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Keywords:

  • Tuberous sclerosis complex;
  • Epilepsy;
  • EEG-fMRI;
  • Spikes;
  • Irritative zone;
  • Children

Summary

  1. Top of page
  2. Materials and Methods
  3. Results
  4. Discussion
  5. Conclusion
  6. Acknowledgments
  7. References
  8. Supporting Information

Purpose: Ninety percent of patients with tuberous sclerosis complex (TSC) have epilepsy. Identification of epileptogenic areas can be difficult and studies are needed to characterize the epileptogenic network in more detail.

Methods: Five children with TSC and focal epilepsy were studied using simultaneous EEG and functional MRI recordings. Tubers were marked by a neuroradiologist on the anatomical MRI. Spike-associated BOLD (blood oxygenation level-dependent) responses were superimposed with lesions.

Results: Thirteen different types of interictal epileptiform discharges (IED) were analyzed with 12 showing a BOLD response, all involving more than one tuber.

Five studies had tubers with activations exclusively within the lesion, three studies had lesional activations extending to perilesional areas, and two studies had activations involving exclusively perilesional areas of at least one tuber. Deactivations exclusively within a tuber were found in six studies, lesional deactivations extending to perilesional areas were found in four studies, and tubers with exclusively perilesional deactivations were found in five studies.

A BOLD response was found in at least one tuber in the lobe of IED generation and presumed seizure onset (according to telemetry) in all patients. In four patients, the same tubers were involved following different IED localizations. The observed changes were always multifocal, sometimes involving tubers distant from the IED field.

Discussion: These findings suggest extended epileptogenic networks in patients with TSC, which exceed networks described in PET and SPECT studies. It was possible to identify specific interictally active tubers. EEG-fMRI provides a noninvasive method to select tubers and areas at their borders for further presurgical investigations.

Tuberous Sclerosis (TS) is a multisystem disorder that is inherited as an autosomal dominant trait and occurs in 1 in 6,000–10,000 live births (Osborne et al., 1991). Mutations in the TSC1 and TSC2 genes lead to abnormal tissue growth and differentiation affecting the brain, eyes, heart, kidneys, and skin (Crino et al., 2006). Findings in the central nervous system consist of cortical tubers, subependymal nodules, and subependymal giant-cell astrocytomas (Christophe et al., 2000). Epilepsy is found in about 90% of all TSC patients and tubers are believed to be epileptogenic. Seizures start in early childhood in most of the cases; complex partial seizures and infantile spasms are the most common seizure types (Guerreiro et al., 1998). Medically refractory seizures are closely linked with mental retardation and behavioral abnormalities (Jozwiak et al., 1998). Despite new anticonvulsants, the proportion of children with medication-resistant epilepsy remains high. In these cases, resection of epileptogenic tubers is the only potential treatment for their seizures (Fujita et al., 1997). In a great number of cases, however, it is difficult to define a target for surgical intervention. Even with new functional neuroimaging techniques and intracranial recordings, it remains challenging to distinguish epileptogenic from nonepileptogenic tubers (Chugani et al., 1998). On one hand, seizures start independently from different tubers in many patients (Bauman et al., 2005). In other patients, more than one tuber is involved in the epileptogenesis (Harvey et al., 2004). On the other hand, a widespread epileptogenicity that is not limited to the structural abnormalities seen on MRI may be responsible for difficulties in finding a candidate region for surgery. Functional neuroimaging studies provided evidence that epileptogenicity is not restricted to cortical tubers but can also affect functionally associated areas (Perreson et al., 1998; Asano et al., 2000). Intracranial EEG recordings revealed that, in particular, the tissue around and at the border of the tuber might be highly epileptogenic (Otsubo et al., 2005).

Simultaneous EEG-fMRI is a noninvasive tool to evaluate epileptogenic networks in the brain. This method allows identification of areas with Blood Oxygenation Level Dependent (BOLD) signal changes correlated with the interictal epileptic discharges (IED). Positive BOLD responses as well as negative BOLD responses therefore can delineate the irritative zone (Gotman et al., 2006). Improvement in data acquisition, using high-field scanners (3T), short recording times, and good artifact correction made it possible to obtain results with a sufficient quality using this technique in sedated children (Jacobs et al., 2007).

