Journal of Chemical Neuroanatomy 22 (2001) 127– 137

www.elsevier.com/locate/jchemneu

D1 and D2 dopamine receptor mRNA expression in whole

hemisphere sections of the human brain
Yasmin L. Hurd *, Michio Suzuki 1, Göran C. Sedvall
Psychiatry Section, Department of Clinical Neuroscience, Karolinska Institutet, Karolinska Hospital, SE-171 76 Stockholm, Sweden

Received 25 January 2001; received in revised form 9 May 2001; accepted 9 May 2001

Abstract

Understanding dopamine signaling in human behavior requires knowledge of the distribution of all molecular components
involved in dopamine pathways throughout the human brain. In the present study, the relative distributions of D1 and D2
dopamine receptor mRNAs were determined by in situ hybridization histochemistry in whole hemisphere sections from normal
human post mortem brains. The findings confirmed information documented from single structure examination that the highest
expression of both the D1 and D2 mRNAs were localized to the striatum. The cerebral cortex expressed moderate D1 mRNA in
all regions with the highest signal in the medial orbital frontal area (Brodmann areas 11, 14), the paraterminal gyrus (Brodmann
area 32) and the insular cortex (Brodmann areas 13–16), whereas the D2 mRNA expression had very low cortical expression. The
bed nucleus of the stria terminalis and islands of Calleja had high expression of the D1 mRNA and moderate D2 mRNA levels.
Moderate to high expression of the D2 mRNA was evident in the hippocampal formation, parafascicular and paraventricular
thalamic nuclei, geniculate bodies, subthalamic nucleus, and pineal gland, all of which were devoid of, or showed only faint, D1
mRNA expression. Brainstem regions, e.g. substantia nigra, red nucleus, inferior colliculus, medial lemniscus, and pontine nuclei
expressed D2, but not D1, mRNA. These results emphasize the differential anatomical localization of D1 and D2 dopamine
receptor mRNA neuronal populations in the human brain. The restricted expression of the D1 mRNA to the cortical mantle and
to a few forebrain structures indicates a strong involvement of the D1 system in cognitive function. © 2001 Elsevier Science B.V.
All rights reserved.

Keywords: Dopamine receptor; In situ hybridization; Neocortex; Striatum; Islands of Calleja

Abbre6iations: ac, anterior commissure; aCg, anterior cingulate
gyrus; BNST, bed nucleus stria terminalis; Cb, cerebellum; cc, corpus
callosum; Cl, claustrum; CN, caudate nucleus; CNt, caudate nucleus,
tail; Cr, corona radiata; F, frontal lobe; GPe, globus pallidus, exter-
nal; gR, gyrus rectus; Hipp, hippocampus; Hyp, hypothalamus; I,
insula; ia, internal capsule, anterior limb; IC, inferior colliculus; ICj,
islands of Calleja; ip, internal capsule, posterior limb; Lg, lateral
geniculate; Mg, medial geniculate; MTg, medial temporal gyrus;
NAc, nucleus accumbens; O, occipital lobe; Org, orbital gyrus; P,
parietal lobe; Pf, parafasciular thalamus; PH, posterior hypothala-
mus; Phg, parahippocampal gyrus; Pi, pineal; Pn, pontine nuclei;
PTg, paraterminal gyrus; Pu, putamen; Pul, pulvinar; RN, red nu-
cleus; S, subiculum; SC, superior colliculus; SN, substantia nigra;
STg, superior temporal gyrus; T, temporal lobe; Th, thalamus; Ug,
uncal gyrus.
* Corresponding author. Tel.: +46-8-517-72379; fax: +46-8-346-
563.
E-mail address: yasmin.hurd@ks.se (Y.L. Hurd).
1
Present address: Department of Neuropsychiatry, Toyama Medi-
cal and Pharmaceutical University, 2630 Sugitani, Toyama 930-0194,
Japan.
1. Introduction
The importance of dopamine signaling in the brain
for the mediation of motor, endocrine, cognitive, cona-
tive and affective functions and their coordination has
been well acknowledged (Fibiger, 1995; Graybiel, 1997;
Goldman-Rakic, 1998; Berke and Hyman, 2000). The
diversity of dopamine functions is partly related to the
vast and highly variable distribution of dopamine fibers
and terminals to most regions of the brain and also to
the multitude of receptor proteins mediating different
types of effects in response to dopamine release
(Sokoloff and Schwartz, 1995; Jaber et al., 1996).
For practically every major neuropsychiatric disease
group, dopamine has been implicated to play a primary
or secondary role in the pathophysiology or in the
mechanisms of drug action (Knable and Weinberger,
1997; Tarazi et al., 1997). This is particularly true for

