Vagus Nerve Stimulation in Refractory Epilepsy:SPECT Activation StudyKoenraad Van Laere, Kristl Vonck, Paul Boon, Boudewijn Brans, Tom Vandekerckhove, and Rudi Dierckx

Division of Nuclear Medicine; Epilepsy Monitoring Unit, Department of Neurology; and Department of Neurosurgery,Gent University Hospital, Gent, Belgium

Left-sided vagus nerve stimulation (VMS) is an efficacious treatment for patients with refractory epilepsy. The exact mechanismof action remains to be elucidated. This study investigated theacute effects of initial VNS in patients with refractory complexpartial epilepsy with or without secondary generalization (complex partial seizures [CPS] ±SG) by means of a perfusionactivation study with SPECT. Methods: Twelve patients (meanage, 32.2 ±10.2y; age range, 12-47 y) with a mean duration ofCPS ±SG of 19.8 ±10.0 y (range, 5-33 y) received VNS. All

patients were considered unsuitable candidates for resectivesurgery because of nonlocalizing findings on presurgical evaluation. VNS efficacy was evaluated for patients with at least 4-mofollow-up. VNS-induced regional cerebral blood flow alterationswere studied by a 99mTc-ethylcysteinate dimer activation studywith a single-day split-dose protocol before and immediately afteran initial stimulation. Images were acquired on a triple-head

camera with fanbeam collimators. After coregistration to a standardized template, both a semiquantitative analysis using predefined volumes of interest and a voxel-by-voxel analysis of theintrasubject activation (statistical parametric mapping) wereperformed. Results: Seizure-frequency changes ranged from

100% decrease to 0% after VNS. The semiquantitative analysisrevealed a consistent decrease of activity in the left thalamus(ratio stimulator on/off = 0.94 ±0.04; P = 0.005). These resultswere concordant with the voxel-by-voxel analysis in which asignificant deactivation in the left thalamus was found withspread to the ipsilateral hippocampus. There was no statisticallysignificant correlation between initial VNS-induced thalamichypoperfusion and seizure reduction at maximum follow-up.Conclusion: Our findingsare consistentwiththe hypothesisthatacute VNS reduces seizure onset or propagation through inhibition of the thalamic relay center. Differences with limited H215O

PET data may be associated with temporal effects caused by astimulation-Induced local hemodynamic response and need further investigation. SPECT allows study of cerebral physiopatho-

logic effects of vagus nerve electrostimulation in complex partialepilepsy.

Key Words: epilepsy;SPECT; thalamus;vagusnervestimulationJ NucÃ-Med 2000; 41:1145-1154

Received Jul. 6,1999; revision accepted Oct. 4,1999.For correspondence or reprints contact: Koenraad Van Laere, MD, DSc,

Division of Nuclear Medicine, P7, Gent University Hospital, De Pintelaan 185,9000 Gent, Belgium.

refractory complex partial seizures with or withoutsecondary generalization (CPS ±SG) are present in approximately one third of epilepsy patients. Apart from newanticonvulsant drugs, seizures can be controlled neurosurgi-

cally by partial resection, subpial trans section, or corpuscallosotomy (7). However, up to 40% of these patients showinsufficient response or are unsuitable candidates for thesetherapies (2). For a number of them, intermittent electricalstimulation of the vagus nerve can provide a new form oftherapy (3-5). Although the effect of vagus nerve stimula

tion (VNS) of cats was reported in 1952 (6), the idea ofpreventing human epileptic insults by VNS has been introduced only recently. Five-year follow-up studies haveshown that left-sided VNS is a safe and effective treatmentfor refractory epilepsy (7). About 30%-40% of patients

experience a reduction of >50% (the cutoff defined in moststudies on anticonvulsant drug efficacy) from VNS (7-9).However, only a small number (2%-3%) of patients become

completely free of seizures in the long term (/). Side effectsof the treatment are limited primarily to initial hoarseness,throat pain, paresthesia, or dyspnea (7,10), but vocal cordparesis associated with the surgical procedure (7), intraoperative ventricular asystole (3 cases) (77), and an unexplainedmyocardial infarction (5) have been reported.

