Reduced Gray Matter Brain Volumes Are Associated with Variants of The

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Molecular Psychiatry (2008) 13, 1093–1101 & 2008 Nature Publishing Group All rights reserved 1359-4184/08 $30.00 www.nature.com/mp ORIGINAL ARTICLE Reduced gray matter brain volumes are associated with variants of the serotonin transporter gene in major depression T Frodl1, N Koutsouleris1, R Bottlender1, C Born2,MJa¨ger1,MMo¨rgenthaler1, J Scheuerecker1, P Zill1, T Baghai1, C Schu¨le1, R Rupprecht1, B Bondy1, M Reiser2, H-J Mo¨ller1 and EM Meisenzahl1 1Department of Psychiatry and Psychotherapy, Ludwig-Maximilians-University of Munich, Munich, Germany and 2Department of Radiology, Ludwig-Maximilians-University of Munich, Munich, Germany

The serotonergic system is involved in the pathophysiology of major depression as well as in the early central nervous system development and adult neuroplasticity. The aim of the study was to examine in 77 patients with major depression and 77 healthy controls the association between the triallelic polymorphism in the promoter region of the serotonin transporter gene (5-HTTLPR) and gray matter (GM) brain volumes measured with 1.5 T magnetic resonance imaging. Voxel-based morphometry were estimated on magnetic resonance images and genotyping was performed. We found that healthy controls have a strong association between the 5-HTTLPR and GM volumes of the dorsolateral prefrontal cortex, left anterior gyrus cinguli, left amygdala as well as right hippocampus, whereas there is no such association in patients

with major depression. Healthy subjects carrying the S- or LG-allele have smaller GM volumes than those with the LA-allele, indicating that 5-HTTLPR contributes to the development of brain structures. Patients with depression show reduced GM volumes, particularly when they

are homozygous for the LA-allele, suggesting that these patients are more vulnerable for morphological changes during depressive episodes. Molecular Psychiatry (2008) 13, 1093–1101; doi:10.1038/mp.2008.62; published online 1 July 2008 Keywords: major depression; VBM; serotonin; imaging genetics; MRI; morphometry

