Genetically Determined Measures of Striatal D2

Signaling Predict Prefrontal Activity during Working

Memory Performance
Alessandro Bertolino1,2*, Paolo Taurisano1, Nicola Marco Pisciotta3, Giuseppe Blasi1, Leonardo Fazio1,2,
Raffaella Romano1, Barbara Gelao1, Luciana Lo Bianco1, Madia Lozupone1, Annabella Di Giorgio1, Grazia
Caforio1, Fabio Sambataro4, Artor Niccoli-Asabella3, Audrey Papp5, Gianluca Ursini1,6, Lorenzo
Sinibaldi6,7, Teresa Popolizio2, Wolfgang Sadee5, Giuseppe Rubini3
1 Psychiatric Neuroscience Group, Department of Neurological and Psychiatric Sciences, University of Bari, Bari, Italy, 2 Department of Neuroradiology, IRCCS ‘‘Casa
Sollievo della Sofferenza’’, San Giovanni Rotondo, Italy, 3 Nuclear Medicine Unit, Department of Internal Medicine and of Public Medicine, University of Bari, Bari, Italy,
4 Clinical Brain Disorders Branch, Genes, Cognition and Psychosis Program, National Institute of Mental Health, National Institutes of Health, Bethesda, Maryland, United
States of America, 5 Program in Pharmacogenomics, Department of Pharmacology, College of Medicine, and Division of Biostatistics, College of Public Health, The Ohio
State University, Columbus, Ohio, United States of America, 6 IRCCS CSS-Mendel Institute, Rome, Italy, 7 Department of Experimental Medicine, Sapienza University, Rome,
Italy

Abstract
Background: Variation of the gene coding for D2 receptors (DRD2) has been associated with risk for schizophrenia and with
working memory deficits. A functional intronic SNP (rs1076560) predicts relative expression of the two D2 receptors
isoforms, D2S (mainly pre-synaptic) and D2L (mainly post-synaptic). However, the effect of functional genetic variation of
DRD2 on striatal dopamine D2 signaling and on its correlation with prefrontal activity during working memory in humans is
not known.

Methods: Thirty-seven healthy subjects were genotyped for rs1076560 (G.T) and underwent SPECT with [123I]IBZM (which
binds primarily to post-synaptic D2 receptors) and with [123I]FP-CIT (which binds to pre-synaptic dopamine transporters,
whose activity and density is also regulated by pre-synaptic D2 receptors), as well as BOLD fMRI during N-Back working
memory.

Results: Subjects carrying the T allele (previously associated with reduced D2S expression) had striatal reductions of
[123I]IBZM and of [123I]FP-CIT binding. DRD2 genotype also differentially predicted the correlation between striatal dopamine
D2 signaling (as identified with factor analysis of the two radiotracers) and activity of the prefrontal cortex during working
memory as measured with BOLD fMRI, which was positive in GG subjects and negative in GT.

Conclusions: Our results demonstrate that this functional SNP within DRD2 predicts striatal binding of the two radiotracers
to dopamine transporters and D2 receptors as well as the correlation between striatal D2 signaling with prefrontal cortex
activity during performance of a working memory task. These data are consistent with the possibility that the balance of
excitatory/inhibitory modulation of striatal neurons may also affect striatal outputs in relationship with prefrontal activity
during working memory performance within the cortico-striatal-thalamic-cortical pathway.

Citation: Bertolino A, Taurisano P, Pisciotta NM, Blasi G, Fazio L, et al. (2010) Genetically Determined Measures of Striatal D2 Signaling Predict Prefrontal Activity
during Working Memory Performance. PLoS ONE 5(2): e9348. doi:10.1371/journal.pone.0009348
Editor: André Aleman, University of Groningen, Netherlands
Received September 15, 2009; Accepted January 29, 2010; Published February 22, 2010
Copyright: ß 2010 Bertolino et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits
unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.
Funding: Part of this work was supported by a National Institutes of Health (NIH) grant DA022199 (W.S.). The funders had no role in study design, data collection
and analysis, decision to publish, or preparation of the manuscript.
Competing Interests: The authors have declared that no competing interests exist.
* E-mail: [email protected]

Introduction Indeed, all antipsychotics available on the market block dopamine

D2 receptors (even though other receptors may also be involved).
Susceptibility to schizophrenia is explained for the largest Phenomenologically, schizophrenia is characterized by cognitive
fraction by genetic variation [1]. While the specific genes deficits, in particular in the working memory domain [4]. Working
conferring risk for schizophrenia are still undetermined, several memory deficits in schizophrenia have been associated with
studies and meta-analyses point to the potential involvement of the dysfunction of the prefrontal cortex [7,8] and of the dopamine
gene for dopamine D2 receptors (DRD2) [2,3]. Moreover, several system [4,6]. Indeed, several authors have hypothesized that
lines of evidence suggest involvement of the dopamine system and altered working memory performance and related prefrontal activ-
of D2 signaling in the pathophysiology of schizophrenia [4–6]. ity can be part of a systems level pathophysiological mechanism

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 DRD2 and Prefrontal Activity

