; Levy et al., 2001). Wittenberg’s (2003) study described with the use of PET a decrease of ipsilesional primary sensorimotor cortex (SMC) activation. Johansen-Berg’s (2002) fMRI study documented an increased activation in the ipsilesional secondary somatosensory cortex, bilateral cerebellum and ipsilesional dorsal premotor cortex which was associated with improved hand function in a group analysis of seven patients. In Schaechter’s (2002) fMRI study – a group analysis of four patients – showed a shift in laterality of SMC activation toward the unaffected hemisphere after CI. Levy et al. (2001) used fMRI in two patients and found increased perilesional activation in both patients and increased bilateral SMC activation in one patient. These divergent findings suggest, e.g., an interindividual variability or that different findings reflect different reorganization mechanisms in different patients. Here, we focussed on individual

711

subjects, and related fMRI activation changes (increase or decrease) within ipsilesional primary motor cortex to TMS measurements of intracortical inhibition and facilitation in training induced improvement of altered hand motor function.

Methods Patients 29 patients were screened (their data are not presented). Six patients (one female and five males mean age: 70.3 years; range: 63 – 80 years) completed both fMRI and TMS investigation. They were post stroke at least 1.5 years prior to this study. Table 1 shows the patients’ data and their infarct localization. All patients suffered from an ischemic lesion. All patients were right-handed. Inclusion criteria consisted of at least active 20- extension of the affected wrist and 10- of each finger; onset of stroke more than 1 year before starting CI; a maximum score of 3.0 on the Motor Activity Log (MAL) (to exclude patients whose motor functioning was too close to a ceiling for them to benefit from the therapy); ability to walk without balance problems while wearing the constraint device; no hemodynamically relevant intra- or extracranial artery stenosis in Doppler ultrasound, which may alter the BOLD (‘‘blood oxygenation level dependent’’) response (Hamzei et al., 2003a); no cognitive impairments or aphasia that could compromise the questionnaire’s comprehension or attention deficits (i.e., visual neglect); no serious uncontrolled medical problems; no pace makers or epilepsy; no spasticity (Ashworth scale = 0) and no motor improvement in the last 3 months before starting CI (see below). All patients had received extensive physical therapy, both in the acute and post-acute phase. Behavioral investigation At least 3 months prior to the study, subjects underwent a physical examination to determine their eligibility for the experiment. If subjects met the inclusion criteria, they received an explanation of project procedures and signed informed consent. Motor function tests were applied which have been previously used in clinical CI studies (Miltner et al., 1999; Taub et al., 1993; Wolf et al., 1989; Dettmers et al., 2005; Rijntjes et al., 2005). A Motor Activity Log (MAL) interview was carried out to determine the Amount of Use (MAL-AoU) and Quality of Movement (MALQoM) of the affected arm as rated by the patient (Taub et al., 1993). The physiotherapist performed the Wolf Motor Function Test (WMFT) at this time (Wolf et al., 1989; Taub et al., 1993). Before

Table 1 Pat

Age

Interval

Localization of stroke

Stroke side

‘‘Hand knob’’/fibers lesion

MEP

#1 #2 #3 #4 #5 #6

71 70 71 67 80 63

4 6 5 10 4 1.5

Pons MCA, cortical MCA, cortical MCA, cortical Internal capsule Internal capsule

Left Left Left Left Left Left

No No No Yes Yes Yes

Normal Normal Normal Disturbed Disturbed Disturbed

Patient data: age; interval between stroke and begin of CI in years; localization of stroke and side (ischemic lesion in the middle cerebral artery = MCA or internal capsule); stroke lesions affected the hand area of the primary motor cortex (‘‘hand knob’’) or outflow fibers within the middle third of the posterior limb of the internal capsule (‘‘Yes’’) or spared (‘‘No’’); motor evoked potentials of the affected hand (MEP).

