DOI: 10.1126/science.1223006
, 1550 (2012);337Science
et al.Thomas Agren
Human Amygdala
Disruption of Reconsolidation Erases a Fear Memory Trace in the
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Acknowledgments: D. Bottjer and M. Hogan provided advice
for 15
N2 additions. Water samples were collected with the
help of S. Curless, M. Church, S. Wilson, S. Tozzi, and the captain
and crew of the research vessel Kilo Moana. On-board flow
cytometry was made possible by K. Doggett and D. Karl. Funding
was provided by the Gordon and Betty Moore Foundation (J.P.Z.)
and the NSF Center for Microbial Oceanography: Research and
Education (C-MORE). The Max Planck Society sponsored the
HISH-SIMS analysis. We thank G. Lavik (Max Planck Institute,
Bremen) for advice and suggestions for data analysis.
J. Waterbury provided the scientific name for UCYN-A. D.V.
was supported by PHYTOMETAGENE (JST-CNRS), METAPICO
(Genoscope), and Micro B3 (funded by the European Union,
contract 287589). BIOSOPE metagenome sequencing was
performed at Genoscope (French National Sequencing Center) by
J. Poulain. We thank H. Claustre, A. Sciandra, D. Marie, and all
other BIOSOPE cruise participants. GenBank accession nos.:
JX291679 to JX291804 and JX291547 to JX291678 (see table
S4 for details).
Supplementary Materials
www.sciencemag.org/cgi/content/full/337/6101/1546/DC1
Materials and Methods
Figs. S1 and S2
Tables S1 to S6
References (34–46)
2 April 2012; accepted 20 July 2012
10.1126/science.1222700
Disruption of Reconsolidation
Erases a Fear Memory Trace in the
Human Amygdala
Thomas Agren,1
Jonas Engman,1
Andreas Frick,1
Johannes Björkstrand,1
Elna-Marie Larsson,2
Tomas Furmark,1
Mats Fredrikson1
Memories become labile when recalled. In humans and rodents alike, reactivated fear memories
can be attenuated by disrupting reconsolidation with extinction training. Using functional
brain imaging, we found that, after a conditioned fear memory was formed, reactivation and
reconsolidation left a memory trace in the basolateral amygdala that predicted subsequent
fear expression and was tightly coupled to activity in the fear circuit of the brain. In contrast,
reactivation followed by disrupted reconsolidation suppressed fear, abolished the memory trace,
and attenuated fear-circuit connectivity. Thus, as previously demonstrated in rodents, fear memory
suppression resulting from behavioral disruption of reconsolidation is amygdala-dependent also
in humans, which supports an evolutionarily conserved memory-update mechanism.
A
nxiety disorders are common, and they
cause great suffering and high societal
costs (1). The etiology involves amygdaladependent
memory mechanisms that link stressful
events to previously neutral stimuli (2), and
the amygdala has been demonstrated to be hyperresponsive
across the anxiety disorders (3).
Pharmacological and behavioral treatments of
anxiety reduce symptomatology and amygdala
activity (4) but have limited success because relapses
occur (5). However, fear memories may be
erased by recalling them and preventing their
reconsolidation (6, 7). In rodents, the amygdala
seems vital for fear memory reconsolidation
(7, 8), but this has not been investigated in
humans.
Fear conditioning, in which a previously neutral
stimulus turns into a conditioned stimulus
(CS) through pairings with an aversive stimulus,
forms a memory trace in the amygdala (2). Memory
activation produces behavioral (2, 9) and
autonomic fear reactions, such as skin conductance
responses (SCRs) (10–12), frequently used
to measure fear learning. Studies in animals (13)
and anxiety patients (14) demonstrate that extinction
weakens, but does not erase, fear memories.
In rodents (13) and humans (15) alike,
extinction attenuates conditioned fear expression
through prefrontal inhibition. Fear can return after
stress, be renewed when altering the environmental
context, or reoccur with the passage of
time (16).
By activating memories and disrupting their
reconsolidation, through protein synthesis blockade
local in the amygdala (8) or through systemic
administration of b-adrenergic receptor antagonists
(17, 18), fear memories are inhibited. Fear
memory reconsolidation can also be disrupted
behaviorally (6, 7, 19). In rodents, extinction of
fear conditioning performed 10 or 60 min after
presenting a reminder of the conditioned fear, but
not after 6 or 24 hours, inhibited fear expression
(7). Fear did not return in a new context, after
shock exposure, or with time. Thus, extinction
conducted within, but not outside, the reconsolidation
window resulted in permanent attenuation
of the fear memory (7).
In humans, extinction performed within
the reconsolidation interval also inhibited fear,
whereas extinction training performed outside
of the reconsolidation interval spared the memory
and fear returned (6). In animals, the neural
functions enabling fear memory formation
and reconsolidation are located in the amygdala
(2, 7–9, 20). In humans, lesion (21) and
brain imaging studies (10–12, 22) confirm that
the amygdala is a key area for fear memory encoding.
