- Email: [email protected]
fMRI) in focal epilepsy with reference to benzodiazepine effect
Magnetic Resonance Imaging 22 (2004) 1487 – 1492
Hemodynamic response (BOLD/fMRI) in focal epilepsy with reference to benzodiazepine effect Giovanni Battista Riccia, Diego De Carlib,c,d, Claudio Colonnesee, Giancarlo Di Gennaroe, Pier Paolo Quaratoe, Gianpaolo Cantoree, Vincenzo Espositoa,e, Girolamo Garreffab,c,d,e, Bruno Maravigliab,c,d,* a
Department of Neurological Sciences, University of Rome bLa Sapienza,Q 00185 Rome, Italy b Enrico Fermi Center, 00184 Rome, Italy c Department of Physics, University of Rome bLa Sapienza,Q 00185 Rome, Italy d Centro di Ricerca e Sviluppo bSoft,Q 00185 Rome, Italy e Istituto di Ricovero e Cura a Carattere Scientifico bNeuromed,Q Pozzilli, 86077 Isernia, Italy Received 29 October 2004; accepted 29 October 2004
Abstract We studied a new procedure of BOLD/fMRI acquisition in epilepsy. They use the benzodiazepine effect to achieve a more reliable baseline for statistical analysis. The method works only in the MR domain without EEG correlation. It compares the EPI images during interictal epileptic discharges and the images binactivatedQ by benzodiazepine. The results in five out of eight patients show that this procedure in comparison with the EEG/fMRI method gives a net improvement of spatial definition of BOLD areas. These preliminary results seem to confirm the hypothesis that the better BOLD/fMRI procedure in epilepsy is to make use of physical features of MR that, unlike EEG, is not influenced by the distance of intercerebral sources and consequently allows a more complete and undistorted display of BOLD areas. D 2004 Elsevier Inc. All rights reserved. Keywords: Hemodynamic response; BOLD/fMRI; EEG/fMRI; Benzodiazepine
1. Introduction The BOLD/fMRI localization of interictal epileptic discharges (IEDs) or EEG/fMRI is a promising method in the presurgical study of drug-resistant focal epilepsies. It localizes IEDs sources corresponding to the birritative zoneQ [1] and generally there is agreement with the results of other methodologies such as EEG/MEG, PET, SPECT and MRI. Nevertheless, the IEDs recorded on the scalp usually correspond to a richer activity in depth and even intense epileptic activity cannot be recorded on the scalp [2–5]. This last limitation, which justifies depth recordings, can compromise the EEG/fMRI results. In fact, they come from a statistical analysis by comparing the temporal presence —
* Corresponding author. Dipartimento di Fisica, Universita` di Roma bLa Sapienza,Q 00185 Rome, Italy. Tel.: +39 649913473, +39 64454859; fax: +39 649913484. E-mail address: [email protected] (B. Maraviglia). 0730-725X/$ – see front matter D 2004 Elsevier Inc. All rights reserved. doi:10.1016/j.mri.2004.10.014
bactive stateQ — and the absence — binactive stateQ — of IEDs. A possible uncertainty about the inactive state can make the results unreliable. With the purpose of overcoming this obstacle in BOLD spike-triggered acquisition, the Geneva University Hospital’s team use clonazepam for a bcontrol conditionQ [6,7]. The intravenous injection of clonazepam, like other benzodiazepines, transitorily cancels or drastically reduces any epileptic activity providing a more reliable baseline for statistical analysis. However, the drug, using the EEG/fMRI continuous acquisition procedure [8,9], can give us a second opportunity, namely, to reveal possible BOLD areas not correlated to the surface EEG [10]. We can forecast that these areas can be highlighted by making a comparison between the MR images before and after the intravenous injection of benzodiazepine. Hence, we would be able to take advantage of MR physical features such as the better spatial definition and, above all, the lack of penalization due to the distance from the source that so strongly affects the electromagnetic measurements. Conse-
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quently, we can forecast a richer and a more complete BOLD areas display. The aim of this study is to control this hypothesis.
