REVIEWS
RETINOID SIGNALLING IN THE
DEVELOPMENT OF THE CENTRAL
NERVOUS SYSTEM
Malcolm Maden
Retinoids — a family of molecules that are derived from vitamin A — have been implicated in many
developmental processes. In the embryonic vertebrate central nervous system (CNS), retinoic acid
(RA) has a role in patterning both the anteroposterior and dorsoventral axes. Initially, RA was
thought to be involved in generating the entire anteroposterior extent of the CNS, but more recent
experiments have identified its main sites of action as the hindbrain and anterior spinal cord. RA
also regulates interneuron and motor neuron development along the dorsoventral axis. This review
describes the studies that led to these conclusions, and discusses how understanding the
mechanisms of RA action in the developing CNS might provide insights into neurological disease.
XEROPHTHALMIA
An excessive dryness of the
conjunctiva and cornea.
HYDROCEPHALUS
A condition, marked by an
expansion of the cerebral
ventricles and a compression of
neural structures, that is caused
by a block in the flow of cerebral
spinal fluid or by its
overproduction.
SPINA BIFIDA
Failure of neural tube closure at
the posterior neuropore, which
causes the spinal cord tissue to
protrude through the vertebral
column.
MRC Centre for
Developmental
Neurobiology, 4th Floor New
Hunt’s House, King’s College
London, Guy’s Campus,
London Bridge,
London SE1 1UL, UK.
e-mail:
malcolm.maden@kcl.ac.uk
doi:10.1038/nrn963
Vitamin A was discovered as an essential dietary component at the beginning of the twentieth century.
Deprivation studies soon revealed that, in its absence,
the adult animal shows characteristic changes, including widespread keratinization of epithelia, decreased
immune function, anaemia, XEROPHTHALMIA and blindness1. Interestingly, from the point of view of this
review, there were occasional reports of nerve degeneration in the spinal cord2,3, which resulted in a loss of
coordination and apparent symptoms of motor neuron disease in pigs and chickens. Following the observation that the deprivation of vitamin A in pregnant
pigs resulted in the birth of a litter without eyeballs4,
it became clear that such a dietary deficiency generated characteristic congenital malformations of
the embryo. In the central nervous system (CNS), the
reported malformations were HYDROCEPHALUS, SPINA
BIFIDA, ANOPHTHALMIA and MICROPHTHALMIA, but these
superficial descriptions of abnormalities included no
further anatomical details. Only recently have such
details been described in deprivation studies using
quail and rat embryos, which have generated precise
and interesting CNS patterning abnormalities, as
described in detail below.
The opposite type of experiment, in which an excess
of vitamin A is administered to embryos, did not begin
NATURE REVIEWS | NEUROSCIENCE
until 1953, when the metabolic pathway of vitamin A
had been established and retinoic acid (RA) was identified as the compound that acts within cells. RA — but
not vitamin A itself — was highly teratogenic when
administered in excess to pregnant mammals, and it
affected multiple systems of the body, including the
CNS, causing EXENCEPHALY, ENCEPHALOCOEL, MICROCEPHALY,
microphthalmia and spina bifida5–7. Important though
they were from the teratological point of view, these
observations did not really indicate what RA was doing
in terms of altering the patterning of the CNS. These
discoveries came only recently, after the cloning of genes
(for example, the Hox genes) that could serve as markers of different axial regions of the CNS. The absence
or duplication of these gene-expression domains gave
precise patterning information, as described below.
Perhaps the most interesting teratological observation from the perspective of the CNS is that embryos
treated with RA at TAIL BUD stages produce ectopic neural
tubes within the tail bud8,9. Therefore, RA can induce
neural tissue from mesoderm. Similar results had been
generated in abundance using embryonal carcinoma
(EC) cells. When RA was added to the cultures, neurons
and glia were induced to differentiate10,11. This type of
result has been observed consistently with various EC
cell strains, and with TERATOCARCINOMA CELLS, stem cells
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a
Vitamin A (retinol)
Alcohol dehydrogenases/
retinol dehydrogenases
Retinal
Retinaldehyde dehydrogenases
all-trans-RA, 9-cis-RA
CYP26A1, CYP26B1
4-oxo-RA, 4-OH-RA, 5,8-epoxy-RA
b
CRBP
RA
Retinol
RoDH
Retinol
RA
RXR RAR
RALDH
CRABP
Retinal
ANOPHTHALMIA
Absence of the eyeballs.
MICROPHTHALMIA
RARE
Figure 1 | Pathways for the synthesis and mechanism of
action of RA. a | The metabolic pathway that converts vitamin
A (retinol) into the various forms of retinoic acid (RA), which
involves three classes of enzymes. b | The cellular mechanism
of retinoid action. Retinol is taken up from the blood and bound
to CRBP (cellular retinol-binding protein) in the cytoplasm. The
retinol dehydrogenase (RoDH) enzymes metabolize retinol to
retinal, then retinal is metabolized to RA by the retinaldehyde
dehydrogenases (RALDHs). RA is bound in the cytoplasm by
CRABP (cellular RA-binding protein). RA enters the nucleus
and binds to the RA receptors (RARs) and the retinoid X
receptors (RXRs), which themselves heterodimerize and bind
to a sequence of DNA known as the RARE (RA-response
element). This activates transcription of the target gene.
A reduction in the size of the
eyeballs.
A blastema-like structure at the
posterior end of the embryo that
gives rise to all the structures of
the sacrocaudal body region:
neural tube, notochord, paraxial
mesoderm and gut endoderm.
and NEUROBLASTOMA CELLS from both mouse and human12.
