Neurochemical Effects of Theta Burst Stimulation as Assessed by Magnetic Resonance Spectroscopy

The Neurochemical Effects of Theta Burst Stimulation as assessed by Magnetic Resonance Spectroscopy

*Stagg CJ1,2, Wylezinska M1,3, Matthews PM1,4, Johansen-Berg H1, Jezzard P1,

Rothwell JC5, Bestmann S5,6.

1. Centre for Functional Resonance Imaging of the Brain, University of Oxford,

Oxford, UK

2. Department of Clinical Neurology, University of Oxford, Oxford, UK

3. Biological Imaging Centre, Imaging Sciences Department, MRC Clinical

Sciences Centre, Imperial College London, Hammersmith Campus, London, UK

4. Department of Clinical Neurosciences, Imperial College, London and GSK

Clinical Imaging Centre, Hammersmith Hospital, London, UK

5. Sobell Department of Movement Neuroscience and Movement Disorders,

Institute of Neurology, University College, London, UK

6. Wellcome Trust Centre for Neuroimaging, Institute of Neurology, University

College, London, UK

* Corresponding Author

FMRIB Centre

Department of Clinical Neurology

John Radcliffe Hospital

Oxford, OX3 9DU

Tel: +44 (0)1865 222729; Fax: +44 (0)1865 222717

Email: [email protected]

Running Head: Theta Burst and Magnetic Resonance Spectroscopy

Articles in PresS. J Neurophysiol (April 1, 2009). doi:10.1152/jn.91060.2008

Copyright © 2009 by the American Physiological Society.

Theta Burst and Magnetic Resonance Spectroscopy

Abstract

Continuous Theta Burst Stimulation (cTBS) is a novel transcranial stimulation technique

that causes significant inhibition of synaptic transmission for up to an hour when applied

over the primary motor cortex (M1) in humans. Here we use Magnetic Resonance

Spectroscopy to define mechanisms mediating this inhibition by non-invasively

measuring local changes in the cortical concentrations of the GABA and

glutamate/glutamine (Glx). cTBS to the left M1 led to an increase in GABA compared to

stimulation at a control site without significant change in Glx. This direct evidence for

increased GABAergic interneuronal activity is framed in terms of a new hypothesis

regarding mechanisms underlying cTBS.

Keywords: motor cortex; plasticity; neurorehabilitation; GABA; transcranial

magnetic stimulation


Introduction

Repetitive Transcranial Magnetic Stimulation (rTMS) is an increasingly used non-

invasive method of stimulating neural pathways in the awake human brain. rTMS can

induce significant excitability changes within the cortex which outlast the period of

stimulation (Rossini and Rossi 2007). Of particular interest is the promise of rTMS for

clinical applications, such as neurorehabilitation after stroke, when facilitatory

stimulation to the affected hemisphere or inhibitory stimulation to the unaffected

hemisphere can improve motor function (Di Lazzaro et al. 2008a; Webster et al. 2006,

Hummel and Cohen 2006).

However, many of the reported effects of rTMS are short lasting, often highly variable or

weak, especially when applied to patients with a range of different neuropathologies

(Maeda et al. 2000a; Wassermann 2002). To address this problem, a more novel protocol

has been developed that induces more robust and less variable responses across

populations of subjects (Huang et al. 2005). Theta Burst Stimulation (TBS) uses 50Hz

trains of 3 TMS pulses, repeated every 200ms. When applied intermittently, TBS leads

to a robust increase in cortical excitability for up to 30 minutes. By contrast, continuous

TBS (cTBS) at a frequency of 5 Hz over a period of 40s (cTBS-600) leads to depression

of MEP amplitudes by up to 50% of the baseline amplitude that can be maintained for up

to 60 minutes (Huang et al. 2005). TBS also has robust effects in stroke patients (Di

Lazzaro et al. 2008b).


The mechanisms by which TBS exerts these effects on the cortex are not well defined.

There is some evidence that rTMS alters cortical excitability via changes in synaptic

strength (Chen et al. 1997; Cooke and Bliss 2006; Fitzgerald et al. 2006; Huang et al.

2005; Maeda et al. 2000b; Pascual-Leone et al. 1994). The TBS protocol is designed to

be more similar to paradigms used to induce Long Term Potentiation (LTP) and Long

Term Depression (LTD) in animal models (Hess et al. 1996; Huemmeke et al. 2002;

Larson and Lynch 1986; Vickery et al. 1997), which, in the cortex, depend primarily on

the GABAergic system, with some glutamatergic involvement (Komaki et al. 2007;

Trepel and Racine 2000). However, in humans, the direct effects of TBS on synaptic

transmission are commonly measured indirectly, through administration of neuroactive

compounds (Huang et al. 2007).