also involving dopamine D2 receptors in the striatum [4,6]. The (rs4680). COMT is a key enzyme for dopamine catabolism in
key anatomical and molecular mechanisms regulating the rela- prefrontal cortex but not in striatum [34,35] and this polymor-
tionship between DRD2 genetic risk, dopamine dysregulation phism alters activity of the enzyme in association with prefrontal
of D2 signaling, and working memory dysfunction remain function [36].
undetermined.
A large series of studies in animals indicates that prefrontal Materials and Methods
neuronal activity during performance of working memory tasks is
regulated by dopamine [9]. In the prefrontal cortex both D1 and Participants to [123I]FP-CIT SPECT, [123I]IBZM SPECT, and
D2 receptors are involved in working memory, with the latter BOLD fMRI Studies
more specifically implicated in the response phase [9]. In this Thirty-seven healthy subjects (16 males, mean age6SD
regard, genetically modified mice lacking D2 receptors exhibit 23.563.0 years) participated to [123I]FP-CIT SPECT. Thirty-
behavioral working memory deficits and reduced activity of the two of these subjects underwent [123I]IBZM SPECT and twenty-
prefrontal cortex during treatment with D1 agonists [10]. eight also underwent Blood Oxygen Level Dependent (BOLD)
Moreover, behavioral performance and prefrontal neuronal functional Magnetic Resonance Imaging (fMRI) during perfor-
activity during working memory (WM) performance are also mance of the N-Back task. Exclusion criteria included history of
regulated by dopamine and D2 receptors in the striatum, a key significant drug or alcohol abuse (no active drug use in the past
node within the cortico-striatal-thalamic-cortical circuit [11]. In year), head trauma with loss of consciousness, and any significant
this regard, an experiment inducing developmental over-expres- medical condition. Parental socio-economical status (Hollingshead
sion of striatal D2 receptors demonstrated specific working Scale 40.4614.9), handedness (Edinburgh Inventory 0.7460.4),
memory deficits and altered prefrontal activity to D1 stimulation and total IQ (WAIS-R, 108.1614.8) were measured. The present
[12]. These experiments in animals indicate that genetically study was approved by the local IRB. After complete description
modified striatal D2 signaling has systems-level effects in dopamine of the study to the subjects, written informed consent was
modulation of prefrontal cortical activity and related behaviors. obtained.
Data in humans have been consistent with these studies Genotype determination. DRD2 rs1076560 genotypes were
demonstrating that infusion of D2 receptor agonists or antagonists determined as in [29–31]. SNP rs1076560 was analyzed with
is respectively associated with relative improvement or deteriora- allele-specific PCR primers as described [37] or SNaPshot
tion of working memory performance [13] as well as of prefrontal (Applied Biosciences (ABI), Foster City CA) [38]. Consistent
and striatal activity [14,15]. Other imaging studies in humans have with the distribution observed in earlier studies [38], no DRD2 TT
also demonstrated that striatal dopamine and D2 signaling are subjects were observed in this sample. COMT Val158Met genotype
correlated with prefrontal activity or behavioral performance (rs4680) was determined as a restriction fragment length
during different cognitive operations [16–18]. polymorphism after PCR amplification and digestion with NlaIII
D2 receptors in the striatum are both pre- and post-synaptic and [39,40]. The allelic distribution of DRD2 and COMT was in Hardy
affect the final output of the striatum to the thalamus, providing an Weinberg equilibrium (DRD2 df 1, chi2 = 2.1, p.0.1, COMT df 2,
important regulation of prefrontal cortex stimulation [19]. At chi2 = 1.3, p.0.4).
present, however, it is unknown whether genetic determination of Acquisition of SPECT data. Each subject was injected
pre- and post-synaptic striatal D2 signaling as modulated by intravenously with an average of 150 MBq (range 111–186 MBq)
dopamine is a mechanism contributing to neuronal activity in of commercially available [123I] FP CIT or [123I]IBZM
prefrontal cortex in humans. The D2 receptor gene (DRD2) codes radiotracer (GE Healthcare, Amersham, UK) [41–43]. These
for two isoforms, D2S (short) and D2L (long). D2L receptors two radiotracers bind to dopamine transporters [33] or D2
mainly mediate post-synaptic signaling. D2S receptors mainly receptors [32], respectively. Potassium Iodide solution (Lugol) was
serve as auto-receptors on pre-synaptic neurons [20], even though administered at least 3 hours before and 12 hours after
they are also found on post-synaptic neurons [21]. Moreover, pre- radiopharmaceutical injection to block thyroid uptake of free
synaptic D2 receptors strongly contribute to physically regulate radioactive iodide. Images were acquired 3–6 h after [123I]FP-CIT
density and activity of the dopamine transporter (DAT) [22–29]. injection [44] or 1.5 hours after [123I]IBZM injection [45]. A dual-
An intronic DRD2 polymorphism (rs1076560, G.T) is associated head gamma camera (Infinia, General Electric) equipped with
with mRNA splicing [30]. More specifically, the minor T allele is parallel-hole, low-energy high-resolution collimators was used.
associated with relatively reduced expression of D2S in prefrontal SPECT data were acquired using the following parameters:
cortex and striatum as well as with altered activity of the striato- 1286128 matrix, rotation of 360u, 60 view, 6u view angle, 45 s for
thalamic-prefrontal pathway during WM in healthy subjects [30] projection. Slice thickness was 3.68 mm, acquisition time was 22
and in schizophrenia [31]. However, the effect of this SNP on minutes; total brain counts .1 million were achieved in all
prefrontal cortical activity may be also because of indirect effects examinations. Reconstruction was performed by filtered back-
via the striatum [11]. The effects of this genetic variant on pre- projection with a Butterworth filter (cut-off frequency: 0.3 cycle/
and post-synaptic signaling of dopamine in the striatum are not cm, 10th order) to provide transaxial slices that were attenuation
known. In this study in healthy humans, we hypothesized that this corrected. Attenuation correction was performed according to
functional DRD2 variant, rs1076560, would be associated: with Chang’s method, (attenuation coefficient: 0.12 cm21), after
differential binding of [123I]IBZM measured with SPECT, which manually drawing an ellipse around the head contour [46].
reflects availability of post-synaptic D2 receptors [32]; with System spatial resolution (full width at half-maximum) at a radius
differential binding of [123I]FP-CIT measured with SPECT, of rotation of 15.9 cm is 11 mm, as reported elsewhere [47]. For
which reflects availability of pre-synaptic DAT [33]; and with analysis of striatal radiotracer uptake, slices were reoriented
how a factor score of the two radiotracers identifying striatal D2 parallel to the canthomeatal line.
signaling would predict prefrontal activity measured with BOLD Processing of SPECT data. The irreversible binding
fMRI during performance of a working memory task. Moreover, characteristics and the stability of regional [123I]FP-CIT and of
in an effort to test the relative specificity of these associations, [123I]IBZM uptake have been shown to allow estimation of the
subjects were also genotyped for COMT Val158Met genotype specific-to-nondisplaceable equilibrium partition coefficient (V30),

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 DRD2 and Prefrontal Activity

which is proportional to free transporter or receptor density analysis approach using a principal component analysis (PCA,
(Bmax) [48,49]. V30 can be calculated as indicated earlier [48,50]. total variance used) as a reduction method [64] to combine
Under equilibrium conditions between a compartment with binding data from the two radiotracers into a single factor. We
specific binding and a compartment representing nonspecifically performed a factor analysis using Principal component Analysis
bound and free activity, V30 is proportional to Bmax given that (PCA) with STATISTICA (StatSoft, Tulsa, Oklahoma). [123I]FP-
the dissociation constant and the volume of distribution of the CIT and [123I]IBZM V30 data were extracted in each subject from
nonspecifically bound and free activity compartment (V2) are a basal ganglia ROI, which included bilateral caudate and
relatively invariant. The occipital region was selected as the putamen. ROIs were identified using the WFU PickAtlas software
background region because 1) the density of dopamine D2 and (Functional MRI Laboratory at the Wake Forest University School
DAT proteins is negligible compared with the striatum [51,52]; 2) of Medicine, http://www.rad.wfubmc.edu/fmri) [57–59]. One
this region can be identified with greater reliability than the principal component (PC) that is a linear combination of the two
cerebellum [53]; 3) in humans [123I] IBZM activity in the occipital binding measures was estimated. This PC represents average
region is equal to the nonspecific activity in the striatum [54]. dopamine D2 signaling weighted by the variance of the binding
Therefore, as in earlier studies [53,55,56], the occipital region was measures. For each subject we calculated a factor loading, which
used to model non-specifically bound and free activity indicates the weight of each subject bindings in the PC. PC
compartment. V30 can be calculated in all voxels with the loadings were then used as predictors in random effects analyses to
formula as in [50]: identify potential relationships with the fMRI data in SPM (see
below).
v2 is an effect size measure which estimates the proportion of
ðcounts per minute=voxelÞVT
 variance in a dependent measure accounted for by independent
{ ðcounts per minute=voxelÞV2 ðcounts per minute=voxelÞV2 categorical variables in the population from which the sample was
drawn. Thus, we used v2 to measure the amount of variance
accounted for by DRD2 genotype. v2 is given by the equation:
v2 ~ðSSeffect { ðdf effect ÞðMSerror ÞÞ=ðMSerror zSStotal Þ.
where VT represents specific binding, and V2 the nonspecifically
bound and the free activity compartment. BOLD fMRI Data Acquisition and Processing
Image transformation, calculation of V30 and statistical analysis N-Back working memory paradigm. During fMRI, all
was performed using SPM5 (Wellcome Department of Cognitive subjects completed a blocked paradigm of the N-Back task with a
Neurology, London, UK). V2 was calculated with a ROI of the 2-Back working memory condition and a non-memory guided
occipital lobe from the WFU Pickatlas (http://fmri.wfubmc.edu/ control condition 0-Back [40,65]. This paradigm has been
cms/software#PickAtlas) [57–59]. Since parametric images of extensively used to evaluate activity of prefrontal cortex. ‘‘N-
[123I]FP-CIT and [123I]IBZM V30 lack anatomical detail, an back’’ refers to how far back in the sequence of stimuli the subject
indirect approach was employed for spatial normalization as had to recall. The stimuli consisted of numbers (1–4) shown in
detailed in [50,60]. Briefly, the raw [123I]FP-CIT and [123I]IBZM random sequence and displayed at the points of a diamond-shaped
SPECT data of each subject were normalized on the SPECT box. There was a visually paced motor task which also served as a
template in MNI (Montreal Neurological Institute) space [50] with non-memory guided control condition (0-Back) that simply
a 12-parameter affine transformation of the raw data onto the required subjects to identify the stimulus currently seen. In the
template image followed by estimation of the nonlinear deforma- working memory condition, the task required recollection of a
tions between the applied images. A mean image of the previously stimulus seen two stimuli (2-Back) previously while continuing to
normalized raw data acquisitions was then computed and used as encode additionally incoming stimuli. Performance data were
a template image. For each individual SPECT acquisition, a recorded as the number of correct responses (accuracy) and as
parametric V30 image was calculated. The raw data image was reaction time.
transformed to the template image and the resulting transforma- BOLD fMRI acquisition parameters. Each subject was
tion parameters were then applied to the corresponding subject’s scanned with a 3T MR scanner (GE) with a gradient-echo echo-
parametric V30 image. The spatially normalized parametric planar imaging (EPI) sequence using the following parameters:
images were convolved with a gaussian kernel (66666 mm) for 20 contiguous axial slices, slice thickness = 5 mm, echo time =
smoothing. 30 msec, repetition time = 2000 msec; field of view 24 cm; matrix
Statistical analyses of SPECT data. Two sample T tests 64664, voxel size after normalization = 3.75 mm isotropic
were used within SPM5 to evaluate potential differences between [40,66]. We used a simple block design in which each block
genetic groups with a statistical threshold p,0.005, with further consisted of eight alternating 0-Back and 2-Back conditions (each
correction for multiple comparisons within ROIs in putamen lasting 30 seconds), obtained in 4 min and 8 sec, 120 whole-brain
obtained with the WFU_PickAtlas tool, p = 0.05. fMRI volumes. The first four scans of the time series were
Binding of [123I] FP-CIT and of [123I] IBZM is associated with acquired to allow the signal to reach a steady state and were not
protein density and affinity for the radiotracer as well as with the included in the final analysis.
relative concentration of endogenous dopamine occupying these BOLD-fMRI image analysis. Preprocessing and statis-
proteins [61,62]. Moreover, there is an extensive literature tical analyses. Analysis of the fMRI data was completed using
detailing the interaction between dopamine transporters and D2 statistical parametric mapping (SPM5; http://www.fil.ion.ucl.ac.
receptors [22–29,63]. Therefore, we hypothesized that the effects uk/spm). Images for each subject were realigned to the first
of DRD2 genotype on striatal dopamine D2 signaling would be volume in the time series to correct for head motion (,2.5 mm of
additive and therefore identifiable with a measure reflecting the translation, ,1.5u rotation), spatially normalized into a standard
shared and not the unique variance derived by the two radiotracer stereotactic space (Montreal Neurological Institute, MNI,
binding measures. This index derived by factor analysis would template) using a 12 parameter affine model and smoothed to
reflect individual dopamine D2 signaling and could be used for minimize noise and residual differences in gyral anatomy with a
correlation with fMRI activity. Therefore, we used a factor Gaussian filter, set at 10 mm full-width at half-maximum. Voxel-