In the present study, we evaluated for the first time children with TSC using EEG-fMRI. We hypothesize that the underlying epileptogenic networks in patients with TSC is more wide spread than the tubers delineated on MRI, and that this network can be identified through the BOLD responses at the time of IEDs.

Materials and Methods

  1. Top of page
  2. Materials and Methods
  3. Results
  4. Discussion
  5. Conclusion
  6. Acknowledgments
  7. References
  8. Supporting Information

Five patients with focal epilepsy and diagnosis of TSC (one female, four males, mean age 5.2 ± 5.1) were recruited in the Department of Neuropediatrics of the University of Kiel, Germany and the Northern German Epilepsy Centre, Raisdorf. In all patients, the diagnosis of TSC was established following the current clinical diagnostic criteria for TSC (Roach et al., 1998). Clinical work-up included video-EEG monitoring, echocardiogram, renal ultrasound, ophthalmologic, and dermatologic examination. In addition, in all except one patient, the diagnosis was confirmed by molecular genetics studies (mutations in the TSC1 and TSC2 genes) at the time of the study.

All patients had medically refractory epilepsy; in all but one the epilepsy started before the second year of life. At the time of examination, all but one patient received two medications to control their seizures, but remained with daily seizures. Only patient 1 was seizure free with sulthiame monotherapy for two months at the time of the examination, but started to have seizures again in the clinical follow-up. A clear lateralization of the seizure onset could be seen in only two patients in long-term video EEG monitoring (patients 4 and 5). Seizures were classified as different types if they showed distinguishable EEG onset in regard to topography, pattern and clinical pattern. More than one seizure pattern could be clearly identified in three patients: patient 3, with 3 seizure types and patients 1 and 2, with 2 seizure types. On routine sleep EEG, multifocal interictal epileptiform discharges were seen in all patients. The clinical details and MRI findings are shown in Table 1.

Table 1.  Clinical information
PatientAgeEpilepsy onsetSeizure typeSeizure onset on EEGAEDGenetic diagnosisNeurologic involvementCardiacRenalDermatologicMRI findings
TuSNGCA
  1. This table provides information on the clinical aspects of the patient's epilepsy and the major and minor features of tuberous sclerosis complex found in the patient during clinical evaluation (Roach et al., 1998). On the right part, it presents the MRI changes of all patients as identified by the neuroradiologist.

  2. AED, antiepileptic drugs; bi, bilateral; C, central; CPS, complex partial seizures; F, frontal; GCA: subependymal giant cell astrocytoma; L, left; LTG, lamotrigine; m, months; O, occipital; OXC, oxcarbazepine; P, parietal; R, right; SN, subependymal nodule; SUL, sultiame; T, temporal; TSC, tuberous sclerosis complex; Tu, tuber; VGB, vigabatrin; VPA, valproic acid; y, years.

116 mCPSF R P LSULTSC 2Mild developmental delay1Hypomelanotic nodules2091
242 yCPS Atypical absences Hypermotor seizures (nocturnal)F L T RLTG, SULTSC 1Developmental delay1Hypomelanotic nodules133
326 mCPS Atypical absencesF bi CP R O ROXC, VGBTSC 2Mild developmental delayHypomelanotic nodules, facial angiofibroma1961
4146 yCPS Hypermotor seizures (nocturnal)F RLTG, OXCn.a.Behavioral disorderHypomelanotic nodules, facial angiofibroma51
554 mInfantile spasms, CPS, Myoclonic jerksFP RVPA, VGBTSC 2Severe developmental delay, autistic disorder, left hemipareses11Hypomelanotic nodules, facial angiofibroma, harmar toma186

Informed consent was obtained from the legal guardians of all patients. The study was carried out according to the Declaration of Helsinki and was approved by the Ethics Committee of the University of Kiel, Schleswig, Holstein.

Data acquisition

EEG-fMRI data were only acquired in children who had a clinical indication for a brain MRI, either for presurgical evaluation or as a follow-up examination. The anatomical MRI was acquired in the same scanning session and no additional sedation to that applied for the clinical MRI was needed.

