Longitudinal effects of rTMS on neuroplasticity in chronic treatment-resistant depression

Abstract

Major depressive disorder (MDD) is amongst the most prevalent of psychiatric disorders. Unfortunately, a third of patients will not respond to conventional treatments and suffer from treatment-resistant depression (TRD). Repetitive transcranial magnetic stimulation (rTMS) has been proven effective in treating TRD. The research suggests that rTMS acts via neuroplastic effects on the brain, which can be measured by changes in hippocampal and amygdala volume as well as cortical thickness. This sham-controlled study investigates longitudinal effects of rTMS on the volumes of the hippocampus and amygdala and cortical thickness in patients with chronic TRD. 31 patients received 20 sessions of high-frequency rTMS (N = 15) or sham treatment (N = 16) over the left dorsolateral prefrontal cortex during 4 consecutive weeks. Using structural magnetic resonance imaging, we investigated longitudinal treatment effects on hippocampus and amygdala volume as well as thickness of the paralimbic cortex. We found no clinical differences between the active and sham rTMS group. Longitudinal changes in hippocampal and amygdala volume did not differ significantly, although males showed a significant decrease in left amygdala volume, irrespective of treatment group. Changes in cortical thickness of the paralimbic cortex differed significantly between the active and sham groups. Most notably, the increase in cortical thickness of the isthmus of the left cingulate gyrus was greater in the active as compared to the sham rTMS group. Our data suggest that rTMS can induce neuroplastic changes, particularly in cortical thickness, independent of treatment response. We also found longitudinal changes in amygdala volume in males. For clinical effects to follow these neuroplastic effects, more intensive rTMS treatment might be needed in chronically depressed patients.
Trial registration number: ISRCTN 15535800, registered on 29-06-2017.

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European Archives of Psychiatry and Clinical Neuroscience (2021) 271:39–47
https://doi.org/10.1007/s00406-020-01135-w
ORIGINAL PAPER
Longitudinal effects ofrTMS onneuroplasticity inchronic
treatment‑resistant depression
IrisDalhuisen1,6 · EvelineAckermans1,2· LiekeMartens1· PeterMulders1,7· JoeyBartholomeus4· AlexdeBruijn2,3·
JanSpijker1,2,5· PhilipvanEijndhoven1,6,7· IndiraTendolkar1,6,8
Received: 30 January 2020 / Accepted: 27 April 2020 / Published online: 9 May 2020
© The Author(s) 2020
Abstract
Major depressive disorder (MDD) is amongst the most prevalent of psychiatric disorders. Unfortunately, a third of patients
will not respond to conventional treatments and suffer from treatment-resistant depression (TRD). Repetitive transcranial
magnetic stimulation (rTMS) has been proven effective in treating TRD. The research suggests that rTMS acts via neuro-
plastic effects on the brain, which can be measured by changes in hippocampal and amygdala volume as well as cortical
thickness. This sham-controlled study investigates longitudinal effects of rTMS on the volumes of the hippocampus and
amygdala and cortical thickness in patients with chronic TRD. 31 patients received 20 sessions of high-frequency rTMS
(N = 15) or sham treatment (N = 16) over the left dorsolateral prefrontal cortex during 4 consecutive weeks. Using structural
magnetic resonance imaging, we investigated longitudinal treatment effects on hippocampus and amygdala volume as well as
thickness of the paralimbic cortex. We found no clinical differences between the active and sham rTMS group. Longitudinal
changes in hippocampal and amygdala volume did not differ significantly, although males showed a significant decrease
in left amygdala volume, irrespective of treatment group. Changes in cortical thickness of the paralimbic cortex differed
significantly between the active and sham groups. Most notably, the increase in cortical thickness of the isthmus of the left
cingulate gyrus was greater in the active as compared to the sham rTMS group. Our data suggest that rTMS can induce neu-
roplastic changes, particularly in cortical thickness, independent of treatment response. We also found longitudinal changes
in amygdala volume in males. For clinical effects to follow these neuroplastic effects, more intensive rTMS treatment might
be needed in chronically depressed patients.
Trial registration number: ISRCTN 15535800, registered on 29-06-2017.
Keywords Depression· rTMS· Neuroplasticity· Amygdala· Hippocampus· Cortical thickness· Cingulate gyrus
Introduction
Major depressive disorder (MDD) is a disabling psychiatric
disorder affecting over 300 million people around the world.