Introduction hippocampus and the frontal cortex,4 has neurotrophic effects5 and increases hippocampal neurogenesis.6 More- Dysfunction of neuronal plasticity could contribute to over, serotonergic function is modulated by BDNF.7 the pathophysiology of mood disorders.1 The under- This hypothesis is also supported by preclinical standing of these neuroplastic processes may offer the studies demonstrating that stress and depression lead possibility to define new pathology-related markers. to reductions in total hippocampal volume, and Such markers might be helpful to improve the atrophy and loss of neurons in the adult hippocam- understanding of the neurobiology of major depres- pus.8,9 Many in vivo neuroimaging studies have sion. Moreover, they might offer the opportunity to detected reduced hippocampal volumes in elderly obtain surrogate markers and to find new antidepres- and also in younger patients with major depression. sants, as well as new substances, which may improve The results for other brain regions are rather incon- the clinical outcome. sistent, perhaps because of problems with the exact The neuroplasticity hypothesis of major depression manual measurement of anatomical difficult brain is supported by studies demonstrating that serotoner- regions: enlarged amygdala volumes, as well as gic signaling is an important regulator of early central reduced volumes of the anterior cingulum, prefrontal nervous system development2 and of adult neurogen- cortex and basal ganglia, have been reported in only esis.3 This is supported by findings of reciprocal some of the studies with regions of interest analysis.10 regulations between the serotonergic system and Voxel-based morphometry (VBM) is extremely useful brain-derived neurotrophic factor (BDNF). Anti- because brain regions whose boundaries are difficult depressant treatment, which increases serotonergic to define can be analyzed. VBM has detected smaller neurotransmission, for example, selective serotonin volumes of the medial frontal lobes in patients with reuptake inhibitors, and elevates BDNF in the subthreshold depression,11 as well as of the right hippocampus and the middle frontal gyrus in patients Correspondence: Dr T Frodl, Department of Psychiatry with major depression.12 and Psychotherapy Ludwig-Maximilians-University of Munich, Neuroimaging genetics offer a good possibility to Nussbaumstr. 7, Munich 80336, Germany. E-mail: [email protected] combine the morphological and neurochemical as- Received 19 December 2007; revised 20 March 2008; accepted 12 pects, for example, structural and serotonergic altera- May 2008; published online 1 July 2008 tions. Substantial relative decreases in hippocampal Brain volumes and serotonin transporter in depression T Frodl et al 1094 volume have been found in Met-carriers of the BDNF diagnoses based on DSM-IV criteria and on the polymorphism, in either healthy controls13,14 or structured clinical interview for DSM-IV (SCID) patients with major depression.15 Furthermore, the were determined by a consensus of at least two L-allele of the 5-HTTLPR, which shows increased psychiatrists. Clinical variables were documented 5-HTT expression and increased 5-HT-reuptake using the 21-item Hamilton Depression Rating in vitro and in vivo,16–18 is associated with hippocampal Scale.22 volume reduction in major depression.19,20 On the day of the magnetic resonance imaging, The L-allele of the 5-HTTLPR can be subtyped into patients were receiving the following medication: 17 La and Lg alleles, the latter of which is thought to be patients serotonin reuptake inhibitors (7 sertraline, similar to the S-allele in terms of reuptake efficiency 7 citalopram, 2 paroxetine, 1 fluvoxamine), 8 tricyclic perhaps explaining some of these discrepancies.21 antidepressants (1 amitriptyline, 2 amitriptylinoxide, 4 This polymorphism can then be handled as a triallelic doxepine, 1 trimipramine), 34 other new antidepres- polymorphism. To avoid inconsistencies, we use the sants (11 venlafaxine, 10 reboxetine, 13 mirtazapine), term triallelic for this new polymorphism and 2 maprotiline and 16 patients no antidepressant. diallelic for the older polymorphism. For comparison, 77 healthy control subjects were The aim of this study was to investigate the matched with respect to age (mean age: 43.6±11.3 influence of this functional triallelic 5-HTTLPR/ years), gender and handedness. rs25531 on gray matter (GM) volumes of the brain in A structured interview was used to assess medical patients with major depression and healthy controls history, trauma and other exclusion criteria for all using voxel-based morphology. We hypothesized that, subjects. Exclusion criteria for patients and controls compared to healthy controls, GM volume is particu- were previous head injury with loss of consciousness, larly reduced in the hippocampus, amygdala, anterior cortisol medication in the medical history, previous gyrus cinguli (ACC) and the dorsolateral (DLPFC) and alcohol or substance abuse, neurological diseases or dorsomedial prefrontal cortex (DMPFC) in patients age over 65 years. Comorbidity with other mental with major depression who are homozygous for the illnesses and personality disorders were also ex- L-allele of the 5-HTTLPR. cluded, as were patients with bipolar disorders. Healthy controls had no history of neurological or Materials and methods mental illnesses. No subject received an electroconvulsive therapy before investigation. Handedness was Participants determined by the Edinburgh inventory.23 Seventy-seven inpatients with major depression Written informed consent was obtained from all (mean age: 46.1±11.3 years, Table 1) were subjects subsequent to a detailed description of recruited from the Department of Psychiatry of the the study. The study design was approved by Ludwig-Maximilians-University, Munich. Psychiatric the local ethics committee and was prepared in

Table 1 Demographic and clinical data of patients with an episode of major depression and healthy controls

Patients (n = 77) Controls (n = 77) T-test (mean±s.d.) (mean±s.d.) (P-value)

Age (years) 42.6±12.4 40.5±11.6 0.29 Female/male* 35/42 35/42 1.0 Handedness (R/L)* 65/12 72/5 0.07 Height (cm) 172.7±8.8 175.1±10.3 0.12 Weight (kg) 73.2±15.2 75.1±14.2 0.43 Alcohol (g dayÀ1) 9.8±18.1 8.4±11.3 0.57 Age of onset (years) 37.1±12.1 Duration of illness (years) 5.4±8.2 Number of depressive episodes 2.05±2.1 HDRS—baseline 22.8±6.1 Triallelic 5-HTTLPR*

LALA/LALG/LGLG/LAS/LGS/SS 14/18/11/23/1/10 10/10/1/34/8/14 0.002 Allele frequency

LA/LG/S 0.45/0.27/0.28 0.42/0.13/0.45 0.001

Functional consequence* (LA =L,LG =S) LL/LS/SS 14/41/22 10/44/23 0.6

Abbreviation: HDRS, Hamilton Depression Rating Scale. Mean and s.d.(±) are given. No significant differences were found between patients and controls as measured with t-test or w2 test. *P < 0,05, only w2 test.

