Neurobiology and Genetics of Generalized Anxiety Disorder

Anne Schienle, PhD; John M. Hettema, MD, PhD; Ricardo Cceda, MD, PhD; and Charles B. Nemeroff, MD, PhD
Psychiatric Annals, Volume 41, Issue 2, February 2011
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1. Describe regional brain abnormalities implicated in general-
ized anxiety disorder (GAD).
2. Review the neurotransmitter/neuropeptide system as
involved in GAD.
3. Discuss the complex nature of the genetics of GAD.
EDUCATIONAL OBJECTIVES CME
Anne Schienle, PhD, is with the Department of Clinical Psychology,
University of Graz, Graz, Austria. John M. Hettema, MD, PhD, is with the
Department of Psychiatry, Virginia Institute for Psychiatric and Be-
havioral Genetics, Virginia Commonwealth University, Richmond, VA.
Ricardo Cceda, MD, PhD; and Charles B. Nemeroff, MD, PhD, are with
the Department of Psychiatry and Behavioral Sciences, University of
Miami Miller School of Medicine, Miami, FL.
Address correspondence to: Charles B. Nemeroff, MD, PhD, Depart-
ment of Psychiatry and Behavioral Sciences,University of Miami,Miller
School of Medicine, 1120 NW 14 Street, Suite 1455, Miami, FL 33136;
fax: 305-243-8532; or e-mail: cnemeroff@med.miami.edu.
Dr. Schienle and Dr. Hettema have disclosed no relevant nancial
relationships. Dr. Cceda has disclosed the following relevant nan-
cial relationships: the Arsht Foundation, National Institute on Alco-
hol Abuse and Alcoholism (NIAAA), National Institute on Drug Abuse
(NIDA), and National Institute of Mental Health (NIMH): Grant/re-
search support; and MyHealthWin LLC: Partial Owner. Dr. Nemeroff
has disclosed the following relevant nancial relationships: Ameri-
can Foundation for Suicide Prevention (AFSP) and NovaDel Pharma:
Member of Board of Directors; Takeda and Xhale: Consultant; Agency
for Healthcare Research and Quality (AHRQ) and National Institutes
of Health (NIH): Grant/research support; Method and devices for trans-
dermal delivery of lithium (US 6,375,990B1), and method of assessing
antidepressant drug therapy via transport inhibition of monoamine
neurotransmitters by ex vivo assay (US 7,148,027B2): Patent holder;
Anxiety Disorders Association of America (ADAA), AFSP, CeNeRx
BioPharma, National Alliance for Research on Schizophrenia and De-
pression (NARSAD), NovaDel Pharma, PharmaNeuroBoost, Member
of Scientic Advisory Boards; CeNeRx BioPharma, Corcept Therapeu-
tics, NovaDel Pharma Inc., PharmaNeuroBoost, Revaax Pharma, and
Xhale: Stockholder; and CeNeRx BioPharma, PharmaNeuroBoost:
Other nancial interests.
doi: 10.3928/00485713-20110203-10
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4102NemeroffCS.indd 2 2/11/2011 3:30:59 PM
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This activity has been planned and implemented in accordance with the
Essential Areas and policies of the Accreditation Council for Continuing Medical
Education through the joint sponsorship of Vindico Medical Education and
PSYCHIATRIC ANNALS. Vindico Medical Education is accredited by the ACCME to
provide continuing medical education for physicians.
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with the extent of their participation in the activity.
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In accordance with the Accreditation Council for Continuing Medical Educations
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activity audience the relevant nancial relationships of the planners, teachers, and
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The audience is advised that this continuing medical education activity may
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HOW TO OBTAI N CME CREDI TS BY READI NG THI S I SSUE
The criteria for the diagnosis of generalized anxiety disorder (GAD) have evolved over the years, particularly in each new iteration of the
Diagnostic and Statistical Manual. GAD can be a challenge to diagnose and treat because of the treatment options available, and the myriad
symptom overlap between GAD and other depressive/anxiety disorders.
EDUCATIONAL OBJECTIVES OVERVIEW
88
M C E
Psychotherapy for Generalized Anxiety Disorder
Jrgen Hoyer, PhD; Colin van der Heiden, PhD; and Michael E. Portman, DPhil, LISW-S
95
M C E
Pharmacotherapy for Generalized Anxiety Disorder
Martin A. Katzman, BSc, MD, FRCPC; Alex Copeland, BSc; Larry J. Klassen, MD; Pratap Chokka, MD;
and Olga Brawman-Mintzer, MD
104
M C E
Treatment-Refractory Generalized Anxiety Disorder
Vladan Starcevic, MD, PhD; and Sean Hood, MD, MSc
113
M C E
Neurobiology and Genetics of Generalized Anxiety Disorder
Anne Schienle, PhD; John M. Hettema, MD, PhD; Ricardo Cceda, MD, PhD; and
Charles B. Nemeroff, MD, PhD
TABLE OF CONTENTS
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RESPONSIBILITY FOR STATEMENTS
4102NemeroffCS.indd 3 2/11/2011 3:30:59 PM
PSYCHIATRIC ANNALS 41:2 | FEBRUARY 2011 PsychiatricAnnalsOnline.com | 113
CME
Neurobiology and Genetics
of Generalized Anxiety Disorder
G
eneralized anxiety disorder
(GAD) is one of the most com-
mon anxiety disorders in adults,
affecting approximately 5% of the gen-
eral population. It is often a chronic dis-
order and is associated with substantial
impairment in quality of life. Addition-
ally, GAD is frequently comorbid with
depression and other anxiety disorders.