All patients required sedation, as their age and mental impairment prevented them from remaining quiet in the scanner. All children had been sedated for EEG sleep studies before and their sedative medication for the EEG-fMRI session was chosen according to the tolerance of the individual patient. Thus, patients were sedated with either chloral hydrate (Chloralhydrat; average dose approx. 50 mg/kg) or chlorprotixen (Truxal; average dose approx. 20 mg/kg). During the scan, a pediatrician was present and vital parameters were monitored using MRI compatible machine.

The EEG was continuously recorded inside the MRI scanner (3-Tesla Philips Achieva, 8-channel SENSE head coil, Philips Medical Systems, Best, The Netherlands) from 30 scalp sites (10–20 system plus FC1, FC2, CP1, CP2, FC5, FC6, CP5, CP6, TP9, TP10) with a reference located between Fz and Cz. Sintered Ag/AgCl ring electrodes were attached using a “EasyCap” (Falk-Minow Services, Herrsching-Breitbrunn, Germany), which is part of the MR-compatible EEG recording system “BrainAmp-MR” (Brainproducts Co., Munich, Germany). Electrode impedance was kept below 7 kOhm. Two additional electrodes were placed on the infraorbital ridge of the right eye for recordings of the vertical EOG and on the left perivertebral region for electrocardiogram (ECG) recording. Data were transmitted from the amplifier (5 kHz sampling rate, 250 Hz low-pass, and 0.03 Hz high-pass filters) via an optic fiber cable to a computer located outside the scanner room.

All patients had the following clinical anatomical acquisitions: T2 transversal, coronal, sagittal (FOV = 200 mm, Matrix 400 × 400, 36 slices, 3-mm slice thickness, TR = 4,352 ms, TE = 100 ms, flip angle 90°), T2 FLAIR coronal (FOV = 208 mm, Matrix 208 × 208, 50 slices, 3-mm slice thickness, TR = 12,000 ms, TE = 160 ms, °, Sense factor 2). In addition, a 3D-T1-weighted anatomical acquisition (FOV = 224 mm, Matrix 224 × 224, 150 slices, 1-mm slice thickness, TR = 8.4 ms, TE = 3.6 ms, Sense factor 1.3) was performed and later coregistered with the functional images. At the end of each clinical examination, continuous BOLD fMRI data were acquired for 20 min (T2*-weighted single-shot EPI sequence, FOV = 200 mm, 30 slices, Matrix 64 × 64, 3.5-mm slice thickness, TR = 2,250 ms, TE = 45 ms, flip angle = 90°, 540 dynamics).

EEG processing

EEGs were filtered offline using Brain Vision Analyzer (Brain Products) and BESA (Brain Electrical Source Analysis, Megis Software GmbH, Gräfelfing, Germany). Gradient artifacts were corrected using an averaged artifact subtraction method (Allen et al., 2000). This was followed by pulse artifact removal with an Independent Component Analysis (Srivastava et al., 2005) and spatial filters based on artifact and brain signal topography (Siniatchkin et al., 2007a).

Two experienced neurophysiologists reviewed the filtered EEG to mark the IEDs and spike-wave-bursts. These were classified into distinct IED types according to spatial distribution and morphology for each patient if more than one type was present. Semiautomatic detection of the manually selected IEDs was then performed offline in BESA using a spatiotemporal pattern search and all marked IEDs were visually reviewed. No IED types were excluded.

fMRI processing

The fMRI images were motion corrected and smoothed (Gaussian kernel, FWHM = 8 mm) using SPM 2 software (Statistical parametric mapping, http://www.fil.ion.ucl.ac.uk/spm/). fMRIstat software was used for further statistical analysis (Worsley et al., 2002). Each IED that was noted in the EEG was included in the analysis as an event, with IEDs differentiated into different event types based on their spatial distribution. One general linear model was constructed in which each IED type was entered as a separate contrast of the same analysis. In the case of events occurring in close temporal proximity, the model HRFs were assumed to summate linearly. Thus, a close succession of IEDs would be modeled as a larger and longer BOLD change than an individual IED. We performed four separate analyses each using a different model HRF. Models consisted of a single GAMMA function with a FWHM of 5.4 s, with peaks 3, 5, 7, or 9 s after the event, resulting in four separate statistical maps. A single combined statistical map was then created by taking the highest absolute value of each voxel. This allowed some variation in the latency of the BOLD response (both between patients and within regions of each patient) while retaining information about its expected shape (Bagshaw et al., 2004). Significant responses were defined as 10 or more contiguous voxels with t > 3.1 (p = 0.05, corrected for four analyses). Significant positive and negative BOLD responses were visualized using Anatomist (http://brainvisa.info/).