RA also affects neurons at various stages of differentiation; for example, in dissociated embryonic neurons or
explants of neural tissue, RA causes outgrowth of more
and longer neurites than would normally be observed.
Hundreds of genes have now been shown to be regulated by RA during the processes of neuronal differentiation and neurite outgrowth, including transcription
factors, structural proteins, enzymes, cell-surface glycoproteins, extracellular proteins, neurotransmitters,
neuropeptide hormones, growth factors and cell-surface
receptors12. We would therefore expect RA to be involved
in neuronal differentiation in vivo. This is indeed
the case, as we shall see. In addition, RA is involved in the
patterning of the neuraxis along both the anteroposterior (AP) and dorsoventral (DV) axes. Before describing
these data, we will first consider how RA acts in cells.
TERATOCARCINOMA CELLS
What is RA and how does it act?
A cell line derived from a
malignant germ-cell tumour,
which arises from the ovary or
testis and is composed of
embryonal carcinoma cells.
RA is the most biologically active naturally occurring
member of a family of molecules called retinoids, all of
which are derived from vitamin A. Retinoids are
obtained from the diet in the form of retinyl esters
from animal meat or β-carotene from plants. Cells in
the embryo or adult that require RA obtain it from the
blood, where it circulates as retinol bound to retinolbinding protein. Inside the cell, the sequestered retinol is
EXENCEPHALY
Failure of the cranial component
of the neural tube to close.
ENCEPHALOCOEL
A neural tube defect that causes
the herniation of brain tissue.
MICROCEPHALY
An abnormally small head,
caused by reduced growth of the
brain or skull.
TAIL BUD
NEUROBLASTOMA CELLS
An immortalized cell line
derived from tumours that arise
from the neural crest.
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enzymatically converted, first to retinal by the retinol or
alcohol dehydrogenases (RoDHs or ADHs), and then to
RA by the retinaldehyde dehydrogenases (RALDHs)13
(FIG. 1a). There are several members of each of these
enzyme classes, and the most important RALDHs for
the embryo are RALDH1, RALDH2 and RALDH3. RA is
further metabolized by two cytochrome P450 enzymes
— CYP26A1 and CYP26B1 — to supposedly inactive
products such as 4-oxo-RA, 4-OH-RA, 18-OH-RA and
5,8-epoxy-RA14–16, and is then excreted. Some of these
products have recently been shown to be biologically
active, and can even induce neural tissue17. However, the
lethal phenotype of the Cyp26a1 mutant mouse, which
mimics the effects of excess RA administration, is rescued by the heterozygous disruption of Raldh2, which
simply reduces the amount of RA that is synthesized in
the embryo18. This indicates that the products of CYP
catabolism are not used by the embryo.
There are two isomers of RA, all-trans-RA and
9-cis-RA, which act through different receptors. It is not
known whether they are produced by separate enzymatic
pathways — from all-trans-retinol and 9-cis-retinol,
respectively — or whether they can be interconverted
by isomerization. Cells that require RA also contain
cytoplasmic retinoid-binding proteins. As their names
imply, the cellular retinol-binding proteins (CRBP1 and
CRBP2) bind retinol, whereas the cellular RA-binding
proteins (CRABP1 and CRABP2) bind RA (FIG. 1b).
Once RA has been synthesized, it enters the nucleus
and influences gene activity by binding to ligandactivated nuclear transcription factors. There are two
classes of these transcription factors — the RA receptors
(RARs) and the retinoid X receptors (RXRs). In human,
rat and mouse, there are three RARs (RARα, RARβ
and RARγ)19 and three RXRs (RXRα, RXRβ and
RXRγ)20. Each of these molecules has several isoforms.
The RARs and RXRs act as heterodimers (for example,
RARα–RXRβ), and they recognize consensus sequences
known as RA-response elements (RAREs) in the control
elements of RA-responsive genes (FIG. 1b).
So, there are several points at which a cell’s response
to RA could be regulated: the uptake of retinol from the
blood; the presence of the enzymes to convert retinol to
RA; the presence of the RARs and RXRs; and the presence of further co-regulators/co-repressors that interact
with the RARs and RXRs.
Primary neuron differentiation
The first neurons to differentiate within the neuroepithelium of fish and amphibians are known as primary neurons. The function of these early-developing neurons is
to coordinate escape movements, which are crucial for
survival after hatching. In Xenopus, they develop in three
parallel rows along either side of the midline, from the
posterior hindbrain backwards (FIG. 2a). The three rows
form ventral motor neurons (nearest to the midline),
interneurons and dorsal sensory neurons (the most
lateral). They express N-tubulin and islet 1, and there
are about 120 of them on each side of the embryo21.
The number of primary neurons is controlled by RA
signalling. The addition of RA to the embryo increases
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i
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Figure 2 | Primary neurons in Xenopus and the effect of altered RA signalling. a | Schematic
showing normal primary neurons at the neurula stage, as revealed by N-tubulin staining. There
are three lines of neurons on either side of the midline. The most medial ones, marked ‘m’, are
primary motor neurons. The middle lines, marked ‘i’, are interneurons. The lateral lines, marked
‘s’, are sensory neurons. b | The effect of excess retinoic acid (RA) on primary neurons. There are
far more neurons, and the three discrete lines have fused into one. In addition, the neurons are
also found at the anterior end of the embryo. c | The effect of decreasing RA signalling by treating
embryos with a retinoid receptor antagonist. The anterior border of expression has moved
posteriorly and there are far fewer neurons. Adapted, with permission, from REF. 22 © 1999
Company of Biologists.