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F. Hamzei et al. / NeuroImage 31 (2006) 710 – 720

and after CI the following tests were performed: MAL with QoM and AoU and MFT Wolf Motor Function Test Functional Ability (WMFT-FA), and number of seconds needed for these tests (WMFT-sec), measured with a stop-watch. For the WMFT-sec, the average numbers of seconds for the subtests were calculated. For the WMFT-FA, video sequences were recorded and presented to a 2nd physiotherapist for evaluation who was blinded to the time point of recording (i.e., whether the recordings were made before or after therapy CI) as described previously (Dettmers et al., 2005; Rijntjes et al., 2005). In order to test clinical efficacy, we compared MAL-QoM, MAL-AoU and WMFT (WMFT-FA, WMFT-sec) using nonparametric Wilcoxon test before (‘‘pre’’) and after (‘‘post’’) CI. Threshold was set at P < 0.05. Training The aim of training was to force the patient to use the affected hand. Therefore, the non-affected hand was placed in a splint for 90% of waking hours during training period. Treatment was focussed on housekeeping activities (e.g., eating, opening and closing jars and spring-loaded clothespins). Patients received intensive daily motor activities training for at least 6 h a day under physiotherapeutic supervision. Transcranial magnetic stimulation TMS was performed with a circular coil (Dantec, Germany) to investigate central motor conduction times (CMCT) and maximum motor evoked potentials (MEP) amplitudes in all subjects. Electrical stimulation of the median nerve was applied to measure M response latencies, F wave latencies and M response amplitudes. Recordings were obtained with surface electrodes from abductor pollicis brevis muscle on both sides. CMCT was calculated by: total latency minus (M-response latency plus F-wave latency minus 1)/2. The maximal MEP amplitude was expressed as percentage of the maximal M response amplitude. TMS was performed with a stimulator intensity 25% above the individual motor threshold during tonic preinnervation of the target muscle. Recordings were stored on a Viking IV (Nicolet, Kleinostheim, Germany) and analyzed off-line. According to normal values from our laboratory the upper normal limit of CMCT was 8.5 ms, and the lowest normal MEP amplitude was 15% of M response amplitude. Paired pulse stimulation was performed in all subjects to investigate intracortical inhibition (ICI) and intracortical facilitation (ICF) (Kujirai et al., 1993). TMS was applied with a figure-of-eight coil (The Magstim Comp., Dyfed, UK), which was connected to a Bistim device, which triggered 2 magnetic stimulators (Magstim 200 HP). The interstimulus intervals (ISI) were tested at 2, 3, 10 and 15 ms. The stimulus intensity of the first, conditioning shock was 75% of motor threshold (MT) at rest, the intensity of the second pulse was set at 120% of MT at rest. The coil was held with the grip pointing posteriorly and perpendicular to the central sulcus. For each ISI, 8 stimuli were applied in a randomized order. Recordings were stored on a Viking IV (Nicolet, Kleinostheim, Germany) and analyzed off-line. MEP amplitudes, M responses and F wave amplitudes were measured peak-to-peak. MT was defined as the stimulus intensity that produced MEPs of 50 – 100 AV in 5 out of 10 trials. ICI consisted of MEP amplitudes produced by TMS with 2 and 3 ms ISI for ICF results from ISI of 10 and 15 ms were combined. The values were expressed as a percentage of the mean MEP amplitude after single TMS. This experiment was performed