Remarkably, when compared with other
anxiety disorders, there is a striking lack
of research into the biology of GAD.
1. Describe regional brain abnormalities implicated in generalized anxiety disorder (GAD).
2. Review the neurotransmitter/neuropeptide system as involved in GAD.
3. Discuss the complex nature of the genetics of GAD.
Anne Schienle, PhD, is with the Department of Clinical Psychology, University of Graz, Graz,
Austria. John M. Hettema, MD, PhD, is with the Department of Psychiatry, Virginia Institute
for Psychiatric and Behavioral Genetics, Virginia Commonwealth University, Richmond, VA.
Ricardo Cceda, MD, PhD; and Charles B. Nemeroff, MD, PhD, are with the Department of Psy-
chiatry and Behavioral Sciences, University of Miami Miller School of Medicine, Miami, FL.
Address correspondence to: Charles B. Nemeroff, MD, PhD, Department of Psychiatry and
Behavioral Sciences, University of Miami, Miller School of Medicine, 1120 NW 14 Street, Suite
1455, Miami, FL 33136; fax: 305-243-8532; or e-mail: cnemeroff@med.miami.edu.
Dr. Schienle and Dr. Hettema have disclosed no relevant nancial relationships. Dr.
Cceda has disclosed the following relevant nancial relationships: the Arsht Foundation,
National Institute on Alcohol Abuse and Alcoholism (NIAAA), National Institute on Drug
Abuse (NIDA), and National Institute of Mental Health (NIMH): Grant/research support;
and MyHealthWin LLC: Partial Owner. Dr. Nemeroff has disclosed the following relevant
nancial relationships: American Foundation for Suicide Prevention (AFSP) and NovaDel
Pharma: Member of Board of Directors; Takeda and Xhale: Consultant; Agency for Health-
care Research and Quality (AHRQ) and National Institutes of Health (NIH): Grant/research
support; Method and devices for transdermal delivery of lithium (US 6,375,990B1), and
method of assessing antidepressant drug therapy via transport inhibition of monoamine
neurotransmitters by ex vivo assay (US 7,148,027B2): Patent holder; Anxiety Disorders As-
sociation of America (ADAA), AFSP, CeNeRx BioPharma, National Alliance for Research on
Schizophrenia and Depression (NARSAD), NovaDel Pharma, PharmaNeuroBoost, Member
of Scientic Advisory Boards; CeNeRx BioPharma, Corcept Therapeutics, NovaDel Pharma
Inc., PharmaNeuroBoost, Revaax Pharma, and Xhale: Stockholder; and CeNeRx BioPharma,
PharmaNeuroBoost: Other nancial interests.
doi: 10.3928/00485713-20110203-10
EDUCATIONAL OBJECTIVES CME
Anne Schienle, PhD;
John M. Hettema, MD, PhD;
Ricardo Cceda, MD, PhD; and
Charles B. Nemeroff, MD, PhD
4102NemeroffCS.indd Sec1:113 2/11/2011 3:31:00 PM
114 | PsychiatricAnnalsOnline.com PSYCHIATRIC ANNALS 41:2 | FEBRUARY 2011
Studies on the genetics, structural,
and functional neuroanatomy, as well
as neurochemistry of this syndrome
are available, which are helping to
paint a more coherent picture of the
neurobiology of GAD. The overall
lack of understanding of the underly-
ing pathophysiology of GAD has been
a major impediment in the develop-
ment of novel treatment strategies.
This review is divided into two main
sections: neurobiology and genetics
of GAD. In the neurobiology section,
structural and functional neuroanatomy,
as well as neurochemistry (neurotrans-
mitters) and neuroendocrine ndings in
GAD are described. In the genetics sec-
tion, we summarize family, twin, and
molecular genetic studies.
NEUROBIOLOGY
Structural Neuroanatomy
Only a handful of imaging stud-
ies have specically focused on brain
structural abnormalities in children,
adolescents, and adults suffering from
GAD.
1-5
Findings in pediatric samples
have been very heterogeneous. In one
study,
2
GAD patients exhibited larger
amygdala volumes than healthy con-
trols; Milham et al.
4
described the op-
posite nding. The latter group studied
children with different anxiety disor-
ders, including social phobia, GAD,
and separation anxiety. Relative to
healthy comparison subjects, the pa-
tients had reduced amygdala volumes.
This reduction was most pronounced
in the GAD patients. A third investi-
gation
1
observed increased gray and
white matter volume in the superior
temporal gyrus (STG), as well as a
correlation of lateral STG white matter
asymmetry with anxiety symptoms.
The two morphometric studies in
GAD adults revealed similar results,
namely increased amygdala volume.
3,5

Moreover, Schienle et al.
5
observed
increased volume of the dorsomedial
prefrontal cortex (DMPFC) in GAD pa-
tients. This volume increase was posi-
tively correlated with patients scores on
the Penn State Worry Questionnaire,
6
a
measure of GAD symptom severity.
In addition to the amygdalar abnor-
malities in GAD (increased volume
and disrupted connectivity with cor-
tical and subcortical regions), hippo-
campal alterations have been detected
by magnetic resonance spectroscopy
(MRS) measurements of local concen-
trations of the marker of neuronal vi-
ability, N-acetylaspartate (NAA). This
nding persisted after successful par-
oxetine treatment, suggesting that this
is a trait marker.
7
Functional Neuroanatomy
Several comprehensive reviews and
meta-analyses on the functional neu-
roanatomy of affective processing in
healthy conditions and in anxiety disor-
ders are available.