Evaluation considering the lesion and EEG localization

An experienced neuroradiologist (AR) reviewed all anatomical MRIs and highlighted the contours of all tubers, subependymal nodules, and giant cell astrocytomas on the T1 images. The fMRI results were coregistered to and displayed on these images. We correlated the different lesions with the BOLD responses in each patient, looking at lesional as well as perilesional BOLD changes. A BOLD change was considered perilesional if it extended past the border highlighted by the neuroradiologist. As a second parameter, the concordance between BOLD responses and the EEG focus was examined. A BOLD response was considered to be within the EEG focus if it was in the brain area considered to generate such IED topography on the surface EEG, as determined by BESA through averaged voltage maps of the IEDs.

Results

  1. Top of page
  2. Materials and Methods
  3. Results
  4. Discussion
  5. Conclusion
  6. Acknowledgments
  7. References
  8. Supporting Information

fMRI results

Patient 4 was scanned twice. All patients showed IEDs and different IED types could be identified in four (patients 2, 3 and 4 with 2 and patient 5 with 4). This resulted in a total number of 13 different fMRI studies. BOLD responses were observed in all but one study (92.3%). Ten studies showed positive as well as negative BOLD responses (77%) and two studies only showed negative BOLD responses (15.4%) (Table 2). In the following, the relationship of BOLD responses to structural abnormalities, IED topography and seizure onset will be described. All patients also showed additional smaller distant negative and positive BOLD responses, which were neither lying in the lesions visible on MRI nor in the perilesional areas but far away from any lesions.

Table 2.  BOLD responses and their correlation with tubers and EEG-focus for each study.
PatientStudy #Spike localizationSpike #ActivationDeactivation
LesionalPerilesionalEEG-FocusLesionalPerilesionalEEG-Focus
  1. This table displays the relationship between activations and deactivations and cortical abnormalities. Localizations of the lesions, in which the lesional or perilesional BOLD response were seen, are provided. Besides this, localization of activations and deactivations that correspond with the presumed spike field.

  2. Those BOLD localizations with the highest t-value for activations and deactivations are printed in bold letters.

  3. Bi, bilateral; C, central; F, frontal; GCA, giant cell astrocytoma; L, left; No., number; O, occipital; P, parietal; R, right; T, temporal; Tu, tuber.

1 1T/O L44Tu P LTu P LO biTu F R Tu F LTU F R
2 2T R49Tu F RT RTu F LTu F L
 3O L34Tu F LO L
3 4C/P L27Tu F/P L
 5T/P R14Tu P R Tu F R GCA RTu F RP RTu O R Tu P RTu O R Tu F L
4a 6FT R114 Tu T RTu T RFT RTu F R Tu F L
 7FT L36Tu T RT L (s)—-—-—-
4b 8FT R82Tu T R FT RTu F R
 9F R6
510PR123 Tu ORTu P RP R
11FR626 FRTu P RTu P R
12TR32Tu O L
13CR23Tu F LTu O LCP RTu O RTu F R

Correlation between BOLD responses and tubers

Patient 1

In patient 1, 20 tubers could be identified. BOLD responses were limited to three (Fig. 1). A positive response was observed over the lesional and perilesional area of a left parietal tuber, while negative responses were seen over a right frontal and left frontal tuber. In the right frontal tuber, perilesional areas were involved as well. The maximum t-value for negative BOLD responses was located in this tuber. In addition to the lesional BOLD responses, bithalamic positive BOLD responses were observed in this patient.

image

Figure 1. This figure shows all positive and negative BOLD responses of a 1-year-old patient (patient 1). A left temporal-occipital focus was seen on the EEG obtained in the scanner. The topography of the spike was visualized using an averaged voltage map of the spikes (BESA software). Bilateral occipital positive BOLD responses are observed, which correlate with the EEG focus, but is not lying within a tuber. Three different tubers are part of the irritative zone of this patient, all lying in brain areas distant from the EEG focus. Strong negative BOLD responses were lying within bilateral frontal tubers. Seizures starting in the right frontal lobe correlate well with these negative BOLD responses.All BOLD changes were visualized using the software Anatomist. Borders of cortical tubers were marked in light green and giant cell astrocytomas as well as subependymal nodules were highlighted with purple outlines. Red arrows mark the positive and negative BOLD responses with the highest t-value of the presented study.