0891-0618/01/$ - see front matter © 2001 Elsevier Science B.V. All rights reserved.
PII: S 0 8 9 1 - 0 6 1 8 ( 0 1 ) 0 0 1 2 2 - 3
128 Y.L. Hurd et al. / Journal of Chemical Neuroanatomy 22 (2001) 127–137
schizophrenia where dopamine signaling in neostriatal,
mesolimbic and mesocortical systems has been dis-
cussed but remained controversial for many years
(Laruelle et al., 1996).
The development of models for understanding the
role of dopamine signaling in unitary and coordinated
human behavior requires detailed knowledge on the
distribution of all the molecular components involved
in dopamine pathways throughout the human brain.
D1 and D2 receptors constitute the major dopamine
receptor subtypes in the human brain. These receptors
have been thoroughly characterized with regard to rela-
tive protein levels and mRNA expression in a number
of regions of the human brain (Kessler et al., 1993;
Meador-Woodruff et al., 1994b; Smiley et al., 1994;
Hall et al., 1996; Meador-Woodruff et al., 1996). Most
studies of the dopamine receptors have been focused on
the striatum (caudate nucleus, putamen, and nucleus
accumbens). Recent brain imaging studies have confi-
rmed a high degree of localized functional specificity for
allo- and neocortical brain regions in schizophrenia and
affective disorders (Mayberg et al., 1999; MacDonald et
al., 2000). Although in vivo imaging studies allow the
possibility to visualize multiple systems simultaneously,
they do not provide information at the level of gene
expression. Post mortem studies, which can provide
detailed information about mRNA expression in dis-
crete neuronal populations, have generally been limited
to characterizing single brain structures.
In the present study, the relative expression of the D1
and D2 receptor mRNAs were studied in whole human
hemisphere brain sections using in situ hybridization
histochemistry (ISHH). This allowed the possibility to
study the pattern of the DA receptor mRNAs in multi-
ple brain areas simultaneously which can help to iden-
tify groups of discrete neuronal populations that
express the specific dopamine receptor genes within the
human brain and might be impaired in neuropsychiatric
disorders.
2. Material and methods
2.1. Tissue
Normal human brains were obtained at autopsy from
the Forensic Medicine Department at the Karolinska
Institutet under guidelines approved by the ethics com-
mittee and the Swedish Board of Health and Social
Welfare. Whole brain hemispheres were studied from
three subjects: 2 males and 1 female, average age
(49.3396.56), average post mortem delay time of
29.459 6.75 h. All subjects died from acute cardiores-
piratory failure. The toxicological reports showed no
presence of neuroactive drugs including alcohol, antide-
pressants, antipsychotics, or minor tranquilizers. None
of the brains showed any evidence of macroscopic
pathology. In addition, there were no indices of sub-
stance abuse, psychiatric, or neurological disease in any
of the subjects, as assessed from examination of medical
records gathered from hospitals in the Stockholm re-
gion. The brains were divided along the midsagittal line
and the whole hemispheres were frozen (− 85 °C) on
glass plates with the sagittal plane facing down as
previously described (Brené et al., 1994; Hall et al.,
1994). In preparation for cryosectioning, the brain
hemisphere, oriented to allow for brain sections in the
horizontal or coronal plane, was embedded in a metal
frame mounted to a cooled stage with a carboxymethyl-
cellulose semi-liquid gel and frozen at − 85 °C. Subse-
quently the hemisphere was placed in a cold (−20 °C)
cryomicrotome (LKB 2250, LKB, Stockholm, Sweden)
and, after equilibration to the cryostat temperature, 100
mm thick sections were taken throughout the entire
brain hemisphere. The sections were placed onto poly-
L-lysine-treated glass plates, and stored frozen at −
30 °C until later use.
2.2. In situ hybridization histochemistry
One section from at least eight different brain levels
from each subject was studied simultaneously. D1 and
D2 mRNA in situ hybridization experiments were pro-
cessed on different days. Tissues were prefixed in prepa-
ration for ISHH as previously described (Hurd, 1996)
under conditions to prevent contamination by RNases.
Briefly, for ISHH, the glass-mounted tissue sections
were brought to room temperature and fixed by immer-
sion in 4% formaldehyde in phosphate-buffered saline
(PBS, pH 7.4) for 5 min at 25 °C, rinsed twice in PBS,
treated with 0.25% acetic anhydride in 0.1 M tri-
ethanolamine/0.9% saline, pH 8.0, for 10 min, dehy-
drated in 70, 80, 95, and 100% ethanol, delipidated in
chloroform for 5 min, rinsed in 100 and 95% ethanol,
and air-dried. All solutions were made with autoclaved
0.1% diethylpyrocarbonate-treated water.
For the synthesis of the D1 receptor riboprobe, a 900
bp cDNA fragment covering the first five transmem-
brane regions of the human D1 receptor (Zhou et al.,
1990); provided by Dr O. Civelli) was subcloned into a
pBSKS plasmid vector. The RNA probe for the DA D2
receptor was synthesized from a full length 1.58 kb
cDNA fragment of the gene for the long form of the
human receptor (Grandy et al., 1989); provided by Dr
O. Crivelli), subcloned into a pBSKS plasmid vector. In
vitro transcription was carried out by using [35S]uridine
5%-[a-thio]triphosphate (New England Nuclear, Boston,
MA, USA) to radiolabel the D1 and D2 receptor
probes. For the D2 receptor riboprobe, alkaline hydrol-
ysis (to decrease the size of the transcripts to ensure
better penetrance into the brain sections) was per-