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 DRD2 and Prefrontal Activity

wise signal intensities were ratio normalized to the whole-brain Polynomial regression to identify the correlation between
global mean. Each experimental condition was modeled with a behavior, BOLD fMRI, and SPECT data. Because previous
box car convolved with the hemodynamic response function evidence supports the hypothesis that the relationship between
(HRF) at each voxel. Predetermined condition effects at each voxel working memory behavioral performance with dopamine signaling
were calculated using a t-statistic, producing a statistical image for and prefrontal activity is non-linear [9,70], we performed
the contrasts of 2-Back versus 0-Back (N-Back). All these individual polynomial regression to assess this relationship in our data. A
contrast images were then used in second-level random effects second order (quadratic) polynomial model was fitted to the data
models at the group level. ANOVA was used to evaluate the main separately in the two genotype groups: behavioral performance at 2-
effects of working memory and of DRD2 genotype. Then, we Back was the dependent variable; the dopamine D2 signaling factor
performed separate linear regression analyses for the two groups of score (see above) and BOLD fMRI activity in prefrontal cortex
subjects (GG and GT) within SPM5. First, we entered the single (from clusters correlated with the factor score itself) were the
subject contrasts (2Back.0Back) with [123I] IBZM or [123I] FP- independent variables. The F-statistic was used to compare the
CIT binding (from the anatomical ROIs differentiating the two goodness of fit of quadratic relative to linear model by comparing
genotype groups) as predictors. These analyses do not exclude that the ratio of partitioned variances in the two genotype groups.
the biological effects of DRD2 genotype can result by additive
effects of genetic variants on both pre- and post- synaptic Results
compartments of D2 systems that would be captured by a D2
signaling factor (as detailed above). Thus, we also regressed Demographics and Working Memory Performance
separately for the two genotype groups the single subject contrasts Results
(2Back.0Back) with factor scores from the factor analysis of the The two genotype groups did not differ for any of the
SPECT data of each individual subject as predictors. All statistical demographic measures (See also Supplemental Table S1, all
maps were thresholded at a level of p,0.005 uncorrected, with p.0.1) or for working memory behavioral performance (% correct
further FWE small volume correction for multiple comparisons responses GG = 83.4614.3, GT = 88.1615.4; Reaction Time
with a = 0.05 using a 10 mm radius sphere centered around GG = 512.46219.1 msec, GT = 562.06289.5; all p.0.2) thus
coordinates in prefrontal cortex published in previous studies of allowing us to examine the association of genotypes with brain
working memory. These coordinates included: x 220, y 8, z 56 activity and dopamine signaling independent of behavioral
[65]; x 232, y 14, z 38 [67]; x 252, y 26, z 20 [68]; x 38, y 34, z variation in this sample.
11 [69]. The studies used to perform small volume correction of
our data were selected based on the characteristics of the working SPECT Results
memory task that we have used [65,67,69], as well as on the aim of Two-sample t test in SPM5 demonstrated that subjects
the present study (i.e. to investigate the impact of variability in homozygous for the G allele had significantly greater specific
dopamine genes on modulation of the functional activity of the binding of [123I]IBZM in right putamen (GG N = 21, GT N = 11,
prefronto-striatal network, [68]). Because we did not have a priori Z = 2.68; k = 7; p = 0.05, corrected; x 30, y 6, z 0, Figure 1)
hypotheses regarding the activity of brain regions outside of the compared with GT subjects. DRD2 genotype explained 20% of the
prefrontal cortex, we used a statistical threshold of p = 0.05, variance in [123I]IBZM binding as indicated by v2. Similarly, GG
corrected for multiple comparisons across all voxels, for these subjects also had significantly greater specific binding of [123I]FP-
whole-brain comparisons. For anatomical localization, statistical CIT in left putamen (GG N = 26, GT N = 11, Z = 3.1; k = 13;
maxima of activation were converted to conform to the standard p = 0.03, corrected; x 230, y 9, z 18, Figure 2). DRD2 genotype
space of Talairach and Tournoux. explained 10% of [123I] FP-CIT binding variance.

Figure 1. Association between DRD2 rs1076560 genotype and [123I]IBZM binding. Coronal section of the effect of DRD2 rs1076560 genotype
(GG.GT) on [123I]IBZM specific binding (V30) in right putamen (left) and relative scatterplot of individual data points from the cluster differentiating
the two groups (right).
doi:10.1371/journal.pone.0009348.g001

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 DRD2 and Prefrontal Activity

Figure 2. Association between DRD2 rs1076560 genotype and [123I]FP-CIT binding. Coronal section of the effect of DRD2 rs1076560
genotype (GG.GT) on [123I]FP-CIT specific binding (V30) in left putamen (left) and relative scatterplot of individual data points from the cluster
differentiating the two groups (right).
doi:10.1371/journal.pone.0009348.g002