2. Materials and methods We simultaneously recorded the EEG and the BOLD activity (continuous acquisition) in eight patients with pharmacoresistant focal epilepsy candidates for surgery (Table 1). The EEG/fMRI was included among the routine researches: video — EEG seizures monitoring in wake and sleep state, MRI (T1, T2, IR, 3D, FLAIR), fMRI of eloquent areas, Wada test, MR spectroscopy and psychological test. The EEG/fMRI was the last of the series. Informed consent was obtained for all subjects in accordance with the protocol approved by the ethics committee of our institute. The EEG was recorded using a 10/20 system with 21 Ag/ AgCl electrodes soldered to 12 kV current-limiting resistors [11] applied on the scalp with conductive cream. The EEG device was EBNeuro Mizar 40 (Florence, Italy), 32 channels adapted for MR with a sampling rate of 4 kHz, which allows a suitable time resolution for picking up the switching effect of the readout gradient in the high slew rate condition, and the EEG dynamic range of F65.5 mV to prevent MRI artefact waveforms saturating the EEG/ECG. The MR artefact was filtered online [12]. The pulse artefact was minimized using particular wires locked arrangement and an elastic cap. The EPI acquisition was performed by means of a wholebody 1.5-T MR scanner (NVi, General Electrics Medical Systems, Milwaukee, WI, USA) equipped with a standard
birdcage head coil, 40 mT m 1 and 150 mT m 1 ms 1 of gradients amplitude and slew rate. Echo-planar axial images were collected using a continuous sequence: each volume consisted of 25 slices with a matrix 6464, covering the entire brain with a voxel size of 555 mm. The parameters of sequence were: TR/ TE=3000 ms/50 ms, flip angle=908, slice thickness=5 mm, FOV=2424 cm. We performed, for every subject, four runs; each run consisted of 80 volumes, lasted approximately 4 min and the interrun gap was approximately 1 min. Study duration varied from 60 to 90 min, including the anatomic scan. For each patient the same session was repeated after injection of lorazepam (4 Ag) in seven subjects and of diazepam (10 Ag) in one (S.C.).
3. Analysis of fMRI data Functional MRI images were first motion-corrected and then smoothed (Gaussian kernel; full width at half maximum 6 mm). The first two scans of each run were excluded from the analysis to let the system reach a steady state, that is, without the effect of signal saturation at the beginning of each acquisition. Analysis was within subject; hence, no spatial normalization or coordinate transformation procedures were applied. Subsequently, from these data sets, activation maps (R. W. Cox, AFNI) [13] were generated; functional analysis was carried out in the temporal domain on a voxel-by-voxel approach by deconvolving the time course of each voxel with a step function (activation/ baseline) to assess the hemodynamic response function.
Table 1 Clinical characteristics, MRI, EEG, and BOLD-BZ fMRI findings for eight patients with pharmacoresistent focal epilepsy Patients Sex Age Age of Neurological MRI (years) onset examination (years)
Interictal EEG
Ictal EEG
Epileptogenic zone
M.M.
m
31
12
Normal
f
37
12
Normal
F.A.
f
37
14
Normal
R. temporal theta (5–7 Hz) R. temporal theta (4 Hz) R. posterotemporal discharge (after subdural grid)
R. temporal lobe
L.M.
R. anterior temporal spikes R. anterior temporal spikes R. posterior temporal spikes
C.S.
f
27
0.5
C.A.
f
27
11
B.R.
m
44
12
U.A.
f
32
24
P.E.
f
23
9
Slight global Occipital L. frontal N cognitive atrophy R. frontal plus dysfunction L. occipital plus frontal bisynchronism Slight global Normal L. frontocentral cognitive dysfunction Normal L. hipp. L. temporal sclerosis spikes Normal R. hipp R. anterior sclerosis temporal spikes Normal R. hipp Bilateral sclerosis
R. hipp sclerosis R. hipp sclerosis R. hipp sclerosis
No lateralization
Surgery
R. temporal lobectomy R. temporal lobe R. temporal lobectomy R. temporoR. temporal occipital lobectomy plus temporo-occipital corticectomy L. supplementary Waiting for area (?) intracranial investigation
L. hemisphere
L. frontal (?)
L. temporal theta (7 Hz) discharge R. theta (4–5 Hz) discharge Left theta
L. temporal lobe R. temporal lobe ?
Waiting for intracranial investigation Waiting for surgery R. temporal lobectomy Waiting for surgery
BOLD-BZ fMRI
R. temporal R. temporal R. temporal plus R. parietotemporo-occipital L. frontal mesial
Seizure during acquisition L. temporal No activ. No activ.
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Fig. 1. (C.S.) BOLD-BZ/fMRI activation in a case with left supplementary motor area epilepsy (suspected); slice no. 16 in 3D.
Fig. 2. Comparison of the three analyses, BOLD-BZ/fMRI, EEG-BZ/fMRI, and EEG/fMRI, in the same subject (C.S.) (slice nos. 15–17). Note the better definition of BOLD-BZ/fMRI (first line) and the confused display of EEG/fMRI analysis (third line).