Molecular Psychiatry Brain volumes and serotonin transporter in depression T Frodl et al 1095 accordance with the ethical standards laid down in Hilden, Germany) according to the supplier’s the Declaration of Helsinki. instructions. We used the triallelic polymorphism SNP-rs25531, because the L-allele can be subtyped Magnetic resonance imaging procedures into La and Lg alleles, the latter of which is thought to be similar to the S-allele in terms of reuptake Data acquisition. Magnetic resonance images were efficiency.27 obtained with a Magnetom Vision scanner (Siemens, Genotyping was carried out by applying the PCR Erlangen, Germany) operating at 1.5 T. All subjects amplification in a final volume of 25 ml consisting of were scanned with a T1-weighted 3D-MPRAGE 50-ng DNA, l mmol lÀ1 of each primer, 200-mM deoxy- sequence (repetition time, 11.6 ms; echo time, nucleotide triphosphate, 100-mM 7-deaza-guanosine 4.9 ms; total acquisition time, 9 min; number of triphosphate, 5% dimethyl sulfoxide, 10-mM acquisitions, 1; field of view, 230 mm; matrix, Tris-hydrochloride (pH 8.3), 50-mM potassium 512 Â 512; section thickness, 1.5 mm) yielding 126 chloride, 1.5-mM magnesium chloride and 2.5 U of contiguous axial slices with a defined voxel size of DNA polymerase (AmpliTaq Gold; PerkinElmer, 0.45 Â 0.45 Â 1.5 mm. After manually reorienting and Langen, Germany). The PCR products were separated centering the images on the anterior commissure, on a 3% agarose gel (FMC NuSieve 3:1; Biozym data preprocessing were performed with the SPM5 Diagnostic GmbH, Oldendorf, Germany) and software package (Wellcome Department of Cognitive visualized by ethidium bromide staining. Neurology, London, UK) running under MATLAB 6.5 (The MathWorks, Natick, MA, USA). Statistical analysis An analysis of covariance was designed to investigate VBM preprocessing focal GM volume (GMV) differences between the This study employed the VBM5 toolbox, which patients with major depression and healthy controls utilizes and extends the new unified segmentation as well as between genotypes. Age and gender were approach implemented in SPM5.24 Unified segmenta- entered as covariates of no interest in the statistical tion provides a generative model of VBM preproces- design. First, GMV differences (increases/decreases) sing that integrates tissue classification, image between patients with major depression and healthy registration and magnetic resonance imaging inho- controls were assessed at the whole-brain level using mogeneous bias correction. Thus, the model avoids T contrasts (P < 0.05, family wise error correction the ‘circularity problem’ of the optimized VBM (FWE) corrected). Then, the overall genotype effects procedure, as the initial image registration does not were calculated again at the whole-brain level using T require an initial tissue segmentation and vice versa.25 contrasts (P < 0.05, FWE corrected). The VBM5 toolbox extends the unified segmentation Additionally, VBM analysis was performed to model, as it increases the quality of segmentation by examine the interactions between diagnosis and applying a Hidden Markov Field model on the genotype, with additional masking for higher GMV segmented tissue maps.26 The Hidden Markov Field in the S-allele within the patient group thresholded at algorithm provides spatial constraints based on P < 0.05, uncorrected, as the exclusive mask. neighboring voxel intensities within a 3 Â 3 Â 3 voxel The VBM results were also analyzed to test cube. It removes isolated voxels, which are unlikely differences between patients and controls within to be a member of a certain tissue class and also closes each genotype and to test differences between holes in a cluster of connected voxels of a certain genotypes within the patients and within the controls class, resulting in a higher signal-to-noise ratio of the only. Additionally, small-volume-corrected VBM ana- final tissue probability maps. The VBM5 toolbox lyses were performed for the following regions of offers the possibility to write the estimated tissue interest: hippocampus, amygdala, ACC, DMPFC and probability maps without making use of the respec- DLPFC at P < 0.01 for comparison of cross-sectional tive ICBM tissue priors (International Consortium for GMV differences between patients and controls. For Brain Mapping) from SPM5. these five regions of interest patient subgroups, This study used this option, as it improved the analyses were performed at P < 0.01, small volume delineation of the subcortical structures and the sulci corrected: cross-sectional GMV differences between in the final tissue maps. The final tissue maps of gray the patients compared to controls, between patients matter (GM), white matter and cerebro-spinal fluid with each genotype and controls with the same were modulated with the deformation fields obtained genotype, as well as in the patients and control group by normalization to standard space to analyze volume between each genotype. Coordinates of peak signifi- differences between study populations. Finally, the cant voxels were assigned to anatomical regions by modulated GM partitions were smoothed with a means of automated anatomical labeling.28 12 mm FWHM Gaussian kernel and used for statistical analysis. Results Genetics Age, gender, handedness, height, weight and alcohol DNA was extracted from a 5 ml blood sample using consumption were similar in patients and controls the QIAamp Blood Isolation Kit (QIAGEN GmbH, (Table 1). The 5-HTTLPR distributions for the patients