ANIMAL CAPS
An explant cut from an
amphibian embryo at the
blastula stage, comprising a ‘cap’
of about 60° that is centred on
the animal pole. These explants
consist of uncommitted
ectodermal tissue, and they are
often used to test the activities of
putative neuralizing factors.
their numbers, so that they spread into the anterior part
of the embryo and the three stripes merge21–23 (FIG. 2b).
Similarly, increasing RA signalling, by injecting synthetic
messenger RNA that encodes Xenopus RARα2 and RXRβ
into one cell of the two-cell embryo, increases their number21,24. The additional primary neurons still form within
the boundaries of the neural plate by filling in the gaps
between the three stripes; the neural plate is not expanded
laterally to accommodate these extra numbers (FIG. 2b).
Conversely, the administration of citral (a competitive inhibitor of RA synthesis) decreases the number of
primary neurons21,25. As a result, the embryos no longer
respond to physical stimulation. Similarly, decreasing RA
signalling, by the injection of dominant-negative RARα2
or by using the RARα–RARβ antagonist Ro 41-5253,
reduces their numbers (FIG. 2c), and again the embryos
are unresponsive to touch22,24–26.
These methods of increasing or decreasing the
numbers of primary neurons act directly on the neuroepithelium, and not through the mesoderm. This has
been shown by duplicating the above results on isolated
ANIMAL CAPS. The amphibian animal cap normally
receives signals from the mesoderm, which comes to
underlie it after GASTRULATION. These signals turn the animal cap into neural plate, but if the cap is isolated
before receiving these signals, only epidermis is formed.
When isolated animal caps that have been injected with
noggin and/or XASH-3 to neuralize the tissue (this does
not promote primary neuron formation) are treated
with RA and/or injected with RARα–RXR, tubulin
expression is promoted21,23.
Studies of the neural genes that are altered by
increases or decreases in RA signalling have been very
revealing. The process of primary neurogenesis is under
the control of PRONEURAL GENES27, most of which have
been cloned as homologues of Drosophila genes in
vertebrates. There is no RA signalling in Drosophila, but
several of the proneural genes — including XASH-3,
X-ngnr-1 (neurogenin-related protein), X-delta-1, Xiro2,
X-MyT1, Gli3, Zic2 and X-shh (sonic hedgehog)22,25,28 —
have been found to be controlled by RA in vertebrates,
indicating that they must have come under the control
of RA as the vertebrates evolved. Studies of the relationship between these genes, and where RA fits into the
cascade of PREPATTERN GENES, proneural genes, neurogenic
genes and differentiation genes, have placed RA upstream of all of them22. So, RA seems to be responsible
for upregulating the prepattern genes (X-ngnr-1,
X-MyT1) and the neurogenic genes (X-delta-1), and for
downregulating the genes that inhibit neurogenesis
(Zic2, X-shh). RA is thought to act after neural tissue has
been induced by noggin, follistatin and chordin (FIG. 3).
Naive ectoderm
Neural inducers:
noggin, chordin, follistatin
Anterior neural tissue
GASTRULATION
RA, FGFs, Wnts
The process by which the
embryo becomes regionalized
into three layers: ectoderm,
mesoderm and endoderm.
Neurogenesis
AP patterning
RA
PRONEURAL GENES
Genes that encode transcription
factors of the basic
helix–loop–helix class that
specify neural progenitor cells
and promote their
differentiation.
PREPATTERN GENES
Also known as pre-proneural
genes, these genes seem to
provide a link between the
patterning of the nervous system
and the specification of neurons.
Downregulate:
Zic2 and X-shh
Upregulate:
X-ngnr-1,
X-MyT1, X-delta-1
and N-tubulin
Downregulation of
anterior genes:
Otx2, XCG-1, XAG-1,
XA-1, Emx1, Emx2,
Dlx1 and XINK-2
Upregulation of
posterior genes:
Krox20, Wnt1, En,
Pax2, XIF-3, Xlim-1,
and Hox genes
Posterior hindbrain,
anterior spinal cord
Figure 3 | Chart to show where RA fits into the scheme of neural development. Naive ectoderm is induced to form neural
tissue by the neural inducers, but this tissue is of anterior character. Retinoic acid (RA), along with fibroblast growth factors (FGFs)
and Wnts, simultaneously induces neurogenesis and sets up the anteroposterior (AP) pattern of the central nervous system. In the
right-hand pathway, RA upregulates a series of posterior genes and downregulates a series of anterior genes, thereby generating
pattern in the posterior hindbrain and anterior spinal cord. In the left-hand pathway, RA acts to induce neurogenesis by
downregulating and upregulating several identified genes.
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Anteroposterior patterning
c
As the above studies show, RA regulates the cascade of
genes that ultimately leads to neuronal differentiation,
but this neuronal differentiation must be patterned
along both the AP and DV axes. AP patterning takes
place concurrently with neuronal differentiation, and it
transpires that RA also switches on genes that are
responsible for the patterning of the neural plate along
the AP axis (FIG. 3). This can readily be appreciated by
looking at the Xenopus primary neurons, which stretch
posteriorly from the posterior hindbrain and are not
present at the anterior end of the embryo (FIG. 2a). Excess
RA causes the primary neurons not only to fill in the
gaps between the three lateral stripes, but also to expand
into the anterior end of the embryo (FIG. 2b). Conversely,
decreasing signalling causes the border of primary
neuron differentiation to move posteriorly (FIG. 2c).