during complete muscle relaxation. The absence of muscle activity in the target muscle was controlled auditory by a loudspeaker that was connected to the EMG channel. The TMS investigator was not aware of the fMRI results. Functional MRI The following conditions were investigated the day before starting CI (‘‘pre’’) and the day after CI (‘‘post’’): rest scans with eyes closed served as a low-level baseline condition (REST). Two active conditions included right and left passive wrist joint movements. Passive hand movements have been found to induce almost identical patterns of activation compared to active movements in healthy subjects and stroke patients (Weiller et al., 1996; Lee et al., 1998; Loubinoux et al., 2001; Tombari et al., 2004). Its activation pattern is reliable over a time period of several weeks to months (Loubinoux et al., 2003; Tombari et al., 2004; Nelles et al., 2001). We chose passive instead of active hand movements because its performance is identical during follow-up and it is independent of individual performance and possible clinical improvements. All conditions were presented in pseudo-randomized order. There were four epochs/cycles, each containing the experimental conditions in alternation with rest conditions (no gap between epochs). Each condition lasted 31 s. Right and left hands were manually moved with an examiner within the scanner room. Dorsal extensions and plantar flexions of the wrist (0 – 50-) were performed three times (with acoustical signals) in 3.1 s, 30 times within each cycle. The amplitude of the passive hand movement was limited by the physiological dorsal extension of the hand (50-) in the absence of spasticity. The forearm was fixated and only a dorsal extension of the hand was performed. Before scanning, subjects underwent passive hand movements in the MR environment to learn to avoid active movements of the hand and movement artefacts of the head. Surface EMG electrodes were positioned on the dorsal interosseus muscle I and on the extensor digitorum communis muscle. Contraction of arm or hand muscles activated an acoustic signal. MRI acquisition was performed when patients were able to relax their arm and hand. Data acquisition BOLD sensitive T2*-weighted functional magnetic resonance images were acquired on a 1.5 T Magnetom VISION whole-body MRI system (Siemens, Erlangen, Germany) equipped with a standard head coil. Contiguous multi-slice echo planar images (TE 60 ms) were obtained in axial orientation. 30 slices (3 mm thickness) were acquired every 3.1 s. Voxel size was 3.28  3.28 mm (64  64 pixel), which gave a total field of view of 210  210 mm. A total of 160 image volumes were acquired in the first experiment over a time period of approximately 9 min. Data processing and statistics All volumes were realigned to the first volume (Friston et al., 1995a). Residual motion effects were eliminated by regress the time course of each voxel on a periodic function of estimated movement parameters. The estimated movement did not exceed 2 mm. A mean image was created using realigned volumes. An individual 3-D T1weighted MRI (1  1  1 mm voxel size) was coregistered to this mean image. This ensured that functional and structural images were spatially aligned. The functional images and the structural T1-

F. Hamzei et al. / NeuroImage 31 (2006) 710 – 720 Table 2 Pat

#1 #2 #3 #4 #5 #6 Mean

MAL-QoM

MAL-AoU

WMFT-sec

WMFT-FA

Pre

Post

Pre

Post

Pre

Post

Pre

Post

2.6 2.5 2.5 2.0 2.8 0.1 2.08

3.5 3.0 2.6 3.0 3.8 2.1 3.0

2.2 2.0 0.8 1.1 2.4 0.3 1.47

3.3 2.7 2.8 2.7 4.0 1.5 2.83

4.1 11.1 5.3 14.4 5.45 12.9 8.88

2.5 6.5 2.8 4.2 2.75 11.9 5.11

2.9 2.5 2.9 2.5 3.8 1.45 2.68

3.4 3.0 3.4 3.0 4.3 2.3 3.23

713

relative to the anterior commissure, according to the template used for spatial normalization (Evans et al., 1994). We were interested in comparing hand movement (affected and non affected hand) before and after training (‘‘pre’’ versus ‘‘post’’ and ‘‘post’’ versus ‘‘pre’’ comparisons) in individual subjects. BOLD signal intensity during passive movement of the affected and unaffected hand was calculated as session mean. The activation peaks between both investigation days revealed activation changes over time. Activation changes over time were identified within different brain region which were identified as follows: motor cortex as the cortex lying immediately anterior to the central sulcus; somatosensory cortex as the cortex lying immediately posterior to the central sulcus and anterior to the postcentral sulcus; superior parietal lobule as the area lying superior to the intraparietal sulcus and immediately posterior to the postcentral sulcus; inferior parietal lobule as the area lying inferior to the intraparietal sulcus and posterior to the postcentral sulcus; lateral premotor cortex as the cortex lying dorsally to the precentral sulcus; and the supplementary motor cortex as the cortex lying above the cingulate sulcus (Fink et al., 1997; Hamzei et al., 2003b). To test whether BOLD signal changes between both investigation days are contingent on changes on REST condition, we compared REST condition between both investigation days. In the single subject analysis the threshold was set uncorrected for multiple comparisons at P < 0.001.

