1
Supplementary information
2
3
Results conditioning subjective ratings study 1
4
Analyses of variance of the arousal ratings revealed significant main effects of CS-type
5
(F1,58 = 101.56, p < .001) and conditioning phase (F2.62, 152.06 = 18.6, p < .001) as well as a significant
6
phase x CS-type interaction (F2.38, 138.00 = 31.14, p < .001). Follow-up paired t-tests displayed arousal
7
differentiation after acquisition 1 (t = 10.49, p < .001), acquisition 2 (t = 13.68, p < .001) as well as
8
after extinction (t = 4.32, p < .001). Analyses of variance of the valence ratings revealed significant
9
main effects of CS-type (F1,58 = 192.78, p < .001) and conditioning phase (F2.41, 139.62 = 13.14, p < .001)
10
as well as a significant phase x CS-type interaction (F2.51, 145.84 = 61.50, p < .001). Follow-up paired t-
11
tests displayed valence differentiation after habituation (t = -2.26, p = .03), acquisition 1 (t = 14.86,
12
p < .001), acquisition 2 (t = 13.74, p < .001) as well as after extinction (t = 5.23, p < .001). Analyses of
13
variance of the contingency ratings revealed significant main effects of CS-type (F1,58 = 236.18,
14
p < .001) and conditioning phase (F3, 174 = 53.21, p < .001) as well as a significant phase x CS-type
15
interaction (F2.47, 143.29 = 86.40, p < .001). Follow-up paired t-tests displayed contingency awareness
16
after habituation (t = -2.06, p = .04), acquisition 1 (t = 14.30, p < .001), acquisition 2 (t = 22.33,
17
p < .001) as well as after extinction (t = 3.57, p = .001). Interactions with between-subjects factor CS-
18
duration were not significant for phase or CS-type nor was the three-way interaction phase x CS-type
19
x CS-duration for any type of rating, indicating no influence of the differing CS-durations.
20
21
22
1
1
2
Results morphological correlations without exclusion of subjects without acquisition SCR
(n = 10) study 1
3
4
Table 1:
5
Correlations between cortical thickness and the differential SCR of the first extinction block without
6
exclusion of subjects that show no SCR during acquisition
7
cluster
anatomical label
Brodman area
MNI*
# vertices Tpeak pclus
coordinates
[2, 18, -7]
376
4.29 < .001
1
right subgenual area
25
2
left pars orbitalis
47
-
-
-
n.s.
3
right orbitofrontal
25
[9, 8, -18]
10
4.77
.016
8
9
10
Results morphological correlations alternative extinction measures study 1
11
12
To ensure that the significant correlations are not restricted to the specific extinction measure we chose
13
(subdivision into three blocks), we repeated the analyses (GLM with age, gender and differential SCR
14
during acquisition as nuisance factors) with alternative measures of extinction learning.
15
Extinction learning index 2 covered the summed differential SCRs (CS+unpaired minus CS-) from the
16
first two to up to nine trials of extinction learning separately (See Table 2). Results show the stability
17
of the association of the right subgenual cluster with extinction learning over a varying number of
18
trials.
