Receptorarchitecture and Neural Systems
Karl Zilles
Institute of Neuroscience and Medicine INM-1
Research Centre Jülich
and
University Hospital of Psychiatry, Psychotherapy and
Psychosomatics
RWTH Universität Aachen
Receptors are protein complexes in the cell
membrane, to which transmitters selectively bind.
ionotropic receptors have an integrated ion channel
and regulate ion fluxes between extra- and intracellular space
metabotropic receptors are linked to intracellular
second messenger systems, and control metabolic
pathways, ion channels, gene activity, etc.
What does receptor mapping provide?
 It provides insight into the molecular basis of the resting state condition.
 Most of the brain‘s energy consumption is used for maintaining ion
channel functions during resting state. Only a minority of the energy is
required during stimulation conditions.
 Receptor mapping overcomes limitations of unimodal brain mapping
approaches
 It provides brain maps based on molecules which are key elements of
signal transmission within and between neural systems
 Areas within a given functional system have similar multireceptor
expression patterns. Different functional systems differ by those
expression patterns
Example: Ionotropic Transmitter Receptors
presynaptic
membrane
postsynaptic
membrane
postsynaptic
ionotropic
receptor
R
ion channel
axonal bouton
transmitter
presynaptic
ionotropic
receptor
postsynaptic
Spine
AMPA
PSD
NMDA NR1
presynaptic
Dendrite
Postsynaptic density PSD
Dendrite
NMDA NR1
AMPA
Joachim Lübke, Research Center Jülich, Germany (unpublished)
NMDA and AMPA are the most abundant
glutamate receptor types in the human
cerebral cortex.
Method: Cryo-Electron-Microscopy and
Immunocytochemistry with
specific antibodies labeled with
gold nanoparticles of different size
(for AMPA 9 nm, for NMDA 12 nm)
5-HT1A, 5-HT2
GABAB
AMPA, NMDA, kainate
Adenosin A1
D1, D2, D4
α1, α2
AMPA, NMDA, kainate
AMPA, NMDA, kainate
GABAA, bz.binding site
nicotinic, M1, M2, M3
GABAA, bz.binding site
D1, D2, D4
GABAB
5-HT1A, 5-HT2
GABAA, bz.binding site
Glutamate
GABA
Acetylcholine
α1, α2
nicotinic, M1, M2, M3
GABAB
AMPA, NMDA, kainate
GABAA, bz.binding site
GABAA, bz.binding site
Dopamine
Noradrenaline
Serotonin
How to map receptor distributions:
• in vivo receptor PET
• in vitro quantitative receptor autoradiography
• immunohistochemistry of receptor proteins and
subunits
• transcriptomics
Quantitative in vitro Receptor Autoradiography: Method (1)
Gyrus
praecentralis
Sulcus
centralis
slab 2
HG05/00
slab 2
native
slab 2
frozen
at -70° C
Quantitative in vitro Receptor Autoradiography: Method (2)
Cryostate serial sections (10 – 20 µm) – mounting on glass slides
 Incubation of alternating serial sections with tritium-labeled ligands
 Labeling of different receptors of all classical transmitter systems
(Glutamate, GABA, Acetylcholine, Dopamine, Serotonin, Noradrenaline)
 Exposition against β-sensitive film together with radioactive scales
 Digitization, quantification and colour coding of the film
 Identification of cytoarchitectonic areas by comparison with neighbouring cell-body
stained sections
Examined receptor binding sites
Neurotransmitter
Glutamate
GABA
Acetylcholine
Receptor
AMPA
Kainate
NMDA
GABAA
GABAB
BZ
M1
M2
M3
Noradrenaline
Histological staining:
Cell bodies
Myelin
Serotonin
Dopamine
N
α1
α2
5-HT1A
5-HT2
D1
D2
[3H] Ligand
AMPA
kainate
MK-801
muscimol,
SR 95531
CGP 54626
Flumazenil
pirenzepine
oxotremorine-M,
AF-DX 384
4-DAMP
epibatidine
prazosin
UK 14,304
RX-821002
8-OH-DPAT
ketanserin
SCH 23390
Fallypride or
Raclopride
V2
V1
V1
V2
Sulcus
calcarinus
vertical
vertical
meridian
meridian
V1
V1
V2
Gennari‘s stripe
Myelin
V2
vertical
vertical
meridian
meridian
GABAA Receptor
Zilles, K., Palomero-Gallagher, N. , Schleicher, A.: Transmitter receptors and functional anatomy
of the cerebral cortex. J. Anat. 205: 417-432 (2004)
Entorhinal-Hippocampal
System
Why are architectonic studies essential for understanding the
organization of neural systems?