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The fMRI event slice number (read on the EEG traces) was also used as input for an event-related analysis of time course of BOLD activation. F-parametric maps were obtained with P value threshold at Pb.001 (Bonferroni correction was used to eliminate false positives derived from multiple comparisons); further restriction of the interpretation of the results was applied by considering only those regions where more than 10 neighboring pixels were significantly correlated. However, in the analysis referred to the inactive state estimated by the examiner, we were compelled to lower this coefficient. The results on EPI images were overlaid on the T1weighted images to anatomically illustrate activation sites. For each patient the areas of activation were delimited by repeating the same analysis three times: 1. IEDs with reference to the intervals estimated inactive by the examiner: EEG/fMRI. 2. IEDs with reference to the period of benzodiazepine (BZ) effect: EEG-BZ/fMRI. 3. Correlation between MR images of active state and the EPI images of inactive state caused by benzodiazepine and without reference to EEG: BOLD-BZ/fMRI.
4. Results A comparison between the three analyses was performed in five out of eight patients. One (A.C.) had a seizure during the acquisition and the movements artefacts did not allow data analysis. In two (U.A., P.E.), we did not record IEDs
during the acquisition. Consequently, our results regard five patients (M.M., L.M., F.A., C.S., B.R.). We found an agreement with BOLD-BZ/fMRI analysis in five of five patients using the EEG-BZ/fMRI analysis and in four of five patients with the EEG/fMRI. However, it was an agreement with qualitative differences. In fact, the BOLD-BZ/fMRI showed the better spatial definition (Figs. 1 and 2) while the EEG/fMRI gave less definite and blurred BOLD areas. In one case (L.M.) with a low number of IEDs, the EEG/fMRI analysis did not show activation. Note that the results of EEG/fMRI analysis were obtained by lowering the threshold coefficient. The EEGBZ/fMRI with the same coefficient threshold of BOLD-BZ/ fMRI analysis gave a satisfactory even if poorer spatial definition (Fig. 2). In particular, in temporal lobe epilepsy the BOLD areas were clearly defined by BOLD-BZ/fMRI, less so by EEGBZ/fMRI and were almost imperceptible to EEG/fMRI. Briefly, this comparison seems to show that the BOLD-BZ/ fMRI without EEG correlation gave better results with respect to the EEG-BZ/fMRI and particularly to the EEGfMRI analysis. These results if confirmed in a wider number of cases show for the first time that the better procedure of BOLD/ fMRI in epilepsy is comparing the active EPI images with the images inactivated by benzodiazepine and without EEG correlation. The use of the simultaneous EEG is only for checking the active state. Furthermore, the reference to inactive state due to benzodiazepine seems to improve the EEG/fMRI too. Finally, this new method is less dependent on the examiner opinion.
Fig. 3. (L.M.) Right temporal lobe epilepsy; right hippocampal sclerosis. The BOLD-BZ/fMRI analysis shows a right temporal BOLD area associated with other BOLD areas (secondary?) (slice nos. 10–13, 16 and 17).
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the analysis. Such drawback happens in a third of our patients and justifies some cases without BOLD activation (20–40%) published in the literature [8,9,11,12].
Fig. 4. (M.M.) Right temporal lobe epilepsy; right hippocampal sclerosis (slice no. 10). Note the right temporal BOLD response and the associated left frontal lobe activation (slice no. 13).