formed as follows: The probe was incubated for 30 min
 Y.L. Hurd et al. / Journal of Chemical Neuroanatomy 22 (2001) 127–137
with an equal volume of 0.2 M carbonate buffer (pH
10.2) and neutralized with 3 M sodium acetate (pH 6.0)
and 10% (v/v) glacial acetic acid. Ethanol precipitation
was subsequently carried out in the presence of 1 ml
yeast tRNA; − 80 °C for 60 min. The pellets were
rinsed with 70% cold ethanol for 5 min and redissolved
in DEPC-treated water containing 20 mM dithiothrei-
tol (DTT).
The hybridization was carried out with 20× 104 cpm-
labeled probe per mm2. The whole hemisphere sections
were covered with 2000 ml of heat-denatured hybridiza-
tion solution (4X standard sodium citrate (SSC; 1X=
3M sodium chloride, 0.3M sodium citrate, pH 7.0),
10% (w/v) dextran sulfate, 1X Denhardt’s (0.02% Fi-
coll, 0.2% polyvinylpyrrolidone, 0.2 mg/ml bovine
serum albumin), 0.5 mg/ml sheared, single-stranded
salmon sperm DNA, 250 mg/ml yeast tRNA, 200mM
DTT and 50% formamide (the chemicals used in the
hybridization procedure were purchased mainly from
Sigma, MO, USA)). The tissue sections were cover-
slipped and placed in an incubation oven overnight (at
55°C for D1 ISHH and 60 °C for D2 ISHH) with 4X
SSC/50% formamide humidifying environment. The
coverslips were removed and the tissue sections were
placed in 2X SSC diluted in 1 mM DTT solution and
were carried through the following washes: 10 min in
RNase A buffer (5 M NaCl, 1 M Tris (pH 8.0), 0.5 M
EDTA (pH 8.0)) at 37 °C, 30 min in RNase A (20
mg/ml) plus RNase A buffer at 37 °C, several 5– 10 min
series of washes in decreasing concentrations of SSC in
1 mM DTT at room temperature, 60 min in 0.5X
SSC/1 mM DTT/50% formamide (at 48 °C for D1
receptor ISHH; at 60 °C for D2 receptor ISHH) and 60
min in 0.1X SSC/1 mM DTT (at 53 °C for D1 receptor
ISHH; at 55 °C for D2 receptor ISHH). Subsequently
tissue sections were rinsed for 1 min in 0.1X SSC/1 mM
DTT at room temperature and dehydrated by ascend-
ing 1 min ethanol rinses containing 300 mM ammo-
nium acetate, 100% ethanol, and dried. The slides were
then apposed to Hyperfilm b-Max (Amersham, Buck-
hinghamshire, UK) in X-ray cassettes for 7–18 days
with 14C standards, and developed (D19, Kodak). Near
adjacent sections were stained with cresyl violet for
gross histological examination of Nissl substances.
Digitized images were made from the film autoradio-
grams of the whole hemisphere sections. The films were
scanned (ScanMarker III; Microtek Electronics, Düs-
seldorf, Germany) at a resolution of 300 dpi. Densito-
metric readings within specific brain structures were
taken using a Macintosh-based image analysis software
system (IMAGE, Wayne Rasband, NIMH). Anatomi-
cal sites were identified using histochemical landmarks
in conjunction with information obtained from pub-
lished sources (Paxinos, 1990; Parent, 1996) as well as
the Mai et al. (1997), Duvernoy (1991) human brain
atlases. Riboprobe hybridization signals for the D1 and
129
D2 receptor mRNA were measured as grey level (digi-
tally quantified into 255 subunits). The Nissl-stained
sections were used for identification of the brain areas.
In situ hybridization was also carried out on near
adjacent brain sections to localize tyrosine hydroxylase
and serotonin transporter mRNA expression, which
were used to identify aminergic neuronal populations in
the brainstem, and preprotachykinin (Hurd et al.,
1999), and prodynorphin (Hurd, 1996) mRNA expres-
sion in order to dissociate various forebrain neuronal
populations. In the whole hemisphere brain sections,
measurements were taken throughout the rostral-to-
caudal extent of each structure of interest, and in the
dorsal and ventral subregions of any structure that was
visible at different levels of the brain sections studied.
Measurements obtained for each brain region were
averaged. Light transmittance values were converted to
dpm/mg values by means of the 14C standards. Back-
ground labeling in the surrounding white matter was
subtracted from dpm/mg values obtained in gray mat-
ter structures.
All figures were prepared digitally by using the IM-
AGE and the PhotoShop (Adobe, version 6.5) software
programs and contrast was adjusted for some images.
3. Results
Hybridizations carried out with the D1 and D2 anti-
sense riboprobes resulted in a discrete heterogenous
distribution pattern in the human brain specimens
(Figs. 1–4). The specificity of the hybridization signal
was evidenced by the different distribution patterns
exhibited by the D1 and D2 probes and from other
transcripts that had been previously studied on the
same brain specimens (see, e.g. Suzuki et al., 1998;
Hurd et al., 1999). The distinct D1 and D2 hybridiza-
tion patterns were also observed in other brain speci-
mens that were processed using a similar in situ
hybridization procedure (data not shown). In addition,
no positive hybridization signals above background
(white matter) were observed with the sense riboprobe
(Fig. 1).
The distribution pattern of the mRNA expression is
described below relative to the expression levels de-
tected in the highest (striatum) and lowest (cerebellum)
mRNA-expressing brain regions. Fig. 2 shows the rela-
tive dpm/mg levels (as a percentage of the caudate
nucleus) measured in different structures in the whole
human hemisphere cryosections.
3.1. D1 Dopamine receptor mRNA expression in the
human brain
Telecephalon: low to moderate hybridization signals
were detected in the cerebral cortex. All cortical areas
130 Y.L. Hurd et al. / Journal of Chemical Neuroanatomy 22 (2001) 127–137