8
These investigations
have implicated a limited number of cor-
tical and subcortical regions involved in
the detection of threat and in the regula-
tion of the associated fear states.
One central structure within this
context is the amygdala. The amyg-
dala decodes stimulus valence and co-
ordinates attention-vigilance aspects
of affective processing. During threat
processing, hyperreactivity of this
structure and associated hyperarousal/
hypervigilance has been suggested to
form a central abnormality in anxiety
disorders, including posttraumatic
stress disorder (PTSD), social anxiety
disorder, and specic phobia.
8
In addition to hyperresponsivity to
disorder-relevant cues, problems in ef-
fectively regulating negative affect elic-
ited by such cues have been described
in anxiety disorder patients. They are
aficted by less efcient cognitive con-
trol mechanisms, which are needed to
reduce, and nally terminate, an aver-
sive emotional state. Cognitive emo-
tion regulation has been attributed to
different prefrontal cortex regions, in-
cluding the DMPFC and ventromedial
prefrontal cortex (VMPFC), and parts
of the anterior cingulate cortex (ACC),
as well. Altered activation within these
regions is considered a second distinc-
tive feature of anxiety disorders.
9
A step beyond nding discreet
changes in brain activity in the amyg-
dala and ACC in GAD is to understand
this disorder in the context of brain
circuitry. In the resting state, GAD
patients showed distinct patterns of
connectivity between the basolateral
and central nucleus of the amygdala
with a frontoparietal executive control
network and the insula- and cingulate-
based salience network compared with
TABLE 1.
Summary of Select Neurotransmitter Abnormalities in
Generalized Anxiety Disorder
Neurotransmitter Abnormality Effect
GABA
Decreased GABA-A receptor density in GAD; GABA-A
agonists are anxiolytic; afnity for GABA-A predicts ef-
cacy of benzodiazepines
Serotonin Decreased CSF 5HIAA concentrations
Serotonin transporter
Platelet, PET, postmortem brain studies show density cor-
relates negatively with anxiety symptoms in GAD
5HT
1A
Agonists are anxiolytic
5HT
2A
Agonists are anxiolytic
Source: Schienle A, et al.
4102NemeroffCS.indd Sec1:114 2/11/2011 3:31:01 PM
PSYCHIATRIC ANNALS 41:2 | FEBRUARY 2011 PsychiatricAnnalsOnline.com | 115
controls.
3
This is further illustrated by
failure to regulate emotional conict
and altered functional connectivity
between the amygdala and pregenual
ACC in patients with GAD.
10
This
may represent a decit in regulation of
amygdala negative emotional drive by
the ACC that in turn would translate
into anxiety symptoms.
Symptom Provocation and
Reduction in GAD
The central feature of GAD is path-
ological worry. There are only two
brain imaging studies that have di-
rectly addressed this mental process in
symptom provocation experiments.
11,12

The patients were confronted with dis-
order-relevant stimuli (worry-related
sentences) and their brain activation
was simultaneously recorded.
Hoehn-Saric et al.
11
presented GAD
patients with worry and neutral
sentences (eg, Today is a nice day;
I may lose my job). After pharma-
cological treatment with a selective
serotonin reuptake inhibitor (SSRI),
citalopram, for 7 weeks, worry-in-
duced activation decreased in an ex-
tended cortical network, including
medial prefrontal areas. This decrease
was accompanied by a signicant re-
duction in self-reported symptom se-
verity (degree of worrying). Because
the study recorded brain activation in
only six patients and did not include a
control group, the ndings can only be
considered preliminary.
Besides exaggerated amygdalar
responses in specic contexts, GAD
patients showed decreased localized
prefrontal activation, especially in the
DMPFC.
11
This region is involved in
the reection of ones own mental state,
the integration of outcome alternatives,
prospective memory and outcome an-
ticipation.
12
Although the precise role
of the medial prefrontal group (DMP-
FC, dorsal ACC), as termed by Kober
et al.,
13
is not fully understood, its rel-
evance for monitoring and appraising
emotions is no longer in question.
Moreover, activation of this area has
been reliably associated with amygda-
lar coactivation. In a controlled func-
tional magnetic resonance imaging
(fMRI) study, Paulesu et al.
12
observed
comparable DMPFC and ACC activ-
ity to worry-related sentences in GAD
patients and in healthy controls. Thus,
prefrontal activation was observed in
normal and pathological worrying.
However, these two groups differed in
the postworry resting state; the patient
groups elevated activation level in
the aforementioned regions persisted,
whereas the controls showed a reduc-
tion. It would appear from these data
that GAD patients were unable to ter-
minate the worry process.
A second paradigm addressing a
central problem in GAD patients re-
fers to anticipatory anxiety. This con-
cept directly relates to worrying that
can be conceptualized as a strategy to
cope with potentially negative events
in the future. A typical paradigm to in-
vestigate anticipatory anxiety has been
applied by Nitschke et al.
14
This design was composed of a
warning cue that preceded aversive
pictures (eg, attack scenes) and a
safety signal that preceded neutral
pictures (eg, household items). Rela-
tive to healthy controls, patients with
GAD showed greater amygdala acti-
vation in the anticipatory interval pre-
ceding aversive and neutral pictures.
Moreover, patients anticipatory ACC
activation could be used to predict an
8-week treatment response to venla-
faxine, a serotonin-norepinephrine
reuptake inhibitor (SNRI).
Greater pretreatment activation
was associated with a better treatment
outcome, namely a reduction of expe-
rienced anxiety and worrying when
controlling for depression. Interest-
ingly, the groups did not differ in their
neural responses to the actual adverse
stimuli (negative pictures). In another
study, the magnitude of symptomatic
TABLE 2.