Motor scores before (‘‘pre’’) and after (‘‘post’’) CI. Mean of the Motor Activity Log (MAL) for Quality of Movement (MAL-QoM) and Amount of Use (MAL-AoU) and Wolf Motor Function Test Functional Ability (WMFT-FA), and numbers of seconds needed for these tests (WMFT-sec) are presented before and after CI.

volume were spatially normalized (Friston et al., 1995b) to templates in a space defined by a template from the Montreal Neurological Institute (Evans et al., 1994) using 12 affine parameters and a set of non-linear basis functions. Since normalization in patients with large lesions might lead to incorrect normalization, we made a mask of the lesion and included it in this procedure. Functional data were smoothed using a 9 mm FWHM (full-width at half maximum) filter in individual subject analyses to compensate for residual variability after spatial normalization. This also facilitates the application of Gaussian random field theory to provide for corrected statistical inference (Friston et al., 1995b). Data analysis was performed by modeling the different conditions as reference waveforms (i.e., box-car functions convolved with a hemodynamic response function) in the context of the general linear model as employed by SPM99 (Friston et al., 1995b, 1997). A high-pass filter was applied to the data to minimize effects of aliased physiological noise. We tested for significant effects using voxelwise t statistics assembled into a statistical parametric map. Co-ordinates are reported in millimeters,

Results Behavioral investigation The motor hand function improved in all subjects after CI. Amount of daily use (MAL-AoU; P < 0.018), Quality of movement (MAL-QoM; P < 0.018) and Wolf Motor Function Test Functional Ability (WMFT-FA; P < 0.031), and number of seconds needed for

Table 3 BOLD Signal intensity within primary sensorimotor cortex Pat

‘‘pre’’ vs. ‘‘post’’ CI x

#1 #2 #3 #4 #5 #6

‘‘post’’ vs. ‘‘pre’’ CI y

18 27 42

21 30 15

z

T

72 60 51

4.58 5.45 4.51

x

y

39 36 39

24 9 9

z

T

63 57 63

3.92 4.21 2.88

BOLD Signal intensity within motor association cortices after CI Pat

#1 #2 #3 #4 #5 #6

Affected hemisphere

Non-affected hemisphere

PC

SMA

Decreased Decreased Decreased Decreased

Decreased Increased

PMC

PC

PMC

Increased Decreased

Decreased

Increased

Increased

Increased

Decreased

BOLD signal intensity changes in individual subjects demonstrate decreased activation within affected primary sensorimotor cortex in patients 1, 2 and 3 after CI. Patients 4, 5 and 6 display an increased activation in the affected SMC after CI. Activation changes within the motor association cortices are individually different without a regular manner (parietal cortex = PC; supplementary motor area = SMA; premotor cortex = PMC).

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Fig. 1. fMRI activation of patients 1, 2 and 3 are presented. In these patients, BOLD signal decreased after CI. Stroke localization in patient 3 is presented on different transversal slices (from left to right in higher z-level) of T1-weighted MRI. Decreased BOLD signal intensity (also decreased number of voxels that is not presented here) after CI was paralleled to increased amplitude of intracortical inhibition within affected SMC, presenting a decreased intracortical inhibition after CI. Changes in BOLD signal and amplitude ICI are presented in percent (normalized to the investigation day before starting CI). CS for central sulcus.