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Patient 2

T2-images showed 13 tubers, two with BOLD responses. Two different types of interictal discharges were observed. The analyses for the first IED type over the right temporal region showed one tuber with a positive BOLD response in the right frontal lobe and another with a negative response in a left frontal lobe (E-Fig. 1). In the latter, perilesional areas were involved as well. The same left frontal tuber showed a negative BOLD response related to the second IED type, which was located over the left occipital lobe. The maximum t-value of the negative response was located within this tuber. No other tuber showed BOLD responses for this second IED type.

Patient 3

The tuber count of patient 3 was 19. Five showed BOLD responses following either of the two IED types of this patient. The first IED type was over the left centroparietal area. Only one tuber located in the left parietal area showed a negative BOLD response following this IED type. Four tubers showed BOLD responses following the second right temporoparietal IED type (Fig. 2). A positive BOLD response was observed in a right parietal tuber, and within and around a right frontal tuber. Additionally, a positive response was observed in the Giant cell astrocytoma located close to the right ventricle. Negative BOLD responses were seen in a right occipital tuber as well as a left frontal tuber. The maximum t-value for positive BOLD responses was located in the right parietal tuber and for negative BOLD responses in the left occipital tuber.

image

Figure 2. This figure displays all positive and negative BOLD responses observed in patient 3 (study no. 5). The maximum t-value for positive responses was seen within a right parietal tuber and the maximum t-value for negative responses within a right occipital tuber. One positive response is observed inside the subependymal giant cell astrocytoma. In total, six different lesions were involved with the epileptogenic network of this patient. BOLD responses of tubers in the right occipital lobe, right parietal lobe, and right frontal lobe following interictal discharges in this patient may be the origin of his seizures, which started over the right occipital, right parietal, and bilateral frontal areas. The results confirm the multifocal interictal discharges and seizure onsets seen in the long-term telemetry of this patient. Visualization was performed as described in Fig. 1.

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Patient 4

Patient 4 was scanned in two independent sessions. Five tubers could be identified, and three showed BOLD responses in one of his spike types. Two IED types could be identified in each scan. During both scans, positive BOLD responses were seen over the right temporal tuber and perilesionally following right temporofrontal IEDs (E-Fig. 2). Perilesional negative BOLD responses over a left and right frontal tuber could be observed, as well. The second IED type was different for the two scans. During the first, left frontotemporal IEDs were seen, which resulted in a positive perilesional BOLD response around the same right temporal tuber that was involved following the right temporal IED. In the second scan, very focal right frontal IEDs were observed, but no corresponding BOLD response was seen. In summary, a right temporal tuber was involved in three of the four studies of this patient, and the maximum t-value for positive BOLD responses was observed inside this tuber. A right frontal tuber showed negative BOLD responses in 2 of the patient's studies.

Patient 5

Eighteen tubers were identified in patient 5. Four studies were analyzed and BOLD responses were found in 6 different tubers. A positive BOLD response in right occipital and a negative one in a right parietal tuber were observed following the first IED type, right parietal (E-Fig. 3). The same right parietal tuber showed a negative BOLD response in the second IED type, right frontal. This time, the perilesional area was involved as well, but no other tuber showed BOLD responses. The third IED type was over the right temporal region and resulted in a negative BOLD response in a left occipital tuber. The fourth IED type, over the right central area, resulted in positive BOLD responses inside a left frontal and around a left occipital tuber; negative BOLD responses were found in a right frontal and right occipital tuber. In summary, the same right parietal tuber showed negative BOLD responses following two different IED types and the maximum negative t-value of the study was inside this tuber in both cases. The maximum t-value for positive responses in the first study was seen in a right occipital tuber that was also involved in study four, although it showed a negative response in the latter case.