So, which part of the neural tube does RA pattern?
As we shall see, initial experiments implied that RA acts
throughout the AP extent of the neural tube through its
posteriorizing activity, but subsequent experiments have
instead focused on a more localized role of RA in the
hindbrain and anterior spinal cord.
mb
Eye
mb
hb
1
2
3
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Endogenous
gradient
of RA
RA concentration
Increased
signalling
Decreased
signalling
fb
mb
hb
sc
Anterior
BLASTULA
An embryo before the
gastrulation stage, consisting of a
hollow ball of epithelial cells that
surround a fluid-filled cavity.
NEURULA
The stage of development that
follows gastrulation, when the
neural plate starts to develop
from the ectoderm.
PRIMITIVE STREAK
An elongated depression of
reptile, bird and mammalian
embryos, through which
mesodermal and endodermal
cells migrate into the interior of
the embryo. The most anterior
tip of the primitive streak forms
Hensen’s node. The streak is
functionally homologous to the
amphibian blastopore.
846
Posterior
Figure 4 | The effects of altered RA signalling on Xenopus
embryos. a | The normal central nervous system (CNS) of an
embryo, showing the main subdivisions. 1–7, rhombomeres
1–7; fb, forebrain; hb, hindbrain; mb, midbrain; sc, spinal cord.
b | The effect of retinoic acid (RA) treatment on Xenopus
embryos. The eyes, forebrain and most of the midbrain are
missing, and the remaining hindbrain and spinal cord are
enlarged. c | The effect of injecting a dominant-negative RA
receptor (RARα1; decreased signalling) into the left cell of a
two-cell embryo. At the neural plate stage, the right (control)
side shows the normal expression domains of neural markers,
including Otx2 (brown; forebrain), En2 (blue; midbrain–hindbrain
border), Krox20 (green; two stripes, one in rhombomere 3 (r3)
and the other in r5) and Hoxb9 (red; spinal cord). On the left
side of the embryo, decreased signalling has resulted in the
posterior expansion of Otx2 and En2, loss of the posterior
stripe of Krox20, and downregulation of Hoxb9. d | The effect
of injecting a constitutively active RARα1 (increased signalling)
into the left cell of a two-cell embryo. At the neural plate stage,
the right (control) side shows the normal expression domains of
neural markers, as in c. On the left side of the embryo,
increased signalling has resulted in the anterior compression of
the Otx2, En2 and Krox20 domains, with no change in Hoxb9.
e | Explanation of the effects of increasing and decreasing RA
signalling, based on an endogenous whole-body gradient of
RA with a high point at the posterior end. Different parts of the
CNS develop at different concentration thresholds. When
signalling levels are changed, parts of the CNS are deleted
because the thresholds are too low (red line) or are expanded
(green line).
| NOVEMBER 2002 | VOLUME 3
Effects of RA on the whole neuraxis. If whole embryos
are treated with an excess of RA, anterior structures such
as the forebrain and eyes are lost, and the hindbrain and
spinal cord seem to expand to compensate (FIG. 4a,b).
This has been observed most commonly in Xenopus
when RA is administered at late BLASTULA /early NEURULA
stages29–42, but it also occurs in urodeles43 and zebrafish44,
and when RA is administered at mid/late PRIMITIVE STREAK
stages in rat and mouse45–47. In addition to the loss of
anterior structures, anterior genes, such as Otx2, XCG-1,
Emx1 and Dlx1, are repressed. At the same time, more
posterior structures, such as hindbrain and spinal cord,
are expanded, and posterior genes such as Krox20, Pax2
and various Hox genes are upregulated. The cells at the
anterior end of the embryo do not just die after RA
treatment, but are respecified48, which explains why the
remaining hindbrain and spinal cord seem to be
enlarged31 (FIG. 4b). As in the case of the induction of
excess primary motor neurons, RA acts directly on the
neuroepithelium rather than through the mesoderm to
cause this effect23,35,49–52.
Alteration of RA signalling produces the same
phenotypes as excess RA in Xenopus. RAR- and RXRselective ligands cause anteriorization and upregulation
of posterior genes. Injection of a dominant-negative
RARα1 enlarges anterior structures, shortens the tail
and reduces the overall length of the embryos, and also
results in an expansion of the anterior gene-expression
domains26 (FIG. 4c, left side versus right side). Conversely,
a constitutively active RARα1 reduces anterior structures, and the anterior gene-expression domains
become compressed26 (FIG. 4d, left side versus right
side). Overexpression of Xenopus CRABP produces
abnormalities that are typical of excess RA administration; namely, reduced eyes, reduced forebrain and
midbrain, and enhancement of Hoxb4 and Hoxb8
expression53.
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4
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Figure 5 | The effects of increasing and decreasing RA signalling on the hindbrain of chick
and mouse embryos. a | Normal embryonic central nervous system (CNS). 1–7, rhombomeres
1–7; fb, forebrain; hb, hindbrain; mb, midbrain; sc, spinal cord. b | Typical effect of an excess of
retinoic acid (RA) on a mouse embryo. The anterior hindbrain is deleted, leaving behind one large
rhombomere (r4), with a loss of segmentation in the posterior rhombomeres. c | Typical effect of a
lower dose of RA than that used in b on a mouse embryo. Here, r2 is transformed to a r4 identity,
and r3 is partially transformed to r5. d | Typical effect of removing all RA from quail or rat embryos.