By contrast, Magnetic Resonance Spectroscopy (MRS) measures GABA and glutamate

levels directly and non-invasively. For example, previous MRS studies have

demonstrated reductions of GABA in the human motor cortex while subjects learned a

motor skill (Floyer-Lea et al. 2006), after facilitatory stimulation of the motor cortex

using an electrical stimulation technique, transcranial direct current stimulation (tDCS)

(CJ Stagg, unpublished observations), after acute deafferentation (Levy et al. 2002) or in

a number of neurological conditions (Levy and Hallett, 2002).

Here, we wish to test whether the effects of cTBS are mediated by changes in the local

activity of inhibitory interneuronal cortical pathways in the primary sensorimotor cortex

using GABA optimised MRS. We measured changes in concentrations of GABA and


glutamate/glutamine (Glx) following cTBS over the “hand” area of the motor cortex

(verum cTBS) relative to those with stimulation over the vertex of the cranium (control

cTBS).

Materials and Methods

Subjects 16 male subjects (mean age 27.5 years; range: 21-44) gave their informed consent to

participate in the study in accordance with local ethics committee approval. All subjects

were right-handed as assessed by the Edinburgh Handedness Inventory (Oldfield 1971).

TMS Paradigm All TMS was given using a hand-held figure-of-eight coil (70mm standard coil, Magstim

Co., Whitland, Dyfed, UK), connected to Magstim Rapid2 stimulator. The cortical

representation of the right first dorsal interosseous muscle (FDI) within the left primary

motor cortex (M1) was identified using single biphasic pulse TMS and the scalp position

overlying this marked – referred to as the “hotspot”. The active motor threshold (AMT)

was then established for each subject, defined as the minimum intensity of stimulation

required to produce an MEP of greater than 200 μV on five out of ten trials from the

contralateral FDI when the subject was maintaining a voluntary contraction of about 20%

of maximum. Electromyographic responses were recorded using Ag/AgCl electrodes in a

belly-tendon montage. The pattern of TBS consisted of bursts containing 3 pulses at

50Hz and an intensity of 80% AMT repeated at 200ms intervals for 40s (i.e. 600 pulses).


Verum stimulation was targeted to the previously identified hotspot, with the coil held

tangentially to the skull, orientated at 45° the midsagittal axis, inducing lateromedial

current flow. Control stimulation was given to the control-site at the vertex (Cz).

Subjects were instructed to relax their hand at all times during the experiment, once the

AMT had been recorded and were blinded as to which TMS condition they received.

GABA MRS

A 3T Siemens/Varian MRI system was used. Sagittal and axial T1 weighted scout

images were acquired and used to place a 2x2x2cm voxel of interest by hand over the left

precentral knob, a landmark previously described to identify the hand motor

representation in the human brain (Yousry et al. 1997). The placement of a typical voxel

is shown in Figure 1B. To assess the creatine and NAA linewidths, a standard PRESS

sequence was used to acquire an unedited spectrum with TE = 68ms and 64 averages.

Water was suppressed at 4.7ppm using a technique similar to WET (Ogg et al. 1994).

The MEGA-PRESS (Mescher et al. 1998) sequence was then used to allow simultaneous

spectral GABA editing, three-dimensional voxel localisation and water suppression, as

reported previously (Floyer-Lea et al. 2006). With the MEGA-PRESS scheme, GABA at

3 ppm was detected through J difference spectral editing and use of the frequency

selective double-banded 180o pulse.

A selective double-banded 180° pulse was created from 20-ms Gaussian pulses. The

frequency of the first band of this pulse was set to 4.7 ppm to suppress water. The second

band was alternated between 1.9 ppm, the resonance frequency of C3 protons (strongly


coupled to the observed C4 GABA protons) and 3.0 ppm (odd-numbered acquisitions),

and 7.5 ppm (even numbered acquisitions), which is symmetrically disposed about the

water resonance to equalize off resonance effects. During the odd-numbered acquisition

the GABA C4 (triplet) resonance at 3.0 ppm was fully refocused, whereas during the

even-numbered acquisitions this peak was not refocused, but phase-modulated so that the

outer triplet signals were inverted at echo time TE = 68 ms. The difference between

spectra collected during even and odd acquisitions (at TE = 68 ms) revealed the edited

GABA spectrum without the larger overlapping creatine resonance.

A metabolite-nulled spectra was acquired using a modified MEGA-PRESS editing

sequence with pre-inversion module consisting of inversion recovery, pulse and recovery

time (TI) adjusted to minimize the creatine peak (TI = 0.720s). In addition, a T1-

weighted anatomical image was acquired with each spectral set to allow for correction for

varying grey matter volumes within the voxel (3D Turbo Flash, 128x 2mm axial slices,

TR =12ms, TE = 5ms, TI = 200ms, flip angle = 8˚, FOV = 256x256, matrix = 128x128).