It greatly impacts quality of life and has severe economic
and societal consequences [1]. Up to 35% of MDD patients
do not respond sufficiently to first-line treatments with anti-
depressant medication or psychotherapy [2]. Patients with
treatment-resistant depression (TRD) have a high risk for
chronicity and often also suffer from comorbid disorders and
suicide attempts, emphasizing the need for more effective
treatment options [3, 4]. From a neurobiological perspective,
decreased neuroplasticity is thought to be the most important
underlying mechanism to explain treatment resistance and
chronicity [5, 6]. This is reflected on a macroscopic level by
for example decreased hippocampal and amygdala volume
Communicated by Sebastian Walther.
Eveline Ackermansand Lieke Martens Shared second author.
Philip van Eijndhoven and Indira Tendolkar Shared last author.
Electronic supplementary material The online version of this
article (https ://doi.org/10.1007/s0040 6-020-01135 -w) contains
supplementary material, which is available to authorized users.
* Iris Dalhuisen
iris.dalhuisen@radboudumc.nl
Extended author information available on the last page of the article
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40 European Archives of Psychiatry and Clinical Neuroscience (2021) 271:39–47
1 3
in patients with MDD, which was associated with duration
of illness [7, 8]. A decrease in hippocampal volume was also
observed in patients with recurrent episodes as compared
to first-episode patients and controls [9], and post mortem
analyses of brains of MDD patients showed decreases in
cortical thickness and neuronal density, which correlated
with duration of illness [10].
New treatment options such as brain stimulation could
be of added value in the treatment of depression because of
their effects on neuroplasticity. Repetitive transcranial mag-
netic stimulation (rTMS) is a form of non-invasive brain
stimulation that is increasingly used for the treatment of
depression and has been shown to be effective, with most
studies focusing on patients with TRD [11, 12]. Treatment
with rTMS consists of a coil, placed against the head, that
induces a magnetic field in the targeted cortex that modu-
lates neuronal activity. In accordance with the neurotrophic
hypothesis of depression and its treatment, a neuroplastic
component in the effects of rTMS has been suggested by sev-
eral studies. In a study of TRD patients, response to rTMS
was associated with an increase in left amygdala volume
and unchanged hippocampal volumes, while non-response
was associated with a decrease in left hippocampal volume
[13]. An increase in hippocampal volume on the side of the
brain targeted with rTMS has also been found [14]. These
findings indicate remote neuroplastic effects of rTMS that
are not limited to the targeted stimulation site.
Besides these volumetric changes in subcortical struc-
tures, neuroplastic effects of rTMS have also been observed
in the cortex. Increased cortical thickness after rTMS treat-
ment has been found in regions of the left rostral and caudal
anterior cingulate cortex (ACC), which correlated with clini-
cal response [15, 16]. Moreover, after rTMS treatment in
patients with TRD increases in structural grey matter volume
were found in the left ACC, left insula, left superior tem-
poral gyrus and right angular gyrus [17]. Since volume is a
function of cortical surface area and cortical thickness, these
increases could also be the result of an increase in cortical
thickness. In line with these findings, the most relevant neu-
roplastic effects could be expected in the paralimbic cortex
[18].
The aim of this work is to further describe longitudinal
changes in chronic treatment-resistant depression. In this
study, we describe the neuroimaging results in a pre–post
design of a randomized controlled trial in which 31 patients
with chronic TRD were treated with either 20 sessions of
high-frequency stimulation of the left dlPFC or sham. The
clinical results of this trial are reported separately, in which
it was concluded that this rTMS protocol was not an effec-
tive treatment option for chronic TRD [19]. Here we aimed
to investigate the rTMS treatment-induced neuroplastic
effects in this group of patients with chronic treatment-
resistant depression, despite the lack of clinical response.
We specifically studied the longitudinal effects of rTMS
on hippocampal and amygdala volumes, and thickness of
the paralimbic cortex, using structural magnetic resonance
imaging (sMRI). As a secondary question, we wanted to see
if these possible volumetric changes or changes in cortical
thickness are related to treatment response, as measured by
Hamilton Depression Rating Scale (HDRS) [20]. Thirdly, we
were interested in whether the degree of treatment resistance
is related to neuroplastic changes in the brain.