The hindbrain region now comprises only the anterior three rhombomeres, each of which is
enlarged. This effect is also seen in retinaldehyde dehydrogenase 2 (Raldh2)-knockout mice,
RA receptor (RAR)-antagonist-treated mice and chicks, and RARα/RARγ double-knockout mice.
e | The hindbrain in a RARα/RARβ double-knockout mouse. The r6–r7 border has gone, and r5
is expanded.
The concept that emerged to explain these findings is
that there is an endogenous gradient of RA within the
developing CNS with a high point at the posterior end
(FIG. 4e). Shifting the gradient anteriorly (increasing signalling) eliminates the forebrain and midbrain, whereas
shifting it posteriorly (decreasing signalling) enlarges
the forebrain and midbrain (FIG. 4e).
OTOCYST
An ectodermal invagination that
constitutes the primordium of
the internal ear.
Effects of RA on the hindbrain. The whole-neuraxis gradient model fits well with the experimental data above.
However, several other Xenopus experiments in which
RA signalling was altered produced results in which
only the hindbrain was affected. For example, CRABP
overexpression caused loss of segmentation in the hindbrain53, and a dominant-negative RARβ that was
injected into Xenopus resulted in a shorter and thicker
hindbrain, with no rhombomere boundaries and
increased numbers of Mauthner neurons (the neurons
that regulate the escape response) in the posterior hindbrain54. As Mauthner neurons are normally present only
in rhombomere 4 (r4), and Krox20 (which is normally
expressed in r3 and r5) was ectopically expressed in r4,
r6 and r7, it seemed that the posterior hindbrain had
been partially anteriorized. Ectopic expression of
Cyp26a1 (FIG. 1a) in Xenopus induces an anterior hindbrain duplication, which is reflected in a duplicated
trigeminal ganglion and a posterior shift of Krox20,
Pax6 and Hoxb3 expression55.
NATURE REVIEWS | NEUROSCIENCE
Another example of the effects of RA signalling on
the hindbrain came from studies by Godsave et al.56,
who showed that there was a concentration-dependent
effect of excess RA on rhombomere-specific gene markers. Anterior rhombomere markers were induced at low
concentrations, and more posterior markers required
progressively higher concentrations of RA for their
induction. Spinal cord genes, on the other hand, were
virtually unresponsive.
This rhombomere-specific effect of high levels of
excess RA was already well established in mammalian
embryos. Embryos that have been treated in this way have
a shortened preotic hindbrain, which is reflected externally in an abnormally rostral position of the OTOCYST. It
results from the loss of a section of CNS tissue from the
anterior hindbrain, as well as a loss of posterior rhombomere segmentation (FIG. 5b). Most vertebrate embryos
show this phenotype, from zebrafish44,57 through
Xenopus58,59 and chick58,60, to rat and mouse46,47,61–65. The
effect on Hox gene expression was used to establish
what tissue had been lost. After RA treatment, it was
shown that there is an anterior spread of the Hox-geneexpression domains into the anterior hindbrain and
midbrain, followed by a retraction, leaving behind an
aberrant expression pattern66. The interpretation that is
most consistent with the gene-expression data65 and the
neuronal architecture67 is that the single large rhombomere is r4, so r1, r2 and r3 have been lost, in addition to
the posterior boundaries (FIG. 5a,b).
Another effect of excess RA on the hindbrain is
striking, because it involves not a loss of tissue, but a
transformation of anterior hindbrain rhombomeres
from one state to another. It occurs with a lower dose
of RA, or by treating with RA at a slightly later
stage47,61,65,68, and it has been characterized mainly in
zebrafish and mouse57,69, although it also occurs in
Xenopus 70. In the mouse, Hoxb1–lacZ transgenic
embryos were generated, which showed the restriction
of Hoxb1 to r4 and the facial nerve that derives from it.
After treatment of late-streak-stage embryos with a
relatively low dose of RA, the expression of Hoxb1
shifted anteriorly and then became restricted to two
stripes, rather than the normal single stripe. The second
stripe is in r2, and the nerve that usually emerges from
r2 — the trigeminal nerve — comes to resemble the
facial nerve of r4 (REFS 69,71). Rhombomere 3 changes
in some of its neuronal characteristics to resemble r5,
but not in terms of its Hox-gene-expression profile.
So, two rhombomeres are at least partially transformed
by the RA treatment (FIG. 5c).
So, excess RA signalling seems to eliminate or alter
the anterior hindbrain. Conversely, decreased signalling,
as we shall now see, eliminates or alters the posterior
hindbrain.
Decreased RA signalling and the hindbrain. A decrease
in RA signalling can be brought about in several ways;
for example, by removing RA from the diet, by treating
embryos with RAR antagonists, by knocking out the
RARs or by knocking out the enzymes that make RA.
The overall effect of decreased signalling is a sequential
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Figure 6 | The hindbrain structure after gradually
decreasing RA signalling. a–f | Retinoic acid (RA) signalling
is decreased from normal (f) to no RA signalling at all (a). At
each decreased level, one rhombomere boundary is lost.
g | Graphical representation of the level of RA signalling at each
of these stages. This data was gathered by decreasing the
levels of RA, by treating with a RA receptor antagonist at
successively earlier stages, or by decreasing RA signalling.
NODE
A major organizing centre in
primitive-streak-stage embryos
that regulates pattern formation.
It is known as Hensen’s node in
chick and the Spemann
organizer in frog.