The exact mechanism by which VNS exerts its antiepilep-

tic effect is poorly understood. Elucidation of this mechanism is an important first step in the development ofstrategies to improve VNS efficacy. The vagus nervecommunicates with the nucleus tractus solitarius in the brainstem. From this location, it may influence various parts ofthe brain. Regions including the medulla, cerebellum, para-

brachial nucleus, locus coeruleus, hypothalamus, thalamus,amygdala, hippocampus, cingulate gyrus, and contralateralsomatosensory cortex have been implicated in electrophysi-ologic and PET imaging studies (9,77-75). A recent study by

Henry et al. (73) suggested a correlation between increasedthalamic blood flow during stimulation and improvements inseizure control. Other data indicate that stimulation decreases global brain excitability, increases seizure threshold,and can abort seizures (16).

In view of the limited and conflicting knowledge on theunderlying mechanism of action of VNS, it was our objective to study the short-term changes in regional cerebral

BCD SPECT IN VAGUSNERVESTIMULATION•Van Laere et al. 1145

blood flow (rCBF) caused by the initial stimulation in aseries of VNS patients who had undergone recent implantation. Functional imaging, including SPECT, allows themeasurement of rCBF, which reflects primarily changes intrans-synaptic neurotransmission. Because of inherent tech

nical difficulties of functional MRI (fMRI) activation studiesin patients with electrode implants (17), to our knowledge,only PET studies with a limited number of patients havebeen published (12-15,18).

Analysis of SPECT activation studies can be done inseveral ways. One conventional approach is evaluation ofstatistical differences in activity using volumes of interest(VOIs). This approach allows automated statistical comparison of individual patient data in predefined regions afteraccurate image coregistration. On the other hand, thesecoregistration techniques have also allowed the development of robust voxel-based approaches, in which small

volume elements corresponding to gray matter are comparedregardless of anatomic localization. Increasingly, statisticalparametric mapping (SPM) (79) is becoming the goldstandard for PET, fMRI, and SPECT group activationstudies (20). Because both VOI and SPM analyses havespecific advantages for SPECT, results obtained by bothmethods are presented.

MATERIALS AND METHODS

PatientsTwelve patients (4 men, 8 women) with refractory epilepsy

(CPS ±SG) participated in this prospective study. The meanpatient age at the time of study was 32.2 ±10.2 y (range, 12-47 y),

and the mean duration of the disease before VNS implantation was19.3 ± 11.0 y (range, 5-33 y). All patients underwent VNS astreatment for medication-resistant CPS ±SG (Table 1) but were

considered unsuitable candidates for resective surgery because ofnonlocalizing findings on presurgical evaluation. This presurgicalevaluation consisted of thorough neurologic and neuropsychologicinvestigation, electroencephalography (EEC), ictal video EEGMRI and spectroscopy, FDG PET, and ictal SPECT (1 case).Detailed patient characteristics with the main presurgical evaluations are given in Table 1. For 11 patients, it was the first surgicaltreatment for their disease and the first neurosurgery they underwent. For patient 12 (Lennox-Gastaut syndrome), anterior callo-

sotomy had been performed 5 y before VNS without clinicallysignificant long-term improvement.

All patients were under chronic treatment with antiepilepticdrugs (AEDs). On average, 3.3 AEDs were taken before VNS. OneAED was tapered after VNS in only 1 patient (patient 1). AEDdosages were not changed during the weeks before the operationand the SPECT study. Patients took their medication on the day ofthe study.

All patients gave informed consent on the basis of the SPECTstudy protocol approved by the local ethics committee. Noindustrial firms sponsored any patients. At this time, no financialreimbursement by the government health insurance is granted forVNS.

Electrode Implantation and Clinical ImprovementA neurocybernetic prosthesis (NCP) (model 100 NCP; Cyberon-

ics, Webster, TX), composed of a pulse generator and a bipolar

helical lead, was implanted during a surgical procedure undergeneral anesthesia that required 2 incisions. After the pulsegenerator was implanted in the subclavicular area, the lead wastunneled subcutaneously and wound around the left vagus nerve.By means of an externally applied device (model 200 NCPProgramming Wand; Cyberonics), the model 250 NCP Programming Software, and a personal computer, this generator wasprogrammed with standard parameters (output current, 0-3 mA;

pulse width, 500 MS:frequency, 30 Hz; on/off, 30 s/600 s). Allpatients received the initial stimulation at the time of the SPECTstudy. Depending on each patient's characteristics, a 0.25- or

0.5-mA pulse intensity was programmed (Table 2).

