Participants

All actively participating children from the Nijmegen Longitudinal Study on Child and Infant Development (n = 116) were approached to take part in this imaging study. Anatomical scans were obtained from participants at 14 (M = 14.6, SD = 0.17) and 17 years of age (M = 17.09, SD = 0.15). Forty-nine at the first imaging time-point and ninety-six at the second imaging time-point agreed to participate. Participants who could not undergo magnetic resonance imaging (MRI) or who had missing data at one of the two time-points were excluded from these analyses. The final sample consisted of 37 adolescents (15 boys). Participants did not have a history of psychiatric disorders or neurological illness (as indicated by parent/guardian report). Table 3 presents the characteristics of the sample. Written informed consent was obtained from parents and participants during each measurement wave. The study was approved by the local ethics committee (CMO region Arnhem – Nijmegen) and was conducted in compliance with these guidelines.

Table 3 Sample characteristics. Full size table

Life events

The experience of early-life events (before age 5) and current life events (between age 14 and 17) were assessed via parent report. All life event reports were collected within one to two years after the event had taken place. This meant that for early-life events, reports were taken at 15 months, 28 months, and 5 years. For current adolescent life events, the report was taken at age 17 for reports until age 14. The life events questionnaire consisted of items selected from Sarason, Johnson, and Siegel’s Life Experiences Survey70 and Coddington’s Life Events Scale for Children71 based on the likelihood they would have an aversive influence on the child’s development72. Both measures have been widely used in international research73,74. The life events questionnaire remained the same at all measurement times and has previously been used in this longitudinal study75,76. Items require a ‘yes’ or ‘no’ response. The score represents the total number of negative personal life events in the given assessment period and was calculated for events that took place until early childhood (until age 5) and during late adolescence (i.e., between ages 14 and 17).

Social environment

We used age-relevant measures of the individual’s social environment (SE). The quality of parent-child interactions was used as an index of early SE. Poor parent-child interaction quality has previously been shown to be related to elevated childhood cortisol levels in the NLS cohort29. Parent-child interactions were assessed at 15 months, 28 months, and 5 years of age during a home visit29,75. Video recordings of these interactions were rated by four trained observers on five 7-point scales: Supportive Presence, Respect for Child’s Autonomy, Structure and Limit Setting, Quality Instruction, and Hostility. An average score for SE before age 5 was taken across all scales (with reversed coding for hostility scores) and time-points for each child77. In case of a missing assessment (n = 2 cases) the average was computed based on the two remaining time points.

Peer environment (social preference) between age 14 and 17 was assessed in the classroom with a well-established sociometric measure previously used in this cohort78,79. Children were asked to nominate classmates who they liked (“Who do you like the most?”) and disliked (“Who do you like the least?”). Students were asked to nominate at least one classmate, excluding self-nominations. There was no maximum number of nominations. For each question, the number of nominations that a child received was counted and standardized within the classroom, to control for differences in classroom size. A score for social preference was calculated by subtracting the liked least from the liked most score. This difference score was again standardized within classrooms.

Socioeconomic Status

SES scores were computed based on education (7-point scale) and occupation (6-point scale) levels for both parents in line with previous reports on this cohort75. The levels of education and occupation for the two parents were first standardized and then summed to create a single score per parent. The final SES score was derived by taking the average score of the mother and father.

Behavioral measures of psychopathology

Internalizing symptoms at age 17 were measured using the Child Behaviour Checklist (CBCL)80. The CBCL is a parent-report questionnaire used to assess the frequency of emotional and behavioral problems exhibited by the adolescent in the past six months. The parent rated each behavior or symptom on a three-point Likert scale (not true, somewhat or sometimes true, very true or often true). Items from the scales anxious/depressive, withdrawn/depressive, and somatic complaints were summed to provide a score for internalizing symptoms.

Specific aspects of socialization during adolescence was measured with the Inventory of Callous Unemotional Traits 81. Self-report and parent-report versions of the questionnaire were used to assess the occurrence and intensity of affective features of callousness such as lack of empathy, disregard for others, and shallow affect. It consisted of 24 items scored on a four-point Likert scale (not at all true, somewhat true, very true, definitely true). The self-report and parent-report versions were significantly correlated with each other (r = 0.42, p = 0.013). A mean score was created from both versions to increase consistency of the measure. For two participants with missing data (1 self-report, 1 parent-report) the available score was used for further analysis.

The statistical threshold for correlations of GMV with psychopathology measures were set to p < 0.025 (multiple correction for number of regions tested for each measure).