F. Hamzei et al. / NeuroImage 31 (2006) 710 – 720

715

Fig. 2. fMRI activation of patients 4, 5 and 6 are presented. These patients showed a BOLD signal increase after CI. Stroke localization in patient 5 is presented on different transversal slices from left to right in a higher z-level presented on a T1-weighted MRI. Increased BOLD signal intensity (also increased number of voxels that is not presented here) was associated with decrease amplitude of intracortical inhibition within affected SMC after CI, which presents an increased intracortical inhibition after CI. Changes in BOLD signal and amplitude ICI are presented in percent (normalized to the investigation day before starting CI). CS for central sulcus.

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these tests (WMFT-sec; P < 0.027) improved significantly after CI (see Table 2).

Table 4 Pat

Functional MRI No patient had to be excluded because of active hand movements during session time or head movement artefacts (the estimated movement did not exceed 2 mm). The comparison ‘‘pre’’ versus ‘‘post’’ and ‘‘post’’ versus ‘‘pre’’ revealed activation changes (increased and decreased) within primary sensorimotor cortex (SMC) and motor association cortices. Activation changes within motor association cortices of the affected and non-affected hemisphere were individually different and showed no general rule (see Table 3) whereas activation changes within SMC revealed two groups of patients: group one (patients 1, 2 and 3) showed a decreased activation within ipsilesional SMC after CI and another group (patients 4, 5 and 6) demonstrated an increased activation in the ipsilesional SMC when comparing both investigation time points (‘‘pre’’ and ‘‘post’’ CI, see Table 3). BOLD signal decreased significantly after CI ( P < 0.02) in patients 1, 2 and 3. Patients 4, 5 and 6 demonstrated a significant increase in BOLD signal intensity within ipsilesional SMC after CI ( P < 0.015) (Figs. 1 and 2). Individuals with decreased SMC activation after CI showed reduced number of voxels within ipsilesional SMC after CI. This demonstrates not only decreased activation peaks, but also a reduction in the extent of activation. Individuals with increased SMC activation after CI demonstrated increased number of voxels within ipsilesional SMC after CI. Movement of the non-affected hand of all patients showed a decreased SMC activation in the non-affected SMC after CI, probably due to the immobilization of this hand during the training period. There were no differences of REST conditions between both investigations days within the motor network. Attribution of functional activation to ICI and ICF and stroke localization Patients (patients 4, 5 and 6) with increased SMC activation (BOLD signal and number of voxels) after CI suffered from a lesion of M1 (‘‘hand knob’’) or of the outgoing fibers from the hand area of M1 within the internal capsule according to Fries et al. (1993), who have shown that fibers from primary motor cortex pass through the middle third of the posterior limb of the internal capsule. This group of patients showed delayed MEPs and reduced amplitudes recorded from the affected abductor pollicis brevis muscle (CMCT of patient 4: 11.8 ms, patient 5: 11.0 ms and patient 6: 11.2 ms; see Table 4). They also increased ICI and decreased ICF of the affected SMC after CI. In the group of patients with increased BOLD signal, percent amplitude ICI significantly decreased ( P < 0.03). Percent amplitude of ICF was also significantly different between pre and post CI in this group of patients (patients 4, 5 and 6; P < 0.03). Patients (patients 1, 2 and 3) with decreased ipsilesional SMC activation in functional MRI (BOLD signal and number of voxels) showed a lesion which spared the ‘‘hand knob’’ or corresponding descending fibers within the internal capsule. These patients demonstrated normal MEPs of the abductor pollicis brevis muscle of the affected hand (CMCT of patient 1: 5.2 ms, patient 2: 6.4 ms and patient 3: 5.8 ms; see Table 4). The

#1 #2 #3 #4 #5 #6

Single pulse response pre CI

post CI

0.289 0.069 0.285 0.337 0.193 0.280

0.374 0.081 0.419 0.242 0.135 0.260

M-response

F-wave

CMCT

4.0 3.9 4.1 4.0 4.1 3.8

27.6 33.8 30.2 33.4 29.2 27.8

5.2 6.4 5.8 11.8 11.0 11.2

Single pulse response are presented before (pre) and after (post) CI. Mresponse, F-waves and MEP (in milliseconds) were measured from the affected abductor pollicis brevis muscle of individual subject to calculate CMCT. Single pulse response was not significantly different between both investigation days.