Family Studies of Generalized Anxiety Disorder
Study
Diagnostic
Criteria
Blinded Cases Controls
Odds Ratio
(95% CI)
Source N
P
N
R
MR
a
Source N
P
N
R
MR
a
Noyes, et al.
101
DSM-III No Volunteer 20 123 19.5 Nonanxious 20 113 3.5 6.6 (2.2-19.7)
Mendlewicz, et al.
102 b
DSM-III Yes Clinical 25 102 8.9 NMI 25 130 1.9 5.0 (1.0-24.3)
Newman and Bland
103 c
DSM-III Yes Community 160 430 27.0 Community 764 1956 16.9 1.6 (1.22.1)
Coelho, et al.
104 d
DSM-IV Yes
Obstetrics
clinics
22 72 8.3 Obstetrics clinics 41 146 6.8 1.2 (0.433.6)
Abbreviations: NP = number of probands, NR = number of rst-degree relatives, MR = morbidity risk, CI = condence interval, NMI = never mentally ill
a MR = morbidity risk (%) in rst-degree relatives (age-corrected by Strmgren method in Mendlewicz, et al.)
b Four diagnostic groups included in study: panic disorder, GAD, MDD, and NMI controls.
c Statistics listed are those adjusted for age, sex, and comorbid disorders in relatives.
d This was a study in pregnant women comparing familial aggregation of pure GAD to that of pure generalized social phobia (GSP) and comorbid GAD/GSP. Statistics listed are those reported for pure
GAD in cases and relatives.
Source: Schienle A, et al.
4102NemeroffCS.indd Sec1:115 2/11/2011 3:31:01 PM
116 | PsychiatricAnnalsOnline.com PSYCHIATRIC ANNALS 41:2 | FEBRUARY 2011
response to venlafaxine treatment in
GAD patients was predicted by greater
pretreatment reactivity to fearful faces
in the rACC and lesser reactivity in
the amygdala.
15
Furthermore, success-
ful treatment with SSRIs or cognitive
behavioral therapy (CBT) has been
shown to be associated with increase
rostral ventrolateral prefrontal cortex
(VLPFC) activity.
16
In an analogue study of healthy indi-
viduals, anticipatory threat processing
recruited the amygdala and prefrontal
cortical areas.
5
The participants were
presented with warning cues indicating
that an aversive consequence (negative
pictures) might occur with a 50% prob-
ability or safety signals that always pre-
ceded neutral pictures. Similar to the
GAD patients, the healthy individuals
recruited an extended prefrontal net-
work and the amygdala during the an-
ticipatory interval. Habitual worrying,
as assessed with the Penn State Worry
Questionnaire,
6
was positively corre-
lated with amygdala activity during ex-
perienced uncertainty while waiting for
the picture presentation.
The data suggest that patients suffer-
ing from GAD and healthy individuals
use similar brain networks when trying
to cope with future uncertainties. For
example, Kalisch et al.
17
induced an-
ticipatory anxiety in healthy volunteers
by signaling that they may receive an
electric shock. This instruction pro-
voked activation of the DMPFC as was
effectively seen in GAD patients.
Recruitment of a frontolimbic cir-
cuit in response to expected aversive
consequences, of course, is not spe-
cic for GAD. In addition to healthy
individuals, it is also seen in other psy-
chiatric disorders. Thus, Abler et al.
18

demonstrated that patients suffering
from major depression showed height-
ened amygdala activation during the
anticipation of aversive stimuli.
Finally, it should be noted that sev-
eral brain imaging studies in GAD pa-
tients did not use disorder-specic ma-
terial, but, rather, presented generally
aversive stimuli, such as pictures with
negative facial emotional expressions.
The ndings are inconsistent with
some studies reporting amygdalar hy-
perresponsivity while others nd hy-
poresponsivity to angry and fearful ex-
pressions.
19
These discordant ndings
might be due to the limited disorder
relevance of this paradigm. An im-
proved experimental design has been
applied by Etkin et al.
3
who identied
a decit in ACC recruitment in GAD
patients during emotional conict. The
patients were presented with emotional
facial expressions together with either
congruent or incongruent affective la-
bels. The ACC hyporesponsiveness to
incongruence was interpreted to reect
a dysfunction in emotion regulation.
NEUROTRANSMITTER
DYSREGULATION
Amino Acid Neurotransmitters
The observed limbic abnormalities,
mainly in the ACC-amygdala circuit-
ry, in patients with GAD could result
from decreased inhibitory neurotrans-
mission, increased excitatory neu-
rotransmission, or a combination of
these two processes (see Table 1, page
114). Dysregulation of GABA inhibi-
tory neurotransmission has been docu-
mented in several anxiety disorders.
20

For instance, benzodiazepine receptor
binding in platelets is decreased in pa-
tients with GAD; it is normalized with
diazepam treatment and after cessation
of treatment, it decreases again.
21
Benzodiazepine receptor binding
is decreased in the left temporal pole
among drug -nave patients with GAD
compared with controls.
22
GABA-A
receptor downregulation is observed
in GAD patients and has been hypoth-
esized to play a role in the etiology of
this illness.
23,24
In support of this hy-
Figure. Brain regions with functional and structural abnormalities implicated in GAD.
Photo courtesy A. Schienle.
4102NemeroffCS.indd Sec1:116 2/11/2011 3:31:02 PM
PSYCHIATRIC ANNALS 41:2 | FEBRUARY 2011 PsychiatricAnnalsOnline.com | 117
pothesis is the nding that symptoms
of GAD, including excessive worry,
hypervigilance, and psychomotor
agitation, are treated effectively with
agents that increase GABA-A signal
transduction, including benzodiaze-
pines and barbiturates.