Experimental Protocol

The motor hotspot was identified prior to the baseline scan. A 256 acquisition-baseline

GABA-optimised MRS was acquired for each subject. The subjects were then removed

from the scanner and TBS performed in a separate room. Following TBS the subject was

asked to keep their hand relaxed at all times and interactions with them were kept to a

minimum. The subject was then re-positioned into the scanner, with assistance from the


experimenters to avoid physical activity of the hand and arm (Huang et al., 2008). A

voxel as close as possible to the baseline voxel was then visually identified, from which a

512-acquisition MRS spectrum was obtained. The MR-operator was naïve with respect

to the TMS stimulation condition. MRS acquisition started approximately 20 minutes

after TBS was completed (figure 1A). Participants watched a nature DVD whilst in the

MR scanner to maintain a constant level of wakefulness.

MRS Analysis

In order to assess the overlap between the pre- and post-stimulation voxels, the voxels

were registered in standard space using FLIRT, part of the FMRIB Software Library

(www.fmrib.ox.ac.uk/fsl) (Jenkinson et al. 2002; Smith et al. 2004). The overlap

between the two voxels was then compared with the volume of the pre-stimulation voxel

to ensure meaningful co-localisation. This was confirmed (see results) for all subjects.

Quantitative analyses of the spectra were performed using the jMRUI software package

version 2.2 (http://www.mrui.uabes/mrui). The free induction decay signal (FID) was

first corrected for any non-zero DC offset and the signal was smoothed using a 2Hz

Lorentzian filter. The residual water signal was then filtered out by fitting and removing

Gaussian peaks around the water frequency using Singular Value Decomposition

techniques (van den Boogaart 1997; van den Boogaart et al. 1994; Vanhamme et al.

1997). The spectrum was then phased with respect to both the 0th and 1st order phase.


All spectra were analysed using AMARES, a non-linear least square fitting algorithm

operating in the time domain (Vanhamme et al. 1997). Peak fitting for the GABA and

Glx (a combined measure of glutamate and glutamine) spectra was performed using

Lorentzian curves with the linewidth constrained to that of the Creatine resonance in the

non-edited spectrum. The GABA and the glutamate/glutamine resonances were both

fitted with 2 Lorentzian peaks. The linewidth of the inverted NAA resonance was

constrained to the linewidth of NAA in the unedited spectrum and a single Lorentzian

curve was fitted to this peak. The amplitudes of both GABA peaks were summed to give

a signal intensity for GABA, likewise summing was performed for the Glx peak.

To correct for the expected contribution from mobile brain macromolecules (MM; which

include cytosolic proteins) the GABA-nulled spectrum was analysed as above, and

amplitude of the peak resonating at 3ppm calculated. This was then deducted from the

signal intensity of the GABA resonance.

All results presented for both GABA and Glx given here are as a ratio to NAA, the

simultaneously acquired reference peak. Because we are looking for percentage changes

within subject, no correction has been made for editing efficiency, nor number of

equivalent protons.

Results

Data for the structural scan from one subject was corrupted and spectra from that subject

were discarded. All spectral acquisitions were of adequate quality, defined a priori based


on an NAA linewidth of <10 Hz. Analyses were performed on the remaining 7 subjects

in the verum group and 8 subjects in the control group. There was no significant

difference between the ages of subjects in the two groups (Wilcoxon Two Sample Test,

W = 67.5, p < 0.99). The average active motor threshold was 48.7 ± 5.9% of maximal

stimulator output (MSO). The average stimulation intensity was 40 ± 5% MSO, with no

significant difference between groups (Wilcoxon Two Sample Test, W = 55, p < 0.18).

Good registration of voxels was achieved within all subjects. On average the pre- and

post-cTBS voxels within individual subjects overlapped in volume by a mean 77%.

There were no significant difference between grey matter volume contained within the

pre- compared to the post-cTBS voxels (Wilcoxon Two Sample Test, W = 122, p < 0.95)

and the grey matter content for pre- versus post-cTBS voxels was highly correlated

across subjects (r = 0.93, p < 0.01). A representative pair of voxels is shown in figure 2A

and an overall summary in figure 2B.

