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Post Mortem Brain Studies
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Characterization of receptor and transporter
changes in schizophrenia as studied using post mortem autoradiography.
Summary
Schizophrenia is characterized by abnormal
behavior, which not always can be correlated to microscopical changes
in the brain. The main aim of this project is to characterize the
distribution of several receptor subtypes connected to schizophrenia,
in the control and pathologic brain using whole hemisphere autoradiography.
The dopamine (D1-, D2- and D3-) and serotonin
(5-HT1A, 5-HT2A) receptor subtypes are the main targets of this
project. In addition the GABA-A/benzodiazepine and the glutamate
receptors as well as the dopamine and serotonin transporters are
studied. The quantification of densities is performed on several
brain regions for all receptors and transporters studied by the
use of calibrated scales. Comparison between densities between the
normal and diseased brain will be performed on a high number of
brains. Cluster analyses and data mining analyses will then be performed
to obtain an overall comparison of differences between subjects
with and without brain diseases.
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Figure 1
(for larger image, click on it)
Dopamine transporter visualization
in human brain using [125I]PE21. To the left, horizontal section,
and to the right, coronal section.
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Project Aims
Of the more than hundred neurotransmitter
receptor subtypes known, we have focussed our studies to the dopamine
and serotonin receptor systems. The receptor subtypes of these systems
have been considered to be involved in several aspects of higher
mental functions, and are therefore the main focus of the present
study. The transporters (dopamine, serotonin and norepinephrine)
are furthermore characterized in brains from alcoholics.
Of crucial importance is the collection of
brain tissue (whole hemispheres, preferably both hemispheres) from
patients with psychiatric and neurological diseases as well as control
hemispheres from subjects without brain related diseases. During
the previous studies the brain bank developed mainly in the nineteeneighties
has been used. However, new material is now required, especially
with regard to the pathological brains.
The cryosectioning technique has been developed
and optimized at the Department. In connection to the cryosectioning
each section is photographed using a video camera and a large number
of sections are stained with cresyl violet using the Nissl staining
technique. These two visualization techniques are used for as anatomical
correlates to the autoradiographical technique.
Methodology
In previous studies (see below) we have found
suitable ligands for the D1-, D2- and D3-dopamine receptor, several
serotonergic receptor subtypes (5-HT1A, 5-HT1B, 5-HT2A and 5-HT4
receptors), adenosine receptor subtypes, the GABA-A/benzodiazepine
receptor and the dopamine and serotonin transporters. These receptors,
and also receptors for which radioligands are under optimization
for whole hemisphere autoradiography (like the glutamate receptors),
are used for the extensive quantitative characterization of the
brain distribution in control and pathological brain.
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Figure 2
(for larger image, click on it)
5-HT2A receptors in post mortem human
brain, labeled with [3H]MDL 100907. Horizontal section.
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Normally a number (normally around 10 levels)
of levels of the hemisphere, covering every region of interest,
and radioligand autoradiography is perform on these levels. A more
extensive, and labor-intensive method is to cryosection the entire
hemisphere and perform autoradiography every ten sections per radioligand.
This latter technique results in a receptor mapping that can be
transformed into a 3D image of the receptor distribution (LINK coming
to 3D project).
The quantification of densities is performed
on several brain regions for all receptors and transporters studied
by the use of calibrated scales commercially available.
Results
The D1-dopamine receptors have been of interest
in the discussions on neuroleptics, as the effects of this receptor
is counteracting many of the effects of the D2-dopamine receptor
in animal studies. However, no D1-dopamine receptor antagonist or
agonist has proven useful in any aspects of treatment of the psychotic
patient. The characterization of the D1-dopamine receptors have
been done mainly with [3H]SCH 23390 and with [3H]NNC 112. The D1-dopamine
receptor is found in the basal ganglia of the human brain, but to
a certain extent (the density is approximately 30 % of that in the
caudate and putamen) also in the neocortex. The cellular localization
of this receptor indicates that it can have a role in Huntington’s
disease rather than in schizophrenia. However, due to its opposing
behavioral effects to those of the D2-dopamine receptor, we have
found it important to include in our studies.