Molecular Psychiatry Brain volumes and serotonin transporter in depression T Frodl et al 1096 and controls were in Hardy–Weinberg equilibrium. observed between heterozygous patients and patients

For each genotype, age, weight and height did not homozygous for either the S-(LG or S) or LA-allele differ between patients and controls. There was (Table 4). no significant difference in illness duration (F(2/74) = 0.6; P = 0.55) or depression severity Genotype effects within healthy controls (Hamilton Depression Rating Scale, F(2/74) = 1.5; Healthy controls homozygous for the LA-allele had P = 2.3) between patients with a specific genotype. significantly larger GMVs in the ACC, DLPFC right Interestingly, with respect to the allele frequencies pronounced, right DMPFC, left amygdala and left

of the 5-HTTLPR-rs25531 the LG-allele was more hippocampus than those homozygous for the S-allele frequent in our patients sample, whereas the S-allele (Table 4). was less frequent compared to the healthy controls 2 (X = 13.5; P = 0.001). As the LG-allele is considered Discussion to be functionally comparable to the S-allele, functional triallelic LL/LS/SS genotypes can be used. The main finding from this study is that healthy Frequencies of these were not significantly different controls show a strong association between between patients and controls (Table 1). 5-HTTLPR polymorphism and GMVs in all Using w2 test, there was no significant difference investigated brain regions, with the smallest volumes between genotypes with respect to different medica- being found in healthy subjects homozygous for the

tion (no medication, selective serotonin reuptake S-allele (LGLG,LGS, SS), whereas in patients with inhibitor, tricyclic, newer antidepressants). major depression this association is only seen in In the overall VBM analyses, patients with major subregions of the hippocampus. The explanation for depression had significantly smaller GMVs in the left this discrepancy may be that patients with depression

DLPFC, bilaterally in the supplementary motor area, who are homozygous for the LA-allele have prominent in the thalamus and the precentral area, and showed GM reductions compared to controls with the same trends bilaterally in the DMPFC, the right orbitofron- genotype. Patients have the largest volume reductions tal cortex as well as the left insular cortex compared compared to controls in cortical brain regions such as to healthy controls (Figure 1). the DLPFC, DMPFC, ACC as well as in the amygdala Small volume correction for regions of interest and anterior hippocampus when they are homo-

revealed that patients had significantly reduced zygous for the LA-allele. Patients homozygous for the GMVs bilaterally in the ACC, DLPFC and in S-allele (LGLG,LGS, SS) or heterozygous patients the DMPFC compared to controls (Table 2). The (LAS, LALG) only differ in small regions from controls hippocampus and amygdala GMVs were not smaller with the same genotype. For example, only the in patients. middle hippocampal volume was found to be reduced in patients carrying the S-allele compared to controls Interaction between genotype and diagnosis with the same genotype. The interaction between diagnosis and genotype was This finding is very interesting because the normal significant in the DLPFC right pronounced, left ACC, influence of 5-HTTLPR on the brain may be dimin- left amygdala as well as right hippocampus: the ished in patients with major depression, indicating difference between controls homozygous for the that other factors may have a greater influence on