20
Furthermore,
an open trial with riluzole, an anti-glu-
tamatergic agent, resulted in improve-
ment in GAD symptoms.
25
Monoamines
It is important to note that GAD of-
ten occurs comorbid with other mood
and anxiety disorders, including major
depression, panic disorder, and social
anxiety disorder, all responsive to cer-
tain antidepressant treatments.
26
Al-
though SSRIs and SNRIs are effective
and FDA approved for the treatment of
GAD, the drug most frequently stud-
ied in anxiety disorders is the SSRI
paroxetine, which decreases symp-
toms of harm avoidance. Serotonin is
widely distributed in the brain, par-
ticularly in regions associated with
anxiety. Serotonin concentrations in
cerebrospinal uid (CSF) of patients
with GAD have been reported to be
decreased compared with controls.
27

Daeken and Graeff have proposed two
pathways originating from the dorsal
raphe nucleus: an ascending pathway
to the amygdala and PFC involved in
anticipatory anxiety and avoidance
(conditioned fear), pertinent to GAD;
and a second pathway projecting to the
periaqueductal gray matter associated
with panic (unconditioned fear).
28
Additional evidence supporting a
role for serotonin (5-HT) circuitry
in GAD includes challenge with the
5-HT
2c
/5-HT
3
receptor agonist m-
chlorophenypiprazine (mCPP), which
elicits anxiety and anger in GAD
patients.
29
Similar to what has been
observed in depressed patients, [
3
H]-
paroxetine platelet binding, a measure
of serotonin transporter binding sites,
is decreased in patients with GAD.
30

Further evidence for a serotoner-
gic component of GAD is provided
by functional brain imaging studies,
which have found that midbrain se-
rotonin transporter density correlates
negatively with symptom severity,
although no difference in serotonin
transporter density was found in GAD
patients compared with controls.
31
Evidence for a pathological in-
volvement of the adrenergic system
in GAD is more limited. Study of
baseline plasma norepinephine (NE)
levels, a measure of sympathetic ner-
vous system activity, in GAD yielded
mixed results.
32-34
However, challenge
with clonidine, an alpha-2 agonist,
induced a blunted growth hormone
response in GAD patients compared
with controls.
35
Lastly, a trial with yo-
himbine, an alpha-2 receptor antago-
nist, induced no behavioral changes in
GAD patients compared with controls
but led to a normalization of plasma
3-methoxy-4-hydroxyphenylglycol
(MHPG), a major NE metabolite, in
GAD patients.
36
Neuropeptides
Patients with GAD are hypersensitive
to the anxiogenic effects of exogenously
administered cholecystokinin (CCK) ag-
onists.
37
This nding has led to the study
of CCK receptorselective antagonists
as a putative novel class of anxiolytics.
One such drug was developed but was
not demonstrated to possess anxiolytic
efcacy.
38-40
Additional research and de-
velopment of unique CCK antagonists
may be an important step in clarifying
the role of CCK in anxiety and its poten-
tial as a therapeutic target.
To the authors knowledge, no stud-
ies have specically examined the role
of neuropeptide Y (NPY) in GAD.
NPY does possess anxiolytic effects in
laboratory animals.
41
These anxiolytic
effects may be caused by NPYCRF
interactions. These two neuropeptides
are colocalized in numerous limbic re-
gions and exert opposing effects on the
amygdala, locus coeruleus, and periaq-
ueductal gray matter, the latter region
is responsible for the motor output for
the behavioral stress response.
42
Corticotropin-releasing Factor
(CRF) and the Hypothalamic-
pituitary-adrenal axis (HPA)
Although very few studies have spe-
cically examined HPA axis reactivity
in GAD patients, there is some evi-
dence of hypercortisolism, dexameth-
asone nonsuppression, or increased
CSF/CRF concentrations.
43
CRF
1
re-
ceptor antagonists have been demon-
strated to possess anxiolytic proper-
ties.
44
Furthermore, there is a 30% rate
of nonsuppresion after dexamethasone
suppression testing in GAD patients
45

compared with approximately 50% in
major depression,
46
a rate which is sig-
nicantly greater than that observed in
normal volunteers.
Additionally, successful treatment
with escitalopram in a sample of older
adults with GAD was associated with
normalization of salivary cortisol lev-
els.
47
Additional studies combining
brain imaging, genetics, and treatment
approaches may shed further light on
the association between the CRF/HPA
axis and GAD.
The HPA and hypothalamic-pitu-
itary-gonadal (HPG) axes are inter-
linked; HPG axis activity is inhibited
by CRF and glucocorticoids during the
response to stress, and promoter ele-
ments in the CRF gene are regulated by
sex steroids.
48
Some evidence suggests
that the HPG axis activity is also de-
creased in GAD patients.
49,50
In men,
No studies have specically
examined the role
of NPY in GAD.
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testosterone supplementation might
be beneficial in the treatment of anxi-
ety associated with hypogonadism.
51

However, HPG axis alterations neither
predicted treatment outcome nor cor-
related with anxiety symptoms in pa-
tients with GAD, suggesting a lack of
causal relationship between GAD and
HPG axis alterations.