848
loss of posterior rhombomeres. After removing RA
from quail embryos by dietary deficiency, the posterior
hindbrain fails to develop. Rhombomeres 4, 5, 6 and 7
are missing, and the remaining rhombomeres (r1, r2
and r3) expand in size to compensate72,73 (FIG. 5d). This
outcome was assessed by the use of various hindbrain
markers, including Hoxb1, Fgf3, MafB and Krox20, and
by examining the structure of the hindbrain, which
now consisted of three distinct rhombomeres attached
to a spinal-cord-like structure (FIG. 5d). Essentially the
same result was seen in RA-deficient rat embryos74,75,
although as this spinal-cord-like tissue expressed
Hoxb1, it was interpreted as being an enlarged r4.
Decreasing signalling by knocking out RALDH2, one
of the enzymes that synthesizes RA (FIG. 1a), produces
the same phenotype; namely, two or three rhombomeres that are expanded in size and attached to a thinner spinal-cord-like structure76,77. A similar repertoire
of markers, including Hoxb1 and other Hox genes,
MafB, Wnt8c and Eph (ephrin receptor) genes, was
used to reach this conclusion. However, a zebrafish
mutation in Raldh2 only has a very mild hindbrain
phenotype, possibly because of the recent genome
duplication that has occurred in the fish lineage,
which means that there might still be one perfect copy
of Raldh2 (REFS 78,79). These all-or-nothing experiments showed that the entire posterior hindbrain
segment (the myelencephalon) depends on RA for its
development. So, the next question is, what happens
when RA signalling is decreased gradually rather than
removed entirely?
| NOVEMBER 2002 | VOLUME 3
This question was addressed by studying receptorknockout embryos, and by using a pan-RAR antagonist. Culturing chick embryos in high concentrations of
a pan-RAR antagonist caused the loss of posterior
rhombomeres, with r1, r2 and r3 remaining attached to
a smooth neural tube, the anterior portion of which
was again interpreted as being an enlarged r4 due to its
expression of Hoxb1 (REF. 80). Gradually decreasing
signalling (increasing concentrations of the antagonist),
or treating with a high concentration of antagonist at
successively earlier stages (from stage 10 to stage 5),
produced a sequential loss of posterior rhombomeres.
Each loss was preceded by a rhombomere boundary
loss and then by an expansion of that domain in the
following order: first, the r7–r8 border was lost and r7
was expanded; then the r6–r7 border was lost and r6
and r7 were expanded then lost; then the r5–r6 border
was lost and r5 was expanded then lost. Then, the r4–r5
border was lost and r4 was expanded. The fact that the
rhombomere boundaries are lost one by one as RA signalling is experimentally decreased indicates that RA
might function in the sequential formation of these
boundaries.
Receptor-knockout mice also show loss-of-boundary
phenotypes. Although embryos in which a single RA
receptor is knocked out have normal hindbrains,
RARα–RARβ double knockouts have fused r6 and r7
(due to loss of the r6–r7 boundary) and an expanded r5
(REF. 81) (FIG. 5e). RARα–RARγ double-knockout embryos
have a similar phenotype to RA-deficient embryos and
Raldh2-knockout embryos, with missing posterior
rhombomeres and expanded anterior rhombomeres82
(FIG. 5d). Both of these defects can be mimicked by the use
of a pan-RAR antagonist. Treating mouse embryos at
day 7 with the antagonist phenocopies the RARα–RARγ
phenotype (FIG. 5d), and treating at day 8 phenocopies the
RARα–RARβ phenotype, which consists of an expanded
r5 and fused r6–r7 (FIG. 5e). This indicates that RARα
and/or RARβ function later to form the r6–r7 border,
after RARα and/or RARγ have acted to form the r3–r4,
r4–r5 and r5–r6 boundaries.
From these findings, we can propose a model for
the function of RA in building the posterior hindbrain
(FIG. 6). Similar ideas have recently been described by
Gavalas83. The ground state in the absence of RA is
enlarged r1, r2 and r3 (FIG. 6a). RA could be released
from a source that is posterior to the developing hindbrain, forming a gradient that decreases in concentration towards the anterior end of the hindbrain.
Alternatively, RA could be generated in a population of
cells akin to a NODE, the progress zone of the limb bud,
or a root tip of a plant, and when cells leave this population they no longer receive RA signals. Earlier-leaving
cells would have received a lower level of RA signalling
than later-leaving cells. The sequence of concentrationdependent RA-induced events (low-concentrations
events first, high-concentration events last) would
then be as follows. First, make a r3–r4 border (FIG. 6b).
Then, make a r4–r5 border (FIG. 6c); make a r5–r6 border (FIG. 6d); make a r6–r7 border (FIG. 6e); and finally,
make a r7–r8 border (FIG. 6f).
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Does endogenous RA fit either theory?
PARAXIAL MESENCHYME
So, we have two ideas about how RA might organize AP
patterning. One is that there is a head-to-tail gradient
that extends over the whole neuraxis, with a high point
at the posterior end (FIG. 4e). The second is that there is a
localized source of RA at the posterior end of the hindbrain that is responsible for patterning the posterior
hindbrain (FIG. 6g). Do endogenous measurements
provide support for either of these ideas?
Endogenous measurements of RA in Xenopus have
been contradictory. Some data support the idea of an
AP gradient at the neural plate stage, with a high point
at the posterior end84, but other measurements have
found higher levels at the anterior end of the embryo36.
A region of the mesoderm
adjacent to the notochord,
which becomes segmented
rostrocaudally to give rise to the
somites early in development.