A typical localised spectrum from the sensorimotor cortex is shown in Figure 3A. A

Mann-Whitney U-test confirmed that there was a significant increase in GABA

concentrations between verum and control stimulations (verum pre-stimulation corrected

GABA:NAA ratio [median (IQ range)] 0.23 (0.16 - 0.27), post-stimulation 0.26 (0.16 -

0.30); control pre-stimulation corrected GABA:NAA ratio 0.18 (0.14 - 0.26), post-

stimulation 0.17 (0.14 – 0.19); median change after verum stimulation = 11.0% (2.4 -

16.44); median change after control stimulation, 2.9% (-4.17 - 9.33); Wilcoxon Two

Sample Test, W = 46, p < 0.04) (Figure 3B) and no significant difference in Glx


concentrations (verum pre-stimulation Glx:NAA ratio [median (IQ range)] 0.06 (0.05 -

0.09), post-stimulation 0.08 (0.05 - 0.08); verum median change = 16.1% (-25.1 - 39.1);

control median change 6.8% (-13.7 - 87.8); Wilcoxon Two Sample Test, W = 36, p <

0.68).

To further rule out confounding changes in tissue water, we also measured the absolute

concentration of Cr pre and post- stimulation. There was no significant change in the Cr

concentration seen after stimulation (Figure 3C) (verum Cr concentration pre-stimulation

[median (IQ range)] 112 (92 – 123) [arbitrary units], post-stimulation 106 (83 – 117)

median change = -0.02 % (-0.1 – 0.03) control Cr concentration pre-stimulation 106 (100

– 114), post-stimulation 96 (91 – 100) median change –0.15% (-0.26 - 0.04); Wilcoxon

Two Sample Test, W = 33, p < 0.61).

Discussion

Continuous TBS to the M1 increases GABA concentration when compared with control

stimulation, without any significant effect on Glx levels. GABAergic activity therefore

may be a mechanism by which long-lasting after-effects of TBS on corticospinal

excitability are generated. This is consistent with pre-clinical data suggesting the

importance of the GABAergic mechanisms in LTD-like phenomena within the neo-

cortex in freely moving animals (Hess et al. 1996; Komaki et al. 2007; Trepel and Racine

2000). More indirectly, it may be related to the decreases in local sensorimotor cortex


GABA concentrations during successful motor learning in humans (Floyer-Lea et al.

2006).

GABA is produced in neurons by decarboxylation of glutamate by the enzyme glutamic

acid decarboxylase (GAD65). The GAD65 isoform is associated with synaptic vesicles and

is likely to be involved in synthesising GABA for neurotransmission (Martin et al. 1991a;

Martin et al. 1991b; Martin and Rimvall 1993). In contrast, the GAD67 isoform

distributed more widely in the cytoplasm and is thought to be important in synthesising

GABA for cytosolic use. GAD is the rate-limiting step in production of GABA. In vitro,

neuronal activity is associated with increases in the active isoform of GAD65 (de Graaf et

al. 2003). In neural stem cell derived cultures, depolarization of the neurones also leads to

an increase in the active isoform of GAD65 (Gakhar-Koppole et al. 2008). Induction of

synthesis therefore likely enables enhanced GABAergic activity in vivo.

It is important to note that MRS is sensitive only to changes in the overall concentration

of a neurotransmitter and cannot inform our understanding of changes within the

receptors at the synapses. Specifically, the lack of a change in this measure of Glx does

not contradict previous pharmacological studies that show abolition of the effects of

cTBS after NMDA-receptor antagonism (Huang et al., 2007), but instead strongly

suggests that these changes are due to changes in the NMDA receptors themselves.

The after-effects of cTBS on motor cortex are commonly assessed from their effects on

the amplitude of EMG responses (MEPs) evoked in a small intrinsic hand muscle by a


standard single TMS pulse (Huang et al. 2005). At intensities commonly used for TBS,

the effect of TMS is cortical, and thus results from stimulation of either excitatory or

inhibitory synaptic inputs to layer-V pyramidal neurones (Di Lazzaro et al. 1998). A

reduced MEP after cTBS therefore indicates either a reduction in the net efficiency of

excitatory stimulation by the TMS pulse or an increase in intracortical inhibition.

Our results here suggesting increased GABAergic inhibition contributes to the after-

effects of TBS were unexpected. Previous work has shown that the effects of cTBS are

abolished by administration of memantine at concentrations sufficient to antagonise

NMDA function in the human brain (Huang et al. 2007; Teo et al. 2007). Thus, it has

usually been assumed that the after-effects are due to an action on glutamatergic

transmission, which leads to a reduction in cortical excitability via an LTD-like action on

the excitatory synapses activated by the TMS test pulses.

Integration of the neuropharmacological results with the MRS data suggests a new

hypothesis regarding cTBS action. cTBS is applied at a low intensity (80% active motor

threshold, AMT), which is below the threshold for activating the excitatory inputs to

pyramidal neurons (Ziemann et al. 1996). The intensity is the same as that used in a

common double pulse paradigm to assess short interval intracortical inhibition (SICI) in

the motor cortex (Kujirai et al. 1993). To elicit SICI, two TMS pulses are applied

through the same coil with the first pulse being subthreshold intensity (usually 80%

AMT) and the second pulse large enough on its own to elicit an MEP. If the interval

between the pulses is between 1-5 ms then the MEP is suppressed. Pharmacological


studies suggest the effect is due to activation of the GABAA-ergic neurons by the first

pulse (Muller-Dahlhaus et al. 2008; DiLazzaro et al. 2008b; Werhahn et al. 1999;

DiLazzaro et al. 2000). Whether the GABAA neurons are activated directly by the TMS

pulse or indirectly via an excitatory synaptic input is unclear; there is some evidence that

it may be the latter (Bestmann et al. 2003).