Factor Analysis demonstrated that both data from [123I]IBZM Relationship between fMRI and SPECT data. Regression
and [123I]FP-CIT basal ganglia ROIs load into one factor (GG analyses in SPM5 demonstrated a direct correlation between [123I]
N = 21, GT N = 11, Factor loading for [123I] FP-CIT = 0.76, IBZM binding in right putamen with prefrontal cortex in GG
factor loading for [123I] IBZM = 0.76, Eigenvalue 1.16, 58% Total subjects (x 238, y 10, z 37, k = 3, Z = 3.1, p = 0.02, corrected) and
Variance) which presumably reflects dopamine D2 signaling. a negative correlation in GT subjects (x 260, y 26, z 16, k = 12,
In an effort to test the specificity of the association with DRD2, Z = 3.1, p = 0.05, corrected). However, no correlation was found
SPECT data were also grouped and analyzed for COMT in either group between [I123] FP CIT binding and BOLD activity
Val158Met genotype (rs4680). Because of the small number of during performance of the N-Back working memory task. Also,
Met homozygous subjects, Met/Met and Met/Val subjects were regression analyses in SPM5 demonstrated a positive relationship
grouped together. Two sample t tests in SPM5 demonstrated no between the factor score obtained from SPECT data and BOLD
significant association of COMT genotype with both [123I]IBZM activity in prefrontal cortex in GG subjects (N = 18; x 226, y 10, z
(Met carriers N = 19; Val/Val N = 8) and [123I]FP-CIT data (Met 54, BA 6, Z = 3.6, k = 5, p = 0.007 corrected; BA 9 x 234, y 16, z
carriers N = 20; Val/Val N = 10), even after lowering the statistical 30; Z = 2.98, k = 8, p = 0.03, corrected, Figure 3)) and a negative
threshold to p,0.01, uncorrected (data not shown). one in GT subjects (N = 10; BA 46, Z = 3.36, k = 12, p = 0.03,
corrected, x 259, y 30, z 12; BA 46 Z = 2.77, k = 5, p = 0.09,
fMRI Results corrected, x 37, y 34, z 16, Figure 4; see also Supplemental Table
Effect of the 2-Back working memory task. As expected S2 for further results).
from previous studies with the N-Back task [7,29,31,39,40, In GG subjects, there was a strong statistical trend for a
65,71,72], ANOVA within SPM5 demonstrated that perfor- quadratic relationship between 2-Back percent correct responses
mance of the 2-Back working memory condition was associated with the striatal dopamine D2 signaling factor score and prefrontal
with activity in a distributed network of brain regions including the activity during working memory as measured with fMRI
prefrontal cortex, the parietal cortex, and the striatum bilaterally. (R2 = 0.45, F = 2.67, df = 4,13, p = 0.07; F-statistic for quadratic
Regions that also survived correction for multiple comparisons vs. linear model (4,13) = 6.44, p = 0.004; Univariate Results: factor
include the left middle frontal gyrus (x 229, y 13, z 54, BA 6/8 score2 t = 2.5, p = 0.02; BOLD activity in BA9 x 234, y 16, z 302
and 9, Z = 6.79, k = 159, corrected p = 0.000), the left inferior t = 1.9, p = 0.07, Figures S1 and S2). In GT subjects, no such
frontal gyrus (x 251, y 15, z 19, BA 44/45, Z = 5.55, k = 83, relationship was evident (R2 = 0.22, F = 0.3, df = 4,5, p = 0.8).
p = 0.000) and the right middle frontal gyrus (x 40, y 40, z 15, BA
10/46, Z = 5.42, k = 66, p = 0.000). Discussion
Effect of DRD2 genotype. Consistent with earlier studies
[29–31], ANOVA within SPM5 indicated that, despite similar The results of the present study with multimodal imaging in
behavioral performance, GT subjects tend to have greater activity humans demonstrate that DRD2 rs1076560 genotype predicts
in prefrontal cortex (x 33, y 21, z 45, middle frontal gyrus, BA 6, striatal [123I]IBZM and [123I]FP-CIT binding and the direction of
Z = 3.13, k = 41, p = 0.001; x 63, y 8, z 7, inferior frontal gyrus, BA the correlation between a factor score reflecting striatal D2
44, Z = 3.36, k = 16, p = 0.0001) and anterior cingulate (x 10, y 16, signaling with prefrontal activity during performance of working
z 40, anterior cingulate, BA 32, Z = 3.27, k = 21, p = 0.001) memory. More specifically, carriers of the T allele of rs1076560,
compared with GG subjects. However, none of these results known to be associated with relatively reduced D2S, had reduced
survived correction for multiple comparisons. [123I]IBZM and [123I]FP-CIT binding. Consistent with its

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 DRD2 and Prefrontal Activity

Figure 3. Correlation between BOLD fMRI activity in prefrontal cortex and the striatal dopamine D2 signaling factor in GG subjects.
3D rendering of the correlation between the striatal dopamine D2 signaling factor and BOLD fMRI activity during the 2-Back WM task in GG subjects
(left) with the relative scatterplot of the correlation in prefrontal cortex showing individual data points (right).
doi:10.1371/journal.pone.0009348.g003

anatomical distribution, with a relatively specific effect of the correlation which is also weakly predictive of behavioral
DRD2 genotype and with an earlier study which was published performance, whereas in GT subjects the correlation is negative.
while the present paper was being reviewed [73], COMT rs4680 Earlier studies from our group had provided in vitro, post mortem and
genotype did not demonstrate any association with SPECT data. in vivo evidence for the functionality of this SNP and for its
Importantly, the present data also indicate that DRD2 genotype is association with activity of the working memory network [30,31].
associated with qualitatively different relationships between In the present study we provide evidence for a systems level
[123I]IBZM binding and the striatal factor score with prefrontal genetically determined mechanism correlating striatal D2 signal-
activity during working memory. GG subjects have a positive ing with alteration of prefrontal activity during working memory.

Figure 4. Correlation between BOLD fMRI activity in prefrontal cortex and the striatal dopamine D2 signaling factor in GT subjects.
3D rendering of the correlation between the striatal dopamine D2 signaling factor and BOLD fMRI activity during the 2-Back WM task in GT subjects
(left) with the relative scatterplot of the correlation in prefrontal cortex showing individual data points (right).
doi:10.1371/journal.pone.0009348.g004

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 DRD2 and Prefrontal Activity