LA-allele and those homozygous for the S-allele these processes, for example psychosocial factors or 29 (LGLG,LGS, SS) was larger than the difference between early life events. patients homozygous for the LA-allele and those Interestingly, patients had higher LG-allele frequen- homozygous for the S-allele (Table 3). cies and less S-alleles (LGLG,LGS, SS) compared to healthy controls. Unfortunately, our sample is too

Effects of diagnosis within genotypes small to conclude that the LG-allele may be a risk

Patients homozygous for the LA-allele had signifi- factor for major depression or to analyze the different cantly smaller GMVs in the hippocampus, amygdala, influences between LG-allele and S-allele (LGLG,LGS, ACC, DLPFC and DMPFC than healthy controls with SS) on the brains of patients with major depression the same genotype. Heterozygous patients differed and healthy controls. significantly from controls with the same genotype These data suggest that the main GM alterations are with respect to the DLPFC, and patients homozygous seen in patients homozygous for the functional

for the S-allele (LGLG,LGS, SS) had significantly LA-allele of the 5-HTTLPR-rs25331. In our earlier smaller subregions within the hippocampus DLPFC study, the homozygous L-allele of the 5-HTTLPR was and DMPFC compared to controls with the same found to be associated with reduced hippocampal genotype (Figure 2). volumes in patients with major depression and not in controls.19 Moreover, patients with a late-onset Genotype effects within the depressive group geriatric depression who were homozygous for the

Patients homozygous for the LA-allele had signifi- L-allele of the 5-HTTLPR had smaller hippocampal cantly reduced bilateral hippocampal volumes volumes than other groups.30 than those patients being homozygous for S-allele One possible explanation may be that the higher

(LGLG,LGS, SS). No significant differences were reuptake of serotonin in subjects homozygous for the

Molecular Psychiatry Brain volumes and serotonin transporter in depression T Frodl et al 1097

Figure 1 Overall brain gray matter (GM) volume reduction in patients compared to controls. Regions where GM density is reduced in patients with major depression compared to healthy controls. P-values are corrected for multiple comparisons (FDR, P < 0.01). Numbers below slices represent the vertical distance in millimeter to the anterior commissure.

Molecular Psychiatry Brain volumes and serotonin transporter in depression T Frodl et al 1098 Table 2 Gray matter volume reduction in patients with major depression compared to healthy controls with SVC for ACC, hippocampus, amygdala, DLPFC, DMPFC (P < 0.01, uncorrected)

kk(%) FWE FDR TPxyz

Left ACC 219 1.96 0.057 0.028 3.21 0.001 2 36 28 Right ACC 1200 11.42 0.031 0.028 3.44 < 0.001 4 42 28 Left DLPFC 12667 32.56 0.006 0.004 4.34 < 0.001 À25 5 61 Right DLPFC 13570 33.23 0.038 0.004 3.78 < 0.001 43 4 51 Left DMPFC 6302 26.33 0.004 0.002 4.26 < 0.001 0 32 46 Right DMPFC 2886 16.9 0.004 0.002 4.24 < 0.001 1 32 46 Left amygdala 9 0.51 0.017 0.126 3.27 0.001 À16 À2 À17 Right amygdala NS Left hippocampus 7 0.09 0.041 0.166 3.33 0.001 À15 À2 À16 Right hippocampus 1 0.01 0.053 0.166 3.24 0.001 26 À40 À3

Abbreviations: ACC, anterior gyrus cinguli; DLPFC, dorsolateral prefrontal cortex; DMPFC, dorsomedial prefrontal cortex; FDR, false discovery rate; FWE, family wise error correction; k, number of suprathreshold voxels in anatomical region; k (%), percentage of suprathreshold voxels in anatomical region, T-test; NS, not significant; SVC, small volume correction.