No intraoperative or immediately postoperative adverse eventsoccurred for any patients in this study. The mean time interval(±SD) between surgery and VNS initialization was 4.1 ±3.2 wk,depending on seizure frequency and severity, wound healing, andlogistic aspects. All patients recorded their seizures on calendars.During the course of the follow-up, it was necessary to increment

the pulse intensity for most patients to improve or stabilize seizurereduction. Estimates of the reduction in seizure severity were madeby the patients, taking into account postictal alertness, length ofseizures, injuries, and severity of the ictal state. Fractional reduction was calculated for each individual as [(number of CPS duringVNS - number of CPS during baseline)/number of CPS during

VNS]. Because of difficulties in accountability, simple partialseizures (SPS) were not included. We prospectively assessedchanges in seizure frequency and adverse events in relation tooutput current in all patients.

Activation Paradigm and Data Acquisition"Tc-ethyl cysteinate dimer (BCD) (Neurolite; Du Pont Pharma

ceuticals Ltd., Brussels, Belgium) was used to obtain rCBFestimates. Each subject underwent 2 consecutive SPECT scans. Asplit-dose SPECT technique was used to study the situation with

and without stimulation. Figure 1 schematizes the activationparadigm. For the first measurement, the patient was injected with a555-MBq (15 mCi) bolus dose under standard conditions (rest,

supine position, quiet surroundings, dimly lit room, eyes open). Acomplete EEG was obtained during the entire study time; injectiontime was marked on the EEG. No seizures occurred during SPECTor during the previous 12 h (as determined by the subject's report).

The first scan was obtained 10 min after injection.Before the second scan, the electrode was switched on to 0.25 or

0.5 mA for a total duration of 30 s. During the first stimulation train,all patients reported left-sided cervical paresihesia (primarily throat

irritation), which often was accompanied by coughing. Becausestimulation lasts only 30 s and BCD uptake is a weighted integralover the first 2-3 min after injection, possible transient temporal

effects caused by the stimulation itself were avoided by starting theinjection immediately after the stimulus sequence. The second dosewas injected under the same standard conditions at exactly at theend of the 30-s stimulus. The time interval between the first andsecond injections was 35-45 min after the first injection. Further

stimulation (normally occurring every 10 min) was interruptedtemporarily until after the SPECT session to avoid motion artifactsduring the course of obtaining the second scan.

SPECT images were acquired using a GCA-9300 triple-head

system (Toshiba, Tokyo, Japan). The same acquisition parameterswere used for both serial SPECT scans. A 360°acquisition was

performed over 90 angles (continuous mode) with 40 s perprojection on a 128 X 128 matrix. A low-energy, superhigh-

1146 THEJOURNALOFNUCLEARMEDICINE•Vol. 41 •No. 7 •July 2000

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TABLE 2VNS Characteristics and Initial Results

Patientno.123456789101112Stimulus

intensityat SPECT(mA)0.250.250.250.50.250.50.250.50.50.50.50.25Follow-up(mo)271997238665221Stimulus

intensityat maximum

follow-up(mA)2.251.52.252.252.51.751.252.01.750.750.50.5Seizure

frequency(CPS/mo)Before

VNS16200

(clusters)1220831023054120After

VNS4091220.56130(D*(4)*(100)*%change-75-100-25-40-75-83-40-500(-80)*(0)*(-17)*

•Resultsfor insufficient follow-up (incomplete ramping up) are given in parentheses.

resolution fanbeam collimator was used, resulting in a transaxialresolution of 7.8-mm full width at half maximum. Images wereacquired in 3 windows (20% at 140 keV, 7% adjacent upper- andlower-scatter windows). The average counts obtained were 2.5

million counts for the first study and 5 million counts for the secondstudy.