Concordance with the EEG topography and seizure onset

Concordance with the topography of the IED was seen as positive in eight (61.5%) and as negative BOLD response in two studies (15.4%). These BOLD responses were those with the maximum t-value in two studies for positive and in one for negative BOLD responses. In five studies, the BOLD responses correlating with the IED topography were also lying within tubers. Different IEDs in one patient seemed to result in BOLD responses within the same tubers independently of where the IED was generated.

Four of our patients had relatively high tuber counts; in one patient three and in three patients two different seizure onset zones were described on long-term Video-EEG monitoring. Because of the high tuber count, we could only correlate whether the tuber was lying in the same lobe as the assumed seizure onset. In each patient, at least one tuber in the assumed lobe of seizure onset showed a BOLD response. In five studies positive (38.5%) and in ten studies negative BOLD responses (77%) were observed in these tubers or perilesional areas.

Discussion

  1. Top of page
  2. Materials and Methods
  3. Results
  4. Discussion
  5. Conclusion
  6. Acknowledgments
  7. References
  8. Supporting Information

In this study, we applied simultaneous EEG-fMRI for the study of epileptogenic networks in a pediatric series of TSC patients. In all patients, multiple, but not all, tubers were involved following the interictal discharges. Sometimes they were located distant to the IED topography, possibly over tissue that was normal on the MRI, suggesting that extended brain areas might be involved during interictal spiking in patients with TSC. In all cases, BOLD responses were limited to parts of the lesions, but extension beyond tuber boundaries and even exclusive involvement of perilesional areas was noted. Different IED types in one patient often lead to involvement of the same tubers.

Our EEG-fMRI study demonstrated a sufficient sensitivity. All but one IED type were associated with BOLD responses. These results correspond with other studies in children (Jacobs et al., 2007; DeTiege et al., 2007). In adult EEG-fMRI studies, positive BOLD responses strongly correlate with the irritative zone, i.e., the EEG-focus or a lesion, while negative BOLD responses are more likely to be observed in distant brain areas (Kobayashi et al., 2005; Salek-Haddadi et al., 2006). In this study, we analyzed both positive and negative BOLD responses and their concordance with the EEG topography and cortical tubers. We could not find different patterns for positive and negative BOLD responses as both were found in lesional areas, areas of IED topography, and distant brain areas in similar amounts. This corresponds with our previously published study, which found positive and negative BOLD responses encompassing parts of the irritative zone equally (Jacobs et al., 2007). This relative increased prevalence of negative BOLD responses in children may be related to sleep, sedation, or age, i.e., conditions that cause changes in baseline activity (Siniatchkin et al., 2007b). There has been evidence that negative BOLD responses may result from different baseline states while still representing the same activity as positive BOLD responses (Shulman et al., 2007). Therefore, in the following part of the article, we will discuss BOLD responses without differentiation between positive and negative.

Correlation between BOLD responses and lesions

There is a need for functional neuroimaging studies in TSC patients to identify epileptogenic brain areas and describe targets for a surgical intervention. PET can sometimes be highly sensitive in distinguishing epileptogenic from nonepileptogenic tubers (Chugani et al., 1998). However, this technique is insufficient and provides inconclusive results in some patients (Sood & Chugani, 2006). Interictal SPECT studies alone show a very low sensitivity in localizing focal epileptogenic areas (Harvey & Berkovic, 1994) but a combination with ictal studies has demonstrated sensitivity as high as 95% (Véra et al., 1999). However, ictal SPECT can be misleading if an insufficient number of seizures is evaluated or if there is a rapid seizure propagation as is often seen in children. EEG-fMRI may represent an alternative to other functional neuroimaging techniques.

All of our patients had multiple tubers and presented with multifocal interictal discharges and seizure patterns, suggesting that more than one tuber could be involved in the epileptogenic process. High tuber counts were found in all but one of our patients, indicating the severity of their disease; patients with a high tuber count are likely to suffer from severe impairment and epilepsy (Goodman et al., 1997). Especially in patients with high tuber counts, the identification of epileptogenic tubers with surface EEG is very difficult due to insufficient spatial resolution. These patients may benefit from combining the temporal resolution of the EEG with the spatial resolution of a high field MRI. A recent study showed that EEG-fMRI may be useful for presurgical evaluations in epilepsy patients (Zijlmans et al., 2007).