To test the hypothesis that reconsolidation
in humans is amygdala-mediated and that
disruption of reconsolidation inactivates a memory
trace in the basolateral amygdala, we performed
a study combining brain imaging with a
physiological measure of fear.
On day 1, twenty-two subjects (11 women)
aged 24.0 T 0.48 (mean T SEM) underwent fear
conditioning to establish an associative fear
memory (Fig. 1A and fig. S1). On day 2, the fear
memory was reactivated by presenting the cue
previously paired with the shock (CS+) for 2 min.
Subjects were randomized into two groups. One
group received extinction, consisting of repeated
CS presentations with the shock withheld, 10 min
after reactivation and thus within the reconsolidation
interval. The other group received extinction
6 hours after the reactivation—i.e., outside of
the interval. Fear expression was measured using
SCRs (6, 19). On day 3, a renewal session was
performed in a new environment, a magnetic resonance
scanner, where shock electrodes were attached,
although no shocks were delivered. SCRs
were not measured for technical reasons. On day
5, subjects were exposed to unsignaled shocks
and then re-exposed to CS+. Return of fear was
defined as the increase in SCR from the last extinction
trial on day 2 to the first reinstatement
trial on day 5 (Fig. 1B) (6).
First, we evaluated if the predicted behavioral
reinstatement effect was present on day 5. Confirming
this, increased fear responding was observed
in the 6 hours, but not the 10 min group
1
Department of Psychology, Uppsala University, SE-751 42
Uppsala, Sweden. 2
Department of Radiology, Oncology and
Radiation Science, Uppsala University, SE-751 42 Uppsala,
Sweden.
*To whom correspondence should be addressed. E-mail:
thomas.agren@psyk.uu.se
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(Fig. 1B, right panel). Groups were indistinguishable
in acquisition and extinction (Fig. 1B and
fig. S1).
Next, we tested the hypothesis that the fear
memory representation is localized to the amygdala.
Significantly greater activity was evident
bilaterally in the basolateral amygdala in the
6 hours group as compared with the 10 min
group (Fig. 1B).
We then tested if the amygdala-localized memory
predicted return of fear. Positive correlations
were present between return of fear and blood
oxygen level–dependent (BOLD) activity bilaterally
in the basolateral amygdala in the 6 hours
group (Fig. 2A). In the 10 min group, a cluster in
the right claustrum extending into the amygdala
correlated significantly with SCRs (Fig. 2A).
BOLD activity reflecting the amygdala-localized
memory trace also correlated with fear recall
during extinction the previous day in the 6 hours,
but not the 10 min, group (Fig. 2B).
Fig. 1. Extinction during reconsolidation blocks
reinstatement of fear and abolishes a memory
trace in the amygdala. (A) After fear conditioning
on day 1, when 16 shocks were paired with a visual
cue, a memory reminder was given on day 2, and
extinction was performed after 10 min or 6 hours,
by exposure to eight conditioned cues with no
shocks. On day 3, amygdala activity was assessed
with functional magnetic resonance imaging
(fMRI) during renewal-induced fear. On day 5,
return of fear was evoked by presenting unpaired
shocks before CSs were again presented. (B)
Groups were equivalent in acquisition [t(20) =
0.66, P = 0.51] and extinction [t(20) = 1.03, P =
0.31]. Return of fear was confirmed in the 6 hours
group [blue bar; t(10) = 2.72, P = 0.02] but not in
the 10 min group [red bar; t(8) = 0.23, P = 0.82].
fMRI demonstrated a remaining fear memory
representation in the amygdala after reactivation
and normal reconsolidation but not after reactivation
followed by disrupted reconsolidation. The
voxels reflecting the bilateral memory trace, encompassing the basolateral
amygdala, indicate superior memory representation in the 6 hours as compared
with the 10 min group (brain coordinates: x, y, z = 27, 5, –17; Z-score =
2.46;P=0.007;378mm3
;x,y,z=–15,–1,–14;Z=2.22;P=0.013;162mm3
).
The autonomic nervous system measure of fear is the SCR. The CNS measure
of amygdala activity is BOLD activity. Brain coordinates are according
to the Montreal Neurological Institute (MNI). Error bars are standard error
of means.
Renewal
Day 3
Reinstatement
Day 5
Reactivation
Day 2
24 h
10 min
24 h
Renewal
Day 3
Reinstatement
Day 5
6 h
48 h
A
B
Fear conditioning
Day 1
Extinction
Day 2
Extinction
Day 2
24 h
0
0,1
0,2
0,3
0,4
6 h
10 min
-0,2
-0,1
0
0,1
0,2
0,3
0,4 *
ExtinctionAcquisition
SCR
SCR
Return of fear
48 h
6 h
10 min
Fig. 2. Amygdala activity predicts return of fear
and correlates with recall of fear. (A) In the 6 hours
group (top), activity bilaterally in the basolateral
amygdala predicted return of fear 2 days later (x, y,
z = 21, –1, –17; Z = 2.06; P = 0.002; 999 m3
; x, y,
z = –21, –4, –14; Z = 2.38; P = 0.009; 1107 m3
).