paired pulse stimulation revealed a decreased ICI and an increased facilitation in the affected SMC after CI. Percent amplitude ICI in this group of patients with decreased BOLD signal intensity (patients 1, 2 and 3) showed a significant increase after CI ( P < 0.014). Percent amplitude of ICF showed a trend between pre and post CI ( P < 0.078). BOLD Signal intensity differences between both investigation days (‘‘pre’’ and ‘‘post’’) showed a close correlation to ICI differences (r = 0.93; P < 0.002) and ICF differences (r = 0.76; P < 0.042) in the affected SMC between ‘‘pre’’ and ‘‘post’’ CI. Difference in the number of voxels within SMC between ‘‘pre’’ and ‘‘post’’ CI revealed also a close correlation to ICI differences (r = 0.92; P < 0.003) (Fig. 3).

Discussion Single subject analysis revealed two groups of fMRI activation pattern: increased ipsilesional SMC activation in patients (4, 5 and 6) with affected M1 and disturbed MEP which was closely correlated to a decreased intracortical excitability. Patients with intact M1 and MEP showed a decreased SMC activation associated with an increased intracortical excitability. Decreased activation in the SMC is not the result of habituation, since all subjects in this group showed an increased activation in the association cortices (Table 3). Imaging signals (rCBF in PET and BOLD in fMRI) have been shown to be relatively stable over time when identical active and passive hand movements are repeated within a time interval of weeks to months (Loubinoux et al., 2003; Tombari et al., 2004; Nelles et al., 2001). We also excluded BOLD signal changes due to differences in activation pattern in rest time between both investigation days. Thus, activation changes over time should represent ongoing changes of brain organization and thus plasticity. We hypothesize two different cortical reorganization patterns in our chronic patients with motor improvement induced by CI. In case of intact M1 (‘‘hand knob’’) or descending fibers (intact MEP) we suggest the following mechanisms of reorganization schema: decreased BOLD signal may reflect a decreased neuronal activity (or decreased synaptic input) (Logothetis et al., 2001). In conjunction with improved performance, decreased neuronal activity may be the effect of improved intersynaptic communication which is induced by increased intracortical excitability (probably as an effect of decreased g-aminobutryic acid – GABA-activity; Ziemann et al., 1996a, 1996b). This effect might be a consequence of an increased effective synaptic interaction or an increase in

F. Hamzei et al. / NeuroImage 31 (2006) 710 – 720

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Fig. 3. On top, the correlation is demonstrated between BOLD signal intensity difference (arbitrary units) and difference of ICI (measured by paired pulse stimulation, in percent) in the affected SMC between ‘‘pre’’ and ‘‘post’’ CI (r = 0.93; P < 0.002). Difference in number of voxels between both investigations days were also close correlated to difference of ICI (in percent) between ‘‘pre’’ and ‘‘post’’ CI (r = 0.92; P < 0.003).

effective connectivity within SMC at a synaptic level after massed practice (Bu¨chel et al., 1999). The main effect in all forms of CI appears to be the behavioral change, to massed practice for many hours a day. The effect of CI may be partially related to the greater intensity of training of the affected arm compared to conventional physiotherapy (Van der Lee, 2001; Taub et al., 1999). CI applies massed practice to obtain an increase in the use of the more impaired limb and thereby induces a use-dependent functional reorganization. Repetitive arm, thumb and leg training has been shown to improve clinical outcome and induce cortical reorgani-

zation (Classen et al., 1998; Hesse et al., 1995; Nelles et al., 2001; Liepert et al., 1998, 2000, 2001; Liepert and Weiller, 1999). Patients with affected M1 or outgoing fibers showed a different cortical reorganization pattern: daily motor training improved hand function, which may have induced an expansion of the synaptic interaction, because M1 is affected and an improved intersynaptic communication (or increased synaptic efficiency) within M1 is not possible (MEP disturbed from the affected hand). This effort leads to an extension of neuronal activity (increased BOLD signal) in the presence of decreased intracortical excitability.