Imaging parameters

Structural T1 images were acquired at 3 Tesla using Siemens MAGNETOM Trio or PRISMA systems (acquired at the same site; 18 participants at age 17) with a 32-channel coil. Images were acquired using the same MPRAGE sequence (TR = 2300 ms; TE = 3.03 ms; 192 sagittal slices; 1.0 × 1.0 × 1.0 mm voxels; FOV = 256 mm). To ensure that there were no differences in the quality of T1 images acquired on the TRIO and PRISMA scanners at age 17, these normalized and smoothed GM images were checked using the “Check sample homogeneity” function in CAT12 (Computation Anatomy Toolbox). One participant was identified as a potential outlier for manual inspection. After manually checking the data for artefacts, it was included in the analyses.

Voxel Based Morphometry

Magnetic resonance images were processed using the Matlab toolbox SPM12 [Statistical Parametric Mapping (www.fil.ion.ucl.uk/spm)]. Each MR image was checked for artifacts or anatomical abnormalities and alignment to the anterior commissure. Using a pairwise longitudinal registration approach82 a Jacobian difference map was generated as well as a “halfway space” image, which was subsequently segmented into white matter, grey matter (GM), and cerebrospinal fluid (CSF). Diffeomorphic anatomical registration through exponentiated lie algebra (DARTEL) was used for inter-subject registration of the GM “halfway” images to a group average template image83. The GM “halfway” image was multiplied by the Jacobian difference map for each participant. This subsequent GM difference map was transformed and resampled at an isotropic voxel size of 1.5 mm, resulting in spatially normalized, Jacobian scaled, and smoothed (8 mm FWHM Gausian kernel) images in Montreal Neurological Institute (MNI) space. Data quality of normalized and smoothed difference images was checked with CAT12 using the “Check sample homogeneity” function. This function did not indicate any potential outliers - based on a mean correlation of the sample below 2 standard deviations. The GM images were entered into a multiple regression analysis with standardized scores of life events and social environment as early (0–5 years) and current (14–17 years) stressors entered as covariates. Gender and average (age 14 and 17) grey and white matter total brain volume (TBV) were entered as covariates of no interest. To minimize boundary effects, a binary mask of the group template was used to exclude voxels outside of the brain. Statistical significance was assessed using non-parametric permutation tests using the Threshold Free Cluster Enhancement (TFCE) Toolbox in SPM12 (Version 90; http://dbm.neuro.uni-jena.de/tfce/) with 5000 permutations. After TFCE, the statistical threshold was set to p < 0.05 adjusted for family wise error at a whole brain level. TFCE suppresses random noise that may have a similar intensity as the real signal, but lacks spatial continuity (smoothness). The TFCE values at each voxel represent a combination of spatially distributed cluster size and height information. In other words, the TFCE statistic summarizes the cluster-wise evidence at each voxel. There is no initial threshold for voxel level inference. Statistical inference is based on the distribution of TFCE values - derived from the non-parametric permutations. This type of approach is particularly beneficial for VBM data84,85. Anatomical inference was drawn by superimposing images on a standard SPM single-subject T1 template, the group-specific average template (created in DARTEL), and subject-specific T1 scans standardized in MNI space.

Behavioral relevance of longitudinal GMV changes

To relate the longitudinal GMV changes observed in adolescents to psychopathology, that is callous unemotional traits and internalizing symptoms, GMV changes were extracted from the relevant significant clusters. To achieve anatomical specificity, an overlap was taken between the significant cluster and the brain area, based on the Automated Anatomical Labeling (AAL) Atlas86. As such, the grey matter estimates reflected only the significant changes in an anatomically defined area. To test the association between adolescent-social-stress-related volumetric changes and callous unemotional traits, we identified regions significantly modulated by adolescent social stress that have previously been also identified as part of the callous unemotional neuro-profile in adolescent samples59,60,87, namely the anterior cingulate cortex (only the right hemisphere in our study) and bilateral orbital frontal cortex. Parameter estimates of grey matter volume changes of these two regions were entered into SPSS and JASP for correlational analysis.

To test the association between early-life stress-induced volumetric changes and internalizing symptomology, we identified regions modulated by negative personal early-life events that have previously been suggested as developmental targets for internalizing disorders, namely the amygdala-prefrontal circuit and anterior cingulate cortex4,67,68. GMV changes were extracted from these relevant clusters and analyzed for associations with internalizing symptoms, following the same procedure described for callous unemotional traits.

Finally, two post-hoc analyses were conducted to rule out the effects of additional stressors. The first model included standardized SES scores as an additional regressor in the previously described multiple regression analysis. The second model explored the interaction between early and current stressors. The standardized interaction scores of personal early-life events and adolescent peer environment (the two significant predictors of GMV changes) were entered into the original multiple regression analysis. For both analyses, all other parameters were kept the same as in the original model.

Data Availability

The data that support the findings of this study are available from the corresponding author upon request.

Code Availability

The code used to analyze the data is available from the corresponding author upon request.