Materials andmethods
Participants
Eligible participants were patients with a diagnosis of unipo-
lar MDD without psychotic features, with a chronic course
during the last two years and treatment resistance, defined
as inadequate response to at least two adequate trials of
antidepressants and one trial of psychotherapy. Exclusion
criteria for participation included the presence of a current
or past relevant somatic or neurological disorder; a comorbid
diagnosis of bipolar disorder, schizophrenia or substance
dependence disorders; epilepsy; serious head trauma or
brain surgery; large or ferromagnetic metal parts in the head
(except for a dental wire); implanted cardiac pacemaker or
neurostimulator; and pregnancy. Previous treatment with
electroconvulsive therapy (ECT) was not considered a rea-
son for exclusion. None of the participants had received ECT
within 6months before entering the trial. The concomitant
use of antidepressants and psychotherapy was allowed, as to
not endanger the safety of the participant due to aggravation
of depressive symptoms. Detailed information regarding the
previous and current treatments are presented in Supplemen-
tary Tables1 and 2.
Study overview
The study was approved by the local ethics committee (CMO
region Arnhem–Nijmegen, The Netherlands). Participants
were recruited through the outpatient clinics of the depart-
ment of psychiatry of Radboud University Medical Centre
and Pro Persona Mental Health Care. All participants gave
written consent prior to participation. The randomized con-
trolled trial was registered in the ISRCTN registry (ISRCTN
15535800). The purpose of the current study is to report
a secondary outcome of this trial, namely investigating
potential brain changes. The main results will be reported
elsewhere.
Patients were randomized to receive either active or sham
rTMS treatment. Treatment consisted of 20 sessions in a
period of four weeks. Within one week before and within
one week after treatment, an MRI-scan was performed, as
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41European Archives of Psychiatry and Clinical Neuroscience (2021) 271:39–47
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well as a baseline and post-treatment clinical assessment.
A follow-up clinical assessment was scheduled six months
after start of the intervention.
Evaluation andoutcome measures
MDD was diagnosed by administration of the Structured
Clinical Interview for DSM-IV (SCID) by a trained psy-
chiatrist (P.v.E). Assessment of comorbid psychiatric disor-
ders was done through the Mini Internal Neuropsychiatric
Interview (M.I.N.I.). To assess treatment resistance, the
Dutch Method for quantification of Treatment Resistance in
Depression (DM-TRD) was used [21]. The 17-item Hamil-
ton Depression Rating Scale (HDRS-17) was used to assess
severity of depressive symptoms [20]. Response is defined
as a ≥ 50% reduction in score on the HDRS-17, whereas
remission is defined as a total score ≤ 7 post-treatment.
rTMS procedure
rTMS was administered using a Magstim Super Rapid2 mag-
netic stimulator (Magstim Company, Whitland, UK) with a
92-mm figure-of-eight coil. Resting motor threshold (rMT)
was determined at the beginning of each treatment week.
The rMT was defined as the minimal stimulation intensity
evoking a visual observation of thumb or finger movement
in ≥ 5 out of 10 trials. Stimulation was set at 110% of the
rMT. High-frequency (10Hz) rTMS was administered
five days a week for four consecutive weeks. Participants
received 60 trains of 50 stimuli each, with a duration of 5s
and an inter-train interval of 25s, resulting in a total of 3000
pulses per session. Treatment was applied to the left dlPFC,
which was located using electrode position F3 of the 10–20
EEG system [22]. The use of F3 for the localization of the
dlPFC has been shown to be equally effective as localization
based on neuronavigation [23]. For the sham rTMS group,
the same parameters were used, the exception being that the
orientation of the coil was tilted 45° away from the cortex.
MRI data acquisition andcortical reconstruction
High-resolution anatomical images of the whole brain were
acquired on a 1.5-T Siemens Sonata whole-body scanner
(Siemens, Erlangen, Germany) using a three-dimensional
T1-weighted magnetization prepared rapid acquisition gradi-
ent echo sequence (MPRAGE) with the following acquisi-
tion parameters: T1 850 [ms], TR 2250 [ms], TE 3.68 [ms],
flip angle 15 [deg], FoV 256 × 256 × 176 [mm], voxel-size
1.0 × 1.0 × 1.0 [mm]. Due to unavailability of the MRI scan-
ner, three patients did not undergo MRI-scanning and could
therefore not be included in our analyses. Two patients only
had an MRI-scan before the first rTMS session but not after
the last rTMS session. These patients were only included in
our analyses concerning pre-treatment volumetric measures.