Image ProcessingImages were scatter corrected using the commercially available

triple-energy window method; Butterworth order 8 filtering was

used with a cutoff of 0.14 cycle/cm for the main window and 0.07cycle/cm for the scatter windows. Scatter-corrected projectionswere reconstructed using filtered backprojection with a Butter-

worth filter of order 8. The filter cutoff was increased from 0.15cycle/cm for the first study to 0.20 cycle/cm because of the highercount statistics on the second scan. Uniform attenuation correctionwas applied with automatic definition of the sinogram border(Sörensencorrection: mean attenuation coefficient, 0.12/cm). Reconstructed images were transferred in Interfile format onto acentral personal computer-based image-processing system (Pen

tium II, HERMES; Nuclear Diagnostics, Stockholm, Sweden). Thereconstructed data were fitted automatically onto an in-house-constructed database template positioned in Talaraich co-ordinates

(21). This template consisted of 30 healthy volunteers with the

same age distribution as the patients in this study and was measuredwith the same equipment and total dose, under the same standardconditions, and reconstructed as above. The fitting procedure wasperformed with 9 parameters (scale, shift, and rotation) using aprincipal axis transform and a count-difference minimization

algorithm (fit threshold, 0.50; BRASS, Nuclear Diagnostics) (22).

VOI ActivityTo evaluate the magnitude of activity differences over extended

predefined regions, the first approach consisted of calculation of theactivity changes for 23 predefined VOIs, including the majorneocortical and subcortical gray matter structures throughout thebrain. This set of VOIs was delineated on the standard perfusiontemplate described above by direct reference to the Talaraich atlas(27). For each individual subject and scan, the VOI activity countswere calculated per voxel and normalized onto the total number ofcounts in the complete VOI set of the scan.

SPMApart from the conventional VOI approach, images were

analyzed on a voxel-by-voxel basis. To apply SPM within SPM96

(Wellcome Department of Cognitive Neurology, Institute of Neurology, London, UK), images were converted into ANALYZE formatby means of an in-house conversion program (MedCon). All SPM

FIGURE 1. SPECT activation paradigmfor studying acute effects of VNS on rCBF.Second injection takes place immediatelyafter 30-s stimulus. tx = time in minutesrelative to first radioligand injection.

555 MBq555MBqT

SPECT 1 T SPECT2tlO-30t45-65to

i:RestAmtmmmm I35p#pStimulator

ON30sec, 30 Hzt:::':::':-:':-:':-:';::':::':::-

*•ffiVÄVVVA^vVh.

1148 THEJOURNALOFNUCLEARMEDICINE•Vol. 41 •No. 7 •July 2000

calculations were performed in Matlab 4.2 (The MathWorks,Natick, MA) on a SUN SPARC 10 computer (Sun MicrosystemsEurope Inc., Brussels, Belgium). Normalization was performed byapplying the transformation parameters of the SPECT perfusiontemplate onto a PET SPM96 template smoothed isotropically witha 14-mm Gaussian kernel. This normalization was done using a9-parameter rigid body transformation (shift, scale, and rotation).

No affine (shear) parameters were used because of the relativelypoor spatial resolution of SPECT data. Differences in globalactivity between scans were removed by scaling the activity in eachpixel proportionally to the global activity. The mean global activityof each scan was adjusted to 50. The threshold value for gray matterwas set at 0.60, and a resulting voxel size of 3 X 3 X 3 mm wasused. The normalized studies were smoothed with an isotropie14-mm kernel to account for individual variability in the structure-function relationship and to improve the signal-to-noise ratio (23).

For determination of significant sites of increased or decreasedperfusion, a categorical multisubject, multiple condition model wasused with a design matrix shown in Figure 2. Planned comparisonsbetween conditions were performed on a pixel-by-pixel basis by t

statistics, generating SPM(t) maps subsequently transformed to theunit normal distribution SPM(Z) maps. We investigated areas usinga height threshold corresponding to an uncorrected P < 0.01.Without a correction for multiple comparisons, findings can only begiven descriptively. Surviving areas were considered to be meaningful in those areas for which preexisting anatomophysiologic

JS 03 3 3W W W «

CO ffl ffi 0)

8980

14

FIGURE 2. Design matrix for SPM analysis: multiple conditions, multisubject analysis. Paired analysis includes regionaldifferences between individual subjects. In design matrix, this isexpressed as extra column (effect) for each subject (1-12) nextto 2 scanning conditions (scan_1-scan_2). In horizontal rows,white rectangles show combinations that are considered for eachobservation.

hypotheses were present, such as the areas implied by previousstudies (20,24).

