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DORSAL NUCLEUS RAPHÉ (DR)
Josh J. Royall, Lydia L. Ng and John A. Morris
Introduction
This report contains a gene expression summary of the dorsal nucleus raphé (DR), derived from the Allen Brain Atlas
(ABA) in-situ hybridization (ISH) mouse data set. The structure’s location and morphological characteristics in the
mouse brain are described using the Nissl data found in the Allen Reference Atlas. Using an established algorithm, the
expression values of the DR were compared to the values of the macro/parent-structure, in this case the midbrain, for
the purpose of extracting regionally specific gene expression data. The highest ranking ratios were then manually cu-
rated and verified. The 50 Select Genes were compiled for expression characterization. The experimental data for each
gene may be accessed via the links provided; complementary sagittal data may also be accessed using the ABA. Corre-
lation between gene expression in the DR and the rest of the brain, across all genes in the coronal dataset (~4300
genes), were derived computationally and are presented below. A gene ontology table (derived from DAVID Bioinfor-
matics Resources 2007) is also included, highlighting possible functions of these 50 Select Genes.
To read more about how the 50 Select Genes list is derived, please refer to the Fine Structure Annotation white paper.
Allen Reference Atlas Coronal Levels: 94-105
Allen Reference Atlas Sagittal Levels: 18-20
Shown below is a plate from the Allen Reference Atlas, depicting the dorsal nucleus raphé (level 100):
ANNOTATION REPORT
Nature Precedings : doi:10.1038/npre.2008.2054.1 : Posted 7 Jul 2008
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Description of Structure:
LOCATION and STRUCTURAL ANATOMY:
The hierarchical relationship within the brain is depicted below in the structure legend. The Allen Reference Atlas (based on Nissl-
stained sections scanned at 10X) was the primary resource for the following descriptions.
For additional information please refer to the Allen Reference Atlas white paper.
BrainInfo houses a search engine that allows searches for structure name aliases.
The dorsal nucleus raphé (DR) is a cell rich sub-region of the central midbrain. It is located
on the midline (raphé means “seam”), within the ventral aspect of the periaqueductal gray (PAG).
Ventrally, it is bounded by two white matter tracts, the superior cerebellar peduncle decussation
and the medial longitudinal fascicle (dscp, mlf), as well as the large, dark staining cells of the ocu-
lomotor nucleus (III). The caudal aspect is delineated by the less cell dense pontine central gray
(PCG) of the pons and lobule II (CENT2) of the cerebellum. The rostral border, with the more co-
lumnar appearance of Edinger-Wesphal nucleus, is difficult to perceive.
While broad and flat in the sagittal aspect, the DR maintains a tall, thin appearance coron-
ally, with a slightly widened dorsal region. The DR consists primarily of medium to large sized
cells, which exhibit dark Nissl staining that contrast readily with neighboring regions. Nissl and
ISH expression reveal cells of the ventral DR in a vertical orientation that is best observed in the
sagittal plane. Some internal cell density and staining intensity differences suggest dorsal and
ventral subdivisions (see coronal level 97 and sagittal level 20 below).
The appearance and location of the DR can be appreciated on the following two pages. Nissl-
stained sections and Allen Reference Atlas plates reveal the cytoarchitecture and extent of the
DR, and its location in relation to surrounding structures.
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In-Situ Hybridization Expression Patterns of 50 Select Genes:
This survey of 50 Select Genes revealed a modest range of expression patterns for Dorsal
Nucleus Raphé (DR). While the majority of genes show uniform expression throughout the DR, the
level of intensity varies from gene to gene. The density of cells expressing also varies, albeit to a
lesser degree. Several genes exhibit regional expression patterns that suggest dorsal and ventral
partitions. With the exception of Pdyn, all genes that display a regional expression pattern show en-
richment of the ventral aspect of the DR. Characteristics in the sagittal plane also indicate possible
caudal and rostral regionalization as well (e.g. Clcn5), although further corroboration is required.
Maob clearly demarcates the rostral border of the DR with the Edinger-Westphal nucleus.
To view heat map at brain-map.org, right click on an ISH image and select “Show Expression Analysis.”