Scans were analyzed using FreeSurfer software (ver-
sion 5.3, https ://surfe r.nmr.mgh.harva rd.edu/). FreeSurfer
includes skull stripping, B1 bias field correction, gray-white
matter segmentation and reconstruction of cortical surface
models (gray-white boundary surface and pial surface). In
cases where dura was included in the gray matter, a manual
correction was applied. The software enables automatic
labeling of subcortical structures using a probabilistic algo-
rithm. Initially, each image is a rigid body registered to a
probabilistic atlas based on manually-labeled images. Then,
the image is morphed to the atlas by a non-linear transform
and a Bayesian segmentation procedure is employed. Each
voxel in the MRI volume is automatically assigned to a
neuro-anatomical label based on probabilistic information
estimated from a manually-labeled training set. The labeling
procedure is not biased by anatomical variability. The seg-
mentation procedure is based on three types of probabilities
to disambiguate labels: the likelihood that a given structure
occurs at a specific atlas location; the likelihood of the image
intensity given that tissue class; and the probability that a
voxel belongs to a given tissue class based on likelihood of
the spatial configuration of labels. This automated segmenta-
tion and labeling procedure has been shown to be of equal
accuracy to manual tracing methods and relatively insensi-
tive to changes in acquisition parameters [24, 25].
A longitudinal pipeline implemented in FreeSurfer was
used in which the subjects were their own controls [26]. In
the longitudinal pipeline, for every patient a template vol-
ume is created from the baseline and post-treatment MRI
after rTMS treatment, reducing random variation in the pro-
cessing procedure and improving robustness and sensitiv-
ity of the overall longitudinal analysis. The volumes of the
amygdala and hippocampus were corrected for brain size,
by dividing these volumes by the estimated total intracranial
volume (TIV), obtained from FreeSurfer.
Statistical analysis
All statistical analyses were performed using SPSS Statis-
tics 22.0 (IBM Corp., Armonk NY, USA) and procedures
were 2-tailed with significance set at an alpha-level of 0.05,
unless stated otherwise. T-tests and chi-squared tests were
used to examine differences on demographic and clinical
variables between the active and sham groups. To assess the
effect of time (pre/post-rTMS) and treatment group (sham/
active) on HDRS score, a repeated measures analysis of
covariance (ANCOVA) was performed with age and gender
as covariates.
To assess if there was a difference in volume change of
amygdala and hippocampus between the active and sham
group, a repeated measures ANCOVA was performed
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42 European Archives of Psychiatry and Clinical Neuroscience (2021) 271:39–47
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with time (pre/post-rTMS) and hemisphere (left/right) as
within-subject factors, and treatment group (sham/active)
as between-subjects factor. Gender and age at baseline were
added as covariates. ANCOVAs were done separately for
normalized total amygdala and hippocampal volume change.
To assess whether volumetric changes were related to treat-
ment response, Pearson correlation analyses were performed
between change in HDRS score and change in hippocam-
pal and amygdala volumes. To assess whether volumetric
changes are related to degree of treatment resistance, Pear-
son correlation analyses were performed between baseline
DM-TRD score and change in hippocampal and amygdala
volumes. For the hippocampus a 1-tailed design was used
based on the earlier findings that show an increase in hip-
pocampal size in effective antidepressant treatment [27–29].
Change in cortical thickness between the active and sham
group was assessed using QDEC, FreeSurfer’s graphical
interface for analyzing group data. The main effect of the
treatment group was estimated for the whole brain (in ver-
tex-wise statistical difference maps) using the FreeSurfer
question ‘’Does the average longitudinal cortical thickness
symmetrized percent change, accounting for gender, differ
between active and sham treatment?’’ and including the nui-
sance factor age.
On the basis of the previous literature, we a priori hypoth-
esized that we would find changes in cortical thickness in
the paralimbic cortex, as a result of rTMS treatment. We
therefore report differences as significant below an uncor-
rected p value of 0.001 (two-tailed) and at least 100 verti-
ces, which is considered an appropriate threshold when an
a priori hypothesis is present [18, 30].
Results
Demographic andclinical characteristics
The demographic and clinical variables of the patients are
shown in Table1. Patients in the active and sham group
did not differ in these variables except for current use of
antidepressants. The trial was discontinued after 31 patients
for futility reasons. HDRS-17 scores did not differ from pre-
to post-treatment for the whole group (F(1, 27) = 0.547,
p = 0.466), nor between the active and sham group (F(2,
26) = 0.120, p = 0.731).