StatisticsThe Wilcoxon signed rank test, which considers both direction

and size of differences among paired observations, was used toanalyze seizure occurrence preoperatively and after stimulation andVOI uptake differences before and after stimulation. The Mann-

Whitney U test was used to try to differentiate VOI data betweeneffective and ineffective stimulation (responders versus nonre-

sponders). Statistics were calculated with SPSS (version 7.5 forWindows; SPSS Inc., Heverlee, Belgium).

RESULTS

Clinical ImprovementOn the basis of the results of the preoperative findings

(Table 1), all patients were unsuited for resective surgery. Aprecise epileptic focus could not be identified in 7 patients.Lateralization was considered to be consistent, and anintracarotid amobarbital procedure (Wada test (25)) wasconducted in 4 patients (patients 1, 6, 7, and 11). The optionof resective surgery had to be abandoned because of the riskof severe memory impairment in these patients. One patient(patient 10) with a dysplastic parahippocampal lesion didnot undergo surgery because of fear of damaging the visualcortex.

The baseline SPECT studies are also included in Table 1.These were compared on a voxel-by-voxel basis with theSPECT ECD perfusion template (n = 30). Significant

perfusion differences are given with a threshold > 2 SDs(BRASS; Nuclear Diagnostics).

The average follow-up for the whole series of patientswas 9.4 mo (range, 1-27 mo) (Table 2). Complete stimula

tion had been ramped up to therapeutic output current levelsin 9 of 12 patients (Table 2). In Table 2, the clinicaleffectiveness is given as reduction in occurrence of epilepticseizures after the onset of stimulation for those patients withat least 4-mo follow-up. VNS significantly reduced (meanreduction, 54%; P = 0.012) the number of typical seizures in

these patients.Seizure-frequency changes ranged from 100% decrease to

0%. Four patients (37%) achieved successful improvement—

i.e., >50% seizure reduction. One patient became free ofCPS for >2 y. For 3 patients, VNS resulted in moderatereduction (30%-50%), whereas 2 patients showed only a

minor effect of VNS (reduction < 30%).

rCBF AnalysisTable 3 shows results of the activity analysis for each of

the 23 defined regions of interest. The total voxel size isgiven in Table 3 for each defined region because it isimportant for the extent of averaging over the VOI. Asignificant perfusion decrease was found for the left thalamus (activity difference averaged over VOI, 5.4%; Wilcoxon P = 0.005) and the left parietal cortex (activitydifference, 1.7%; P = 0.023). No significantly increased

perfusion areas were present.Figure 3 shows SPM96 glass brain maps of the clusters

ECD SPECT IN VAGUSNERVESTIMULATION•Van Laere et al. 1149

TABLE 3VOI Comparison (Study Stimulator On/Study Stimulator Off)

for 12 VNS-Stimulated Epilepsy Patients

VOI

ActivityVOI size ratio(voxels) (on/off) SD

L dorsolateralprefrontalLlateralfrontalL

centralLparietalL

lateraltemporalLmedialtemporalL

occipitalLcerebellumL

nucleuslentiformiLthalamusL

headcaudatePonsR

dorsolateralprefrontalRlateralfrontalR

centralRparietalR

lateraltemporalRmedialtemporalR

occipitalRcerebellumR

nucleuslentiformisRthalamusR

head caudate67353825442560117742595910711874130673538254425601177425959107118740.9911.0020.9720.9830.9880.9810.9781.0131.0150.9461.0211.0161.0091.0201.0061.0021.0080.9720.9761.0121.0251.0130.9810.0510.0370.0500.0250.0320.0710.0510.0420.0710.0390.0810.1050.0580.0270.0540.0570.0490.0720.0470.0280.0590.0700.071-0.39-0.36-1.65-2.27-1.33-0.82-1.84-1.29-0.94-2.83-0.78-0.39-0.27-1.87-0.08-0.43-0.82-1.57-1.49-1.73-1.14-0.94-1.020.590.720.100.023*0.180.410.060.190.350.005*0.430.690.790.060.940.670.410.120.140.080.260.350.31

*Two-tailedsignificance level; P< 0.05.