Whatever the precise mechanism, these data imply that cTBS (600 stimuli) at 80% AMT

activates a population of cortical GABAA-ergic interneurons. The increase in GABAergic

activity is then sustained by induction of GAD65 and subsequent increased GABA

concentration within the cytoplasm of the GABAA-ergic interneuron.

However, this simple formulation does not account for the decrease in SICI (a TMS

measure of relative GABAergic transmission) found with cTBS (Huang et al. 2005,

2008). To account for this, we propose an extension of this simple formulation into a

mixed model in which effects are mediated by changes in both glutamatergic and

GABAergic signaling within local excitation-inhibition networks (Logothetis, 2008). The

excitability of the cortical output neurons (as reflected by the MEP) is controlled by

associative mechanisms of LTD, whereby both NMDA-receptor dependent mechanisms

and GABA input control excitability (Tsumoto, 1992).

There is one more important aspect that may help to explain our present findings. LTD of

GABAA synapses is predominantly presynaptic, being mediated through

endocannabinoids (Tsumoto, 1992). On the other hand, LTD at glutamatergic synapses is


more likely to involve postsynaptic changes (Hess & Donoghue, 1996). Previous work

shows that cTBS reduces the effectiveness of excitatory inputs to MEP generators in

motor cortex, as well as the effectiveness of the circuits mediating short-intracortical

inhibition (SICI) (Huang et al., 2008). In both cases, cTBS is likely to provoke activity in

GABAergic and glutamatergic circuits that may be followed by an increased synthesis of

both transmitters. However, upon termination of cTBS, presynaptic LTD at GABAergic

neurons prevents further GABA release. This leads to increased pre-synaptic GABA

levels that in turn increase the MRS signal. On the other hand, presynaptic release of

glutamate is unchanged and no accumulation of glutamate should occur. Consequently,

no changes in Glx should occur.

This explanation would make the previous pharmacological studies suggesting that the

after-effects of cTBS are NMDA-receptor dependent (Huang et al. 2007), consistent with

the results from this study showing an increase in GABA activity. The reduction in SICI

can be accounted for as this phasic test of relative GABAA activity in the paired pulse

paradigm would be less effective in the context of a baseline of increased, ongoing,

inhibition. Although this cannot be proven from this data, this model provides a

framework for further testing in future experiments.

There are limitations to our experiment. The measure of total GABA within the voxel

gives no information concerning subcellular localisation, and therefore our interpretation

can only be speculative. As sensitivity did not allow a full relaxation time

characterisation, it remains possible that the increases in apparent concentration arise


simply from redistribution of GABA from pools in which it is relatively immobilized,

e.g., by association with macromolecules (MM) and therefore MR “invisible” (Matthews

et al. 1986). However, such large relaxation time changes with short-term changes in

functional state would be unprecedented as far as we are aware. It also is possible that

the results reflect a change in the volume of the cells within the voxel. However, we

have controlled for this by considering the GABA: NAA ratio rather than using GABA

absolute quantification. In addition, there is no significant change in the creatine

concentration after stimulation.

There are some questions raised by the current study that should be addressed in future

investigations. First, due to the constraints of the technique we have only acquired data

from the brain tissue in the stimulated SMC and therefore a future study is needed to

clearly distinguish the local and more general effects of tDCS on neurotransmitter levels.

In addition, we are unable to determine the direct relationship between neurotransmitter

changes as assessed by MRS and neurophysiological changes as assessed by TMS.

However, at this point the experiments required to investigate this relationship remain

technically challenging. Subjects were moved out of the scanner for stimulation and

there was a delay of approximately 20 minutes between end of stimulation and the start

of MRS measurements, which then demanded a further 20 minutes. Shorter-lived

neurochemical changes would not be able to be defined. However, the

neurophysiological after-effects of 600 pulses of cTBS, as applied in the present study,

are relatively stable for up to an hour (Huang et al. 2008; Huang et al. 2005), suggesting

that the dynamics of the underlying neurochemical changes also occur over a similar


period. In addition it would be of interest to investigate the effects of intermittent TBS in

a similar study. There is evidence that iTBS and cTBS affect different intracortical

circuits (Di Lazzaro et al. 2005; 2008a) so different effects might be expected.

