Supplementary Materials and Methods (doc 554K)

Exclusion criteria
During screening, participants were checked for psychiatric, neurological, cardiovascular or endocrine
disease, irregular sleep/wake rhythm, non-admissibility to the MRI scanner, smoking (>5 cigarettes
weekly), alcohol consumption (>21 beverages weekly), use of recreational drugs (>weekly), psychotropic
medication, and hepatic, cardiovascular, or renal impairments. Athletes were excluded because of a
possible positive doping test result after spironolactone intake. All participants had normal or correctedto-normal vision and normal hearing. They had to refrain from any medication other than paracetamol
for acute pain and recreational drugs for 72h, alcohol for 24h, and coffee for 3h before testing.
Stress measurements
For negative mood, the Positive and Negative Affect Scale (PANAS, Watson et al, 1988) was administered
either on paper or presented on the screen in the MRI scanner, programmed in Presentation®
(Neurobehavioral Systems, Inc.). Sum scores for negative mood were calculated per time point. Vital
signs were measured using an automatic blood pressure monitor with arm cuff (Intellisense®, OMRON,
the Netherlands) outside the MRI scanner. Heart rate during scanning was acquired using the heart rate
device of the MRI scanner, peak-scored using in-house software and averaged over four time bins of
1min to cover the task. To measure cortisol levels, saliva samples were taken using Salivettes® (Sarstedt,
Germany). For each sample, participants were asked to chew the cotton swab gently for 1min. The
samples were stored at -24°C until they were analyzed by the Dresden LabService (Germany). After
thawing, the samples were centrifuged and analyzed using a commercially available chemiluminescence
immunoassay with high sensitivity (IBL Inc.). The cortisol levels were not normally distributed and
normalized using log-transformation.
fMRI preprocessing
For spatial realignment, head motion was estimated on the first echo using least-squares estimation and
applied to the 5 echoes acquired for each excitation using 6 rigid-body transformation parameters. The
echo images per volume were then combined into a single volume using an optimized echo weighting
procedure (Poser et al, 2006). The functional images were coregistered to the structural image using
rigid-body transformations. The T1-image was segmented into cerebral spinal fluid (CSF), white matter
(WM) and gray matter and used to normalize functional and structural scans to MNI space with the
unified segmentation procedure. Finally, the images were spatially smoothed using an 8mm FWHM
Gaussian kernel.
Stress measures in the experiment phase
Negative Mood. We found main effects of stress (F(1,91)=10.907, p=0.001) and time (F(2.4,218.4)=12.954,
p<0.001), and a time-by-stress interaction (F(2.4,218.4)=9.812, p<0.001, Figure 1). Post-hoc tests revealed
stronger negative mood in the stress groups at 5min (trend level, p=0.096) and 45min after stress
induction (p<0.001) compared to the control groups. No other significant group differences were found.
However, we found a trend for a time-by-stress-by-MR-blockade interaction (F(2.4,218.4)=2.692, p=0.060,
Figure 1). Post-hoc tests revealed a time-by-stress interaction only in the MR-available groups
(F(2.2,93.5)=10.269, p<0.001), but no significant stress-related increase in negative mood over time in the
MR-blocked groups. It has been reported before, mostly in rodents, that the MR is involved in reactivity
to novel situations, behavioral flexibility, and coping strategies (Berger et al, 2006; de Kloet et al, 1999;
Oitzl & Dekloet, 1992; Oitzl et al, 1994). However, in humans this effect of MR-blockade on appraisal is
not supported yet. Thus, we interpret our finding as first evidence supporting a role of the MR in
appraisal of novel or stressful situations in humans, but a replication of this finding is needed.
Cortisol. One participant from the Control/MR-blocked group was removed from this analysis because of
excessive cortisol levels (both cortisol at 100min after stress and the increase of cortisol from 70 to
100min exceeded the mean + 3SD of the group). We found main effects of stress (F(1,92)=13.004, p=0.001)
and MR-blockade (F(1,92)=15.013, p<0.001), time-by-stress (F(2.5,229.5)=8.927, p<0.001), and time-by-MRblockade (F(2.5,229.5)=6.217, p=0.001, Figure 1) interactions, but no time-by-stress-by-MR-blockade
interaction. Post-hoc tests showed time-by-stress interactions in both drug groups (MR-available:
F(2.5,114.4)=3.018, p=0.041; MR-blocked: F(2.3,105.6)=6.933, p=0.001). In the MR-available groups, the stress
group had higher cortisol levels than the control group at 45min (p=0.044) and at trend level 70min after
stress (p=0.085). Within the MR-blocked groups, stressed individuals had higher cortisol levels from
15min after stress onwards (all p<0.05). Drug effects were found from 5 to 100min, with the MR-available
groups having lower cortisol levels than the MR-blocked groups (all p<0.05) in line with an impaired
negative feedback in the MR-blocked group. No other significant group differences were found.
Heart rate and blood pressure. We found main effects of stress (F(1,88)=4.665, p=0.033) and time
(F(2.9,252.3)=14.721, p<0.001) on heart rate. Stressed participants had higher heart rates than control
participants, indicative of heightened NE levels. No significant main effect of or interaction with MRblockade was present. Both systolic and diastolic blood pressure were unaffected by stress and MRavailability.
Importantly, results did not change when raw data (log-transformed in the case of cortisol) and not
difference scores were used.
Figure S1: Brain areas showing connectivity to the basolateral amygdala (BLA, left) or the centro-medial
(CMA, right) during the emotional face-matching task, plottet on a template brain. All p<0.05, FWE
corrected (peak level).
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