showed a similar laminar distribution pattern of the D1
mRNA expression with labeling concentrated in two
bands consistent with layers II and V/VI. The highest
hybridization signals were observed in the medial, or-
bital frontal areas of the gyrus rectus (BA area 11, 14)
and subcallosal/paraterminal gyrus (BA 25, 32; Figs.
1A, 2 and 3D) as well as in the insular cortex (BA
13 – 16; Figs. 1A, 2, 3 and 4A). Moderate to high levels
were also detected in the striate cortex (BA 17). The
lowest expression levels were evident in the inferior (BA
44, 45, 47) and middle (BA 8– 10, 46) frontal cortices
(Fig. 4). As evident in Fig. 3, the striate cortex showed
the most distinct band of D1 mRNA labeling perhaps
due to the narrow deep layers in the striate as com-
pared to other cortices.
The striatum expressed the highest levels of the D1
mRNA in the human brain (Figs. 1– 4). A heteroge-
nous pattern with islands of high intensity against a
background of somewhat lower homogenous signal was
characteristic of this region. There were no significant
rostrocaudal differences in the striatal expression levels
of the signal, but there tended to be a medial-to-lateral
decreasing gradient in the D1 mRNA expression. The
bed nucleus of the stria terminalis (Figs. 2, 3B and 4A)
also showed very high expression levels of the D1
mRNA as well as the islands of Calleja (Figs. 1A, 2 and
4A). No positive signal was detected in the globus
pallidus (Figs. 2, 3B and 4A). Low expression levels
were apparent in the claustrum, but only at the mid-
dorsal level (Fig. 3B).
The hippocampal formation including the subicular
region showed no positive D1 hybridization signals
(Figs. 1A, 2, 3B–D and 4D). The amygdaloid complex
also showed very low expression levels with weak sig-
nals evident in the lateral amygdala nuclei (Figs 1A and
2).
Diencephalon: no level of the thalamus studied ex-
pressed the D1 mRNA above background (Figs. 2,
3A–C and 4B). The hypothalamus was also devoid of
any positive hybridization signal in the in situ hy-
bridization conditions carried out in this study.
Mes- and metencephalon: the brainstem regions ex-
amined appeared devoid of the D1 mRNA expression
and no positive hybridization signals were detected in
the cerebellum (Figs. 2, 3D and E and 4B).