The ISH data presented below presents the anatomical and cytoarchitectural characteristics of the DR in the context of actual gene
expression. In addition to presenting molecularly defined borders, ISH gene expression patterns also aid in phenotyping cell popula-
tions that otherwise can not be differentiated on purely morphological grounds. The 50 genes in this section were selected based on
a mathematical algorithm to identify gene expression patterns that allow selective identification of the DR. The gene expression pat-
terns were then verified manually. As such, these genes do not represent the only genes found in this structure, genes specific to this
structure, or genes expressing at the highest level within this structure.
The ISH protocol is described in the Data Production Processes white paper.
To read about heat map conversion, refer to the Informatics Data Processing white paper.
The expression data subsequently presented can be further explored, in coronal and sagittal planes, at brain-map.org.
ISH DATA The Allen Institute ISH images below were selected to highlight various expression patterns of the dorsal nucleus raphé
ISH
Slc22a3
Coronal:
A widespread expression pattern in the DR.
Heat map
Slc22a3
Coronal:
The DR shows high density and intensity expression based on a
heat map analysis of Slc22a3.
Dorsal Nucleus Raphé Cerebral Aqeduct
Dorsal Nucleus Raphé
Cellular density expression key Cellular intensity expression key
None No expression No color Very low intensity
Sparse Very few cells expressing Blue Low intensity
Scattered Less than 10% of cells expressing in scattered pattern Green Medium intensity
Medium 10-80% of cells expressing Yellow High intensity
High Greater than 80% of cells expressing Red Very high intensity
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ISH
Gchfr
Sagittal:
Widespread expression in the DR as seen in the medial sagittal
view.
Heat map
Gchfr
Sagittal:
Gchfr expression shows high density and intensity
characteristics.
ISH
Pcsk5
Coronal:
A ventral zone enrichment can be seen from gene Pcsk5, in the
coronal view.
ISH
Pcsk5
Sagittal:
Pcsk5 expressions appears to differentiate a rostral and caudal
zone of the DR in the sagittal aspect.
Dorsal Nucleus Raphé Pontine Central Gray
Dorsal Nucleus Raphé
Cerebral Aqueduct
Dorsal Nucleus Raphé
Dorsal Nucleus Raphé Cerebral Aqueduct
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Dorsal Nucleus Raphé Cerebral Aqueduct
Dorsal Nucleus Raphé
Dorsal Nucleus Raphé Periaqueductal Gray
Dorsal Nucleus Raphé Periaqueductal Gray
ISH
Pdyn
Coronal:
Pdyn expression shows enhanced dorsal expression as seen in
the coronal plane.
Heat map
Pdyn
Coronal:
Pdyn expression appears high density, very high intensity in the
dorsal aspect, and medium density, medium intensity in the
ventral aspect.
ISH
Peg10
Coronal:
A widespread expression pattern appears to diffuse into the
neighboring regions of midbrain.
Heat map
Peg10
Coronal:
Peg10 expression is presented as high density, very high inten-
sity in the DR, and the nearby ventral periaqueductal gray.
Nature Precedings : doi:10.1038/npre.2008.2054.1 : Posted 7 Jul 2008
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Number Gene Symbol Gene Name Expression Pattern
1 Maob monoamine oxidase B High density, very high intensity
2 Cpne7 copine VII High density, very high intensity
3 Cdyl2 chromodomain protein, Y chromosome-like 2 High density, high intensity
4 Gchfr GTP cyclohydrolase I feedback regulator High density, high intensity
5 C130021I20Rik Riken cDNA C130021I20 gene High density, high intensity
6 Ttyh3 tweety homolog 3 (Drosophila) High density, high intensity
7 Id4 inhibitor of DNA binding 4 High density, high intensity
8 Tph2 tryptophan hydroxylase 2 Medium density, very high intensity
9 Gch1 GTP cyclohydrolase 1 Medium density, very high intensity
10 Dnajc12 DnaJ (Hsp40) homolog, subfamily C, member 12 Medium density, very high intensity
11 Stard13 serologically defined colon cancer antigen 13 High density, medium intensity
12 Cntnap3 contactin associated protein-like 3 High density, medium intensity
13 Slc22a3
solute carrier family 22 (organic cation trans-