Wilcoxon signed rank test statistics were used.

derived from the deactivation contrast. A significant clusterof decreased perfusion was found in the ipsilateral thalamus,with spread to the ipsilateral hippocampus and the left sideof the brain stem. The spatial resolution of the SPECTimages did not permit designation of this deactivation toparticular nuclei within the thalamus, but most of thesignificant voxels were located in the medioposterior regions of the thalamus. Table 4 shows the standard coordinates of the maximal significant voxel in the identifiedclusters, the cluster volume size, and the cluster Z values.Figure 4 shows a scatter plot of individual adjusted rCBFvalues in both states at the SPM maximum for the leftthalamus at Talaraich co-ordinates x = -18, y = -30, andz = 6. Adjusted rCBF values are obtained by normalization

of global flow to 50 mL/min/100 g. The adjusted rCBFchange for the maximum voxel was 10.6%.

The activation contrast showed a perfusion differencebilaterally in the frontal superior gyri (BA 10 left and BA 8right) at a height threshold of 0.01 and uncorrected extentthreshold of 0.05 (Table 4). No other activation clusters wereidentified at these thresholds.

No significant difference in VOI perfusion was foundfor the ipsilateral thalamus (P = 0.46) (Mann-Whitney's U

test) between the 2 groups defined as good responders(reduction > 50% in seizure frequency; n = 4) and

SPM{Z}

FIGURE 3. Significantly reduced rCBF by acute initial VMSstimulation, shown as SPM projections with height threshold of0.008 and spatial extent P = 0.05. L = left; R = right.

nonresponders (reductionn = 5).

< 50% in seizure frequency;

DISCUSSION

Clinical ResultsThe preliminary clinical follow-up results of this group of

patients are concordant with other series in which an efficacyof 30%—40%is described (defined as seizure reductions >

50%) (1,4,7). In this series, only 1 patient with preoperativecluster-type seizures became completely free of insults. This

is more or less in agreement with published percentages aslow as 2%-3% (7). Therefore, our results in this patient

TABLE 4Areas Showing Activity Differences Caused by Acute Vagus

Nerve Electrostimulation in VNS Patients

Co-ordinate(mm)

Clustersize Anatomic location cluster l'UlÃ-lÃ- rCBF

(voxels) (Brodmann's area) score x y z change*

Deactivation514 L thalamus

Activation312 L gyrus frontalis

superior (BA10)311 R gyrus frontalis 3.49 15 33 39 +5.7

superior (BA 8)

3.65 -21 -26 3 -10.6

3.53 -18 60 -6 +6.5

*Mean rCBF decrease in adjusted response for maximally signifi

cant voxel.Analysis was by SPM96 (multisubject,dual-condition design).

1150 THEJOURNALOFNUCLEARMEDICINE•Vol. 41 •No. 7 •July 2000

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o0)§58l56

3W5250••ibaseline.•••VNS

FIGURE 4. Scatter plot of individualadjusted rCBF values inboth states at SPM maximum for left thalamus (Talaraich coordinates: x = -18, y = -30, z = 6). Adjusted rCBF values areobtained by normalization of global flow to 50 mL/min/100 g. Zscore = 3.45; P < 0.001 (uncorrected).

group confirm that VNS seems to be an effective treatmentoption in the case of CPS ±SG. Overall, a significantreduction in the number of CPS was noticed for mostpatients.

In this series we found that the (documented) interictalbaseline study showed significant thalamic perfusion differences in several patients when compared on a voxel-by-voxel basis with an age-matched normal template. Although

the epileptogenic origin was unclear in most patients, thisobservation confirms the central role that the thalamus playsin epilepsy.

Activation ResultsThe mode of action by which VNS suppresses epileptic

seizures is not fully understood and is currently underinvestigation by various approaches such as human electro-

physiologic studies (3,9,26), animal experimentation involving drugs (27), surgery (28), anatomopathologic techniques(29), and imaging studies with PET and SPECT (12,13,15).Two general theories on the operative mechanism of VNShave emerged from these research efforts (JO). First, thedirect connection theory hypothesizes that the anticonvul-sant action of VNS is caused by a threshold-raising effect of

the connections to the nucleus of the solitary tract and toother structures. The second is the concept that chronicstimulation of the vagus nerve increases the amount ofinhibitory neurotransmitters and decreases the amount ofexcitatory neurotransmitters.