Even though the specificity for the two D2 isoforms is not known, schizophrenia. Other recent family-based and case control studies
[123I]IBZM is believed to bind to post-synaptic D2 receptors have also implicated these SNPs [89,90]. All these SNPs are linked
[17,32]. [123I]IBZM binding can be modulated by endogenous with rs1076560 in various degrees of LD. Given these earlier
dopamine and by receptor density, affinity, and internalization studies in the literature and the present findings, it is possible to
[53,74,75]. The rs1076560 G allele has been associated with greater speculate that risk for schizophrenia may involve the effects of
D2S expression, but not with total density of D2 receptors [30]. rs1076560 on striatal dopamine signaling and its relationship with
Thus, reduced [123I]IBZM binding in GT subjects can reflect prefrontal activity. This speculation is also consistent with the
reduced D2S receptors, which are also found post-synaptically [21]. above cited animal studies indicating that developmentally
Our [123I]-IBZM results are also consistent with earlier studies regulated over-expression of striatal D2 receptors is associated
investigating the association between DRD2 polymorphisms and D2 with behavioral working memory deficits and with altered activity
binding with different radiotracers [76–78]. These polymorphisms of the prefrontal cortex [12].
are in high linkage disequilibrium with rs1076560 and these earlier A limitation of the present study needs to be addressed. The
studies have reported an average 30% reduction in D2 binding time interval between the two SPECT scans was two weeks.
based on genotype. Our data indicate that GT subjects have an Although it would have been preferable that the SPECT and
average 34% reduction in [123I]-IBZM binding compared with GG fMRI scans were performed closely in time, this was not always the
subjects. Resting state [123I]FP-CIT measures pre-synaptic binding case and the time interval between the two SPECT scans and the
to DATs [33]. Given the robust literature providing indication of fMRI scan was variable. On average, 25.663.6 months elapsed
the physical and functional relationship between DATs and pre- between these scans. Earlier studies have demonstrated good
synaptic D2 receptors [28,29], reduced [123I]FP-CIT binding in GT reproducibility for [123I] IBZM (as good as 6.565.2 [91,92] and
subjects may be because of reduced pre-synaptic D2S. An for [123I] FP CIT (5.5364.12%/0.89, [93]). As for fMRI,
alternative interpretation of both SPECT datasets is also possible. reproducibility has been investigated in several studies using
Because GT subjects have reduced pre-synaptic D2S which inhibits different working memory paradigms. Studies have reported good
dopamine release [19], reduced [123I]IBZM and [123I]FP-CIT reproducibility of Dorsolateral Prefrontal cortex activation in
binding may be because of greater steady-state levels of dopamine healthy subjects (Intra Class Correlation (ICC = 0.81) coefficient
competing with the radiotracer. Consistent with these contentions [94]). Similar results have also been reported by other investigators
and with knowledge that the main factor contributing to a [95,96]. These earlier studies suggest that it is less likely that the
relationship between pre-synaptic DATs and post-synaptic D2 time interval between scans has significantly influenced the results.
receptors is dopamine itself, factor analysis of both SPECT datasets Moreover, it is important to underline that the relationships
identified a factor that may reflect dopamine D2 signaling (the identified in this study reflect known neurobiological pathways.
major element in common likely affecting binding of the two Also, there was no statistically significant difference between the
radiotracers). two genotype groups in time elapsed between SPECT scans and
DRD2 rs1076560 genotype was also differentially associated fMRI (t = 0.09, p = 0.9) which may have unduly influenced the
with the correlation between putative striatal dopamine D2 differential correlations. Finally, SPECT and PET studies of DAT
signaling (as identified with the factor score) and prefrontal activity and of D2 receptors have demonstrated between 3 and 7%
during working memory, which was positive in GG and negative decrease of radiotracer binding per decade, especially after 40
in GT subjects. A plausible cellular mechanism for these effects is years of age [97–101]. Since our healthy subjects are definitely
the differential contribution of dopamine D2S and D2L receptors younger than 40 (mean age6SD 23.563.0), we believe physio-
in modulation of glutamate and GABA transmission on striatal logical dopamine signaling decline did not significantly affect our
output. Both D2 isoforms participate in pre-synaptic inhibition of analyses over the time interval of the study. Thus, we interpret
striatal GABA transmission, while D2S is preferentially involved in these findings as being consistent with stability of fMRI and
modulation of glutamate release [21]. Therefore, modulation of SPECT measures and as suggesting that they reflect neurobiolog-
excitatory and inhibitory transmission in the striatum is tightly ical mechanisms.
linked to balance of the two isoforms [21]. In fact, GABA spiny Another point that needs some discussion is that, at the chosen
neurons, which account for the large majority of neuronal statistical threshold corrected for multiple comparisons, we failed
populations in striatum, only fire action potentials when to find a main effect of rs1076560 genotype on prefrontal activity
depolarized by glutamate released by afferents from the cortex during performance of working memory in the present dataset. In
[79]. Since DRD2 rs1076560 genotype alters the relative three earlier studies, we had already demonstrated association of
expression of the two isoforms, our data are consistent with the rs1076560 with prefrontal activity during performance of the N-
possibility that the balance of excitatory/inhibitory modulation of Back working memory task in healthy subjects with sample sizes as
striatal neurons may also affect striatal outputs and the relationship large as N = 142 [29–31]. In the present study, the sample
with prefrontal activity during working memory. included 28 healthy subjects. Despite the smaller sample size, GT
Since the present study is in healthy subjects, no direct subjects had greater activity in prefrontal cortex and anterior
conclusion can be drawn about schizophrenia. However, some cingulate before correction for multiple comparisons. Earlier
speculations as to the relevance of the present findings to studies have demonstrated that larger sample sizes are generally
schizophrenia can be discussed. A long debated issue in the needed for imaging genetics studies using corrected statistical
pathophysiology of schizophrenia is whether subcortical dopamine thresholds [102]. Therefore, the present results suggest that the
dysregulation is a primary phenomenon or rather it is associated lack of a statistically significant main effect of rs1076560 is likely
with prefrontal cortical dysfunction [4,80–82]. Indeed, heritability associated with the relatively small sample size of the present fMRI
of both these phenotypes has been demonstrated [83–87]. Several study. Moreover, the main objective of the present fMRI data was
studies and meta-analyses point to potential involvement of DRD2 to evaluate statistically significant DRD2 modulation of the
in susceptibility to schizophrenia. Genome-wide linkage meta- relationship between striatal dopamine D2 signaling with
analyses have pointed to involvement of 11q [88]. Metanalyses of prefrontal activity during performance of working memory which
case-control studies indicate SNPs rs1801028 (Cys311Ser), should not be affected by the corrected significance of the above
rs2283265, and rs6277 (C957T) [2,3] as implicated in risk for mentioned DRD2 effect.

PLoS ONE | www.plosone.org 7 February 2010 | Volume 5 | Issue 2 | e9348

 DRD2 and Prefrontal Activity

Our data are consistent with earlier studies about genetic effects and the factor score extracted from both SPECT data sets in
on modulation of the relationship between mesencephalic striatum.
dopamine and prefrontal cortex activity [103]. Indeed, the ventral Found at: doi:10.1371/journal.pone.0009348.s003 (0.51 MB
tegmental area in the mesencephalon may be implicated in these TIF)
mechanisms. Our data also provide evidence for a genetic
Figure S2 Relationship between behavioral performance and
signature that affects striatal dopamine D2 receptor signaling
and its relationship with WM prefrontal activity, a mechanism prefrontal activity. Scatterplot of the non-linear relationship in GG
potentially involved in schizophrenia and other different brain subjects between working memory behavioral performance and
disorders [4]. prefrontal activity during working memory as measured with
BOLD fMRI.
Found at: doi:10.1371/journal.pone.0009348.s004 (0.50 MB
Supporting Information
TIF)
Table S1 Demographics of all subjects included in the study
divided by rs1076560 genotype. Acknowledgments
Found at: doi:10.1371/journal.pone.0009348.s001 (0.03 MB
DOC) We would like to acknowledge Riccarda Lomuscio, BA, Rita Masellis,
PhD, and Francesco Roselli, MD, for help with data acquisition and design
Table S2 Results from the correlation between 2-Back WM of the experiment.
activity and the basal ganglia SPECT factor score that were not
corrected for multiple comparisons. No clusters survived the Author Contributions
statistical threshold in the inverse correlations.
Found at: doi:10.1371/journal.pone.0009348.s002 (0.04 MB Conceived and designed the experiments: AB PT NMP GB LF FS ANA
DOC) ACP GU LS WS GR. Performed the experiments: AB PT NMP LF RR
BG LLB ML ADG GC ANA ACP GU LS TP GR. Analyzed the data: AB
Figure S1 Relationship between behavioral performance and PT NMP GB LF RR BG LLB ML ADG GC FS ACP GU LS TP WS GR.
dopamine D2 signaling. Scatterplot of the non-linear relationship Contributed reagents/materials/analysis tools: AB WS. Wrote the paper:
in GG subjects between working memory behavioral performance AB PT NMP GB LF RR BG LLB ML ADG GC FS ANA TP WS GR.