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In normal subjects it was demonstrated that the optimal coil position for stimulation of the hand motor cortex area and the fMRI activation maximum for finger movements are anatomically very close together (Krings et al., 2001). One should bear in mind that intracortical excitability is determined on one measurement point. It was tested with the coil positioned over the most excitable spot. Changes of intracortical excitability next to this spot should be clarified in further studies. However, improved hand function seems to be possible with either increasing or decreasing SMC activation in chronic stroke patients after motor training. The activation pattern depended on lesion localization which affected intracortical excitability. Further investigation of long term patients’ outcome could be of interest to find out whether activation parameters can have a predictive value for persisting motor improvement. It should be emphasized that we did not investigate the role of the interhemispheric inhibition in our patients. Previously it has been shown that ICI was significantly reduced if a conditioning TMS pulse was applied to the opposite hemisphere in healthy subjects (Daskalakis et al., 2002). After large hemispheric strokes intracortical inhibition in the unaffected hemisphere was also reduced, indicating disinhibition (Liepert et al., 2000a,b; Niehaus et al., 2003). One possible mechanism influencing results in our study is an increase in interhemispheric inhibition from intact M1 to the affected M1 in stroke patients, particularly those with more impairment (Murase et al., 2004). Such mechanism could provide information on a possible role of the increased ipsilateral activation in less recovered individuals. Upper limb immobilization can induce an increase of excitability in the motor cortex contralateral to the immobilized limb (Zanette et al., 2004). However, these authors did not find an effect of immobilization on M1 excitability in the ipsilateral hemisphere. Thus, it seems rather unlikely that immobilization of the non-affected hand modulates excitability in the affected hemisphere of our patients. However, this also should be addressed in future investigations. These data for chronic stroke match the observation made in subacute stroke patients, recently described by Feydy et al. (2002). They identified different activation patterns relative to stroke location: decreased activation (‘‘focussing’’) in the ipsilesional SMC was demonstrated in follow up investigations in case of intact primary motor cortex (10 patients), whereas patients with M1 lesion (4 patients) demonstrated a ‘‘recruitment activation pattern’’ involving bilateral SMC, premotor cortex and SMA in a longitudinal fMRI study (1 to 2 months, 2 to 4 months and 3 to 6 months respectively after onset of stroke). However, one should consider that the limited number of patients in the present study might explain missing correlation to clinical differences between two groups with different types of activation and facilitation behavior after rehabilitation programs. The predictive value of intact MEP for functional outcome of stroke patients after months has been previously shown (e.g., Pe´re´on et al., 1995). Patients with a lesion affecting the ‘‘hand area’’ of M1 or its main outflow tract are generally more affected than other stroke patients and may have poor recovery. Still, our findings are supported by the close correlation between fMRI parameters (BOLD signal and number of voxels) and ICI/ICF.

Acknowledgments We are grateful to P.T. Alleyne-Dettmers, PhD for editing the text. We thank the physiotherapist, Heike Kru¨ger for assuming the

responsibility to train the patients, C. Bu¨chel for his comments during preparation of the study’s design and early part of the manuscript and Thomas Wolbers for his statistical support. We are grateful to all individuals who participated in this study. The German Research Foundation ("Deutsche Forschungsgemeinschaft, DFG; Innovationskolleg (Nr. INK 22/B1-1) ‘‘Bewegungssysteme’’, Project D1) funded this project. CW was supported by DFG, BMBF (GFGO 0 123 7301-01GO 0105; DFG: WE 1352/ 13-1) and EU (QLK6 CT 1999 02140). JL and CW were supported by the Competence network stroke (01GI9917).

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