The results this study have shown that in the 2-3 min after

an initial stimulation, the ipsilateral thalamus, hippocampus,and brain stem show reduced rCBF. The thalamic rCBF

decrease was consistently observed in 11 of the 12 patients,resulting in a mean thalamic VOI reduction of 5.6%. Onepatient showed a slightly increased perfusion (2%). No othercerebral regions showed the same marked perfusion change,although a significant change in left parietal perfusion wasnoted with the VOI approach. A posterior parietal perfusiondecrease at the inferior parietal lobule was also noted withthe SPM analysis, but at a less stringent height threshold ofP = 0.06 (uncorrected). These results suggest that decreased

perfusion of the left thalamus, hippocampus, and pons maybe involved in the mechanism of seizure suppression byVNS.

Experiments with somatosensory evoked potentials in1992 showed that VNS alters neuronal networks outside ofthe brain stem vagus system (26). It is believed that cervicalVNS causes increased trans-synaptic neurotransmission in

the nucleus of the tractus solitarius (NTS), nucleus of thespinal trigeminal tract, and other sites of vagus nerveterminals (12). The NTS projects to several nuclei of thethalamus and hippocampus (3,13). Most vagai afférentssynapse in the NTS, where both vagai fibers and axonsoriginating in the NTS project densely to the medullarreticular formation. This formation has polysynaptic ascending projections to the nucleus reticularis thalami, whichitself projects to many thalamic nuclei. The nucleus reticularis thalami can synchronize efferent activities of thalamo-

cortical relay neurons in different thalamic nuclei. A secondconnection between the NTS and the thalamus existsthrough the parabrachial nucleus of the pons, which projectsheavily to thalamic intralaminar nuclei. These nuclei projectdiffusely to cortical neurons with excitatory synaptic terminations and modulate excitability of the cortex (31). Therefore, anatomically underlying connections between thethalamus and NTS are certainly present, through which apossible decrease in cortical excitability may result.

Our findings agree with earlier results from thalamotomystudies in complex partial epilepsy, which showed significant reduction in seizure rate but were abandoned because ofsevere side effects (28). Evoked potential measurementshave also revealed that activity of spinothalamic neurons canbe depressed by vagai afferent stimulation (3). Functionalthalamic inhibition through VNS may induce this decreasedafferent fiber activity with resulting changes at some of thesynapses within particular structures.

The somatosensory response associated with the subjective paresthesia that was present in almost all patients mayconfound the results of VNS. Both the sign of the perfusionchange and the anatomic location (left cervical sensation isprojected to the right thalamus and right postcentral gyrus)form strong arguments against a merely sensory phenomenon in our results. Naritoku et al. (29) have shown in ratsthat the thalamus is involved in the true mechanism of VNSaction rather than through sham sensory stimulation.

Previous rCBF experiments with H215OPET on the effects

of VNS have been conducted by mainly 3 groups. Ko et al.(14) described results in 3 patients with commensurate

ECD SPECT IN VAGUSNERVESTIMULATION•Van Laere et al. 1151

isotope injection and stimulus administration, which resulted in activation of the contralateral thalamus, rightposterior temporal cortex, left putamen, and left anteriorcerebellum. These results were obtained in chronicallystimulated patients, 2 of whom had already undergoneablative surgery. Moreover, only 1 patient showed a reduction in seizure frequency, whereas the frequency increased inthe other 2 (+6% and +30% increase). In an expandedseries of 9 patients, the same authors showed that for chronicVNS (5-16 mo after initiation) reduction in seizure fre

quency best correlated with decreased CBF in the rightfusiform gyrus (75). In total, 13 regions of increased CBFand 36 regions of decreased CBF were identified.