INPUT:
fi
CA3
trisynaptic
glutamatergic system:
CA2
Sub
2
CA4
Dent
Parsub
3
1
CA1
1 perforant path
2 mossy fibers
3 Schaffer collaterals
35
temporal and perirhinal
input to subiculum
n.d.
lateral
36
medial
ventral
general neocortical input
Ent
n.d.
dorsal
Prsub
cingulate, superior temporal,
dorsolateral prefrontal, and
IPL input to presubiculum
OUTPUT:
from the subiculum and CA1 to
temporal, parahippocampal, perirhinal,
cingulate, prefrontal, and orbitofrontal
areas
802
633
CA2
Schaffer collateral
CA2
CA3
fimbria
mossy fibres
464
CA3
295
fmol/mg protein
AMPA
mol
CA1
mol
CA1
sub
perforant path
1061
486
632
417
fmol/mg protein
NMDA
629
Glutamatergic terminals of
the perforant path and the
Schaffer collaterals
493
357
221
fmol/mg protein
Kainate
mol
CA1
Glutamatergic terminals of the perforant path
and the Schaffer collaterals
Glutamatergic terminals of the mossy fibers
Primary Sensory Cortices
96
208
320
432
fmol/mg protein
primary
somatosensory 3b
motor
cingulate
3a
insular
parietal
primary
auditory
unimodal
auditory
Toga AW, Thompson PM, Mori S, Amunts K, Zilles K (2006)
Towards multimodal atlases of the human brain.
Nature Rev. Neurosci. 7: 952-966
Zilles K, Amunts K (2009)
Receptor mapping: Architecture of the human cerebral cortex.
Current Opinion Neurol. 22: 331–339
multimodal
temporal
association
pre/parasubiculum
entorhinal
cholinergic muscarinic M2 receptor
Cholinergic muscarinic M2 receptor
low density
high density
sc
sc
motor cortex
motor
cortex
primary
somatosensory
cortex
primary
somatosensory
cortex
primary
auditory
cortex
primary
auditory
cortex
Human
Brain
Macaque
Brain
154
379
604
829
fmol/mg protein 668
sensorimotor
1084
1499
1915
sensorimotor
cingulate
cingulate
insular
insular
primary
auditory
unimodal
auditory
unimodal
auditory
pre/parasubiculum
multimodal
temporal
association
entorhinal
primary
auditory
pre/parasubiculum
multimodal
temporal
association
AMPA receptor
[3H] AMPA
GLUTAMATE SYSTEM
entorhinal
Kainate receptor
[3H] kainate
Receptor Fingerprints
Multimodal visualization of receptor organization
in human cerebral cortex
Low
denisty
High
density
AMPA
Kainate
NMDA
M1
M2
Glutamate
α1
α2
Noradrenaline
M3
nicotinic
Acetylcholine
GABAA
GABA
5-HT1A
5-HT2
Serotonin
D1
D2
Dopamine
Zilles, K., Schleicher, A., Palomero-Gallagher, N., Amunts, K.:
Quantitative analysis of cyto- and receptorarchitecture of the human brain, pp. 573-602.
In: Brain Mapping: The Methods, 2nd edition (A.W. Toga and J.C. Mazziotta, eds.). Academic Press (2002)
A receptor fingerprint is…
AMPA
D1
3000
kainate
2500
5-HT2
NMDA
2000
1500
1000
5-HT1A
GABAA
500
0
α2
GABAB
α1
BZ
nACh
M1
M3
M2
caudate
putamen
area 4
„Receptor Fingerprints“
AMPA
AMPA
3000
D1
5-HT2
2000
5-HT1A
1000
kainate
NMDA
GABAA
0
α2
GABAB
α1
BZ
N
M1
M3
M2
primary motor cortex
3000
D1
5-HT2
2000
5-HT1A
1000
kainate
NMDA
GABAA
0
α2
GABAB
α1
BZ
M1
N
M3
M2
Hippocampus
CA1-3
What receptors tell us about laminar
segregation and input-output relations
in the cerebral cortex
CYTOARCHITECTURE
molecular layer
I
outer granular layer
II
outer pyramidal layer III
inner granular layer
IV
inner pyramidal layer
V
polymorphic layer
VI
CONNECTIVITY
thalamocortical
input