Table 3 Difference in GM volume between LA/LA and S/S (LGLG,LGS, SS) is greater for controls compared to the difference

between LA/LA and S/S (LGLG,LGS, SS) for patients with SVC for ACC, hippocampus, amygdala, DLPFC, DMPFC (P < 0.01, uncorrected)

kk(%) FWE FDR TPxyz

Left ACC 49 0.44 0.153 0.347 2.79 0.003 À12 31 26 Right ACC NS Left DLPFC 11 0.03 0.591 0.266 2.54 0.006 À34 À150 Right DLPFC 926 2.27 0.175 0.266 3.21 0.001 29 35 29 Left DMPFC NS Right DMPFC NS Left amygdala 21 1.19 0.117 0.128 2.47 0.007 À29 0 À26 Right amygdala NS Left hippocampus NS Right hippocampus 17 0.22 0.269 0.253 2.52 0.006 35 À12 À27

Abbreviations: ACC, anterior gyrus cinguli; DLPFC, dorsolateral prefrontal cortex; DMPFC, dorsomedial prefrontal cortex; FDR, false discovery rate; FWE, family wise error correction; GM, gray matter; k, number of suprathreshold voxels in anatomical region, k (%), percentage of suprathreshold voxels in anatomical region, T-test; NS, not significant; SVC, small volume correction.

LA-allele is an additional factor that aggravates stress- during major depression. Importantly, this may not be induced neurotoxic effects during depression. In a common phenomenon because healthy control

addition to its role as a neurotransmitter, serotonin subjects with the S-allele (LGLG,LGS, SS) have the acts as a trophic factor modulating developmental smallest hippocampal volumes. Therefore, the pre- processes such as neuronal division, differentiation, sence of major depression or other factors that play a migration, synaptogenesis31,32 and adult neurogen- part in the pathophysiology of major depression esis.33 Moreover, several studies found interactions seems to be necessary for the changes seen in patients

between the serotonergic system and BDNF, as the homozygous for the LA-allele. 5-HTT function is modulated by BDNF,34 which, in Another possibility for the association of brain turn, was found to be elevated in the hippocampus volume and the 5-HTTLPR polymorphism comes and the frontal cortex after antidepressant treat- from the studies reporting that the L-allele of the ment.35 Upregulation of the cAMP response ele- 5-HTTLPR may be related to Alzheimer disease.37–39 ment-binding protein due to effective therapy The larger reduction in GMVs in patients homozy- increases the expression of BDNF, which seems to gous for the L-allele may then be explained by a larger have neurotrophic effects.36 susceptibility to neurodegenerative changes. How- One might therefore expect that the high-activity ever, this seems to be unlikely, although not exclu-

LA-allele with its increased number of 5-HTT trans- sive, because our patients were young. porter proteins, concomitant decrease in serotonin The effect of stress on the brain is relevant for the levels and reduced effects on neuroplastic processes discussion of these findings. It is well known from would be more likely to cause structural changes clinical and experimental studies that stress is one of

Molecular Psychiatry Brain volumes and serotonin transporter in depression T Frodl et al 1099

Figure 2 Small-volume-corrected (SVC) gray matter volume (GMV) reduction in patients compared to controls for each genotype pair. Regions where GMV is reduced in patients with the LALA genotype compared to healthy controls with the

LALA genotype (red), LAS, LALG genotypes (green), (LGLG,LGS, SS genotypes (yellow), respectively with SVC (P < 0.01, uncorrected). Numbers below slices represent the vertical distance in millimeter to the anterior commissure. Significantly reduced GMVs in patients compared to controls are detected bilaterally in the hippocampus, bilaterally in the anterior gyrus cinguli (ACC), in the left amygdala and the right dorsomedial prefrontal cortex (DMPFC).