Fig. 1. Distribution of D1 and D2 mRNA expression in whole brain horizontal cryosections of human post mortem brain images using antisense
(A) or sense (B) riboprobes at a level approximately 100 mm from vertex. For abbreviations, see list. Scale bar, 2 cm.
 Y.L. Hurd et al. / Journal of Chemical Neuroanatomy 22 (2001) 127–137 131

Fig. 2. Bar graph representing the relative D1 and D2 mRNA expression levels (expressed as a percentage of the dpm/mg measured levels in the
caudate nucleus; mean 9 SEM) within the specified brain areas in the whole hemisphere human sections. Numbers in parenthesis correspond to
Brodmann cerebral cortex nomenclature.

3.2. D2 dopamine receptor mRNA expression in the
human brain
Telecephalon: the cortical expression of the D2
mRNA was extremely low (Figs. 1– 4). The laminar
distribution pattern of the weak D2 hybridization sig-
nal showed three bands with labeling in both superficial
and deep layers.
The striatum showed the highest expression of the
D2 mRNA in the brain with a homogenous distribu-
tion throughout the region (Fig. 2). Moderate labeling
was also apparent in the bed nucleus of stria terminalis
(Figs. 2B and 4A) and moderate levels in the islands of
Calleja (Figs. 1A, 2 and 4A). The globus pallidus
showed hybridization signals close to background levels
(Figs. 3B and 4A). Relatively weak expression levels of
the D2 mRNA were detected in the claustrum (Figs. 2,
3B and C and 4A).
The hippocampus expressed weak to moderate levels
of the D2 mRNA (Figs. 1A, 2, 3B – D and 4B). The
highest levels were observed in the dentate gyrus. The
uncal gyrus also showed moderate D2 mRNA expres-
sion. No signal above background was, however, de-
tected in the subiculum. In the amygdaloid complex,
the basal and lateral amygdala nuclear group showed
weak mRNA expression levels, but no detectable signal
was evident in the periamygdala cortex, central, or
medial nuclei.
Diencephalon: a heterogenous expression of the D2
mRNA was found in the thalamus (Figs. 3AC and 4B).
The parafasicular and the paraventricular nuclei
showed very high expression levels as well as the
paratenial and paracentral nuclei. Most other nuclei of
the dorsal thalamus showed homogenous low levels of
expression. Moderate to high levels were also evident in
the medial and lateral geniculate bodies (Figs. 3C and
4B) and weak levels were detected in the subthalamic
nucleus. The pineal gland showed very high D2 mRNA
expression (Fig. 3B). Several hypothalamic areas ex-
pressed the D2 mRNA (Fig. 3C and D). At the hypo-
thalamic levels examined, the highest signals were
detected in the premammillary body as well as in the
posterior and lateral nuclei. No positive hybridization
signals were found in the medial mammillary nucleus,
ventromedial, or preoptic hypothalamic areas.
Mes- and metencephalon: a strong hybridization sig-
nal was evident throughout the mes- and meten-
cephalon (Figs. 1A, 2, 3D and E and 4B). The
substantia nigra pars compacta showed one of the
highest levels of the D2 mRNA expression in the brain.
132 Y.L. Hurd et al. / Journal of Chemical Neuroanatomy 22 (2001) 127–137

The mRNA expression pattern was not characterized in
the ventral tegmental area since the midline area was
damaged during dissection of the brain hemispheres.
Low to high expression levels were also detected in the
red nucleus, pontine nuclei, inferior colliculus, and me-
dial lemniscus. Hybridization signals above background
levels were detected in the cerebellar cortex, however,
the specificity of at least part of this signal is questioned
since the levels observed using the antisense probe in
one subject was lower than that obtained using the
sense probe.
4. Discussion
In the present study, whole hemisphere cryosections
of human brains were used to map D1 and D2 do-
pamine mRNAs throughout the brain in a limited
number of subjects. The gene expression levels for the
D1 and D2 dopamine receptors as examined in this
study cannot be directly compared since different hy-
bridization conditions were used to detect the two
mRNAs. However, the relative pattern of the anatomi-
cal distributions of the DA receptor mRNAs can be
commented upon since, in contrast to previous studies,
ISSH on the whole hemisphere sections allowed the
comparison of hybridization intensities in a large num-
ber of brain structures in the same section.
As previously reported by Meador-Woodruff et al.
(1994b, 1996, 1997) studying a limited number of brain
regions, the D2 mRNA was more widely expressed in
high to moderate levels than the D1. However, the D1
mRNA generally had a higher cortical presence as
compared to the D2. One important feature of the D1
mRNA expression was the high level in the cerebral
cortex and striatum, with no evidence for D1 receptor
expression in the diencephalon, the brainstem, or the
cerebellum. There are several implications of these find-
ings. The data indicate that neurons in the human
diencephalon, brainstem, or cerebellum have no major
role in mediating D1 receptor-regulated functions,
which is in marked contrast to the multitude of D2