corticocortical
input
cortico-cortical,
-striatal,
-thalamic,
-bulbar, and
-spinal
output
SYNAPTIC DENSITY
Receptor Fingerprints of the primary visual cortex: layer specificity
L. I
D1
5-HT2
AMPA
3000
kainate
NMDA
2000
1000
5-HT1A
GABAA
0
α2
GABAB
α1
BZ
N
D1
5-HT2
AMPA
3000
GABAA
GABAB
α1
BZ
N
M1
M3
M2
GABAA
0
α2
GABAB
α1
L. VI
NMDA
0
NMDA
D1
5-HT2
M2
NMDA
1000
5-HT1A
GABAA
0
α2
GABAB
α1
BZ
N
M1
M3
M2
NMDA
2000
1000
5-HT1A
GABAA
0
α2
GABAB
BZ
M1
M3
kainate
2000
5-HT2
kainate
N
M1
AMPA
3000
D1
AMPA
3000
α1
BZ
N
kainate
L. IV
kainate
1000
5-HT1A
M3
1000
α2
AMPA
3000
2000
M2
2000
5-HT1A
D1
5-HT2
M1
M3
L. V
Ll. II-III
M2
Layer IV: Thalamo-cortical Input
modality-specific balance between receptors
primary somatosensory (touch)
AMPA
3000
D1
5-HT2
N
GABAA
0
α2
D1
NMDA
1000
5-HT1A
GABAB
α1
BZ
N
M1
M3
M2
M1
M2
parietal association
(visually guided grasping)
kainate
2000
GABAB
BZ
M3
prefrontal association
(working memory)
5-HT2
0
M2
AMPA
3000
GABAA
N
M1
M3
NMDA
α1
4
BZ
kainate
1000
α2
GABAB
α1
AMPA
3000
2000
5-HT1A
GABAA
0
D1
5-HT2
NMDA
1000
α2
D1
kainate
2000
5-HT1A
primary visual
5-HT2
AMPA
3000
kainate
NMDA
2000
1000
5-HT1A
GABAA
0
α2
GABAB
α1
BZ
N
M1
M3
M2
Cluster analysis of the multi-receptor fingerprints of
BROCA‘s region, prefrontal and motor cortex
prefrontal
47
primary motor
cortical area
4
premotor
6r1
6v1
op9
4
op8
language
45
44 6v1
44
45
47
0.2
0.3
0.4
0.5
0.6
0.7
6r1
op8
op9
0.8
Amunts, K., Lenzen, M., Friederici, A.D., Schleicher, A., Morosan, P., Palomero-Gallagher, N., Zilles, K.:
Broca's region: Novel organizational principles and multiple receptor mapping. PLoS Biology 8(9): e1000489.
doi:10.1371/journal.pbio.1000489 (2010)
Whole brain hierarchical cluster analysis based on multi-receptor fingerprints
PFt
PFop
PFcm
PGa
PGp
PFm
PF
32'
v23
24
OP
a24'
6d
v1
FG2
FG1
v4v
Oc3v
3b
BA45
BA44
p24'
BA8
BA4
BA38
5l
7pc
5m
BA2
Te4
Te3
Te2
Te1
3a
BA1
30
29
d23
Oc3d
Oc4d
v2v
v2d
7ma
7la
31
7mp
7lp
CA
ent
subic
dg
BA36
BA20‘
25
p32
s32
10m
BA47
4
3
2
0
1
Euclidean distance
5
6
- Normalization: divide by mean
- Euclidean distances, Ward linkage
- Cophrenetic coefficient: 0.58
entorhinal anterior
inferior sensorimotor visual Broca motor somatoauditory retro- visual superior
-hippocampalcingulate- parietal
limbic
splenial
parietal
circuit
prefrontal multimodal
ventral
-sensory
perilimbic
dorsal stream
cortex
stream
domaine
proisocortex
Cluster analysis of the human cingulate cortex:
A multi-region model based on receptor fingerprints
4.5
4
visuospatial
attention
memory
3.5
emotion
3
2
ACC
1.5
motor
control
decision MCC
autonomic
control
2.5
aMCC
pMCC
limbic
memory
PCC
RSC
1
29l
29m
30
v23
d23
31
23c
23d
24dd
24dv
p24b'
p24a'
24c'v
24c'd
32'
a24b'
a24a'
33
25
32
24cd
24cv
0
24b
0.5
24a
Euclidean distance (Ward linkage)
Palomero-Gallagher, N., Vogt, B.A.,
Schleicher, A., Mayberg, H.S., Zilles, K. (2009)
Receptor architecture of human cingulate cortex:
Evaluation of the four-region neurobiological
model.
Human Brain Mapping 30: 2336-2355
Thanks to:
Institute of Neuroscience and
Medicine (INM-1),
Research Centre Jülich
Katrin Amunts
Svenja Caspers
Joachim Lübke
Hartmut Mohlberg
Nicola Palomero-Gallagher
Axel Schleicher