52
Compared with other neuropsychi-
atric disorders, such as schizophrenia,
major depressive disorder (MDD), bi-
polar disorder and Alzheimers disease,
there is a dearth of postmortem brain
tissue studies, as well as neurotrans-
mitter receptor or transporter positron
emission tomography (PET) studies in
GAD. This is likely due in part to the
prevailing view that despite of the chro-
nicity and functional impairment of
GAD, it is either an uncommon disorder
or a relatively mild one. In fact, Barg-
er and Sydeman
53
assessed GAD and
MDD in a sample of more than 3,000
adults in the US. GAD, but not MDD,
predicted increased risk for coronary
heart disease. Concordant ndings have
been reported in an outpatient cardiol-
ogy clinic (n = 516) in which anxiety
symptom severity predicted subsequent
cardiac events and mortality.
54
Genetics Approach
Similar to most major psychiatric
disorders, GAD is considered a com-
plex genetic condition, the etiology of
which involves the effects of multiple
genetic and environmental risk factors
that combine and interact throughout
the lifespan to produce illness. How-
ever, three related, diagnostic dilem-
mas complicate the design and inter-
pretation of genetic studies of GAD.
First, GAD has undergone signifi-
cant revision as a diagnostic entity
over the past several decades.
55
Recent
Diagnostic and Statistical Manual of
Mental Disorders, fifth edition (DSM-
5) discussions have even questioned
whether GAD should remain classi-
fied as an anxiety disorder or should
instead be grouped with MDD.
56,57

Thus, GAD researchers have had to
work with a moving target, compli-
cating phenotypic definition in indi-
vidual study design and interpretation
of findings across studies.
Second, research has shown GAD
to have fairly low reliability compared
with other anxiety disorders.
58
This
most likely derives from its poor diag-
nostic validity and drives the require-
ment for diagnostic revision in search
of the true GAD. Low reliability, in
genetic studies, translates into large
measurement error, resulting in poor
estimation of genetic parameters, such
as odds ratios in family studies and
heritability in twin studies.
Third, while most anxiety disorders
exhibit high lifetime comorbidity with
other psychiatric conditions,
59
this ap-
pears to be one of GADs dening char-
acteristics.
60
Again, this complicates
attempts to dene it diagnostically,
evoking the application of diagnostic
hierarchy rules to better differentiate
it from other related disorders, such as
MDD and panic disorder. For genetic re-
searchers, this comorbidity further com-
plicates phenotypic denition, because
it is unclear whether it is important (or
even makes sense) to study GAD in iso-
lation, or, alternatively, to examine it in
relationship to its comorbid disorders.
Family Studies
Family studies seek to determine
whether conditions, such as GAD ag-
gregate (run) in families by starting
with two proband groups: those affect-
ed with the condition and controls who
are unaffected. They then compare the
rates of the condition in the relatives of
these two groups, with higher rates in
the relatives of affected probands in-
dicative of familial aggregation. Four
published family studies examined the
relationship of GAD in probands and
their rst-degree relatives, summa-
rized in Table 2 (see page 115).
All report elevated rates of GAD in
relatives of probands with GAD suggest-
ing familial aggregation, although there
is notable heterogeneity of study designs
and estimates of odds ratios (OR). The
largest study, by Newman and Bland, re-
ports a modest familial aggregation (OR
= 1.6), although this appears to be par-
tially driven by fairly high rates of GAD
in relatives of unaffected controls com-
pared with the other studies.
61
Twin Studies
Family studies alone cannot deter-
mine whether familial aggregation is
due to genetics or environmental factors
shared among relatives. As there are no
adoption studies of GAD, researchers
rely on twin studies to disentangle the
effects of genetics and environment and
to estimate its heritability (the propor-
tion of individual differences due to ge-
netic factors). They do so by comparing
the rates of concordance (similarity) for
the condition in genetically identical,
or monozygotic (MZ) twin pairs with
those in dizygotic (DZ) twin pairs, who
are, on average, as similar genetically
as sibling pairs.
62
With a few caveats, higher concor-
dance in MZ than DZ twins indicates
that the condition has a genetic com-
ponent. Several (but not all) of the
smaller, earlier twin studies that as-
sessed anxiety disorders found trends
for higher concordance of GAD be-
tween MZ than DZ twin pairs, sug-
gesting that the etiologic basis for
GAD is, at least in part, genetic.
63-65
Research has shown GAD to
have fairly low reliability
compared with other anxiety
disorders.
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PSYCHIATRIC ANNALS 41:2 | FEBRUARY 2011 PsychiatricAnnalsOnline.com | 119
The largest sources of twin data
for GAD are the Virginia Adult Twin
Study of Psychiatric and Substance
Use Disorders (VATSPSUD); the Viet-
nam Era Twin (VET) Registry, and the
Swedish Twin Registry (STR). The
population-based VATSPSUD con-
sists of about 9,000 twins from male
and female same-sex and opposite sex
pairs,
66
while the VET Registry con-
tains a comparable number of male
twins who served during the Vietnam
War.
67
The population-based STR cur-
rently contains more than 100,000
male and female twins depending
upon the cohort selected for study.
68
The VATSPSUD examined GAD
syndromes in two separate studies:
the first assessed DSM-III GAD in
same-sex female twin pairs
69
and the
second assessed DSM-III-R GAD in
same-sex male and female and oppo-
site-sex pairs.
70
The VET study ana-
lyzed the relationship between panic
and generalized anxiety syndromes in
males.
71
Both studies supported the
findings of familial aggregation from
the family studies. Although the best-
fitting model varied somewhat across
syndromic definitions and interview
waves of the VATSPSUD (likely due
to issues of validity and reliability of
GAD), the overall picture suggests that
GAD may have only modest heritabil-
ity (15% to 30%), with an uncertain
role for common family environment
in its familial aggregation.