The hemodynamic response in epilepsy must be considered a nonexclusive research. In this paper we mention the BOLD localization that is in agreement with the other researches. However, often we have more that one BOLD activation in different and even controlateral regions (L.M., F.A., C.S.) (Figs. 3 and 4). Probably it is a question of secondary activation, but at the moment we have no enough data to express our opinion. 5. Discussion The hemodynamic response referred to IEDs or EEG/ fMRI can nowadays be considered a routine though not exclusive research in the presurgical study of drug-resistant epilepsy. Nevertheless, it has limitations, sometimes without solution: – It localizes the IED sources of irritative zone and not of the bepileptogenic zoneQ that is indispensable for the generation of epileptic seizures and the outcome of surgical treatment. – It works only if the number of IEDs is high enough for a correct statistical analysis. – The time of the acquisition is relatively short and does not permit the study of the whole evolution of the epileptic activity. In particular, the low number of IEDs may make it impossible to distinguish and to localize different morphological aspects of IEDs such as spikes, sharp waves, spike and waves, etc. – As a rule we cannot study the epileptic seizures because of head movement. Even in the rare seizures without head movement there are inherent difficulties for data processing [14–16]. – The hemodynamic response is too slow to define brain activity adequately. – The behavior of the subject during the acquisition can be a determining factor. Often, during the acquisition the patient is tense and anxious. The consequence can be a strong reduction or a block of IEDs that impede
However, in spite of these limitations, the EEG/fMRI in epilepsy is a progress. For the first time we can localize directly on MRI through a harmless procedure the sources of IEDs with a spatial definition otherwise inconceivable. Moreover, we can also study the IEDs relationship with possible pathologies evident on MRI. The aim of our study is to improve the methodology, avoiding the correlation with the simultaneous EEG in order to take the greatest advantage of the physical features of MRI. Till now the sole possible way for BOLD response in epilepsy was considered the EEG correlation as bspike-triggerQ and bcontinuousQ EEG/fMRI acquisitions [9,10,17–19]. The procedure of comparing the EPI images of the active state with the images of the state binactivatedQ by benzodiazepine can be considered a logical evolution of BOLD/fMRI in epilepsy. The EEG and the MR are regulated by different physical laws. Unlike the MR the EEG is penalized by the distance and less by the orientation of the sources within the brain [20]. Furthermore, the intervening tissues modify its spatial definition. The main effect is a reduction of voltage sometimes strong enough to hinder surface measurements. It follows that by using the EEG as point of reference we can reduce the amount of data and compromise the examiner decision because he can never be absolutely sure of an inactive state. The hemodynamic response corresponds to local neuronal activity and can be completely displayed on EPI images without interferences. The simultaneous EEG is necessary because at the state of art it is the only way to ascertain the active state. Our preliminary results seem to confirm these arguments. In comparison with the EEG/fMRI method, the BOLDBZ/fMRI has given more definite and more evident BOLD areas. Thanks to the benzodiazepine effect we have a better baseline (inactive state) for statistical analysis and consequently a more complete display of BOLD activation that is justified by IEDs not recorded on scalp EEG but revealed by BOLD/fMRI. In this study the benzodiazepine method did not show more BOLD areas with respect to areas revealed by EEG correlation. In other words, we did not see BOLD areas when IEDs were lacking during the acquisition (see results). It is likely that in the future and in a wider case study, we shall have the chance to meet BOLD areas highlighted by benzodiazepine method and not shown by EEG/ fMRI aquisitions.
References [1] Rosenow F, Luders H. Presurgical evaluation of epilepsy. Brain 2001;124:1683 – 700.
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[2] Ajmone-Marsan C, Van Buren JM. Epileptiform activity in cortical and subcortical structures in the temporal lobe of man. In: Baldwin P, Bailey P, editors. Temporal lobe epilepsy. Springfield (Ill)7 Charles C. Thomas; 1958. p. 78 – 108. [3] Cooper R, Winter AL, Crow HJ, Walter WG. Comparison of subcortical, cortical and scalp activity using chronically indwelling electrodes in man. Electroencephalogr Clin Neurophysiol 1965; 18:217 – 28. [4] Alarcon G, Guy CN, Binnie CD, Walker SR, Elwes RD, Polkey CE. Intracerebral propagation of interictal activity in partial epilepsy: implications for source localisation. J Neurol Neurosurg Psychiatry 1994;57:435 – 49. [5] Ebersole JS. Defining epileptogenic foci: past, present, future. J Clin Neurophysiol 1997;14:470 – 83. [6] Seeck M, Lazeyras F, Michel CM, Blanke O, Gericke CA, Ives J, et al. Non-invasive epileptic focus localization using EEG-triggered functional MRI and electromagnetic tomography. Electroencephalogr Clin Neurophysiol 1998;106:508 – 12. [7] Lazeyras F, Blanke O, Perrig S, Zimine I, Golay X, Delavelle J, et al. EEG-triggered functional MRI in patients with pharmacoresistant epilepsy. J Magn Reson Imaging 2000;12:177 – 85. [8] Lemieux L, Salek-Haddadi A, Josephs O, AlIen P, Toms N, Scott C, et al. Event-related fMRI with simultaneous and continuous EEG: description of the method and initial case report. Neuroimage 2001; 14:780 – 7. [9] Al-Asmi A, Benar CG, Gross W, Khani YA, Andermann F, Pike B, et al. FMRI activation in continuous and spike-triggered EEG-fMRI studies of epileptic spikes. Epilepsia 2003;44:1328 – 39. [10] Benar CG, Gross DW, Wang Y, Petre V, Pike B, Dubeau F, et al. The BOLD response to interictal epileptiform discharges. Neuroimage 2002;17:1182 – 92.