References
1. Harrison PJ, Weinberger DR (2005) Schizophrenia genes, gene expression, and 18. Volkow ND, Gur RC, Wang GJ, Fowler JS, Moberg PJ, et al. (1998)
neuropathology: on the matter of their convergence. Mol Psychiatry 10: 40–68; Association between decline in brain dopamine activity with age and cognitive
image 45. and motor impairment in healthy individuals. Am J Psychiatry 155: 344–
2. Allen NC, Bagade S, McQueen MB, Ioannidis JP, Kavvoura FK, et al. (2008) 349.
Systematic meta-analyses and field synopsis of genetic association studies in 19. Watanabe K, Kita T, Kita H (2009) Presynaptic actions of D2-like receptors in
schizophrenia: the SzGene database. Nat Genet 40: 827–834. the rat cortico-striato-globus pallidus disynaptic connection in vitro.
3. Glatt SJ, Faraone SV, Tsuang MT (2003) Meta-analysis identifies an J Neurophysiol 101: 665–671.
association between the dopamine D2 receptor gene and schizophrenia. Mol 20. Usiello A, Baik JH, Rouge-Pont F, Picetti R, Dierich A, et al. (2000) Distinct
Psychiatry 8: 911–915. functions of the two isoforms of dopamine D2 receptors. Nature 408: 199–203.
4. Weinberger DR (1987) Implications of normal brain development for the 21. Centonze D, Gubellini P, Usiello A, Rossi S, Tscherter A, et al. (2004)
pathogenesis of schizophrenia. Arch Gen Psychiatry 44: 660–669. Differential contribution of dopamine D2S and D2L receptors in the
5. Kapur S (2003) Psychosis as a state of aberrant salience: a framework linking modulation of glutamate and GABA transmission in the striatum. Neuroscience
biology, phenomenology, and pharmacology in schizophrenia. Am J Psychiatry 129: 157–166.
160: 13–23. 22. Dickinson SD, Sabeti J, Larson GA, Giardina K, Rubinstein M, et al. (1999)
6. Carlsson A, Carlsson ML (2006) A dopaminergic deficit hypothesis of Dopamine D2 receptor-deficient mice exhibit decreased dopamine transporter
schizophrenia: the path to discovery. Dialogues Clin Neurosci 8: 137–142. function but no changes in dopamine release in dorsal striatum. J Neurochem
7. Callicott JH, Mattay VS, Verchinski BA, Marenco S, Egan MF, et al. (2003) 72: 148–156.
Complexity of prefrontal cortical dysfunction in schizophrenia: more than up 23. Kimmel HL, Joyce AR, Carroll FI, Kuhar MJ (2001) Dopamine D1 and D2
or down. Am J Psychiatry 160: 2209–2215. receptors influence dopamine transporter synthesis and degradation in the rat.
8. Manoach DS (2003) Prefrontal cortex dysfunction during working memory J Pharmacol Exp Ther 298: 129–140.
performance in schizophrenia: reconciling discrepant findings. Schizophr Res
24. Lee FJ, Pei L, Moszczynska A, Vukusic B, Fletcher PJ, et al. (2007) Dopamine
60: 285–298.
transporter cell surface localization facilitated by a direct interaction with the
9. Seamans JK, Yang CR (2004) The principal features and mechanisms of
dopamine D2 receptor. Embo J 26: 2127–2136.
dopamine modulation in the prefrontal cortex. Prog Neurobiol 74: 1–58.
25. Meiergerd SM, Patterson TA, Schenk JO (1993) D2 receptors may modulate
10. Glickstein SB, Hof PR, Schmauss C (2002) Mice lacking dopamine D2 and D3
the function of the striatal transporter for dopamine: kinetic evidence from
receptors have spatial working memory deficits. J Neurosci 22: 5619–5629.
studies in vitro and in vivo. J Neurochem 61: 764–767.
11. O’Reilly RC (2006) Biologically Based Computational Models of High-Level
26. Mortensen OV, Amara SG (2003) Dynamic regulation of the dopamine
Cognition. Science %R 101126/science1127242 314: 91–94.
transporter. Eur J Pharmacol 479: 159–170.
12. Kellendonk C, Simpson EH, Polan HJ, Malleret G, Vronskaya S, et al. (2006)
Transient and selective overexpression of dopamine D2 receptors in the 27. Parsons LH, Schad CA, Justice JB, Jr. (1993) Co-adminis0tration of the D2
striatum causes persistent abnormalities in prefrontal cortex functioning. antagonist pimozide inhibits up-regulation of dopamine release and uptake
Neuron 49: 603–615. induced by repeated cocaine. J Neurochem 60: 376–379.
13. Mehta MA, Manes FF, Magnolfi G, Sahakian BJ, Robbins TW (2004) 28. Bolan EA, Kivell B, Jaligam V, Oz M, Jayanthi LD, et al. (2007) D2 receptors
Impaired set-shifting and dissociable effects on tests of spatial working memory regulate dopamine transporter function via an extracellular signal-regulated
following the dopamine D2 receptor antagonist sulpiride in human volunteers. kinases 1 and 2-dependent and phosphoinositide 3 kinase-independent
Psychopharmacology (Berl) 176: 331–342. mechanism. Mol Pharmacol 71: 1222–1232.
14. Kimberg DY, Aguirre GK, Lease J, D’Esposito M (2001) Cortical effects of 29. Bertolino A, Fazio L, Di Giorgio A, Blasi G, Romano R, et al. (2009)
bromocriptine, a D-2 dopamine receptor agonist, in human subjects, revealed Genetically determined interaction between the dopamine transporter and the
by fMRI. Hum Brain Mapp 12: 246–257. D2 receptor on prefronto-striatal activity and volume in humans. J Neurosci
15. Mehta MA, McGowan SW, Lawrence AD, Aitken MR, Montgomery AJ, et al. 29: 1224–1234.
(2003) Systemic sulpiride modulates striatal blood flow: relationships to spatial 30. Zhang Y, Bertolino A, Fazio L, Blasi G, Rampino A, et al. (2007)
working memory and planning. Neuroimage 20: 1982–1994. Polymorphisms in human dopamine D2 receptor gene affect gene expression,
16. Landau SM, Lal R, O’Neil JP, Baker S, Jagust WJ (2009) Striatal dopamine splicing, and neuronal activity during working memory. Proc Natl Acad
and working memory. Cereb Cortex 19: 445–454. Sci U S A 104: 20552–20557.
17. Nyberg L, Andersson M, Forsgren L, Jakobsson-Mo S, Larsson A, et al. (2009) 31. Bertolino A, Fazio L, Caforio G, Blasi G, Rampino A, et al. (2009) Functional
Striatal dopamine D2 binding is related to frontal BOLD response during variants of the dopamine receptor D2 gene modulate prefronto-striatal
updating of long-term memory representations. Neuroimage 46: 1194–1199. phenotypes in schizophrenia. Brain 132: 417–425.