Our study demonstrated that BOLD changes were limited to restricted areas of the tuber (often on one side or at the border of the lesion), as has been described before in patients with polymicrogyria (Kobayashi et al., 2005). The presence of BOLD responses in lesional areas may indicate an intrinsic epileptogenicity, as suggested in many other studies (Koh et al., 2002; Kobayashi et al., 2005; Weiner et al., 2006; Jacobs et al., 2007). Additionally, BOLD responses were found in perilesional areas, sometimes exclusively. This agrees with findings of intracranial recordings and MEG dipole analysis (Perreson et al., 1998; Koh et al., 2002). Most likely, the developmental disorganization of cortical layers extends past the borders of the tuber, which can be seen on MRI. This finding is important for the planning of intracranial electrode placement and all efforts should be made to include all lesional borders in the electrode coverage.

Interestingly, one patient showed a positive BOLD response in a subependymal astrocytoma. These tumors are known to cause neurological complications such as the obstruction of CSF pathways and consecutive hydrocephalus (Goh et al., 2002), but normally not regarded as part of the epileptogenic process and their removal was not associated with seizure improvement (Cuccia et al., 2003). However, these tumors derive from stem cells and consist of proliferating neuronal and glial cells (Sharma et al., 2004), therefore groups of undifferentiated or immature neurons may be part of the epileptogenic network.

Correlation between BOLD response and EEG

BOLD responses in tubers were not limited to the lobe corresponding to the topography of the interictal discharge. In 77% of the studies, a BOLD response was seen in one tuber lying in the brain area that corresponded with the IED topography. Additional tubers were found showing BOLD responses in other brain regions that were located distant from the IED. Patient 1, for example, showed bifrontal lesional negative BOLD responses following a left temporo-occipital IED. This phenomenon is not yet understood, but it is very unlikely that this observation is only a result of the limited spatial resolution of the EEG or a propagation phenomenon.

Therefore, it can be hypothesized that an IED may be associated with BOLD changes not only in the lesional areas corresponding with the discharge, but also in distant lesional areas. These additional lesions might be potential irritative zones and contribute to the spread or generation of IEDs. This might also explain why some patients showed involvement of the same tuber following different types of IEDs. In patient 5, for example, the same occipital tuber showed a lesional positive BOLD response following IEDs over the right parietal region and a negative BOLD response following independent IEDs over the right central region.

In our patients, the seizure onset zone was determined following a long-term telemetry study. Tubers lying in the seizure onset zone showed BOLD changes in all patients, mainly negative responses. This suggests that EEG-fMRI could identify the tubers that are mainly involved in the epileptogenic process. However, intracranial recording would still be needed to confirm that the seizure truly originated from the delineated tuber, as sometimes multiple tubers were localized in one lobe. Nevertheless, in cases where multiple tubers are found in an area representing the seizure onset on surface EEG, EEG-fMRI in combination with other functional studies could help determine depth electrode placement in the correct lesion.

Widespread involvement of different brain structures during interictal discharges

Despite observations in patients with polymicrogyria and heterotopia in whom a single lesion could be delineated as epileptogenic, our patients showed involvement of several tubers in each study. This may represent the multifocal character of the patient's disease. In all but patient 4, we found evidence of bilateral changes of brain activity following the IEDs. Moreover, PET and SPECT studies sometimes showed multifocal epileptic activity in several tubers, which were associated with multifocal interictal discharges on surface EEG, poor surgical outcome, and severe cognitive impairment of some patients with TSC (Rintahaka & Chugani, 1997; Chugani et al., 1998). However, changes extending the lesional areas were not observed in these studies. Our results show an even more widespread involvement of brain tissue following the interictal discharges. All of our patients showed severe neurological impairment as a result of TSC. Neurophysiological changes extending the areas of lesional changes have been described previously in patients with TSC (Jambaque et al., 1993). The severe impairment of our patients may either reflect primary structural changes extending beyond the lesions visible on MRI, or secondary functional changes due to the frequent seizures (Palmini et al., 1991, 1995; Jambaque et al., 1993;). Both factors, lesions and early seizure onset, are known to correlate with reduced intellectual function (O'Callaghan et al., 2004). The multifocality of the BOLD responses in different tubers and the occurrence of distant positive and negative BOLD responses might reflect a large involvement of brain tissue during interictal discharges. The impact of interictal discharges on development and cognition is a controversial issue (Binnie & Martson, 1992; Holmes et al., 2006), but frequent multifocal changes of neuronal activity and blood flow, as we observed, could interfere with both. EEG-fMRI studies in a larger group of patients may help better understand these networks, especially since it could provide a noninvasive method to examine patients with different severities of the disease and even asymptomatic carriers of the gene.