2
1
Table 2:
2
Correlations between cortical thickness and the differential SCR of varying amount of extinction trials
# extinction
Cluster #1 right subgenual area
Cluster #2 left pars orbitalis
trials
(BA25)
(BA47)
1-2
[2, 23 -10]
272 vertices
Tpeak = 4.15, pclus = 0.0082
1-3
1-4
1-5
[3, 18, -8]
124 vertices
[-31, 14, -17]
12 vertices
Tpeak = 3.76, pclus = 0.0179
Tpeak = 5.93,
pclus = 0.0054
[3, 18, -9]
[-31, 14, -17]
11 vertices
65 vertices
Tpeak = 3.73, pclus = 0.032247
Tpeak = 5.78, pclus = 0.0092
[3, 18, -9]
[-31, 14, -17]
245 vertices
Tpeak = 3.9, pclus = 0.0014
1-7
[3, 17, -8]
8 vertices
Tpeak = 5.08, pclus = 0.055 (ns)
144 vertices
Tpeak = 4.51, pclus = 0.0000381
1-8
[3, 18, -8]
13 vertices
Tpeak = 4.83, pclus = 0.00444
1-9
[3, 18, -8]
16 vertices
Tpeak = 4.88, pclus = 0.0017
3
4
Extinction learning index 3 was computed as the difference between early and late extinction SCRs
5
(i.e., the mean differential SCR of the first 9 extinction trials minus the mean differential SCR of the
6
last 9 extinction trials). Note that in line with the earlier extinction measures higher values for this
7
measure represent a smaller amount of fear extinction, as a trial by trial inspection of SCR data (See
8
Figure 1) shows that complete extinction is achieved in the first half of the extinction phase (and
9
amount of acquired fear is already controlled for in the GLM). The GLM revealed a significant
10
negative correlation with a small cluster of 8 vertices (T = 4.53, p = .045) in the right vmPFC (4, 8, -8)
11
overlapping with cluster #1 that emerged with extinction learning index 1.
3
1
Extinction learning index 4 was computed as the difference between peak acquisition and late
2
extinction (i.e., the highest SCR to a CS+ unpaired during any acquisition trial minus the mean CS+ unpaired
3
SCR during the last 9 extinction trials). The GLM (only correcting for age and gender) with this
4
measure revealed no significant correlation with cortical thickness at any given vertex. Adding a mask
5
encompassing the prefrontal regions of interest to lower significance thresholds did not reveal any
6
association either. This might be due to the fact that index 4 does not take into account the pronounced
7
interindividual variance during early extinction. As differentiation of SCRs during late extinction is
8
minimal, this measure might primarily reflect interindividual differences in fear acquisition.
9
0.18
CS+
0.16
CS-
0.14
SCR
0.12
0.10
0.08
0.06
0.04
0.02
0.00
1
2
3
4
5
6
7
8
9 10 11 12 13 14 15 16 17 18
extinction trial
10
11
Figure 1. Single trial SCRs of the extinction phase study 1
12
13
4
1
Correlation between function and structure study 1
2
To identify a potential association between structural properties and functional activation of a region
3
we extracted the functional eigenvalues for the differential contrast CS+ > CS- during extinction for
4
the three prefrontal clusters that emerged from the structural analyses (see Figure 4 main article).
5
Bivariate correlation analyses revealed no association between cortical thickness and strength of
6
activation in these clusters (see Figure 1).
7
8
9
Figure 1. Scatterplot of mean cortical thickness and functional eigenvalues for the contrast CS+>CS-
10
during extinction for the three prefrontal clusters (r = -0.11, p = .41; r = 0.15, p = .26; r = -0.45,
11
p = .74 resp.).
12
13
Table 3:
14
Non-significant correlations of trait anxiety with fear acquisition, extinction learning and amygdalar
15
volume in both samples
Early
Late
Extinction
Right
Left
acquisition a
acquisition a
learning b
amygdala c
amygdala c
Original sample
r=-.14
r = .04
r = .04
r = -.25
r = -.26
STAI
p = .35
p = .80
p = .74
p = .09
p = .08
Replication sample
r = -.02
r = .13
r = .06
r = -.01
r = .14
STAI
p = .88
p = .41
p = .66
p = .95
p = .40
M (SD)
32.9 (6.1)
M (SD)
36.9 (8.9)
16
17
18
a
corrected for differential SCR during habituation
corrected for differential SCR during acquisition
c
corrected for age, gender and intracranial volume.