PLoS ONE | www.plosone.org 8 February 2010 | Volume 5 | Issue 2 | e9348

 DRD2 and Prefrontal Activity

32. Kung HF, Alavi A, Chang W, Kung MP, Keyes JW, Jr., et al. (1990) In vivo 57. Lancaster JL, Woldorff MG, Parsons LM, Liotti M, Freitas CS, et al. (2000)
SPECT imaging of CNS D-2 dopamine receptors: initial studies with iodine- Automated Talairach atlas labels for functional brain mapping. Hum Brain
123-IBZM in humans. J Nucl Med 31: 573–579. Mapp 10: 120–131.
33. Scheffel U, Lever JR, Abraham P, Parham KR, Mathews WB, et al. (1997) N- 58. Maldjian JA, Laurienti PJ, Kraft RA, Burdette JH (2003) An automated
substituted phenyltropanes as in vivo binding ligands for rapid imaging studies method for neuroanatomic and cytoarchitectonic atlas-based interrogation of
of the dopamine transporter. Synapse 25: 345–349. fMRI data sets. Neuroimage 19: 1233–1239.
34. Gogos JA, Morgan M, Luine V, Santha M, Ogawa S, et al. (1998) Catechol-O- 59. Tzourio-Mazoyer N, Landeau B, Papathanassiou D, Crivello F, Etard O, et al.
methyltransferase-deficient mice exhibit sexually dimorphic changes in (2002) Automated anatomical labeling of activations in SPM using a
catecholamine levels and behavior. Proc Natl Acad Sci U S A 95: 9991–9996. macroscopic anatomical parcellation of the MNI MRI single-subject brain.
35. Sesack SR, Hawrylak VA, Guido MA, Levey AI (1998) Cellular and subcellular Neuroimage 15: 273–289.
localization of the dopamine transporter in rat cortex. Adv Pharmacol 42: 60. Rakshi JS, Uema T, Ito K, Bailey DL, Morrish PK, et al. (1999) Frontal,
171–174. midbrain and striatal dopaminergic function in early and advanced Parkinson’s
36. Egan MF, Goldberg TE, Kolachana BS, Callicott JH, Mazzanti CM, et al. disease A 3D [(18)F]dopa-PET study. Brain 122 (Pt 9): 1637–1650.
(2001) Effect of COMT Val108/158 Met genotype on frontal lobe function 61. Laruelle M, Baldwin RM, Malison RT, Zea-Ponce Y, Zoghbi SS, et al. (1993)
and risk for schizophrenia. Proc Natl Acad Sci U S A 98: 6917–6922. SPECT imaging of dopamine and serotonin transporters with [123I]beta-CIT:
37. Papp AC, Pinsonneault JK, Cooke G, Sadee W (2003) Single nucleotide pharmacological characterization of brain uptake in nonhuman primates.
polymorphism genotyping using allele-specific PCR and fluorescence melting Synapse 13: 295–309.
curves. Bio Techniques 34: 1067–1072. 62. Laruelle M, D’Souza CD, Baldwin RM, Abi-Dargham A, Kanes SJ, et al.
38. Zhang Y, Wang D, Johnson AD, Papp AC, Sadee W (2005) Allelic expression (1997) Imaging D2 receptor occupancy by endogenous dopamine in humans.
imbalance of human mu opioid receptor (OPRM1) caused by variant A118G. Neuropsychopharmacology 17: 162–174.
J Biol Chem 280: 32618–32624. 63. Jones SR, Gainetdinov RR, Hu XT, Cooper DC, Wightman RM, et al. (1999)
39. Bertolino A, Blasi G, Latorre V, Rubino V, Rampino A, et al. (2006) Additive Loss of autoreceptor functions in mice lacking the dopamine transporter. Nat
effects of genetic variation in dopamine regulating genes on working memory Neurosci 2: 649–655.
cortical activity in human brain. J Neurosci 26: 3918–3922. 64. HIll T, Lewicki P (2006) Statistics: methods and applications. A comprehensive
40. Bertolino A, Caforio G, Blasi G, De Candia M, Latorre V, et al. (2004) reference for science, industry, and data mining: StatSofr Inc.
Interaction of COMT Val108/158 Met Genotype and Olanzapine Treatment 65. Callicott JH, Mattay VS, Bertolino A, Finn K, Coppola R, et al. (1999)
on Prefrontal Cortical Function in Patients With Schizophrenia. Am J Psychiatry Physiological characteristics of capacity constraints in working memory as
161: 1798–1805. revealed by functional MRI. Cereb Cortex 9: 20–26.
41. Catafau AM, Tolosa E (2004) Impact of dopamine transporter SPECT using 66. Bertolino A, Blasi G, Caforio G, Latorre V, De Candia M, et al. (2004)
123I-Ioflupane on diagnosis and management of patients with clinically Functional lateralization of the sensorimotor cortex in patients with
uncertain Parkinsonian syndromes. Mov Disord 19: 1175–1182. schizophrenia: effects of treatment with olanzapine. Biol Psychiatry 56:
42. Booij J, Hemelaar TG, Speelman JD, de Bruin K, Janssen AG, et al. (1999) 190–197.
One-day protocol for imaging of the nigrostriatal dopaminergic pathway in 67. Curtis CE, Rao VY, D’Esposito M (2004) Maintenance of spatial and motor
Parkinson’s disease by [123I]FPCIT SPECT. J Nucl Med 40: 753–761. codes during oculomotor delayed response tasks. J Neurosci 24: 3944–3952.
43. Meyer PT, Sattler B, Winz OH, Fundke R, Oehlwein C, et al. (2008) Kinetic 68. Tan HY, Chen Q, Goldberg TE, Mattay VS, Meyer-Lindenberg A, et al.
analyses of [123I]IBZM SPECT for quantification of striatal dopamine D2 (2007) Catechol-O-methyltransferase Val158Met modulation of prefrontal-
receptor binding: a critical evaluation of the single-scan approach. Neuroimage parietal-striatal brain systems during arithmetic and temporal transformations
42: 548–558. in working memory. J Neurosci 27: 13393–13401.
44. Seibyl J, Jennings D, Tabamo R, Marek K (2005) Unique roles of SPET brain 69. Manoach DS, White NS, Lindgren KA, Heckers S, Coleman MJ, et al. (2004)
imaging in clinical and research studies. Lessons from Parkinson’s disease Hemispheric specialization of the lateral prefrontal cortex for strategic
research. Q J Nucl Med Mol Imaging 49: 215–221. processing during spatial and shape working memory. Neuroimage 21:
45. Brucke T, Podreka I, Angelberger P, Wenger S, Topitz A, et al. (1991) 894–903.
Dopamine D2 receptor imaging with SPECT: studies in different neuropsy- 70. Williams GV, Castner SA (2006) Under the curve: critical issues for elucidating
chiatric disorders. J Cereb Blood Flow Metab 11: 220–228. D1 receptor function in working memory. Neuroscience 139: 263–276.
46. Tatsch K, Asenbaum S, Bartenstein P, Catafau A, Halldin C, et al. (2002) 71. Bertolino A, Caforio G, Petruzzella V, Latorre V, Rubino V, et al. (2006)
European Association of Nuclear Medicine procedure guidelines for brain Prefrontal dysfunction in schizophrenia controlling for COMT Val(158)Met
neurotransmission SPET using (123)I-labelled dopamine D(2) receptor ligands. genotype and working memory performance. Psychiatry Res 147: 221–226.
Eur J Nucl Med Mol Imaging 29: BP23–29. 72. Callicott JH, Bertolino A, Mattay VS, Langheim FJ, Duyn J, et al. (2000)
47. Soret M, Koulibaly PM, Darcourt J, Hapdey S, Buvat I (2003) Quantitative Physiological dysfunction of the dorsolateral prefrontal cortex in schizophrenia
accuracy of dopaminergic neurotransmission imaging with (123)I SPECT. revisited. Cereb Cortex 10: 1078–1092.
J Nucl Med 44: 1184–1193. 73. Hirvonen MM, Nagren K, Rinne JO, Pesonen U, Vahlberg T, et al. (2009)
48. Laruelle M, Baldwin RM, Innis RB (1994) SPECT imaging of dopamine and COMT Val158Met Genotype Does Not Alter Cortical or Striatal Dopamine
serotonin transporters in nonhuman primate brain. NIDA Res Monogr 138: D2 Receptor. Availability In Vivo Mol Imaging Biol.
131–159. 74. Abi-Dargham A, Rodenhiser J, Printz D, Zea-Ponce Y, Gil R, et al. (2000)
49. Frankle WG, Gil R, Hackett E, Mawlawi O, Zea-Ponce Y, et al. (2004) Increased baseline occupancy of D2 receptors by dopamine in schizophrenia.
Occupancy of dopamine D2 receptors by the atypical antipsychotic drugs Proc Natl Acad Sci U S A 97: 8104–8109.
risperidone and olanzapine: theoretical implications. Psychopharmacology 75. Guo N, Guo W, Kralikova M, Jiang M, Schieren I, et al. (2009) Impact of D2
(Berl) 175: 473–480. Receptor Internalization on Binding Affinity of Neuroimaging Radiotracers.
50. Scherfler C, Seppi K, Donnemiller E, Goebel G, Brenneis C, et al. (2005) Neuropsychopharmacology.
Voxel-wise analysis of [123I]beta-CIT SPECT differentiates the Parkinson 76. Jonsson EG, Nothen MM, Neidt H, Forslund K, Rylander G, et al. (1999)
variant of multiple system atrophy from idiopathic Parkinson’s disease. Brain Association between a promoter polymorphism in the dopamine D2 receptor
128: 1605–1612. gene and schizophrenia. Schizophr Res 40: 31–36.
51. Lidow MS, Goldman-Rakic PS, Rakic P, Innis RB (1989) Dopamine D2 77. Pohjalainen T, Rinne JO, Nagren K, Lehikoinen P, Anttila K, et al. (1998) The
receptors in the cerebral cortex: distribution and pharmacological character- A1 allele of the human D2 dopamine receptor gene predicts low D2 receptor
ization with [3H]raclopride. Proc Natl Acad Sci U S A 86: 6412–6416. availability in healthy volunteers. Mol Psychiatry 3: 256–260.
52. Lewis DA, Melchitzky DS, Sesack SR, Whitehead RE, Auh S, et al. (2001) 78. Ritchie T, Noble EP (2003) Association of seven polymorphisms of the D2
Dopamine transporter immunoreactivity in monkey cerebral cortex: regional, dopamine receptor gene with brain receptor-binding characteristics. Neuro-
laminar, and ultrastructural localization. J Comp Neurol 432: 119–136. chem Res 28: 73–82.
53. Laruelle M, Abi-Dargham A, van Dyck CH, Gil R, D’Souza CD, et al. (1996) 79. Calabresi P, Pisani A, Mercuri NB, Bernardi G (1996) The corticostriatal
Single photon emission computerized tomography imaging of amphetamine- projection: from synaptic plasticity to dysfunctions of the basal ganglia. Trends
induced dopamine release in drug-free schizophrenic subjects. Proc Natl Acad Neurosci 19: 19–24.
Sci U S A 93: 9235–9240. 80. Bertolino A, Knable MB, Saunders RC, Callicott JH, Kolachana B, et al.
54. Seibyl JP, Woods SW, Zoghbi SS, Baldwin RM, Dey HM, et al. (1992) (1999) The relationship between dorsolateral prefrontal N-acetylaspartate
Dynamic SPECT imaging of dopamine D2 receptors in human subjects with measures and striatal dopamine activity in schizophrenia. Biol Psychiatry 45:
iodine-123-IBZM. J Nucl Med 33: 1964–1971. 660–667.
55. Beukers RJ, Booij J, Weisscher N, Zijlstra F, van Amelsvoort TA, et al. (2009) 81. Bertolino A, Breier A, Callicott JH, Adler C, Mattay VS, et al. (2000) The
Reduced striatal D2 receptor binding in myoclonus-dystonia. Eur J Nucl Med relationship between dorsolateral prefrontal neuronal N- acetylaspartate and
Mol Imaging 36: 269–274. evoked release of striatal dopamine in schizophrenia. Neuropsychopharmacol-
56. Verhoeff NP, Kapucu O, Sokole-Busemann E, van Royen EA, Janssen AG ogy 22: 125–132.
(1993) Estimation of dopamine D2 receptor binding potential in the striatum 82. Meyer-Lindenberg A, Miletich RS, Kohn PD, Esposito G, Carson RE, et al.
with iodine-123-IBZM SPECT: technical and interobserver variability. J Nucl (2002) Reduced prefrontal activity predicts exaggerated striatal dopaminergic
Med 34: 2076–2084. function in schizophrenia. Nat Neurosci 5: 267–271.