Conclusion

  1. Top of page
  2. Materials and Methods
  3. Results
  4. Discussion
  5. Conclusion
  6. Acknowledgments
  7. References
  8. Supporting Information

EEG-fMRI was used to observe changes in neuronal activity following IEDs in children with TS and in all patients a few activated tubers could be identified. In many cases, the same tubers were involved in the network following differently localized IEDs. BOLD responses were limited to parts of the lesion, but extended beyond the tuber border in several cases. This finding may be relevant in clinical practice, such as the placement of depth electrodes. Furthermore, the multifocality of BOLD changes even outside the tuber areas might indicate further involvement of brain tissue regarded as normal on MRI. Our results therefore suggest widespread epileptic networks during interictal spike generation. These networks extend those described in PET and SPECT studies, which often suggest that only one or a few tubers show intrinsic epileptogenicity.

Acknowledgments

  1. Top of page
  2. Materials and Methods
  3. Results
  4. Discussion
  5. Conclusion
  6. Acknowledgments
  7. References
  8. Supporting Information

JJ was supported by a grant of the German Research Foundation (Deutsche Forschungsgemeinschaft (JA 1725/1-1)). This research was supported in part by grant MOP-38079 of the Canadian Institutes of Health Research. We confirm that we have read the Journal's position on issues involved in ethical publication and affirm that this report is consistent with those guidelines. The authors have no professional or financial affiliations that might be perceived as having biased the presentation.

Conflict of interest: The authors report no conflicts of interest.

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  3. Results
  4. Discussion
  5. Conclusion
  6. Acknowledgments
  7. References
  8. Supporting Information
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Supporting Information

  1. Top of page
  2. Materials and Methods
  3. Results
  4. Discussion
  5. Conclusion
  6. Acknowledgments
  7. References
  8. Supporting Information

Figure S1. This figure shows positive and negative BOLD responses in patient 2 (study 2). The strongest positive response is seen over the right temporal regions, which corresponds with the spike marked over the right temporal area T4/T8. The strongest negative response is seen in a left frontal tuber, which showed response to the interictal discharge in both studies in this patient and lies in the area of his presumed seizure onset zone. Visualization was performed as described in Fig. 1.

Figure S2. In patient 4 (study 5), a strong positive BOLD response was observed within a tuber in the right temporal lobe. The patient was scanned twice with similar results. The localization of the EEG spike correlated well with the BOLD responses inside the tuber described. The patient showed two types of seizures: typical sleep related hypermotor frontal seizures starting over the right frontal lobe and complex partial seizures with an EEG onset over the right temporal area. Both tubers with BOLD responses therefore correlate well with the area of seizure onset. Visualization was performed as described in Fig. 1.

Figure S3. The strongest negative BOLD response in study 10 of patient 5 was observed within and at the border of a right parietal tuber, which corresponded well with the map of the right parietal spike. A positive BOLD response was found within a right occipital tuber. The seizure onset in these patients was observed over the right fronto-parietal area. Visualization was performed as described in Fig. 1.

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EPI_1486_sm_FigureS1.tif8611KSupporting info item
EPI_1486_sm_FigureS2.tif8576KSupporting info item
EPI_1486_sm_FigureS3.tif8604KSupporting info item

Please note: Wiley Blackwell is not responsible for the content or functionality of any supporting information supplied by the authors. Any queries (other than missing content) should be directed to the corresponding author for the article.