[11] Lemieu L, Allen PJ, Franconi F, Symms MR, Fish DR. Recording of EEG during fMRI experiments: patient safety. Magn Reson Med 1997;38:943 – 52. [12] Garreffa G, Carnı` M, Gualniera G, Ricci GB, Bozzao L, De Carli D, et al. Real-time MR artifacts filtering during continuous EEG/fMRI acquisition. Magn Reson Imaging 2003;21:1175 – 89. [13] Cox RW. AFNI: software for analysis and visualization of functional magnetic resonance neuroimages. Comput Biomed Res 1996;29: 162 – 73. [14] Salek-Haddadi A, Lemieux L, Merschhemke M, Friston KJ, Fish DR. Simultaneous EEG-correlated ictal fMRI. Neuroimage 2002; 16:32 – 40. [15] Krakow K, Baxendale SA, Maguire EA, Krishnamoorthy ES, Lemieux L, Scott CA, et al. Fixation-off sensitivity as a model of continuous epileptiform discharges: electroencephalographic, neuropsychological and functional MRI findings. Epilepsy Res 2000;42:1 – 6. [16] Iannetti GD, Di Bonaventura C, Pantano P, Giallonardo AT, Romanelli PL, Bozzao L, et al. fMRI/EEG in paroxysmal activity elicited by elimination of central vision and fixation. Neurology 2002;58:976 – 9. [17] Ives JR, Warach S, Schmitt F, Edelman RR, Schomer DL. Monitoring the patient’s EEG during echo planar MRI. Electroencephalogr Clin Neurophysiol 1993;87:417 – 20. [18] Warach S, Ives JR, Schlaug G, Patel MR, Darby DG, Thangaraj V, et al. EEG-triggered echoplanar functional MRI in epilepsy. Neurology 1996;47:89 – 93. [19] Krakow K, Woermann FG, Symms MR, Allen PJ, Lemieux L, Barker GJ, et al. EEG-triggered functional MRI of interictal epileptiform activity in patients with partial seizures. Brain 1999;122:1679 – 88. [20] Nunez PL. Neocortical dynamics and human EEG rhythms. New York7 Oxford University Press; 1995.
Hemodynamic response (BOLD/fMRI) in focal epilepsy with reference to benzodiazepine effect Giovanni Battista Riccia, Diego De Carlib,c,d, Claudio Colonnesee, Giancarlo Di Gennaroe, Pier Paolo Quaratoe, Gianpaolo Cantoree, Vincenzo Espositoa,e, Girolamo Garreffab,c,d,e, Bruno Maravigliab,c,d,* a
Department of Neurological Sciences, University of Rome bLa Sapienza,Q 00185 Rome, Italy b Enrico Fermi Center, 00184 Rome, Italy c Department of Physics, University of Rome bLa Sapienza,Q 00185 Rome, Italy d Centro di Ricerca e Sviluppo bSoft,Q 00185 Rome, Italy e Istituto di Ricovero e Cura a Carattere Scientifico bNeuromed,Q Pozzilli, 86077 Isernia, Italy Received 29 October 2004; accepted 29 October 2004
Abstract We studied a new procedure of BOLD/fMRI acquisition in epilepsy. They use the benzodiazepine effect to achieve a more reliable baseline for statistical analysis. The method works only in the MR domain without EEG correlation. It compares the EPI images during interictal epileptic discharges and the images binactivatedQ by benzodiazepine. The results in five out of eight patients show that this procedure in comparison with the EEG/fMRI method gives a net improvement of spatial definition of BOLD areas. These preliminary results seem to confirm the hypothesis that the better BOLD/fMRI procedure in epilepsy is to make use of physical features of MR that, unlike EEG, is not influenced by the distance of intercerebral sources and consequently allows a more complete and undistorted display of BOLD areas. D 2004 Elsevier Inc. All rights reserved. Keywords: Hemodynamic response; BOLD/fMRI; EEG/fMRI; Benzodiazepine
1. Introduction The BOLD/fMRI localization of interictal epileptic discharges (IEDs) or EEG/fMRI is a promising method in the presurgical study of drug-resistant focal epilepsies. It localizes IEDs sources corresponding to the birritative zoneQ [1] and generally there is agreement with the results of other methodologies such as EEG/MEG, PET, SPECT and MRI. Nevertheless, the IEDs recorded on the scalp usually correspond to a richer activity in depth and even intense epileptic activity cannot be recorded on the scalp [2–5]. This last limitation, which justifies depth recordings, can compromise the EEG/fMRI results. In fact, they come from a statistical analysis by comparing the temporal presence —
* Corresponding author. Dipartimento di Fisica, Universita` di Roma bLa Sapienza,Q 00185 Rome, Italy. Tel.: +39 649913473, +39 64454859; fax: +39 649913484. E-mail address: [email protected] (B. Maraviglia). 0730-725X/$ – see front matter D 2004 Elsevier Inc. All rights reserved. doi:10.1016/j.mri.2004.10.014