A significant variability in the Glx measure was seen in both stimulation conditions. In

our experience this is often seen in MRS studies and may represent an interaction

between the resonance and the neighbouring water peak. However, although this

interaction would be expected to add variance to the signal, as seen here, no trend

towards a significant change in Glx measures with cTBS is seen.

cTBS to the contralesional hemisphere is a promising tool for neurorehabilitation in

chronic stroke. In many contexts, inhibitory stimulation to the unaffected hemisphere has

led to more robust increases in cortical excitability and motor function on the affected

side (Di Lazzaro et al. 2008b; Fregni et al. 2005). TBS, in particular, is a stimulation

technique, which employs low intensities and is well tolerated in both healthy controls

(Huang and Rothwell 2004) and stroke patients (Di Lazzaro et al. 2006; Talelli et al.

2007). This study adds direct evidence that TBS induces LTP- and LTD-like plasticity in

the human motor cortex (Huang et al. 2005). It has allowed a more refined hypothesis

regarding the mechanism of action that can be tested in future experiments. With a

stronger neurophysiological rationale, it can be applied with greater confidence in

therapeutic trials (Di Lazzaro et al. 2008a; Fregni et al. 2005), taking advantage of its

excellent tolerability (Di Lazzaro et al. 2006; Huang and Rothwell 2004; Talelli et al.

2007).


Acknowledgements

CJS thanks the Department of Clinical Neurology and the MRC for support through a

departmentally allocated MRC Studentship. SB was funded by the Wellcome Trust

(grant number: 076358/Z/05/Z). JCR was supported by the Medical Research Council

(MRC). HJB was funded by the Wellcome Trust. PMM gratefully thanks the MRC for

personal and laboratory support during most of the period of this study. He currently is an

employee of GlaxoSmithKline.


Figure Legends

Figure 1. A. Experimental Protocol. A baseline MRS acquisition was performed after

the motor hotspot had been identified in each subject. After this was complete, the

subjects were removed from the scanner and a 40s train of TBS pulses were applied,

either to the motor hotspot (M1; verum stimulation) or to the vertex (Cz; control

stimulation). The subjects were then repositioned in the scanner and a post-stimulation

MRS spectrum was acquired. This commenced after a period of approximately 20

minutes of scanner setup. B. Typical 2x2x2cm voxel placed within the hand region of

the left primary motor cortex.

Figure 2. A. Typical locations for the pre-stimulation voxel (yellow) and the post-

stimulation voxel (blue). The substantial overlap between the two shown is shown in

green. B. Plot showing the high correlation between the amount of grey matter within

the voxel of interest before and after stimulation (r2= 0.88, p < 0.01).

Figure 3. A. Typical GABA-optimised spectrum acquired. Insets show representative

GABA and NAA peaks before and after stimulation in the same subject. Glx: composite

measure of glutamate and glutamine. B. Percentage change in GABA:NAA from

baseline. Verum stimulation leads to a significant increase in GABA:NAA ratio,

compared with control. C. Percentage change in Creatine signal for baseline. There is

no significant change with either stimulation condition.


References

Chen R, Classen J, Gerloff C, Celnik P, Wassermann EM, Hallett M, and Cohen

LG. Depression of motor cortex excitability by low-frequency transcranial magnetic

stimulation. Neurology 48: 1398-1403, 1997.

Cooke SF, and Bliss TVP. Plasticity in the human central nervous system. Brain 129:

1659-1673, 2006.

de Graaf R, Mason G, Patel A, Behar KL, and Rothman DL. In vivo 1H-[13C]-NMR

spectroscopy of cerebral metabolism. NMR in Biomedicine 16: 339-357, 2003.

Di Lazzaro V, Dileone M, Profice P, Pilato F, Cioni B, Meglio M, Capone F, Tonali

PA, and Rothwell JC. Direct Demonstration That Repetitive Transcranial Magnetic

Stimulation Can Enhance Corticospinal Excitability in Stroke. Stroke 37: 2850-2853,

2006.

Di Lazzaro V, Restuccia D, Oliviero A, Profice P, Ferrara L, Insola A, Mazzone P,

Tonali P, and Rothwell JC. Magnetic transcranial stimulation at intensities below active

motor threshold activates intracortical inhibitory circuits. Exp Br Res 119: 265-268, 1998.

Di Lazzaro V, Pilato F, Dileone M, Profice P, Capone F, Ranieri F, Musumeci G,

Cianfoni A, Pasqualetti P, and Tonali PA. Modulating cortical excitability in acute

stroke: A repetitive TMS study. Clin Neurophysiol 119: 715-723, 2008a.