The VET Registry study found a
heritability of 38% with little or no
role for family environment. When
the findings from both samples were
combined in a previous meta-analy-
sis, the best-fit model predicted an
additive genetic variance, or heri-
tability, of 31.6% (95% CI: 24-39),
and no significant gender differences
were detected.
72
Analyses in the STR
examining the relationship between
GAD and MDD generally support
these findings.
73
Genetics of GAD in Relation to
Other Internalizing Disorders
Multiple models of comorbidity
have been proposed to explain the high
lifetime rates of co-occurrence of psy-
chiatric illness.
74
These can roughly be
broken down into models of shared etiol-
ogy, causation (ie, one condition causing
the other), or both. Among the models
of shared etiology, the one most relevant
for this discussion is shared genetic eti-
ology. This model attempts to explain
the high rates of GAD seen in subjects
with other psychopathology by testing
whether this is due to genetic factors that
are common across the conditions.
The family studies of GAD cited
earlier produce mixed evidence sup-
porting this hypothesis. Although
Noyes et al. found that relatives of
GAD probands had higher rates of
GAD, specically, and anxiety disor-
ders, in general, compared with rela-
tives of control probands, they did not
detect elevated rates of any other in-
dividual anxiety disorder or groups of
other types (affective or alcohol) of
disorders, suggesting separate familial
aggregation of GAD.
In the Mendlewicz et al. study, mor-
bid risk of GAD was higher in relatives
of probands with GAD (8.9%), MDD
(4.9%), and panic disorder (4.0%) than
controls (1.9%). However, these differ-
ences also did not achieve statistical sig-
nicance. This latter trend suggests par-
tial overlap of familial risk of GAD with
MDD and panic disorder. Two studies
in which relatives of probands were not
interviewed directly (family history
method) generally support the hypothe-
sis that GAD and MDD may share some
portion of their familial risk factors.
75,76
Multivariate analyses from several
large twin registries support the hy-
pothesis of shared genetic susceptibility
between GAD and other internalizing
disorders. Bivariate analyses between
12-month
77
and lifetime
78
MDD and
GAD in female twins from the VAT-
SPSUD showed that MDD and GAD
share the majority of their genetic risk
in common but only a modest propor-
tion of environmental risk factors. This
nding was replicated in younger
79
and
older
73
adult twins from the STR. Sev-
eral studies from the VATSPSUD and
STR tested broader models of comor-
bidity between the anxiety disorders, or
between the anxiety disorders, MDD,
and other psychiatric syndromes.
One such analysis examined the
relationships between GAD, panic
disorder, agoraphobia, social, animal
and situational phobias in more than
5,000 same-sex male and female VAT-
SPSUD twin pairs.
80
In the best-tting
model, the genetic inuences on anxi-
ety for both sexes were best explained
by two additive genetic factors com-
mon across the disorders, with only
signicant specic (non-shared) ge-
netic risk for agoraphobia. The rst
factor loaded most strongly on GAD,
panic disorder, and agoraphobia, while
the second loaded primarily on the two
specic phobias. Social phobia was in-
termediate in that it was inuenced by
both common genetic factors.
A recent analysis in a younger co-
hort of STR that also included obsessive
compulsive disorder (OCD) and post-
traumatic stress disorder (PTSD) found
a latent common liability shared among
all the anxiety disorders with heritability
of 54%.
81
These ndings are generally
consistent with an even broader analysis
still that attempted to identify the risk
structure of a wide range of psychiat-
ric and substance use syndromes. That
study found that genetic risk could be
broadly dened as internalizing (primar-
ily affecting MDD, GAD, and phobias)
and externalizing (primarily affecting
alcohol dependence, illicit drug depen-
dence, adult antisocial behavior, and
conduct disorder).
82
The internalizing genetic factor
was further broken down into one of
anxiety-misery loading primarily
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on MDD and GAD, and fear, load-
ing primarily on animal and situational
phobias, with panic disorder interme-
diate. This was broadly supported by a
smaller adolescent twin study.
83
Analyses from the VET Registry
support the hypothesis of a shared
genetic diathesis between GAD and
other anxiety disorders. They found
signicant genetic correlations be-
tween GAD and panic disorder
71
and
between these syndromes and PTSD,
84

with some role for panic- and PTSD-
specic genetic inuences.
Shared vulnerability across anxiety
and depressive disorders has been as-
cribed to common, predisposing per-
sonality traits, such as neuroticism.
85,86

In the VATSPSUD, Hettema et al. ana-
lyzed the relationship between GAD
and neuroticism as measured by the
short form of the Eysenck Personal-
ity Questionnaire, nding that the
genetic correlation between them is
high (about 0.8). This suggests that the
genetic factors inuencing individual
differences in neuroticism are largely
the same as those that increase suscep-
tibility to GAD.
87
A similar analysis in an older sub-
sample of the STR found somewhat a
smaller but signicant correlation in ge-
netic factors underlying neuroticism and
GAD.
88
A study performed in more than
37,000 twins from the STR examining
the role personality traits may play in
MDD-GAD comorbidity found moder-
ate genetic correlation between neuroti-
cism and both of these disorders.
89
Expanding on this line of investi-
gation, the Virginia group modeled
the relationship between neuroticism
and GAD, MDD, and a range of other
anxiety disorders in the VATSPSUD.
90

That study found that not only is there
a high degree of overlap between the
genetic factors inuencing neuroticism
and genetic risk for these disorders but
also that a substantial proportion of the
comorbidity between the GAD and the
other internalizing disorders can be ac-
counted for by the genetic factors un-
derlying neuroticism.