bactive stateQ — and the absence — binactive stateQ — of IEDs. A possible uncertainty about the inactive state can make the results unreliable. With the purpose of overcoming this obstacle in BOLD spike-triggered acquisition, the Geneva University Hospital’s team use clonazepam for a bcontrol conditionQ [6,7]. The intravenous injection of clonazepam, like other benzodiazepines, transitorily cancels or drastically reduces any epileptic activity providing a more reliable baseline for statistical analysis. However, the drug, using the EEG/fMRI continuous acquisition procedure [8,9], can give us a second opportunity, namely, to reveal possible BOLD areas not correlated to the surface EEG [10]. We can forecast that these areas can be highlighted by making a comparison between the MR images before and after the intravenous injection of benzodiazepine. Hence, we would be able to take advantage of MR physical features such as the better spatial definition and, above all, the lack of penalization due to the distance from the source that so strongly affects the electromagnetic measurements. Conse-
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quently, we can forecast a richer and a more complete BOLD areas display. The aim of this study is to control this hypothesis.
2. Materials and methods We simultaneously recorded the EEG and the BOLD activity (continuous acquisition) in eight patients with pharmacoresistant focal epilepsy candidates for surgery (Table 1). The EEG/fMRI was included among the routine researches: video — EEG seizures monitoring in wake and sleep state, MRI (T1, T2, IR, 3D, FLAIR), fMRI of eloquent areas, Wada test, MR spectroscopy and psychological test. The EEG/fMRI was the last of the series. Informed consent was obtained for all subjects in accordance with the protocol approved by the ethics committee of our institute. The EEG was recorded using a 10/20 system with 21 Ag/ AgCl electrodes soldered to 12 kV current-limiting resistors [11] applied on the scalp with conductive cream. The EEG device was EBNeuro Mizar 40 (Florence, Italy), 32 channels adapted for MR with a sampling rate of 4 kHz, which allows a suitable time resolution for picking up the switching effect of the readout gradient in the high slew rate condition, and the EEG dynamic range of F65.5 mV to prevent MRI artefact waveforms saturating the EEG/ECG. The MR artefact was filtered online [12]. The pulse artefact was minimized using particular wires locked arrangement and an elastic cap. The EPI acquisition was performed by means of a wholebody 1.5-T MR scanner (NVi, General Electrics Medical Systems, Milwaukee, WI, USA) equipped with a standard
birdcage head coil, 40 mT m 1 and 150 mT m 1 ms 1 of gradients amplitude and slew rate. Echo-planar axial images were collected using a continuous sequence: each volume consisted of 25 slices with a matrix 6464, covering the entire brain with a voxel size of 555 mm. The parameters of sequence were: TR/ TE=3000 ms/50 ms, flip angle=908, slice thickness=5 mm, FOV=2424 cm. We performed, for every subject, four runs; each run consisted of 80 volumes, lasted approximately 4 min and the interrun gap was approximately 1 min. Study duration varied from 60 to 90 min, including the anatomic scan. For each patient the same session was repeated after injection of lorazepam (4 Ag) in seven subjects and of diazepam (10 Ag) in one (S.C.).
3. Analysis of fMRI data Functional MRI images were first motion-corrected and then smoothed (Gaussian kernel; full width at half maximum 6 mm). The first two scans of each run were excluded from the analysis to let the system reach a steady state, that is, without the effect of signal saturation at the beginning of each acquisition. Analysis was within subject; hence, no spatial normalization or coordinate transformation procedures were applied. Subsequently, from these data sets, activation maps (R. W. Cox, AFNI) [13] were generated; functional analysis was carried out in the temporal domain on a voxel-by-voxel approach by deconvolving the time course of each voxel with a step function (activation/ baseline) to assess the hemodynamic response function.