The D2-dopamine receptor is the main target
of all neuroleptic drugs, and is therefore also one or the main
targets of our distribution studies. The density of this receptor
is highest in the caudate and putamen, but is also found in low
amounts (a few percent of the caudate and putamen) in the temporal
cortex, in some thalamic nuclei, in the pallidum and in some other
brain regions. In spite of the low densities of the D2-dopamine
receptor it has been suggested that the therapeutic efficacy of
the neuroleptics is due to the blockade in these extra striatal
regions, whereas the motoric side effects are induced by receptor
blockade in the striatum. Using the new very high affinity (Ki is
approximately 20 pM) D2-dopamine (and D3-dopamine) receptor radioligands
[125I]epidepride and [125I]NCQ 298, we have shown that it is possible
to quantitatively study the distribution of the extra-striatal D2-dopamine
receptors using radioligand autoradiography. [125I]Epidepride autoradiograms
have also been used in 3D visualization of D2-dopamine receptor
distribution in the human brain distribution (LINK coming to 3D
project).
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Figure 3
(for larger image, click on it)
D2/D3- dopamine receptors in post mortem
human brain. Whole hemisphere autoradiography using [125I]epidepride.
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Also the D3-dopamine receptor has been suggested
to be of importance for the therapeutic efficacy of the neuroleptic
drugs, mainly due to the localization in limbic regions. The distribution
of D3-dopamine receptor as shown using the selective radioligand
[3H]PD128907 is mainly to the nucleus accumbens and Islands of Calleja.
The studies on the 5-HT1A receptor was performed
with a new selective antagonist, [3H]WAY-100635, has been used for
the characterization of the distribution of this receptor subtype
in the human brain in vitro using whole hemisphere autoradiography
and in vivo using PET. Also for the 5-HT2A receptor a new selective
radioligand has been used M100907. This has been tritiated at the
Department, and a manuscript on the receptor distribution has been
described. In a study performed by a Belgian visiting scientist
during 1998, the distribution of the 5-HT4 receptor was described
in the human brain. A second study, using a more favourable radioligand
for this receptor has also been performed in our group.
As also other serotonin receptor subtypes
are involved in schizophrenia, we have in collaboration with foreign
scientists developed systems for the characterization of these receptor
subtypes in whole human hemispheres.
The distribution of GABA-A/benzodiazepine
receptors in the human brain is very extensive. In one whole hemisphere
autoradiographic study we have used the radioligands [3H]flumazenil
and [3H]Ro15-4513, where the latter was used for a specific visualization
of a6-subunit containing receptors in the granule cells in the cerebellum.
Adenosine is an endogenous neuromodulator
in the human brain and is involved in various conditions like ischemia,
anxiety, sleep disorders etc. There are two main subtypes, A1 and
A2, both studied in our lab in collaboration with Prof. B. Fredholm
and Dr. P. Svenningsson, Karolinska Institutet.
For the studies of the dopamine transporter
a new tropane analogue, PE2I, with much higher selectivity has been
synthesized by Prof. D. Guilloteau in France and studied in our
project This has also been used successfully in whole hemisphere
autoradiography (see image above). The French group has also synthesized
a new selective serotonin transporter radioligand, abbreviated MADAM,
which has been used in whole hemisphere autoradiography.
In collaboration with a group in Kuopio,
Finland (Dr. Erkki Tupala) several studies on the dopamine systems
in alcoholism have been published. In short, these studies have
shown that the D2 receptors and DAT system are impaired in the limbic
regions of the human brain.
More recently, studies on the functional
activity of the dopamine receptors have been performed using a [35S]GTP-gammaS,
an isotope that interacts with the G-protein. A review on this methodology
has been published.
Project leader: Håkan
Hall
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