Molecular Psychiatry Brain volumes and serotonin transporter in depression T Frodl et al 1100 Table 4 Difference in GM volume between patients with LA/LA and those with S/S genotype (LGLG,LGS, SS) as well as

difference in GM volume in healthy controls with LA/LA compared to those with S/S genotype (LGLG,LGS, SS) with SVC for ACC, hippocampus, amygdala, DLPFC, DMPFC (P < 0.01, uncorrected)

kk(%) FWE FDR TPxyz

Major depression Left hippocampus 185 2.48 0.02 0.097 3.59 < 0.001 À26 À38 À4 Right hippocampus 281 3.71 0.012 0.097 3.76 < 0.001 30 À38 À5

Healthy controls Left ACC 40 0.36 0.24 0.144 2.55 0.006 À11 29 25 Right ACC 94 0.89 0.218 0.144 2.61 0.005 11 36 28 Left DLPFC 20 0.05 0.62 0.248 2.5 0.007 À27 7 62 Right DLPFC 867 2.12 0.063 0.248 3.6 < 0.001 28 34 29 Left DMPFC NS Right DMPFC 54 0.32 0.122 0.16 3.08 0.001 12 22 41 Left amygdala 35 1.99 0.078 0.114 2.66 0.004 À17 À6 À18 Right amygdala NS Left hippocampus 65 0.87 0.212 0.22 2.64 0.005 À17 À6 À20 Right hippocampus NS

Abbreviations: ACC, anterior gyrus cinguli; DLPFC, dorsolateral prefrontal cortex; DMPFC, dorsomedial prefrontal cortex; FDR, false discovery rate; FWE, family wise error correction; GM, gray matter; k, number of suprathreshold voxels in anatomical region; k (%), percentage of suprathreshold voxels in anatomical region, T-test; NS, not significant; SVC, small volume correction.

the major factors that contribute to the development certain 5-HTTLPR genotype did not differ with of depression. Several studies have found that the respect to age, gender, illness duration, age of onset, S-allele of 5-HTTLPR increases the vulnerability to medication and origin, it does not seem likely that the depression only in the presence of significant adverse population stratification had any negative effects. life events. Relative to homozygous individuals for Another important factor is medication. There was no the L-allele, young adult carriers of the S-allele were difference in medication between genotypes, how- more vulnerable to major depression when exposed to ever, we cannot exclude from our study an interaction adverse events.40 Young adults homozygous for the S- between medication, genotype and brain volumes, allele also had significantly greater sensitivity to because our sample is too heterogeneous with respect depressogenic effects of stressful life events than to medication and too small, to analyze this effect. carriers of the L-allele.41 An increased risk for major Further studies are therefore necessary.

depression was detected in maltreated children In summary, healthy subjects with the LGLG,LGSor homozygous for the S-allele.42 Moreover, S-alleles SS genotype have smaller GMVs than those being

independently predicted greater depression severity homozygous for the LA-allele. A hypothetical expla- and greater severity of major depression with moder- nation may be that the S-allele may have impact ate-to-severe life events compared with the higher on neurodevelopmental processes, perhaps due to expressing L-allele.43 These studies indicate a role of increased sensitivity to childhood and adolescent the S-allele in predisposing to major depression. stressful life events as suggested from genetic studies. However, one recent study found that the high- Patients with depression show the most reduced activity L-allele was associated with a higher risk of GMVs compared to controls with the same genotype

depression in the presence of adverse life events in a when they are homozygous for the LA-allele, suggest- large sample of 247 young adult female twins from ing that these patients are more vulnerable for Missouri.44 morphological or even neuroplastic changes during Also, controversial is the neurobiological interpre- depressive episodes. tation. It seems puzzling that lower serotonin uptake resulting from a genetic predisposition would be associated with a higher risk of depression because Acknowledgments lowering serotonin uptake by treatment with serotonin selective reuptake inhibitors relieves depression. This study was supported by the German Federal Therefore, more studies are necessary to explore the Research Ministry within the promotion ‘German role of the serotonin transporter in the neurobiology Research Networks in Medicine’ as part of the project of major depression. ‘German Research Network on Depression’ and Maike The primary limitation of this study is its Moergenthaler carried out her doctoral thesis within case–control design, which is sensitive to population this study. Moreover, we thank Mrs Jaquie Klesing for stratification. However, as cases and controls with a native english editing.

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Molecular Psychiatry