In the 10 min group (bottom), an area in the right
temporal claustrum extending into the amygdala
was also related to SCR (x, y, z = 33, 2, –23; Z =
2.49; P = 0.006; 324 mm3
). Because fear did not
return in the 10 min group, the correlation may
reflect individual brain-behavior relations unrelated
to fear and the experimental manipulation.
(B) In the 6 hours group (top), recall of fear
during extinction covaried with the strength of
amygdala activity bilaterally (x, y, z = 24, –1, –20;
Z = 2.35; P = 0.009; 378 mm3
; x, y, z = –15, 4,
–17; Z = 2.27; P = 0.012; 189 mm3
). No covariation
existed in the 10 min group (bottom).
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Amygdala areas harboring the memory trace
(Fig. 1B) and areas covarying with return of fear
(Fig. 2A) overlapped in the 6 hours group only
(Fig. 3A). Moreover, the memory trace was colocalized
to areas involved in fear memory recall
during extinction (Fig. 3B). Finally, all these areas
overlapped with each other only in the 6 hours
group (Fig. 3C). Thus, the localization of the
memory trace in the amygdala overlapped bilaterally
with areas related both to recall of fear
during extinction and return of fear during reinstatement.
The hypothesis that memory was
not erased, but only suppressed, by extinctionmediated
prefrontal inhibition was not supported
because the theoretically predicted (13, 15) negative
coupling between activity in the ventromedial
prefrontal cortex (vmPFC) and return of
fear was absent because vmPFC activity did not
correlate negatively with fear in either group
(Z-scores of <1).
Finally, we evaluated if activation of the fear
memory in the amygdala was linked to activity in
other nodes of the fear network (23) by calculating
the covariation between memory-associated
amygdala activity and activity in the remaining
network. Our amygdala seed of interest correlated
strongly with activity bilaterally in the
insula, hippocampus, and the midline anterior
cingulate cortex and significantly more so in the
6 hours than in the 10 min group (Fig. 4). No
clusters showed a better correlation with the amygdala
seed in the 10 min group. This suggests that
the amygdala could be the primary site of memory
plasticity, but also influence reconsolidation
by affecting other regions of the fear network.
The amygdala could thus play a modulatory, rather
than a solitary, role in human fear reconsolidation
processes.
In summary, whereas the amygdala memory
representation after activation and undisrupted
reconsolidation predicted return of fear and was
functionally coupled to other nodes of the brain’s
fear network, disruption of reconsolidation significantly
weakened the amygdala memory and
its coupling, rendering it unrelated to both recall
and return of fear. We conclude that extinction
training initiated during reconsolidation abolishes
fear expression by erasing a memory trace in the
amygdala. Reactivated fear memories are sensitive
to behavioral disruption (6, 7, 19), and the
amygdala proves to be a key neurobiological
substrate for this process also in humans. This
mechanism holds great clinical promise in anxiety
treatment (6, 17–19) in order to dissociate
fear from cognitive memory.
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Acknowledgments: We thank the Swedish Research Council,
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Life and Social Research for support.
Supplementary Materials
www.sciencemag.org/cgi/content/full/337/6101/1550/DC1
Materials and Methods
Supplementary Text
Fig. S1
References (24–28)
5 April 2012; accepted 26 July 2012
10.1126/science.1223006
Fig. 3. Areas in the amygdala that correlate with recall and return of fear are colocalized with
the memory trace. (A) The amygdala areas reflecting the memory trace (Fig. 1) overlapped with
the areas that predicted return of fear in the 6 hours group (x, y, z = 24, 2, –17; 189 mm3
; x,
y, z = –15, –1, –14; 108 mm3
), illustrated in Fig. 2A (top). (B) The localization of the memory
trace also overlapped with areas involved in recall during extinction (x, y, z = 24, 2, –20; 81 mm3
;
x, y, z = –15, –4, –17; 135 mm3
) illustrated in Fig. 2B (top), in the 6 hours group only. (C) These
three areas overlapped with each other (x, y, z = 24, 2, –20; 135 mm3
; x, y, z = –15, –1, –14;
27 mm3
).
Fig. 4. Amygdala memory trace activity is functionally coupled to the fear network in the brain after
normal memory reconsolidation but not after disrupted reconsolidation. The connectivity analysis, using
BOLD activity in the amygdala from areas representing memory trace activity in the 6 hours group as a
seed of interest, demonstrated stronger functional couplings in the 6 hours than in the 10 min group in
structures forming the fear-circuit of the brain, including the midline anterior cingulate cortex (A and B)
(x, y, z = –9, 47, 13; Z = 3.14; 5994 mm3
), bilateral insula (A and C) (x, y, z = 33, 20, 7; Z = 3.28; 1890 mm3
;
x, y, z = –27, 29, 10; Z = 2.95; 4077 mm3
), and bilateral hippocampus (C), although only the right
hippocampus is illustrated (x, y, z = –27, –13, –14; Z = 2.51; 297 mm3
; x, y, z = 30, –25, –8; Z = 2.51;
total 675 mm3
).
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