Di Lazzaro V, Pilato F, Dileone M, Profice P, Oliviero A, Mazzone P, Insola A,

Ranieri F, Meglio M, Tonali PA, and Rothwell JC. The physiological basis of the

effects of intermittent theta burst stimulation of the human motor cortex. J Physiol 586:

3871-3879, 2008b.


Di Lazzaro V, Oliviero A, Meglio M, Cioni B, Tamburrini G, Tonali P, and

Rothwell JC. Direct demonstration of the effect of lorazepam on the excitability of the

human motor cortex. Clin Neurophysiol 111: 794-799, 2000.

Di Lazzaro V, Pilato F, Saturno E, Oliviero A, Dileone M, Mazzone P, Insola A,

Tonali PA, Ranieri F, Huang YZ, and Rothwell JC. Theta-burst repetitive transcranial

magnetic stimulation suppresses specific excitatory circuits in the human motor cortex.

Journal of Physiology 565: 945-950, 2005.

Fitzgerald PB, Fountain S, and Daskalakis ZJ. A comprehensive review of the effects

of rTMS on motor cortical excitability and inhibition. Clin Neurophysiol 117: 2584-2596,

2006.

Floyer-Lea A, Wylezinska M, Kincses T, and Matthews P. Rapid Modulation of

GABA Concentration in Human Sensorimotor Cortex During Motor Learning. J

Neurophysiol 95: 1639-1644, 2006.

Fregni F, Boggio P, Mansur C, Wagner T, Ferreira M, Lima M, Rigonatti S,

Marcolin M, Freedman S, Nitsche M, and Pascual-Leone A. Transcranial direct

current stimulation of the unaffected hemisphere in stroke patients. Neuroreport 16:

1551-1555, 2005.

Gakhar-Koppole N, Bengtson CP, Parlato R, Horsch K, Eckstein V, and Ciccolini F.

Depolarization promotes GAD 65-mediated GABA synthesis by a post-translational

mechanism in neural stem cell-derived neurons. Eur J Neurosci 27: 269-283, 2008.

Hess G, Aizenmann C, and Donoghue JP. Conditions for the induction of long-term

potentiation in layer II/III horizontal connections of the rat motor cortex. J Neurophysiol

75: 1765-1778, 1996.


Hess G, and Donoghue JP. Long-term depression of horizontal connections in rat motor

cortex. European Journal Neuroscience 8: 658-665, 1996.

Huang Y-Z, Rothwell JC, Edwards MJ, and Chen R-S. Effect of Physiological

Activity on an NMDA-Dependent Form of Cortical Plasticity in Human. Cereb Cortex

18: 563-570, 2008.

Huang YZ, Chen RS, Rothwell JC, and Wen HY. The after-effect of human theta burst

stimulation is NMDA receptor dependent. Clin Neurophysiol 118: 1028-1032, 2007.

Huang YZ, Edwards MJ, Rounis E, Bhatia KP, and Rothwell JC. Theta burst

stimulation of the human motor cortex. Neuron 45: 201-206, 2005.

Huang YZ, and Rothwell JC. The effect of short-duration bursts of high-frequency,

low-intensity transcranial magnetic stimulation on the human motor cortex. Clin

Neurophysiol 115: 1069-1075, 2004.

Huemmeke M, Eysel U, and Mittmann T. Metabotropic glutamate receptors mediate

expression of LTP in slices of rat visual cortex. Eur J Neurosci 15: 1641-1645, 2002.

Hummel F, and Cohen L. Non-invasive brain stimulation: a new strategy to improve

neurorehabilitation after stroke? The Lancet Neurology 5: 708-712, 2006.

Jenkinson M, Bannister P, Brady J, and Smith S. Improved optimisation for the robust

and accurate linear registration and motion correction of brain images. NeuroImage 17:

825-841, 2002.

Komaki A, Shahidi S, Lashgari R, Haghparast A, Malakouti SM, and Noorbakhsh

SM. Effects of GABAergic inhibition on neocortical long-term potentiation in the

chronically prepared rat. Neurosci Lett 422: 181-186, 2007.


Larson J, and Lynch G. Induction of Synaptic Potentiation in Hippocampus by

Patterned Stimulation Involves Two Events. Science 232: 985-988, 1986.

Levy L, and Hallett M. Impaired brain GABA in focal dystonia. Annals of Neurology

51: 93-101, 2002.

Levy LM, Ziemann U, Chen R, and Cohen L. Rapid Modulation of GABA in

Sensorimotor Cortex Induced by Acute Deafferentation. Annals of Neurology 52: 755-

761, 2002.

Maeda F, Keenan J, Tormos J, Topka H, and Pascual-Leone A. Interindividual

variability of the modulatory effects of repetitive transcranial magnetic stimulation on

cortical excitabiliy. Exp Brain Res 133: 425-430, 2000a.