Volumetric results
Total hippocampal volume neither differed from pre- to
post-treatment for the whole group (F(1, 22) = 0.222,
p = 0.642), nor between treatment groups (F(2, 21) = 1.743,
p = 0.200). This was also the case for total amygdala volume
(F(1, 22) = 1.705, p = 0.205 and F(2, 21) = 0.878, p = 0.359,
respectively). Total amygdala volume from pre- to post-
treatment significantly interacted with gender, independent
of treatment group (F(2, 26) = 4.645, p = 0.042). Post hoc
analyses with paired-samples t-tests showed no difference in
total amygdala volume from pre- to post-treatment in males
(t(6) = 2.230, p = 0.067) and in females (t(18) = − 0.009,
p = 0.993). Further post hoc analyses of volumes of right and
left amygdala separately, for males and females separately,
showed a significant decrease in left amygdala volume in
males; see also Table2.
Change in HDRS-17 score from pre- to post-treatment
was not correlated with change in total amygdala volume
(r = − 0.007, p = 0.973) or change in total hippocampal
volume (r = 0.165, p = 0.421). Change in left amygdala
volume also did not correlate with change in HDRS-17
score (r = − 0.027, p = 0.898). Baseline DM-TRD score
Table 1 Demographic and
clinical characteristics of
patients
Values represent mean ± SD or N (%). *indicates a statistically significant result
Active (N = 15) Sham (N = 16) Total (N = 31) P
Female sex 9 (60%) 13 (81%) 22 (71%) 0.193
Age 47.33 ± 11.49 49.69 ± 11.02 48.55 ± 11.12 0.565
Current AD use 7 (47%) 14 (88%) 21 (68%) 0.015*
Previous ECT 6 (40%) 9 (56%) 15 (48%) 0.366
Duration current episode
(months)
54.60 ± 26.19 57.88 ± 54.83 56.29 ± 42.73 0.835
Number of episodes 2.87 ± 1.25 3.50 ± 1.27 3.19 ± 1.28 0.171
Age of onset (years) 28.80 ± 12.07 25.19 ± 10.40 26.94 ± 11.20 0.378
DM-TRD score 18.67 ± 2.37 18.19 ± 2.77 18.42 ± 2.55 0.609
HDRS-17 score pre 24.13 ± 4.29 22.69 ± 3.84 23.39 ± 4.06 0.331
HDRS-17 score post 21.00 ± 5.44 18.56 ± 5.61 19.74 ± 5.57 0.230
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43European Archives of Psychiatry and Clinical Neuroscience (2021) 271:39–47
1 3
was not correlated with change in total amygdala volume
(r = 0.368, p = 0.064) or change in total hippocampal vol-
ume (r = − 0.112, p = 0.293).
Cortical thickness results
The vertex-by-vertex analysis showed significant differ-
ences between the active rTMS and sham condition in
the paralimbic cortex (p < 0.001, uncorrected; adjusted
for age and gender; see Table3). In the left hemisphere,
patients with active rTMS had significantly increased
cortical thickness in the left isthmus cingulate gyrus
(p < 0.000005, see Fig.1) and the pericalcarine cortex
(p < 0.0004). In the right hemisphere, this was the case
for the post-central gyrus (p < 0.0004) and lateral orbito-
frontal cortex (p < 0.0002). In addition, there were areas
of decreased cortical thickness in the left superior pari-
etal lobule (p < 0.0007) and in the right superior tempo-
ral gyrus (p < 0.0007), the post-central gyrus (p < 0.0006)
and the supramarginal gyrus (p < 0.0005).
Discussion
To our knowledge, this is the first sham-controlled study
that investigated the longitudinal effects of rTMS on hip-
pocampal and amygdala volumes and cortical thickness of
the paralimbic cortex in chronic TRD patients. We assessed
possible correlations of volumetric changes with clinical
treatment response, and whether level of treatment resistance
was related to volumetric changes over the course of rTMS
treatment. Active and sham rTMS treatment in patients with
chronic treatment-resistant major depressive disorder had
no clinical effects. Despite the absence of clinical effects,
we identified neuroplastic changes in the cingulate cortex,
which may indicate that a more intensive treatment protocol
could lead to enhanced neuroplastic changes, which in turn
could lead to better treatment response.