Molecular Genetic Studies
There are no published genetic link-
age studies for GAD. Compared with
other anxiety disorders, such as panic
disorder,
91
only a handful of studies
have attempted to identify specic sus-
ceptability genes for GAD. These have
taken the form of small genetic associa-
tion studies for several candidate genes
related to brain neurotransmitters. One
of the earliest was a family-based as-
sociation study in an American sample
of affected children from 125 families
and control children from 53 families.
That study reported modestly signi-
cant associations between a variable
number tandem repeat (VNTR) poly-
morphism in the dopamine transporter
gene and quantitative symptom scales
for GAD, social phobia, OCD, and
Tourettes disorder.
92
A modest association between
GAD (and OCD) and a VNTR poly-
morphism in the serotonin transporter
gene (5-HTT) was reported in a case-
control study of Japanese anxiety-dis-
order patients.
93
A German case-con-
trol study that examined association
between a single nucleotide polymor-
phism (SNP) in a coding region of the
monoamine oxidase (MAO) A gene
and GAD, panic disorder, and MDD
patients and healthy controls reported
a signicant association with GAD but
not panic disorder or MDD.
94
A study in a Polish sample re-
ported a female-specific association
of a functional VNTR polymorphism
in the MAO A gene and anxiety dis-
orders, including GAD.
95
The same
study found no association with the
Val158Met polymorphism of the cat-
echol-O-methyltransferase (COMT)
gene or the 5-HTT promoter 44 base
pair length polymorphism.
However, a small Chinese study re-
ported signicantly higher frequencies
of the short allele of the 5-HTT poly-
morphism in patients with GAD but
no association with 5-HTT VNTR and
tryptophan hydroxylase (TPH) A218C
polymorphisms.
96
A study dedicated
to assessing the effects of exposure to
the 2004 Florida hurricane reported
increased risk of GAD in relation to
polymorphisms in the genes RGS2
97

and NPY.
98
Molecular genetic studies
performed in the VATSPSUD sample
have utilized the nding of shared ge-
netic risk factors between neuroticism
and internalizing disorders to identify
associations between GAD and relat-
ed phenotypes and the glutamic acid
decarboxylase 1 (GAD1) and COMT
genes.
99,100
As should be the case with
all genetic association analyses, cau-
tion is warranted in interpreting the
signicance of these preliminary nd-
ings until they are replicated in inde-
pendent samples.
CONCLUSIONS
Compared with the body of neuro-
biologically and genetically informed
literature on other psychiatric disor-
ders, the literature on GAD is in its
infancy, and there is a significant need
for the expansion and development of
this area of scientific research. The
limited available fMRI data point to a
dysfunction in a brain circuit involv-
ing the amygdala and various prefron-
tal cortical regions (DMPFC, ACC).
Amygdala activity in patients with
GAD or worry proneness is height-
ened in anticipation of aversion but
also to ambiguous stimuli.
11,14,19
Thus, amygdalar hyperreactivity to
anticipatory signals in general, and to
There are no published genetic
linkage studies for GAD.
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PSYCHIATRIC ANNALS 41:2 | FEBRUARY 2011 PsychiatricAnnalsOnline.com | 121
the experience of uncertainty generally,
may be one important pathophysiologi-
cal mechanism of this disorder. Besides
altered amygdalar functionality, struc-
tural changes have been observed. For
adults, GAD has always been associ-
ated with increased amygdala volume,
whereas data for pediatric GAD has
been inconsistent. The latter may be
due to difculties in reliably classify-
ing pediatric GAD. Moreover, neuro-
developmental aspects (eg, individual
differences in brain development) may
contribute to increased variance and
sample-dependent ndings.
Evidence is accruing that besides
the well-known amygdala abnormali-
ties found in GAD, other brain regions,
including the ACC and DMPFC, may
play a crucial role in GAD (see Figure,
page 116). A more coherent view of the
biology of GAD comes with the view
of disruption of brain circuits (ie, ACC-
amygdala; DMPFC-amygdala) involved
in negative emotion regulation.
It still needs to be specified wheth-
er the observed altered changes reflect
trait markers or vulnerability factors
for GAD or directly reflect patho-
physiological mechanisms related to
perseverative worrying and chronic
anticipatory anxiety. Studies with ther-
apeutic interventions coupled with the
previously described paradigms, such
as uncertainty processing and emotion
regulation, will be helpful to further
increase our knowledge about the neu-
ral basis of GAD treatment response.
The available data support familial
aggregation of GAD, and this is like-
ly due to genetic factors. The overall
heritability of GAD is clear but mod-
est, although this might be due to con-
founding by poor diagnostic validity
and reliability. The genetic suscepti-
bility for GAD is likely shared with
that of other anxiety disorders and
MDD, with the genetic factors for
neuroticism explaining some propor-
tion of this. The status of the molecu-
lar genetics of GAD is still too imma-
ture to support the involvement of any
specific genetic loci in its etiology.
It is important to note that some of
the heterogeneous ndings in the biol-
ogy of GAD might be due to comor-
bidity factors. In many, if not most, of
the studies, the patients suffered from
at least one comorbid disorder (pre-
dominantly MDD), and often, patients
were taking antidepressant medica-
tion. This constellation is typical for
GAD, in which additional diagnoses
are common. However, comorbidity
makes a direct attribution of functional
and structural abnormalities to the di-
agnosis of GAD difcult. The disorder
specicity of the observed pathologi-
cal neural signature of GAD, therefore,
needs further investigation, especially
to characterize potential overlap with
MDD and other anxiety disorders.
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