Fig. 3. Distribution of D1 and D2 mRNA expression in whole brain horizontal cryosections of human post mortem brain images using an
antisense probe. The images show the D1 and D2 mRNA expression at five dorsoventral levels throughout the human brain. The levels are
approximately: (A) 62 mm; (B) 80 mm; (C) 85 mm; (D) 90 mm; and (E) 113 mm from vertex. For abbreviations, see list. Scale bar, 2 cm.
Y.L. Hurd et al. / Journal of Chemical Neuroanatomy 22 (2001) 127–137 133

Fig. 3. (Continued)
134 Y.L. Hurd et al. / Journal of Chemical Neuroanatomy 22 (2001) 127–137
receptor expressing neurons in discrete populations in
these brain areas.
Although the same laminar distribution has been
observed in the monkey for all the dopamine receptor
mRNAs (Lidow et al., 1998), the expression of the D1
and D2 mRNAs showed different cortical laminar pat-
terns in the human brain (Meador-Woodruff et al.,
1996). In this study, the D1 receptor mRNA was found
to be predominantly localized to layers II and V/VI.
Most cortical regions showed similar signals, but there
was a tendency for a more intense expression in the
supragranular layer in the frontal and insular cortices,
whereas the infragranular layers showed the most
prominant signal in the occipital cortex. Meador-
Woodruff et al. (1996) showed that the infragranular
expression of the D1 mRNA expression is in fact higher
in the visual cortex as compared to other cortical
regions. In addition to differences in laminar pattern, it
was also apparent in the present study that the total D1
mRNA expression varied between the different neocor-
tical regions. The signal was particularly strong in the
medial frontal (BA 22 and 25), medial orbital (BA 11,
14) as well as insular (BA 13– 16) cortices. Lower
expression levels were evident in the inferior and middle
frontal cortices. Studies in monkeys implicate an impor-
tant role for D1 dopamine receptor mediated signaling
in the prefrontal cortex in short term memory
(Sawaguchi and Goldman-Rakic, 1991; Castner et al.,
2000). Imaging studies also suggest marked functional
Fig. 4. Distribution of D1 and D2 mRNA mRNA expression in whole brain coronal cryosections of human post mortem brain images at a rostral
(A— approximately 75 mm from the frontal pole) and a caudal (B — approximately 105 mm from the frontal pole) level. For abbreviations, see
list. Scale bar, 2 cm.
specificities of anatomical compartments within the
frontal cortical regions (Mayberg et al., 1999; MacDon-
ald et al., 2000). The present results indicate that the
D1 receptor expression differs quantitatively between
subcompartments of the frontal neocortex. Both in
schizophrenia and affective disorders aberrant func-
tional activity in subregions of the frontal cortex have
been implicated (Mayberg et al., 1999). Aberrant levels
of D1 mRNA expressions could be a candidate for
mediating such dysfunctions. However, no significant
alterations of the D1 mRNA levels have been found in
some areas of the cerebral cortex of schizophrenic
subjects (Meador-Woodruff et al., 1997). Additional
studies of discrete subregions of the frontal cortex are
needed to validate these findings.
The striatal complex presented the highest levels of
the D1 mRNA expression in the human brain, a finding
consistent with a number of other studies (Zhou et al.,
1990; Mengod et al., 1991; Meador-Woodruff et al.,
1996, 1997; Lidow et al., 1998). In the striatum, the
pattern was heterogenous implicating compartmental-
ization of D1-regulated neuronal systems in these re-
gions as previously documented in primates
(Rappaport et al., 1993; Brené et al., 1995). The very
high expression of the D1 mRNA signal in the bed
nucleus of the stria terminalis and in the islands of
Calleja is another feature of importance to explore with
regard to functional and neuropsychiatric implications.
The lack of D1 signal in the globus pallidus implicates
 Y.L. Hurd et al. / Journal of Chemical Neuroanatomy 22 (2001) 127–137
that neurons in this region do not mediate D1 responses.
The same holds true for the hippocampal formation and
the thalamus where practically no positive D1 hybridiza-
tion signals were obtained. These findings make it
unlikely that cognitive functions regulated via these
regions involve D1 signaling. This is in marked contrast
to the neocortex were cognitive regulation in all proba-
bility involves the fairly extensive D1 receptor mRNA-
expressing neurons.
The dopamine D2 receptor mRNA expression in the
neocortex of the human brain was detectable but ex-
tremely low, consistent with previous studies (Gandel-
man et al., 1991; Meador-Woodruff et al., 1994b, 1997).
The laminar distribution pattern of the D2 hybridization
signal was more uniform over the cortex with three bands
of labeling in both superficial and deep layers. But also
among the neocortical regions there was evidence of
heterogeneity since relatively high cortical D2 hybridiza-
tion signals were detected in the most rostral part of the
temporal lobe and also in parts of the parietal and
occipital cortices. This is complementary to relatively
higher densities of D2 dopamine receptor binding sites
in the human temporal pole than in other regions of the
temporal cortex (Hall et al., 1996; S. Pauli, H. Hall, Y.
Hurd, C. Halldin, G. Sedvall, unpublished data).
Verifying previous studies (Gandelman et al., 1991;
Huntley et al., 1992; Meador-Woodruff et al., 1996),
many subcortical regions showed very high D2 mRNA
expression. The nucleus accumbens, caudate nucleus and
putamen showed high, fairly uniform D2 mRNA expres-
sion. Similar to the D1 mRNA expression pattern, the
D2 mRNA expression in the bed nucleus of the stria
terminalis and the islands of Calleja was particularly
strong. This points to a substantial potential for do-
pamine signaling via D1, D2, as well as D3 (Suzuki et
al., 1998; Gurevich and Joyce, 1999) receptor mRNA-ex-
pressing neurons in these brain regions. In contrast to the
virtual lack of the D1 signal in the hippocampus there
were moderate levels of the D2 mRNA expression in the
dentate and the uncal gyri of the hippocampus. The D2
mRNA-expressing neurons in the human dentate are
granular cells and those in the CA region are primarily
pyramidal in morphology (Meador-Woodruff et al.,
1994b; Gurevich et al., 1997). Also in the amygdaloid
complex there was significant D2 mRNA expression in
the basal and lateral amygdala which contrasts to the low
D1 expression in these nuclei. D2 mRNA expression has
previously been detected in the basal and lateral nuclei
of the human amygdala, but in contrast to the current
findings, D2 mRNA-expressing cells were also identified
in the central and medial nuclei (Gurevich and Joyce,