porter), member 3 Medium density, high intensity
14 Irs4 insulin receptor substrate 4 Medium density, high intensity
15 Cthrc1 collagen triple helix repeat containing 1 Medium density, high intensity
16 Glra2 glycine receptor, alpha 2 subunit Medium density, medium intensity
17 Zim1 zinc finger, imprinted 1 Medium density, medium intensity
18 Hdac6 histone deacetylase 6 Medium density, medium intensity
19 Idh1 isocitrate dehydrogenase 1 (NADP+), soluble Medium density, medium intensity
20 4930589M24Rik RIKEN cDNA 4930589M24 gene Medium density, medium intensity
21 Kit kit oncogene Medium density, medium intensity
22 Pcsk1 proprotein convertase subtilisin/kexin type 1 Medium density, medium intensity
23 Brunol6 bruno-like 6, RNA binding protein (Drosophila) Medium density, medium intensity
24 Spint2 serine protease inhibitor, Kunitz type 2 High density, low intensity
25 Loh11cr2a
loss of heterozygosity, 11, chromosomal region
2, gene A homolog (human) High density, low intensity
26 2700038N03Rik RIKEN cDNA 2700038N03 gene High density, low intensity
27 Grb10 growth factor receptor bound protein 10 High density, low intensity
28 Fgd5 FYVE, RhoGEF and PH domain containing 5 High density, low intensity
29 Lmx1b LIM homeobox transcription factor 1 beta Medium density, low intensity
30 Gdpd2
glycerophosphodiester phosphodiesterase do-
main containing 2 Medium density, low intensity
31 LOC244958
similar to Chromosome 6 open reading frame
117 Medium density, low intensity
32 Gpr4 G protein-coupled receptor 4 Medium density, low intensity
33 Gata3 GATA binding protein 3 Medium density, low intensity
34 Gpr176 G protein-coupled receptor 176 Medium density, low intensity
35 Dgcr6 DiGeorge syndrome critical region gene 6 Medium density, low intensity
Widespread Expression Pattern
50 SELECT GENES:
This gene list was generated by manual curation of an algorithmically derived list that compared gene expression values
of dorsal nucleus raphé to those of the medulla. Categories of expression are subjectively grouped by
relative expression characteristics.
Curation of 50 Select Genes List: March 2007
Nature Precedings : doi:10.1038/npre.2008.2054.1 : Posted 7 Jul 2008
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Number Gene Symbol Gene Name Expression Pattern
36 Slc17a8
solute carrier family 17 sodium-dependent inor-
ganic phosphate cotransporter, member 8 High density, very high intensity
37 S100a10 S100 calcium binding protein A10 (calpactin)
Medium density, high intensity (dorsal)
High density, very high intensity (ventral)
38 Slc10a4
solute carrier family 10 (sodium/bile acid cotrans-
porter family), member 4
Medium density, high intensity (dorsal)
High density, very high intensity (ventral)
39 Esr2 estrogen receptor 2 (beta) Medium density, medium intensity
40 Cited1
Cbp/p300-interacting transactivator with Glu/Asp-
rich carboxy-terminal domain 1 High density, high intensity
41 Pcdhac1 protocadherin alpha subfamily C, 1 Medium density, high intensity
42 Mgat4c
mannosyl (alpha-1,3-)-glycoprotein beta-1,4-N-
acetylglucosaminyltransferase, isozyme C
(putative) Medium density, medium intensity
43 Cbln4 cerebellin 4 precursor protein Medium density, medium intensity
44 Clcn5 chloride channel 5 Medium density, medium intensity
45 Pcsk5 proprotein convertase subtilisin/kexin type 5 Medium density, low intensity
46 Wrb tryptophan rich basic protein Medium density, low intensity
47 St6galnac6
ST6 (alpha-N-acetyl-neuraminyl-2,3-beta-
galactosyl-1,3)-N-acetylgalactosaminide alpha-
2,6-sialyltransferase 6 Medium density, low intensity
48 Hars histidyl-tRNA synthetase Medium density, low intensity
Ventral Expression Pattern
36.Slc17a8
37.S100a10
38.Slc10a4
39.Esr2
40.Cited1
41.Pcdhac1
42.Mgat4c
43.Cbln4
44.Clcn5
Nature Precedings : doi:10.1038/npre.2008.2054.1 : Posted 7 Jul 2008
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Number Gene Symbol Gene Name Expression Pattern
49 Pdyn prodynorphin
Medium density, very high intensity (dorsal)
Medium density, medium intensity (ventral)
45.Pcsk5
46.Wrb
47.St6galnac6
48.Hars
Dorsal Expression