Hensen’s
node
b
c
RA concentration
a
fb
d
fb
mb
mb
hb
hb
sc
sc
Neural plate
Mesoderm
Anterior
Posterior
fb
e
mb
1–3
Neural plate
Mesoderm
Anterior
Posterior
fb
mb
1–3
Neural plate
Mesoderm
Anterior
Posterior
fb
mb
1–3
Neural plate
Mesoderm
Anterior
Posterior
Figure 7 | Endogenous RA in the chick and mouse embryo. a | Stage 4 chick embryo,
showing that retinoic acid (RA) can be detected in the posterior region (red area), behind Hensen’s
node. This mirrors the distribution of retinaldehyde dehydrogenase 2 (Raldh2) expression at this
early stage. b | Double in situ hybridization of a stage 6 chick embryo, showing the expression of
the cytochrome P450 Cyp26a1 at the anterior end (green) and Raldh2 (red) at the posterior end.
In between is the region that is fated to form the hindbrain, which is 100–200 µm in size.
c | Graphical representation of the levels of RA in the embryo in b, generated by Raldh2 and
catabolized by Cyp26a1. fb, forebrain; mb, midbrain; hb, hindbrain; sc, spinal cord. d | Diagram to
show where the RA in b and c is actually present in the embryo. Raldh2 is expressed in the
posterior mesoderm, whereas Cyp26a1 is expressed in the anterior neural plate, which is fated to
form the forebrain and midbrain. Therefore, RA has to diffuse up into the neural plate and then
turn towards the Cyp26a1 sink. e | An alternative explanation for the part that RA might play in the
generation of the posterior hindbrain. Here, the RA does not form a gradient, but it signals
sequentially to the neural plate above it. As the neural plate grows anteriorly (or the Raldh2expressing posterior mesenchyme moves posteriorly), successively more posterior hindbrain
tissue is exposed to RA. 1–3, rhombomeres 1–3.
NATURE REVIEWS | NEUROSCIENCE
However, in mouse and chick embryos, using a variety
of techniques, including HPLC (high-performance
liquid chromatography)85,86, LacZ reporter cells86–89 or
LacZ transgenic mouse embryos90–94, the consistent
finding has been that the early forebrain, midbrain and
hindbrain have low or undetectable levels of RA, but
that the spinal cord has very high levels. There is a
sharp on–off boundary of RA at the hindbrain–spinal
cord border, which is at the level of the first somite
(FIG. 7a). This distribution does not support the idea of a
whole-body gradient of RA.
The distribution of endogenous RA is validated by
studies of the distribution of the enzyme RALDH2.
From very early stages (stage 4 in the chick), it is
expressed in a butterfly shape (FIG. 7a), with a sharp border where the head and trunk meet, which will later be
at the level of the first somite95,96. At the same stage,
CYP26A1 — one of the enzymes that metabolize RA —
is expressed at the anterior end of the embryo. A double
in situ hybridization using Cyp26a1 and Raldh2 probes
revealed a gap between the two domains, and this is the
region where the hindbrain will develop96,97 (FIG. 7b).
Clearly, there is the potential for a gradient to form
between the presumptive spinal cord (source) and the
presumptive midbrain (sink), which would be responsible for the formation of rhombomere boundaries and
domains of Hox gene expression (FIG. 7c). The size of the
gap at stage 6 (FIG. 7b) is 100–200 µm — an ideal distance over which this gradient could form, and indeed,
such a gradient has been proposed previously to explain
how the expression pattern of the posterior hindbrain
gene MafB might develop98. The expression of Cyp26b1
begins somewhat later than these presomite-stage
events, and is found in r3 and r5, at least in the mouse99.
If it is involved, this form of CYP would serve to sharpen
the proposed gradient.
However, a complicating factor is that Raldh2 is
expressed not in the developing spinal cord, but in the
adjacent PARAXIAL MESENCHYME. Cyp26a1, on the other
hand, is expressed in the anterior neuroepithelium95,96,100. Therefore, the proposed gradient would
have to go from the paraxial mesenchyme (which subsequently forms the somites) into the neuroepithelium
above, and then turn at right angles to enter the anterior neuroepithelium (FIG. 7d). Arguments against this
idea include the view that this mechanism is asking
rather a lot of the cellular machinery, and that the
neural tube itself can generate RA86,101, and can activate
RAR and RXR reporters102, although the enzyme
involved is not known. However, evidence to support
the idea has come from the many experiments showing
that, when cervical somites are grafted rostrally, adjacent to the hindbrain, the expression patterns of Hoxb4,
Hoxa3, CEK8 and MafB are altered 98,103,104, and that
this effect can be mimicked by the implantation of
RA-soaked beads98 and can be inhibited by disulphiram
treatment of the somites105 (disulphiram is an inhibitor
of RA synthesis). In addition, the Raldh2 zebrafish
mutant called neckless, which has a mild hindbrain
phenotype, can be rescued by the transplantation of
wild-type mesodermal cells78. Finally, the gradual loss
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REVIEWS
of this inductive ability of the first four somites from
the two-somite stage to the ten-somite stage104 correlates precisely with the loss of Ralhd2 expression in the
same somites. So, the diffusion of RA from the paraxial
mesoderm into the neural tube to generate the posterior hindbrain seems to be well supported by the
experimental data.
But is a gradient of RA required? Does the RA that comes
from the paraxial mesoderm actually need to form a
gradient across the presumptive hindbrain (FIG. 7c)?