Maeda F, Keenan J, Tormos J, Topka H, and Pascual-Leone A. Modulation of

corticospinal excitability be repetitive transcranial magnetic stimulation Clin

Neurophysiol 111: 800-805, 2000b.

Martin DL, Martin SB, Wu SJ, and Espina N. Cofactor interactions and the regulation

of glutamate decarboxylase activity. Neurochem Res 16: 243-249, 1991a.

Martin DL, Martin SB, Wu SJ, and Espina N. Regulatory properties of brain

glutamate decarboxylase (GAD): the apoenzyme of GAD is present principally as the

smaller of two molecular forms of GAD in brain. J Neurosci 11: 2725-2731, 1991b.

Martin DL, and Rimvall K. Regulation of γ-Aminobutyric Acid Synthesis in the Brain.

J Neurochem 60: 395-407, 1993.

Mescher M, Merkle H, Kirsch J, Garwood M, and Gruetter R. Simultaneous in vivo

spectral editing and water suppression. NMR in biomedicine 11: 266-272, 1998.


Muller-Dahlhaus MFJ, Liu Y, and Ziemann U. Inhibitory circuits and the nature of

their interactions in the human motor cortex a pharmacological TMS study. J Physiol

586: 495-514, 2008.

Ogg RJ, Kingsley RB, and Taylor JS. WET, a T1- and B1-Insensitive Water-

Suppression Method for in Vivo Localized 1H NMR Spectroscopy. J Mag Res B 104: 1-

10, 1994.

Oldfield RC. The assessment and analysis of handedness: the Edinburgh inventory.

Neuropsychologia 9: 97-113, 1971.

Pascual-Leone A, Valls-Sole J, Wassermann EM, and Hallett M. Responses to rapid-

rate transcranial magnetic stimulation of the human motor cortex. Brain 117: 847-858,

1994.

Rossini PM, and Rossi S. Transcranial magnetic stimulation: Diagnostic, therapeutic,

and research potential. Neurology 68: 484-488, 2007.

Smith SM, Jenkinson M, Woolrich MW, Beckmann CF, Behrens TEJ, Johansen-

Berg H, Bannister PR, De Luca M, Drobnjak I, Flitney DE, Niazy RK, Saunders J,

Vickers J, Zhang Y, De Stefano N, Brady JM, and Matthews PM. Advances in

functional and structural MR image analysis and implementation as FSL. NeuroImage

23: S208-S219, 2004.

Talelli P, Greenwood R, and Rothwell JC. Exploring theta burst stimulation as an

intervention to improve motor recovery in chronic stroke. Clin Neurophysiol 118: 333-

342, 2007.

Trepel C, and Racine RJ. GABAergic Modulation of Neocortical Long-Term

Potentiation in the Freely Moving Rat. Synapse 35: 120-128, 2000.


Tsumoto T. Long-term protentiation and long-term depression in the neocortex. Progress in Neurobiology 39: 209-228, 1992. van den Boogaart A. MRUI Manual V. 96.3 A user's guide to the Magnetic Resonance

User Interface Software Package. Delft: Delft Technical University Press, 1997.

van den Boogaart A, van Ormondt D, Pijnappel W, De Beer R, and Ala-Korpela M.

In: Mathematics in Signal Processing III, edited by McWhirter J. Oxford: Clarendon

Press, 1994, p. 175-195.

Vanhamme L, Van den Boogaart A, and Van Huffel S. Improved method for accurate

and efficient quantification of MRS data with use of prior-knowledge. J Mag Res 129:

35-43, 1997.

Vickery R, Morris S, and Bindman L. Metabotropic glutamate receptors are involved

in long-term potentiation in isolated slices of rat medial frontal cortex. J Neurophysiol 78:

3039-3046, 1997.

Wassermann EM. Variation in the response to transcranial magnetic brain stimulation in

the general population. Clin Neurophysiol 113: 1165-1171, 2002.

Webster BR, Celnik PA, and Cohen LG. Noninvasive Brain Stimulation in Stroke

Rehabilitation. NeuroRX 3: 474-481, 2006.

Werhahn KJ, Kunesch E, Noachtar S, Benecke R, and Classen J. Differential effects

on motorcortical inhibition induced by blockade of GABA uptake in humans. J Physiol

517: 591-597, 1999.

Yousry T, Schmid U, Alkadhi H, Schmidt D, Peraud A, Buettner P, and Winkler P.

Localization of the motor hand area to a knob on the precentral gyrus. Brain 120: 141-

157, 1997.


Ziemann U, Rothwell JC, and Ridding MC. Interaction between intracortical inhibition

and facilitation in human motor cortex. J Physiol 496: 873-881, 1996.

Neurochemical Effects of Theta Burst Stimulation as Assessed by Magnetic Resonance Spectroscopy

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