Cortical thickness data
Our results show differences in cortical thickness in the
active rTMS group over the sham group in patients with
chronic TRD. The most pronounced finding to emerge from
the analysis is the increased thickness of the left isthmus of
Table 2 Normalized amygdala
volumes pre- and post-treatment
in males and females. Volumes
represent percentage of total
brain volume
Values represent mean ± SD. *indicates a statistically significant result
Pre-treatment (%) Post-treatment (%) % Change p
Males Total amygdala 0.2078 ± 0.0228 0.2015 ± 0.0215 − 3.03 0.067
Right amygdala 0.1122 ± 0.0134 0.1010 ± 0.0133 − 9.98 0.234
Left amygdala 0.0956 ± 0.0108 0.0916 ± 0.0104 − 4.18 0.045*
Females Total amygdala 0.2138 ± 0.0252 0.2138 ± 0.0233 0.00 0.993
Right amygdala 0.1158 ± 0.0126 0.1160 ± 0.0122 0.17 0.708
Left amygdala 0.0980 ± 0.0134 0.0978 ± 0.0122 0.20 0.837
Table 3 Brain regions with significant differences in cortical thickness between active and sham rTMS treatment
The results were considered significant if p < 0.001 and number of vertices > 100
a. Based on the Talairach coordinates. *indicates a statistically significant result. **indicates a statistically significant results with p < 0.0001
Hemisphere Cortical region Size (mm2) Coordinates maximum p value (mm)a NVtxs Uncorrected p value
xyZ
Active > Sham
Left Isthmus cingulate gyrus 123.98 − 6.8 − 33.0 29.7 415 < 0.001**
Pericalcarine cortex 119.73 − 23.2 − 72.4 5.2 306 < 0.001*
Right Post-central gyrus 78.79 50.9 − 20.3 53.5 161 < 0.001*
Lateral orbitofrontal gyrus 45.26 23.9 13.0 − 18.7 119 < 0.001*
Active < Sham
Left Superior parietal lobule 63.45 − 28.0 − 61.7 43.7 171 < 0.001*
Right Superior temporal lobule 148.28 60.1 − 27.6 1.0 349 < 0.001*
Post-central gyrus 94.68 59.0 − 10.0 33.2 208 < 0.001*
Supramarginal gyrus 69.46 50.3 − 37.9 23.6 159 < 0.001*
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44 European Archives of Psychiatry and Clinical Neuroscience (2021) 271:39–47
1 3
cingulate gyrus in the active rTMS group compared to sham.
Several studies indicate that cortical thickness is affected
in MDD, as regional thinning in the cingulate and orbito-
frontal cortex [18, 31, 32]. Neuroimaging studies show that
in patients with depression, the structure and connectiv-
ity of a subregion of the cingulate cortex, the isthmus, is
altered [33–36]. Additionally, stronger functional connec-
tivity between the ACC and the prefrontal cortex has been
observed after rTMS treatment, but only in responders [37].
The cingulate cortex is a key area within the fronto-limbic
networks involved in emotion, sensory, motor, and cognitive
processes [38, 39] and, as part of the paralimbic cortex, is
interconnected with the orbitofrontal cortex, amygdala, hip-
pocampus and striatum [38]. The connection between the
cingulate cortex and the prefrontal cortex could therefore
play an important role in the antidepressant effect of rTMS.
In addition to the increases in cortical thickness, we also
identified decreases, specifically in the left superior parietal
lobule and in the right superior temporal gyrus, post-central
gyrus and supramarginal gyrus. After treatment with rTMS,
a decrease in the left subcallosal ACC was found, which did
not correlate with clinical improvement [15]. Although the
implications are still unclear, it may be that decreases in
cortical thickness are also relevant for the neurobiological
effects of rTMS treatment in TRD and need to be investi-
gated in future studies.
Volumetric data
Neither the active rTMS treatment group nor the sham
group showed a significant change in hippocampal or
amygdala volumes. This was partly consistent with the
previous findings of treatment-related volume increase
only in the left hippocampus, but no change in amygdala
volume [14]. Notably, in this study treatment response
was much better than in our study, most likely due to less
severe TRD. This suggests that the restricted neuroplastic
effects on amygdala and hippocampus volume in our study
cannot be simply explained as a function of limited treat-
ment response. Three of the participants in our study were
treated with lithium, all of whom were in the sham group.