1999). The apparent discrepancy could be related to the
fact that cellular analysis is more sensitive to detecting
positively labeled cells.
The thalamus serves as a gateway for sensory and
motor processing to the cortex. A marked differential
135
expression of the D1 and D2 mRNAs was evident in the
thalamus. Whereas practically no D1 mRNA expressing
cells were present in any nucleus of this structure, the
parafascicular and paraventricular nuclei showed very
high expression levels of the D2 mRNA with lower levels
in most of the other thalamic nuclei. Gurevich and Joyce
(1999) also showed that there are a large number of the
D2 mRNA-expressing cells throughout the human thala-
mus. A growing body of evidence appears to indicate that
thalamic neurons are not involved in dopamine-regulated
signaling via D1 receptors. Instead, there seems to be a
rather strong role for D2 receptor mRNA-expressing
neurons in this regard.
The distribution of the D2 receptor mRNA expression
in regard to its receptor binding sites in the human brain
has been previously discussed (see e.g. Joyce and
Meador-Woodruff, 1997; Gurevich and Joyce, 1999). In
general, dissimilarities are evident in, e.g. the globus
pallidus where barely any mRNA expression signal
(current results) or some scattered D2 mRNA-positive
neurons (Gurevich and Joyce, 1999) is detected, but the
receptor binding (in particular the external segment) is
dense (Joyce et al., 1991; Hall et al., 1994). These binding
sites most likely arise from striatopallidal projection
neurons, which have high D2 mRNA expression. Both
D2 binding (Joyce et al., 1991; Hall et al., 1994) and
mRNA expression (current results; Hurd et al., 1994;
Meador-Woodruff et al., 1994a) is present in the substan-
tia nigra pars compacta. D1 receptor binding is also
evident in the human substantia nigra pars compacta and
pars reticulata (Besson et al., 1988; Thibaut et al., 1990;
Hall et al., 1994), but there is no expression of the D1
mRNA (current results; Mengod et al., 1991). Overall,
the findings in the substantia nigra pars compacta are
consistent with the D2, and not D1, as a dopamine
autoreceptor. Other notable discrepancies in regard to
the D1 receptor include the presence of the D1 binding
sites in the globus pallidus (primarily internal),
hippocampus, and cerebellar cortex (Cortes et al., 1989;
Mengod et al., 1991; Hall et al., 1994), regions that had
an apparent lack of the D1 mRNA expression. An
absence of the D1 mRNA expression is indicative of a
presynaptic localization of the D1 binding sites in these
structures. Current studies are underway to evaluate
more specifically the correlation between DA receptor
binding and gene expression in discrete subregions of the
human brain.
In summary, the present results clearly indicate
that D1 and D2 dopamine receptor mRNAs are differ-
entially expressed in neuronal populations in various
regions of the human brain. Only the striatum showed
high levels of both transcripts. In most neocortical
regions the D1 expression dominated, with a low D2
expression. In the thalamus, hippocampus, and brain-
stem, major cell populations expressed the D2 mRNA
whereas there is little evidence for D1 mRNA-express-
136 Y.L. Hurd et al. / Journal of Chemical Neuroanatomy 22 (2001) 127–137
ing cells in any of these regions. These findings have
several important implications. From a functional
standpoint the type of dopamine-regulated signaling
from neurons in a specific brain region will be deter-
mined by the relative presence of D1 and/or D2 mR-
NAs-expressing cells. This implicates that functional
responses from a certain brain region may vary due to
the relative presence of neurons expressing the genes for
the two-receptor subtypes. This will also have impor-
tant pharmacological implications. Neuronal circuits
via the thalamus and cerebellum have recently been
implicated in the pathophysiology of schizophrenia
(Andreasen, 1999). Since these relay regions seem to
contain D2 but not D1 mRNA-expressing cells it is
likely that signaling from these regions can be selec-
tively affected by D2 but not by D1 active drugs.
Acknowledgements
This study was supported by grants from the Swedish
Medical Research Council (MRF) 03560, 11252, the
National Institutes of Health (DA08914 and
MH44814), The Wallenberg Foundation, The Sunrise
Medical Research Charitable Trust, and The Human
Brain Informatics Center (Hubin). We thank Mrs Bar-
bro Berthelsson and Ms Pia Eriksson for skilful techni-
cal assistance and image analysis preparation.
References
Andreasen, N.C., 1999. A unitary model of Schizophrenia. Arch.
Gen. Psychiatry 56, 781 –787.
Berke, J.D., Hyman, S.E., 2000. Addiction, dopamine, and the molec-
ular mechanisms of memory. Neuron 25, 515 – 532.
Besson, M.J., Graybiel, A.M., Nastuk, M.A., 1988. [3H]SCH 23390
binding to D1 dopamine receptors in the basal ganglia of the cat
and primate: delineation of striosomal compartments and pallidal
and nigral subdivisions. Neuroscience 26, 101 –119.
Brené, S., Hall, H., Lindefors, N., Karlsson, P., Sedvall, G., 1995.
Distribution of messenger RNAs for D1 dopamine receptors and
DARPP-32 in striatum and cerebral cortex of the cynomolgus
monkey: relationship to D1 dopamine receptors. Neuroscience 67,
37 – 48.
Brené, S., Lindefors, N., Kopp, J., Hall, H., Taubes, T., Ehrlich, M.,
Sedvall, G., Greengard, P., Persson, H., 1994. Expression of
mRNAs encoding ARPP-16/19, ARPP-21 and DARPP-32 in
human brain tissue. J. Neurosci. 14, 985 –998.
Castner, S.A., Williams, G.V., Goldman-Rakic, P.S., 2000. Reversal
of antipsychotic-induced working memory deficits by short-term
dopamine D1 receptor stimulation. Science 287, 2020 –2022.
Cortes, R., Gueye, B., Pazos, A., Probst, A., Palacios, J.M., 1989.
Dopamine receptors in human brain: autoradiographic distribu-
tion of D1 sites. Neuroscience 28, 263 –273.
Duvernoy, H., 1991. The Human Brain: Surface, Three-Dimensional
Sectional Anatomy and MRI. Springer, Vienna.
Fibiger, H.C., 1995. Neurobiology of depression: focus on dopamine.
Adv. Biochem. Psychopharmacol. 49, 1 –17.
Gandelman, K., Harmon, S., Todd, R., O’Malley, K., 1991. Analysis
of the structure and expression of the human dopamine D2a
receptor gene. J. Neurochem. 56, 1024 – 1029.
Goldman-Rakic, P.S., 1998. The cortical dopamine system: role in
memory and cognition. Adv. Pharmacol. 42, 707 – 711.
Grandy, D.K., Marchionni, M.A., Makam, H., Stofkl, R.E., Alfano,
M., Frothingham, L., Fischer, J.B., Burke-Howie, K.J., Bunzow,
J.B., Server, A.C., Civelli, O., 1989. Cloning of the cDNA and
gene for a human D2 dopamine receptor. Proc. Natl. Acad. Sci.
USA 86, 9762 – 9766.