Table 1 Clinical characteristics, MRI, EEG, and BOLD-BZ fMRI findings for eight patients with pharmacoresistent focal epilepsy Patients Sex Age Age of Neurological MRI (years) onset examination (years)
Interictal EEG
Ictal EEG
Epileptogenic zone
M.M.
m
31
12
Normal
f
37
12
Normal
F.A.
f
37
14
Normal
R. temporal theta (5–7 Hz) R. temporal theta (4 Hz) R. posterotemporal discharge (after subdural grid)
R. temporal lobe
L.M.
R. anterior temporal spikes R. anterior temporal spikes R. posterior temporal spikes
C.S.
f
27
0.5
C.A.
f
27
11
B.R.
m
44
12
U.A.
f
32
24
P.E.
f
23
9
Slight global Occipital L. frontal N cognitive atrophy R. frontal plus dysfunction L. occipital plus frontal bisynchronism Slight global Normal L. frontocentral cognitive dysfunction Normal L. hipp. L. temporal sclerosis spikes Normal R. hipp R. anterior sclerosis temporal spikes Normal R. hipp Bilateral sclerosis
R. hipp sclerosis R. hipp sclerosis R. hipp sclerosis
No lateralization
Surgery
R. temporal lobectomy R. temporal lobe R. temporal lobectomy R. temporoR. temporal occipital lobectomy plus temporo-occipital corticectomy L. supplementary Waiting for area (?) intracranial investigation
L. hemisphere
L. frontal (?)
L. temporal theta (7 Hz) discharge R. theta (4–5 Hz) discharge Left theta
L. temporal lobe R. temporal lobe ?
Waiting for intracranial investigation Waiting for surgery R. temporal lobectomy Waiting for surgery
BOLD-BZ fMRI
R. temporal R. temporal R. temporal plus R. parietotemporo-occipital L. frontal mesial
Seizure during acquisition L. temporal No activ. No activ.
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Fig. 1. (C.S.) BOLD-BZ/fMRI activation in a case with left supplementary motor area epilepsy (suspected); slice no. 16 in 3D.
Fig. 2. Comparison of the three analyses, BOLD-BZ/fMRI, EEG-BZ/fMRI, and EEG/fMRI, in the same subject (C.S.) (slice nos. 15–17). Note the better definition of BOLD-BZ/fMRI (first line) and the confused display of EEG/fMRI analysis (third line).
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The fMRI event slice number (read on the EEG traces) was also used as input for an event-related analysis of time course of BOLD activation. F-parametric maps were obtained with P value threshold at Pb.001 (Bonferroni correction was used to eliminate false positives derived from multiple comparisons); further restriction of the interpretation of the results was applied by considering only those regions where more than 10 neighboring pixels were significantly correlated. However, in the analysis referred to the inactive state estimated by the examiner, we were compelled to lower this coefficient. The results on EPI images were overlaid on the T1weighted images to anatomically illustrate activation sites. For each patient the areas of activation were delimited by repeating the same analysis three times: 1. IEDs with reference to the intervals estimated inactive by the examiner: EEG/fMRI. 2. IEDs with reference to the period of benzodiazepine (BZ) effect: EEG-BZ/fMRI. 3. Correlation between MR images of active state and the EPI images of inactive state caused by benzodiazepine and without reference to EEG: BOLD-BZ/fMRI.
4. Results A comparison between the three analyses was performed in five out of eight patients. One (A.C.) had a seizure during the acquisition and the movements artefacts did not allow data analysis. In two (U.A., P.E.), we did not record IEDs
during the acquisition. Consequently, our results regard five patients (M.M., L.M., F.A., C.S., B.R.). We found an agreement with BOLD-BZ/fMRI analysis in five of five patients using the EEG-BZ/fMRI analysis and in four of five patients with the EEG/fMRI. However, it was an agreement with qualitative differences. In fact, the BOLD-BZ/fMRI showed the better spatial definition (Figs. 1 and 2) while the EEG/fMRI gave less definite and blurred BOLD areas. In one case (L.M.) with a low number of IEDs, the EEG/fMRI analysis did not show activation. Note that the results of EEG/fMRI analysis were obtained by lowering the threshold coefficient. The EEGBZ/fMRI with the same coefficient threshold of BOLD-BZ/ fMRI analysis gave a satisfactory even if poorer spatial definition (Fig. 2). In particular, in temporal lobe epilepsy the BOLD areas were clearly defined by BOLD-BZ/fMRI, less so by EEGBZ/fMRI and were almost imperceptible to EEG/fMRI. Briefly, this comparison seems to show that the BOLD-BZ/ fMRI without EEG correlation gave better results with respect to the EEG-BZ/fMRI and particularly to the EEGfMRI analysis. These results if confirmed in a wider number of cases show for the first time that the better procedure of BOLD/ fMRI in epilepsy is comparing the active EPI images with the images inactivated by benzodiazepine and without EEG correlation. The use of the simultaneous EEG is only for checking the active state. Furthermore, the reference to inactive state due to benzodiazepine seems to improve the EEG/fMRI too. Finally, this new method is less dependent on the examiner opinion.