Number Gene Symbol Gene Name Expression Pattern
50 Peg10 paternally expressed 10
High density, very high intensity (expression
blending into PAG)
Diffuse-Boundary Expression
49.Pdyn
50.Peg10
Ventral Expression Pattern
Nature Precedings : doi:10.1038/npre.2008.2054.1 : Posted 7 Jul 2008
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Correlated Expression:
Macro structure: Rostral-> Caudal
Correlation
0.830977
0.777591
0.931932
0.814291
0.945314
0.863267
0.789746
0.848881
0.953663
0.932689
0.915876
0.69521
OLF
(Olfactory areas)
STR (Striatum)
PAL (Pallidum)
CTX (Cerbral cortex)
HY
(Hypothalamus)
TH (Thalamus)
HIP
(Hippocampal region)
RHP
(Restrohippocampal formation)
MB
(Midbrain)
P (Pons)
MY
(Medulla)
CB
(Cerebellum)
Substructure Correlation
Dorsal nucleus raphé (DR) 1
Midbrain raphe nuclei (Ramb) 0.978943
Pons, behavioral state related (Psat) 0.977309
Periaqueductal gray (PAG) 0.97248
Midbrain, behavioral state related (Mbsat) 0.969455
Midbrain trigeminal nucleus (MEV) 0.966099
Superior central nucleus raphé (CS) 0.963406
M idbrain reticular nucleus , magnoc ellular part, general (MR Nmg) 0.96323
Sublaterodorsal nucleus (SLD) 0.962873
Parabrachial nucleus (PB) 0.961998
Pontine central gray (PCG) 0.960545
Midbrain, motor related (Mbmot) 0.957984
Edinger-Westphal nucleus (EW) 0.957505
Pedunculopontine nucleus (PPN) 0.956837
Ventral tegmental nucleus (VTN) 0.956359
Cuneiform nucleus (CUN) 0.956163
Dorsal tegmental nucleus (DTN) 0.95584
Trochlear nucleus (IV) 0.955012
Nucleus incertus (NI) 0.951671
Anterior tegmental nucleus (AT) 0.949758
Nucleus of the solitary tract (NTS) 0.94829
Pos terior hypothalamic nucleus (PH) 0.943703
Locus ceruleus (LC) 0.942764
Hypothalamic lateral zone (LZ) 0.94222
Midbrain trigeminal nucleus (MEV) 0.942159
The ABA coronal set contains the majority of genes of known scientific interest, as well as genes exhibiting marked or unique expres-
sion patterns. A correlation analysis of all available ABA coronal experiments (4376) was performed by comparing an expression
value of the dorsal nucleus raphé (DR) to expression values in other regions of the brain. Following image analysis, the data values
for each experiment were mapped to a 3-D reference brain at 200µm3 voxel resolution. Then, each voxel was assigned a single ex-
pression value based on the product of density and intensity of expression. Values from all 4376 experiments were computed, and
the likelihood of co-expression between any two voxels or regions are reported as a Pearson’s correlation coefficient.
For the purposes of determining correlated expression between the DR and other brain regions, expression values from all voxels
within the DR were aggregated to form a single expression value. Two types of comparisons were then made. First, the aggregate
expression values of the DR and those of other anatomically defined regions (~200 structures) were compared within the 3-D refer-
ence brain (structure vs. structure; table below). Second, a color map was then generated to display the correlation between the DR
and each of the ~53,000 voxels of the reference volume (structure vs. voxel; correlation map on the following page).
STRUCTURE vs. STRUCTURE
The expression value of the DR was compared to expression values for all other defined atlas regions. Degree of correlation is dis-
played as a comparative fraction, with self-correlation = 1.000. Correlation between the DR and macro/parent-structures are pre-
sented, as well as correlation between the DR and the 25 highest ranking substructures. (The most highly correlated macro/parent-
structures do not always contain the 25 top most correlated substructures). Columns match the Allen Reference Atlas palette.
Nature Precedings : doi:10.1038/npre.2008.2054.1 : Posted 7 Jul 2008
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STRUCTURE vs. VOXEL:
Correlation between the DR and all other 200um3 voxels in the brain. Degree of correlation assessed for each voxel is provided visu-
ally (lower value = the correlation value of the 25th ranked substructure reported on the previous page) using the “jet” color scale at
rostrocaudal levels throughout the brain.