Graybiel, A.M., 1997. The basal ganglia and cognitive pattern gener-
ators. Schizophr. Bull. 23, 459 – 469.
Gurevich, E.V., Joyce, J.N., 1999. Distribution of dopamine D3
receptor expressing neurons in the human forebrain: comparison
with D2 receptor expressing neurons. Neuropsychopharmacology
20, 60 –80.
Gurevich, E.V., Kordower, J., Joyce, J.N., 1997. Dopamine D2
receptor mRNA is expressed in maturing neurons of the human
hippocampal and subicular fields. Neuroreport 8, 3605 – 3610.
Hall, H., Sedvall, G., Magnusson, O., Kopp, J., Halldin, C., Farde,
L., 1994. Distribution of dopamine-D1 and dopamine-D2 recep-
tors and dopamine and its metabolites in the human brain.
Neuropsychopharmacology 11, 245 – 256.
Hall, H., Farde, L., Halldin, C., Hurd, Y.L., Pauli, S., Sedvall, G.,
1996. Autoradiographic localization of extrastriatal D2-dopamine
receptors in the human brain using [125I]epidepride. Synapse 23,
115 – 123.
Huntley, G.W., Morrison, J.H., Prikhozhan, A., Sealfon, S.C., 1992.
Localization of multiple dopamine receptor subtype mRNAs in
human and monkey motor cortex and striatum. Mol. Brain Res.
15, 181 – 188.
Hurd, Y.L., 1996. Differential messenger RNA expression of pro-
dynorphin and proenkephalin in the human brain. Neuroscience
72, 767 – 783.
Hurd, Y.L., Keller, E., Sotonyi, P., Sedvall, G., 1999. Prepro-
tachykinin A mRNA expression in the human and monkey brain:
an in situ hybridization study. J. Comp. Neurol. 11, 56 –72.
Hurd, Y.L., Pristupa, Z.B., Herman, M.M., Niznik, H.B., Kleinman,
J.E., 1994. The dopamine transporter and dopamine D2 receptor
mRNAs are differentially expressed in limbic- and motor-related
subpopulations of human mesencephalic neurons. Neuroscience
63, 357 – 362.
Jaber, M., Robinson, S.W., Missale, C., Caron, M.G., 1996. Do-
pamine receptors and brain function. Neuropharmacology 35,
1503 – 1519.
Joyce, J.N., Meador-Woodruff, J.H., 1997. Linking the family of D2
receptors to neuronal circuits in human brain: insights into
schizophrenia. Neuropsychopharmacology 16, 375 – 384.
Joyce, J.N., Janowsky, A., Neve, K.A., 1991. Characterization and
distribution of [125I]epidepride binding to dopamine D2 receptors
in basal ganglia and cortex of human brain. J. Pharmacol. Exp.
Ther. 257, 1253 – 1263.
Kessler, R.M., Whetsell, W.O., Ansari, M.S., Votaw, J.R., De Paulis,
T., Clanton, J.A., Schmidt, D.E., Mason, N.S., Manning, R.G.,
1993. Identification of extrastriatal dopamine D2 receptors in post
mortem human brain with [125I]epidepride. Brain Res. 609, 237 –
243.
Knable, M.B., Weinberger, D.R., 1997. Dopamine, the prefrontal
cortex and schizophrenia. J. Psychopharmacol. 11, 123 –131.
Laruelle, M., Abi-Dargham, A., van Dyck, C.H., Gil, R., D’Souza,
C.D., Erdos, J., McCance, E., Rosenblatt, W., Fingado, C.,
Zoghbi, S.S., Baldwin, R.M., Seibyl, J.P., Krystal, J.H., Charney,
D.S., Innis, R.B., 1996. Single photon emission computerized
tomography imaging of amphetamine-induced dopamine release
in drug-free schizophrenic subjects. Proc. Natl. Acad. Sci. USA
93, 9235 –9240.
 Y.L. Hurd et al. / Journal of Chemical Neuroanatomy 22 (2001) 127–137
Lidow, M.S., Wang, F., Cao, Y., Goldman-Rakic, P.S., 1998. Layer
V neurons bear the majority of mRNAs encoding the five distinct
dopamine receptor subtypes in the primate prefrontal cortex.
Synapse 28, 10 – 20.
MacDonald, A.W. 3rd., Cohen, J.D., Stenger, V.A., Carter, C.S.,
2000. Dissociating the role of the dorsolateral prefrontal and
anterior cingulate cortex in cognitive control. Science 288, 1835 –
1838.
Mai, J.K., Assheuer, J., Paxinos, G., 1997. Atlas of the Human Brain.
Academic Press, San Diego, CA.
Mayberg, H.S., Liotti, M., Brannan, S.K., McGinnis, S., Mahurin,
R.K., Jerabek, P.A., Silva, J.A., Tekell, J.L., Martin, C.C., Lan-
caster, J.L., Fox, P.T., 1999. Reciprocal limbic-cortical function
and negative mood: converging PET findings in depression and
normal sadness. Am. J. Psychiatry 156, 675 – 682.
Meador-Woodruff, J.H., Damask, S.P., Watson, S.J., 1994a. Differen-
tial expression of autoreceptors in the ascending dopamine system
of the human brain. Proc. Natl. Acad. Sci. USA 91, 8297 – 8301.
Meador-Woodruff, J.H., Grandy, D.K., Van Tol, H.H.M., Damask,
S.P., Little, K.Y., Civelli, O., Watson, S.J., 1994b. Dopamine
receptor gene expression in the human medial temporal lobe.
Neuropsychopharmacology 10, 239 – 248.
Meador-Woodruff, J.H., Damask, S.P., Wang, J., Haroutunian, V.,
Davis, K.L., Watson, S.J., 1996. Dopamine receptor mRNA
expression in human striatum and neocortex. Neuropsychophar-
macology 15, 17 – 29.
Meador-Woodruff, J.H., Haroutunian, V., Powchik, P., Davidson,
M., Davis, K.L., Watson, S.J., 1997. Dopamine receptor transcript
expression in striatum and prefrontal and occipital cortex. Focal
abnormalities in orbitofrontal cortex in schizophrenia. Arch. Gen.
Psychiatry 54, 1089 – 1095.
Mengod, G., Vilaro, M.T., Niznik, H.B., Sunahara, R.K., Seeman, P.,
O%Dowd, B.F., Palacios, J.M., 1991. Visualization of a dopamine
137
D1 receptor mRNA in human and rat brain. Mol. Brain Res. 10,
185 – 191.
Parent, A., 1996. Carpenter’s Human Neuroanatomy, 9th ed.
Williams & Wilkins, Baltimore, MD.
Paxinos, G., 1990. The Human Nervous System. Academic Press, San
Diego, NY.
Rappaport, M.S., Sealfon, S.C., Prikhozhan, A., Huntley, G.W.,
Morrison, J.H., 1993. Heterogenous distribution of D1, D2, and
D5 receptor mRNAs in monkey striatum. Brain Res. 616, 242 –
250.
Sawaguchi, T., Goldman-Rakic, P.S., 1991. D1 dopamine receptors in
prefrontal cortex: involvement in working memory. Science 251,
947 – 950.
Smiley, J.F., Levey, A.I., Ciliax, B.J., Goldman-Rakic, P.S., 1994. D1
dopamine receptor immunoreactivity in human and monkey cere-
bral cortex: predominant and extrasynaptic localization in den-
dritic spines. Proc. Natl. Acad. Sci. USA 91, 5720 – 5724.
Sokoloff, P., Schwartz, J.-C., 1995. Novel dopamine receptors half a
decade later. Trends Pharmacol. Sci. 16, 270 – 275.
Suzuki, M., Hurd, Y.L., Sokoloff, P., Schwartz, J.-C., Sedvall, G.,
1998. Anatomical distribution of D3 dopamine receptor is widely
expressed in the human brain. Brain Res. 779, 58 – 74.
Tarazi, F.I., Florijn, W.J., Creese, I., 1997. Differential regulation of
dopamine receptors after chronic typical and atypical antipsy-
chotic drug treatment. Neuroscience 78, 985 – 996.
Thibaut, F., Hirsch, E.C., Raisman, R., Javoy-Agid, F., Agid, Y.,
1990. Microtopography of D1 dopaminergic binding sites in the
human substantia nigra: an autoradiographic study. Neuroscience
37, 387 – 398.
Zhou, Q.-Y., Grandy, D.K., Thambi, L., Kushner, J.A., Van Tol,
H.M., Cone, R., Pribnow, D., Salon, J., Bunzow, J.R., Civelli, O.,
1990. Cloning and expression of human and rat D1 dopamine
receptor. Nature 347, 76 – 79.