The neuroplasticity-facilitating properties of lithium could
have impacted the results; especially because all partici-
pants taking lithium were in the same group. Interestingly,
we did not find any differences between the active and
sham group in volume of the hippocampus and amygdala,
the brain regions where the neuroplastic effects of lithium
are most pronounced [40], so this is unlikely to have influ-
enced the results. We identified a significant decrease in
left amygdala volume in males, independent of treatment
group. The results from the previous studies regarding the
change in amygdala volume after rTMS treatment have
been mixed [13, 14]. Since we did not correct for multiple
comparisons, this result needs to be interpreted with care
and future studies will need to evaluate this finding.
Furthermore, we found no correlation of volumetric
changes with clinical treatment response, which is in
line with the previous results [14]. However, a decline
in left hippocampal volume specifically in treatment non-
responders has also been observed [13]. In line with this,
studies on ECT also do not find a relationship between
volumetric changes and clinical improvement, irrespective
of the large increases in hippocampal volume and strong
clinical effects of ECT [41].
Additionally, we examined whether level of treatment
resistance correlated with volumetric changes. Our find-
ings do not show evidence for this relationship. A recent
review on structural brain characteristics in TRD sug-
gested that volumetric differences in hippocampal vol-
ume are likely subtle and therefore current studies lack
Fig. 1 Isthmus of left cingu-
late gyrusshowing significant
increased difference in cortical
thickness between active and
sham rTMS treatment
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45European Archives of Psychiatry and Clinical Neuroscience (2021) 271:39–47
1 3
the statistical power to detect such changes [42], except
in large samples [9].
Study limitations andstrengths
This study has several limitations. Firstly, the study had a
relatively small sample due to the futility of the trial, which
may also limit the statistical power of the neuroimaging
results. However, other studies investigating the effects of
rTMS on brain structure and volume have used comparable
sample sizes whilst also reporting significant findings [13,
14, 16, 17]. In addition, it is an ethical necessity to stop a
clinical trial when the risks are found to outweigh the poten-
tial benefits [43]. A second limitation is the lack of response
to rTMS treatment. rTMS has been shown to be more effec-
tive in patients with a lower level of treatment resistance
[44], whereas our sample consisted of patients with chronic
depression and a very high level of treatment resistance. To
illustrate, in a sample of nearly 300 patients with recurrent
MDD the average score on the DM-TRD, which can range
from 0 to 27, was 9.8 [21]. In our sample, the average scores
were 18.7 and 18.2 for the active and sham group, respec-
tively. Since a higher number of pulses per session has been
associated with higher response and remission rates [45, 46],
rTMS treatment in this study might have been insufficiently
intense to result in clinical effect in our highly treatment-
resistant chronically depressed sample and an increase in
the number of pulses or treatments might increase treatment
effect in these patients. Furthermore, participants were not
asked to guess their allocated treatment at the end of the
study. Expectations regarding a treatment can affect treat-
ment outcome [47]. However, a review regarding blinding
integrity showed that subjects are unable to distinguish sham
from real rTMS [48]. Moreover, fifteen of the participants
had received ECT. Given the neuroplastic effects of ECT,
this might have confounded the results [41]. However, when
correcting for prior ECT, results remained non-significant.
Finally, the use of angulation as a sham method may have
resulted in residual brain stimulation, which could have been
prevented with the use of a sham coil. However, a true sham
procedure is difficult to achieve since each method has its
limitations [49].
Nevertheless, by including a sham-controlled rTMS
group, we extended on the previous findings since we were
able to dissociate whether volumetric changes or changes in
cortical thickness are related to the rTMS treatment itself or
more global clinical changes during rTMS treatment.
Future directions
Future research should investigate the neuroplastic effects
of rTMS treatment in a larger sample of patients with
MDD, ideally in a group with different levels of treatment
resistance, to provide more substantial evidence for the influ-
ence of rTMS treatment on brain structure and function. For
this group of severe treatment-resistant and chronic depres-
sion, future TMS studies should use more intensive treat-
ment protocols, by increasing the number of sessions and
pulses per session or shift to different TMS protocols with
presumed larger neuroplastic effects such as bilateral rTMS
or theta-burst stimulation.