Fig. 3. (L.M.) Right temporal lobe epilepsy; right hippocampal sclerosis. The BOLD-BZ/fMRI analysis shows a right temporal BOLD area associated with other BOLD areas (secondary?) (slice nos. 10–13, 16 and 17).
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the analysis. Such drawback happens in a third of our patients and justifies some cases without BOLD activation (20–40%) published in the literature [8,9,11,12].
Fig. 4. (M.M.) Right temporal lobe epilepsy; right hippocampal sclerosis (slice no. 10). Note the right temporal BOLD response and the associated left frontal lobe activation (slice no. 13).
The hemodynamic response in epilepsy must be considered a nonexclusive research. In this paper we mention the BOLD localization that is in agreement with the other researches. However, often we have more that one BOLD activation in different and even controlateral regions (L.M., F.A., C.S.) (Figs. 3 and 4). Probably it is a question of secondary activation, but at the moment we have no enough data to express our opinion. 5. Discussion The hemodynamic response referred to IEDs or EEG/ fMRI can nowadays be considered a routine though not exclusive research in the presurgical study of drug-resistant epilepsy. Nevertheless, it has limitations, sometimes without solution: – It localizes the IED sources of irritative zone and not of the bepileptogenic zoneQ that is indispensable for the generation of epileptic seizures and the outcome of surgical treatment. – It works only if the number of IEDs is high enough for a correct statistical analysis. – The time of the acquisition is relatively short and does not permit the study of the whole evolution of the epileptic activity. In particular, the low number of IEDs may make it impossible to distinguish and to localize different morphological aspects of IEDs such as spikes, sharp waves, spike and waves, etc. – As a rule we cannot study the epileptic seizures because of head movement. Even in the rare seizures without head movement there are inherent difficulties for data processing [14–16]. – The hemodynamic response is too slow to define brain activity adequately. – The behavior of the subject during the acquisition can be a determining factor. Often, during the acquisition the patient is tense and anxious. The consequence can be a strong reduction or a block of IEDs that impede
However, in spite of these limitations, the EEG/fMRI in epilepsy is a progress. For the first time we can localize directly on MRI through a harmless procedure the sources of IEDs with a spatial definition otherwise inconceivable. Moreover, we can also study the IEDs relationship with possible pathologies evident on MRI. The aim of our study is to improve the methodology, avoiding the correlation with the simultaneous EEG in order to take the greatest advantage of the physical features of MRI. Till now the sole possible way for BOLD response in epilepsy was considered the EEG correlation as bspike-triggerQ and bcontinuousQ EEG/fMRI acquisitions [9,10,17–19]. The procedure of comparing the EPI images of the active state with the images of the state binactivatedQ by benzodiazepine can be considered a logical evolution of BOLD/fMRI in epilepsy. The EEG and the MR are regulated by different physical laws. Unlike the MR the EEG is penalized by the distance and less by the orientation of the sources within the brain [20]. Furthermore, the intervening tissues modify its spatial definition. The main effect is a reduction of voltage sometimes strong enough to hinder surface measurements. It follows that by using the EEG as point of reference we can reduce the amount of data and compromise the examiner decision because he can never be absolutely sure of an inactive state. The hemodynamic response corresponds to local neuronal activity and can be completely displayed on EPI images without interferences. The simultaneous EEG is necessary because at the state of art it is the only way to ascertain the active state. Our preliminary results seem to confirm these arguments. In comparison with the EEG/fMRI method, the BOLDBZ/fMRI has given more definite and more evident BOLD areas. Thanks to the benzodiazepine effect we have a better baseline (inactive state) for statistical analysis and consequently a more complete display of BOLD activation that is justified by IEDs not recorded on scalp EEG but revealed by BOLD/fMRI. In this study the benzodiazepine method did not show more BOLD areas with respect to areas revealed by EEG correlation. In other words, we did not see BOLD areas when IEDs were lacking during the acquisition (see results). It is likely that in the future and in a wider case study, we shall have the chance to meet BOLD areas highlighted by benzodiazepine method and not shown by EEG/ fMRI aquisitions.
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