b
5
1
Replication study
2
Methods
3
Sixty-five subjects, likewise being paramedics in training from the same longitudinal study underwent
4
the identical conditioning paradigm while the same subjective ratings and skin conductance responses
5
were recorded and preprocessed in the same manner. Structural magnetic resonance imaging data were
6
acquired on a 1.5 T MAGNETOM Vision whole body scanner (Siemens Medical Solutions, Erlangen,
7
Germany) equipped with a head volume coil using a T1-weighted three-dimensional fast low angle
8
shot (FLASH) sequence (TR 15 ms, TE 5 ms, field of view 220 x 220 mm2, 170 sagittal slices, voxel
9
size, 0.86 x 0.86 x 0.86 mm3)). A non-local means filter (Coupe et al. 2006) was applied in order to
10
minimize noise. Exclusion of subjects without skin conductance reactions (n = 12) resulted in a total
11
of 53 healthy subjects (18 female, mean age = 21.83, SD = 2.98) in the replication study. As in
12
study 1, we excluded participants with no SCR during either acquisition phase (n = 11) from
13
extinction analyses.
14
Results
15
Identical to study 1 successful conditioning was observed in skin conductance and subjective ratings
16
(data not shown). In line with the results of the main study, partial correlation analyses of the
17
differential SCR during late acquisition and amygdala volume correcting for intracranial volume, age,
18
gender and years of education yielded a significant positive correlation of fear acquisition with right
19
amygdala volume (r = .36, p = .04) (See Figure 2). Linear regression analysis on the relative
20
contribution of the SCRs to the CS+unpaired and the CS- during late acquisition again revealed a
21
significant result (R2 = .102, F = 2.94, p = .04). Again the model gained its significance by including
22
both CS+unpaired and CS-; the single individual predictors were not significant (CS+ unpaired: b = 0.27,
23
t49 = 1.32, p = .132; CS-: b = -0.08, t49 = .392, p = .70).
24
Only one additional subcortical structure correlated with the differential SCR of any acquisition phase
25
(right caudate nucleus correlated negatively with acquisition 1 (r = -.29, p = .03).
26
6
1
2
3
4
Figure 2. Scatterplot depicting the correlation of differential late fear acquisition with right amygdala
5
volume (r = .30, p = .04) (n = 53)
6
7
1
We used a similar vertex-wise general linear model controlling for age, gender and level of acquired
2
fear with an additional mask encompassing the prefrontal region of interest from study number one,
3
build up by the following regions: medial orbital frontal, rostral anterior cingulate and frontal pole
4
(Desikan et al. 2006). Again we found a significant negative correlation of the differential SCR during
5
the first extinction learning block with the thickness of a cluster within the medial prefrontal cortex of
6
the right hemisphere containing 87 vertices located in the medial frontal gyrus (Brodman area 11)
7
(peak: x = 5, y = 42, z = -16), T = 3.80, pclus = 0.02 (See Figure 3). Neither differential SCR of block 2
8
or 3, nor differential SCR of the whole extinction phase exhibited any morphological correlate on the
9
cortex. Without exclusion of subjects showing no SCRs during acquisition these parameters changed
10
slightly (peak: x = 5, y = 42, z = -16, T = 3.94, pclus = 0.015, 124 vertices). Independent consideration
11
of the extinction SCR to the CS+ and CS- of the first extinction block revealed no significant
12
association between cortical thickness at any vertex and SCR to the CS+ or the CS-.
13
14
A)
15
16
17
Figure 3. Cortical thickness cluster correlating with extinction learning index; controlled for
18
age, gender and level of acquired fear (n = 42). A) Significant cluster on the cortical surface (FWE-
19
corrected for vmPFC mask) B) Correlation plot of extinction learning index and mean cortical
20
thickness of this cluster (r = -.49, p < .001)
B)
21
22
23
8
1
2
3
4
Coupe P, Yger P, Barillot C (2006) Fast non local means denoising for 3D MR images Med
Image Comput Comput Assist Interv 9:33-40
Desikan RS et al. (2006) An automated labeling system for subdividing the human cerebral
cortex on MRI scans into gyral based regions of interest Neuroimage 31:968-980
9