Gameti et al. (78) studied the effect of left VNS in 5patients and found ipsilateral changes (undefined as decreaseor increase) in the thalamus and anterior cingulate gyrus. Inthis study, patients were also examined after several monthsof intermittent stimulation. However, one can argue thatchanges in rCBF measured early in the course of VNStherapy more accurately reflect acute VNS-induced changes

in regional synaptic activity and may better correlate withactivity in central pathways that have not been modified bylong-term adaptations as in chronic VNS.

Henry et al. (72) studied blood flow alterations in thebrain induced by VNS within 24 h of intermittent initialVNS. In their initial study of high- and low-intensity VNS(high-intensity is comparable with our study; low-intensityis 70-fold lower than our study), they found that rCBF

increases in the medulla and the right postcentral gyrus andincreases bilaterally in the hypothalami, thalami, insularcortex, and cerebellar hemispheres. Decreased rCBF wasfound in the hippocampus, amygdala, and posterior cingulate gyri. No values of activity changes were given. In afollow-up study, which correlated acute VNS-induced rCBF

alterations with chronic (3 mo) therapeutic response, thesame authors showed that bilateral increased thalamic CBFcorrelated with decreased seizures. In their series, only 2patients showed a seizure reduction of >50% and 6 showedseizure reduction of

be overconservative, possibly leading to false-negative results (20).

Generally, without a correction for multiple comparisons,SPM findings should only be reported descriptively because,otherwise, type I errors (false-positives) may be introduced.For this investigation, because preexisting studies haveshown involvement of medullar, thalamic, and limbic structures, the use of probability values that were not correctedfor multiple comparisons over the whole brain volume canbe justified in reporting the observed significant left thala-mar and hippocampal deactivation (12,13,15,18,29,37). Theactivation that was noted in bilateral frontal regions at theheight threshold corresponding to an uncorrected P = 0.01should be interpreted cautiously and can merely identify therequirement for further study because no a priori hypothesiswas anticipated. Possible explanations for frontal activationmay be of a stress-induced nature because for all patients itwas the initial onset of their stimulation.

Furthermore, in single-day, split-dose SPECT activationprotocols, the actual intensity of activity changes is underestimated because the second study in the split-dose designconsists of the sum of the baseline study (decayed over thetime between the 2 scans) and the actual second study. For a1:1 injection ratio with proportional scaling as in our study,this means that the actual intensity of the activation isapproximately 2 times higher.

Perfusion tracers such as "Tc-ECD show a swift blood-

brain barrier passage and a relatively large extraction in thebrain. For activation studies by a short-interval, split-dosetechnique, some caution is necessary because of washout(38,39). Corrections for inhomogeneous neuronal washoutare necessary only when scan-to-scan intervals surpass a fewhours (39). Because washout is less in the central brainregions (38,39), such an effect would increase relativeuptake values in the second scan, thereby damping theobserved thalamic inhibition.

Because of the limited SPECT resolution, the applicationof rigid body transformations in the analysis, and theintersubject differences in anatomy, it was not feasible toproject the obtained standard co-ordinates of significantlychanged voxels to the level of specific thalamic subregionsor regions in the brain stem, where the NTS and other nucleithat may be involved in VNS action are located (29). Futurenuclear medicine PET equipment, with resolution reachingthe order of 2 mm, may enable study of these areas (40).

CONCLUSION

VNS produces an effective decrease in seizure frequencyin patients with intractable epilepsy. rCBF measurementsbefore and immediately after initial stimulation indicate thatVNS produces acute effects, mainly by suppressing theipsilateral thalamus. Differences between this finding andprevious PET findings can be explained by a local temporalhemodynamic response during and after the stimulus. SPMresults were in agreement with results obtained by VOIanalysis and nonparametric testing but can give more

detailed spatial information. Finally, notwithstanding itslimitations of resolution and inability to perform single-day,multiple, repeated stimulation measurements, SPECT maybe suited to study both the acute and long-term efficacy ofVNS in larger groups of implanted patients.

ACKNOWLEDGMENTS

The authors gratefully acknowledge the logistical supportobtained from Nuclear Diagnostics Ltd., Sweden, and SunComputers, Belgium. This work was supported by 3 independent special research grants from Gent University and theFlemish Government (Byzonder Onderzoeks Fonds01104699, 01104495, and 011A099) and by grant 1.50.236.99 from the Fund for Scientific Research-Flanders.

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