0.94 1.00
Nature Precedings : doi:10.1038/npre.2008.2054.1 : Posted 7 Jul 2008
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Gene Ontology (GO) Analysis:
GO TABLE: Below is an ontological analysis of the 50 Select Genes, using DAVID Bioinformatics Resources.
The functional terms that follow were returned using these constraints:
Date of table completion: June 2007
Category Definition Constraints
P-value Probability that the term is over-represented in this 50 Select
Genes list relative to the mouse genome
when p ≤ 0.05
Gene Count The minimum number of genes that must fall into an onto-
logical category to be considered a group
5 genes per term group
GO Level The level of functional specificity for GO functional catego-
ries: Molecular Function (mf), Biological Process (bp) and
Cellular Components (cc)
Level GO_All
# of DAVID IDs Number of unique DAVID gene IDs from user’s input list 48 DAVID gene IDs/ 50 input genes
Glynn Dennis Jr., Brad T. Sherman, Douglas A. Hosack, Jun Yang, Michael W. Baseler, H. Clifford Lane, Richard A. Lem-
picki. “DAVID: Database for Annotation, Visualization, and Integrated Discovery.” Genome Biology. 2003 4(5): P3.
GO Category GO Term Gene Count % of Genes p-value
GOTERM_BP_ALL synaptic transmission 4 8.33% 0.003881
GOTERM_BP_ALL transmission of nerve impulse 4 8.33% 0.006153
GOTERM_BP_ALL cell-cell signaling 4 8.33% 0.017951
GOTERM_BP_ALL aromatic compound metabolism 3 6.25% 0.019359
GOTERM_MF_ALL anion transporter activity 3 6.25% 0.020489
GOTERM_BP_ALL inorganic anion transport 3 6.25% 0.039213
Nature Precedings : doi:10.1038/npre.2008.2054.1 : Posted 7 Jul 2008
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Other Tools:
NEUROBLAST:
Many of the 50 genes listed in this report can be used to explore the NeuroBlast tool. This unique mining tool works
seamlessly from within brain-map.org to produce a list of genes that share similar expression patterns to any gene in the
coronal data set. Search for and select any gene, then select one of several brain regions from the NeuroBlast drop-tab
to explore a ranked list of similarly expressed genes for that region.
To learn more about this function, please refer to the NeuroBlast white paper.
BRAIN EXPLORER:
To compare gene expression levels across anatomical structures in 3-D detail, download the Brain Explorer desktop
application. This program is used to view gene expression in 3-D view (coronal, sagittal, horizontal and everywhere in
between) across all brain structures and allows for simultaneous viewing of multiple expression profiles.
The NeuroBlast spatial homology function and an anatomic search tool are also available from within Brain Explorer to
allow the user to search for and visualize genes with similar expression patterns.
Dorsal Nucleus Raphé (DR) Summary:
Anatomy
In the coronal plane, the DR is found on the midline, embedded within the ventral periaqueductal gray
and below the cerebral aqueduct. In the sagittal plane, the DR is located at the caudal portion of the mid-
brain, below the cerebral aqueduct and above the medial longitudinal fascicle.
The dark, medium-sized cells within the DR pack in non-uniform densities and exhibit somewhat vertical
orientation.
The cytoarchitecture of the DR exhibits high contrast with neighboring regions.
Expression Patterns of the 50 Select Genes
Widespread expression patterns across the DR were common but varied in density.
Ventrally dominant expression was the next most common pattern, yet dorsal enrichment was rare.
A few diffuse-boundary expression patterns were observed, with Peg10 as the most obvious.
Taken together, these results suggest that further analysis may reveal sub-regions of the DR.
The rostral border with the Edinger-Westphal nucleus is best delineated with Maob gene expression.
Expression Correlation with the DR
Midbrain and hypothalamus correlated most highly with the DR.
Cerebellum and striatum were the least correlated regions.
Of the top ranking 25 sub-structures highly correlated with the DR, most reside in the midbrain, although
the lateral zone of the hypothalamus provides an interesting exception.
Please send comments or questions by email to !Annotation@alleninstitute.org. To further explore the gene expression data and
analytical tools referred to in this report, please access the genome-wide data set at brain-map.org.
Nature Precedings : doi:10.1038/npre.2008.2054.1 : Posted 7 Jul 2008