Conclusions
In sum, this is the first sham-controlled study that investi-
gates neuroplastic effects of rTMS treatment on amygdala
and hippocampus volume as well as cortical thickness in
patients with chronic treatment-resistant depression. We did
not find clinical improvement within our sample. However,
we did show neurobiological effects of rTMS treatment in
the form of changes in cortical thickness in the paralim-
bic cortex, an area that plays an important role in mood
disorders. For clinical effects to follow these neuroplastic
effects, more intensive rTMS treatment might be needed in
this group of chronically depressed patients. Our results sup-
port earlier findings that indicate rTMS-induced neuroplas-
ticity in the cortex. We suggest that rTMS asserts network
effects that could potentially be the underlying mechanism
responsible for clinical effects. However, findings have so
far been mixed and the exact mechanism and meaning of the
volumetric changes we found remain unclear and subject for
further studies.
Acknowledgements We would like to thank Jan Leijtens for his sup-
port with setting up and conducting this study.
Author contributions All authors contributed to the study conception
and design. Material preparation, data collection and analysis were per-
formed by Eveline Ackermans, Lieke Martens, Peter Mulders and Iris
Dalhuisen. The first draft of the manuscript was written by Iris Dalhu-
isen and all authors commented on previous versions of the manuscript.
All authors read and approved the final manuscript.
Funding None.
Compliance with ethical standards
Conflicts of interest The authors declare that they have no conflict of
interest.
Ethics approval This study was performed in line with the principles
of the Declaration of Helsinki. The study was approved by the local
ethics committee; CMO region Arnhem–Nijmegen, The Netherlands.
Consent to participate All participants gave written consent prior to
participation.
Consent for publication Not applicable.
Content courtesy of Springer Nature, terms of use apply. Rights reserved.

46 European Archives of Psychiatry and Clinical Neuroscience (2021) 271:39–47
1 3
Availability of data and material The imaging data used to support
the findings of this study are available from the corresponding author
upon request.
Code availability Not applicable.
Open Access This article is licensed under a Creative Commons Attri-
bution 4.0 International License, which permits use, sharing, adapta-
tion, distribution and reproduction in any medium or format, as long
as you give appropriate credit to the original author(s) and the source,
provide a link to the Creative Commons licence, and indicate if changes
were made. The images or other third party material in this article are
included in the article’s Creative Commons licence, unless indicated
otherwise in a credit line to the material. If material is not included in
the article’s Creative Commons licence and your intended use is not
permitted by statutory regulation or exceeds the permitted use, you will
need to obtain permission directly from the copyright holder. To view a
copy of this licence, visit http://creat iveco mmons .org/licen ses/by/4.0/.
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Aliations
IrisDalhuisen1,6 · EvelineAckermans1,2· LiekeMartens1· PeterMulders1,7· JoeyBartholomeus4· AlexdeBruijn2,3·
JanSpijker1,2,5· PhilipvanEijndhoven1,6,7· IndiraTendolkar1,6,8
Eveline Ackermans
Eveline.Ackermans@radboudumc.nl
Lieke Martens
Lieke.Martens@student.ru.nl
Peter Mulders
PeterCR.Mulders@radboudumc.nl
Joey Bartholomeus
JBartholomeus@rijnstate.nl
Alex de Bruijn
alexdebruijn@gmail.com
Jan Spijker
J.Spijker@propersona.nl
Philip van Eijndhoven
Philip.vanEijndhoven@radboudumc.nl
Indira Tendolkar
Indira.Tendolkar@radboudumc.nl
1 Department ofPsychiatry, Radboud University Medical
Center, Huispost 961, PO Box9101, 6500HBNijmegen,
TheNetherlands
2 Pro Persona Mental Health Care, PO Box7049,
6503GMNijmegen, TheNetherlands
3 Fundacion Salud Mental Respaldo, Caya Punta Brabo 17,
Oranjestad, Aruba
4 Department ofPsychiatry, Rijnstate Hospital, PO Box9555,
6800TAArnhem, TheNetherlands
5 Radboud University Behavioural Science Institute, PO
Box9104, 6500HENijmegen, TheNetherlands
6 Donders Institute ofBrain Cognition andBehavior, Centre
forNeuroscience, PO Box9104, 6500HENijmegen,
TheNetherlands
7 Donders Institute forBrain Cognition andBehavior,
Centre forCognitive Neuroimaging, PO Box9104,
6500HENijmegen, TheNetherlands
8 Department ofPsychiatry andPsychotherapy, University
Hospital Essen, Virchowstraße 174, 45147Essen, Germany
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