PLoS ONE | www.plosone.org 9 February 2010 | Volume 5 | Issue 2 | e9348

 DRD2 and Prefrontal Activity

83. Huttunen J, Heinimaa M, Svirskis T, Nyman M, Kajander J, et al. (2008) 93. Tsuchida T, Ballinger JR, Vines D, Kim YJ, Utsunomiya K, et al. (2004)
Striatal dopamine synthesis in first-degree relatives of patients with schizo- Reproducibility of dopamine transporter density measured with 123I-FPCIT
phrenia. Biol Psychiatry 63: 114–117. SPECT in normal control and Parkinson’s disease patients. Ann Nucl Med 18:
84. Hirvonen J, van Erp TG, Huttunen J, Aalto S, Nagren K, et al. (2005) 609–616.
Increased caudate dopamine D2 receptor availability as a genetic marker for 94. Manoach DS, Halpern EF, Kramer TS, Chang Y, Goff DC, et al. (2001) Test-
schizophrenia. Arch Gen Psychiatry 62: 371–378. retest reliability of a functional MRI working memory paradigm in normal and
85. Howes OD, Montgomery AJ, Asselin MC, Murray RM, Valli I, et al. (2009) schizophrenic subjects. Am J Psychiatry 158: 955–958.
Elevated striatal dopamine function linked to prodromal signs of schizophrenia. 95. Caceres A, Hall DL, Zelaya FO, Williams SC, Mehta MA (2009) Measuring
Arch Gen Psychiatry 66: 13–20. fMRI reliability with the intra-class correlation coefficient. Neuroimage 45:
86. Callicott JH, Egan MF, Mattay VS, Bertolino A, Bone AD, et al. (2003) 758–768.
Abnormal fMRI response of the dorsolateral prefrontal cortex in cognitively 96. Wei X, Yoo SS, Dickey CC, Zou KH, Guttmann CR, et al. (2004) Functional
intact siblings of patients with schizophrenia. Am J Psychiatry 160: 709–719. MRI of auditory verbal working memory: long-term reproducibility analysis.
87. Seidman LJ, Thermenos HW, Poldrack RA, Peace NK, Koch JK, et al. (2006) Neuroimage 21: 1000–1008.
Altered brain activation in dorsolateral prefrontal cortex in adolescents and 97. Ishikawa T, Dhawan V, Kazumata K, Chaly T, Mandel F, et al. (1996)
Comparative nigrostriatal dopaminergic imaging with iodine-123-beta CIT-
young adults at genetic risk for schizophrenia: an fMRI study of working
FP/SPECT and fluorine-18-FDOPA/PET. J Nucl Med 37: 1760–1765.
memory. Schizophr Res 85: 58–72.
98. Booij J, Bergmans P, Winogrodzka A, Speelman JD, Wolters EC (2001)
88. Lewis CM, Levinson DF, Wise LH, DeLisi LE, Straub RE, et al. (2003)
Imaging of dopamine transporters with [123I]FP-CIT SPECT does not suggest
Genome scan meta-analysis of schizophrenia and bipolar disorder, part II:
a significant effect of age on the symptomatic threshold of disease in Parkinson’s
Schizophrenia. Am J Hum Genet 73: 34–48. disease. Synapse 39: 101–108.
89. Betcheva ET, Mushiroda T, Takahashi A, Kubo M, Karachanak SK, et al. 99. Volkow ND, Ding YS, Fowler JS, Wang GJ, Logan J, et al. (1996) Dopamine
(2009) Case-control association study of 59 candidate genes reveals the DRD2 transporters decrease with age. J Nucl Med 37: 554–559.
SNP rs6277 (C957T) as the only susceptibility factor for schizophrenia in the 100. Chen PS, Yang YK, Lee YS, Yeh TL, Lee IH, et al. (2005) Correlation
Bulgarian population. J Hum Genet 54: 98–107. between different memory systems and striatal dopamine D2/D3 receptor
90. Glatt SJ, Faraone SV, Lasky-Su JA, Kanazawa T, Hwu HG, et al. (2008) density: a single photon emission computed tomography study. Psychol Med
Family-based association testing strongly implicates DRD2 as a risk gene for 35: 197–204.
schizophrenia in Han Chinese from Taiwan. Mol Psychiatry. 101. Volkow ND, Wang GJ, Fowler JS, Logan J, Gatley SJ, et al. (1996) Measuring
91. Catafau AM, Bullich S, Danus M, Penengo MM, Cot A, et al. (2008) Test- age-related changes in dopamine D2 receptors with 11C-raclopride and 18F-
retest variability and reliability of 123I-IBZM SPECT measurement of striatal N-methylspiroperidol. Psychiatry Res 67: 11–16.
dopamine D2 receptor availability in healthy volunteers and influence of 102. Meyer-Lindenberg A, Nicodemus KK, Egan MF, Callicott JH, Mattay V, et al.
iterative reconstruction algorithms. Synapse 62: 62–69. (2008) False positives in imaging genetics. Neuroimage 40: 655–661.
92. Kegeles LS, Zea-Ponce Y, Abi-Dargham A, Rodenhiser J, Wang T, et al. 103. Meyer-Lindenberg A, Kohn PD, Kolachana B, Kippenhan S, McInerney-
(1999) Stability of [123I]IBZM SPECT measurement of amphetamine-induced Leo A, et al. (2005) Midbrain dopamine and prefrontal function in humans:
striatal dopamine release in humans. Synapse 31: 302–308. interaction and modulation by COMT genotype. Nat Neurosci 8: 594–596.

PLoS ONE | www.plosone.org 10 February 2010 | Volume 5 | Issue 2 | e9348