There are two arguments against the gradient hypothesis. First, it has been exceedingly difficult to demonstrate
the existence of an RA gradient. Second, it is difficult to
imagine how rescuing RA-deficient embryos by systemically administering RA could result in the recreation of a
gradient72,76,106. Third, as CYP26A1 functions as the sink
to break down the RA that is generated by RALDH2,
knocking it out should destroy the gradient and produce the same phenotypes as an excess of RA (loss of
anterior hindbrain and the transformation of anterior
rhombomeres). However, although Cyp26a1-knockout
embryos were found to have severe posterior (tail bud)
abnormalities, no significant growth or segmentation
abnormalities in the hindbrain were reported107,108. The
only minor abnormalities in the hindbrain were in the
expression domains of Hoxb1 (slightly larger), Meis2
(slightly smaller) and Krox20 (slightly smaller).
An alternative to a gradient would be a constant supply of RA from the paraxial mesoderm over time, but
where the neuroepithelium grows and moves away from
the source of RA. More posterior rhombomeres and
rhombomere boundaries that develop later than the
more anterior ones would have been exposed to more
total RA, so more posterior genes (requiring a higher
concentration of RA) would be induced (FIG. 7e). A similar conclusion regarding RA activity has recently been
presented83.
a
rp
D
b
BMPs
V0
V1
V2
mn
V3
c
Shh
fp
d
LMCs
RALDH2
850
| NOVEMBER 2002 | VOLUME 3
Dorsoventral patterning
Two experiments have provided evidence for further
roles for RA in the CNS, and these both concern neuronal populations along the DV axis of the spinal cord
(FIG. 8a). In the first experiment, naive neural plate tissue
was cultured in the presence of retinol. As a result, certain subsets of interneurons, which were characterized
by the expression of the homeobox genes Dbx1, Dbx2,
Evx1, Evx2 and En, were induced109. This is consistent
with studies of the DV expression domains in the spinal
cord of the RA-deficient quail embryo, in which these
populations of interneurons are missing (L. J. Wilson
and M.M., unpublished data). In this work, other populations along the DV axis were also studied. Ventral populations (Shh, Nkx6.1) had expanded at the expense of
dorsal populations (Bmp4, Bmp7, Pax3 and Wnt1),
which had shrunk, and the interneuron populations in
the middle were abolished (FIG. 8b,c). This indicates a role
for RA in repressing ventral neuronal genes and inducing dorsal genes, so that interneurons can develop in the
centre of the spinal cord. Interestingly, these DV abnormalities were only present at the anterior spinal cord
level, and not at posterior levels, which accords perfectly
with the proposed role of RA in anterior spinal cord
determination110. This also indicates that specification of
the DV axis occurs at the same time as that of the AP
axis in this region. As no RALDHs have been identified
in the early neural tube, the source of RA for this inductive/repressive action must once again be the somites.
Cyp26a1 is expressed in the dorsal third of the neural
tube96, so this might act as a sink.
A second and later event in the DV axis of the spinal
cord concerns the appearance of subsets of motor
neurons. The enzyme RALDH2 begins to be expressed
in the motor neurons of the spinal cord at the limb levels
at stage 19 in the chick and at day 12.5 in the
mouse95,111,112. These limb levels of the spinal cord had
previously been identified as ‘hot spots’ of RA synthesis
along the AP axis113. Here, there is a special class of
Figure 8 | Role of RA in the dorsoventral axis of the spinal
cord. a | Classes of neurons that can be identified along the
dorsoventral (DV) axis of the normal embryonic spinal cord.
D, dorsal sensory neurons; fp, floor plate; mn, motor neurons;
rp, roof plate; V0, V1, V2, interneurons; V3, ventral neurons.
These classes of neurons are distinguished by their unique
gene-expression profiles, many of which are characterized by
combinations of homeobox transcription factors. b | Dorsal
patterning is controlled by a gradient of bone morphogenetic
proteins (BMPs) that arises from the dorsal roof plate, and
ventral patterning is controlled by a gradient of sonic hedgehog
(Shh) that arises from the floor plate. c | The pattern of dorsal
and ventral genes in the retinoic acid (RA)-depleted quail spinal
cord indicates that there is increased ventral signalling and
decreased dorsal signalling. d | The role of RA in generating a
subset of motor neurons in the spinal cord. Retinaldehyde
dehydrogenase 2 (Raldh2) is expressed in motor neurons at
limb levels (red circles). A subset of motor neurons known as
lateral motor column neurons (LMCs) originates close to the
midline of the cord (green circles) and then migrates through
the Raldh2-expressing motor neurons to differentiate at the
edge of the cord (arrow). During this journey, these cells are
exposed to RA released by the motor neurons (red circles),
and as a result, are induced to form LMCs.
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BRACHIAL
At the level of the forelimbs.
LUMBAR
At the level of the lower back.
SUBSTANTIA NIGRA
A part of the midbrain that
contains dopamine-producing
neurons, the axons of which
innervate the striatum and
thereby control body
movements.
MORRIS WATER MAZE
A learning task in which an
animal is placed in a pool filled
with opaque water and has to
learn to escape to a hidden
platform that is placed at a
constant position. The animal
must learn to use distal cues, and
the spatial relationship between
them and the platform. Learning
in this task involves the
hippocampus.
motor neurons — the lateral motor column neurons
(LMCs) — which are induced earlier in development by
the presence of BRACHIAL somites (at the brachial eminence) and LUMBAR somites (at the lumbar eminence).
When these somites were grafted up or down the spinal
cord, ectopic LMCs were induced114, and a similar result
was obtained when Raldh2 was virally misexpressed at
thoracic non-limb levels112. Furthermore, the specific
LMCs that were generated by this viral misexpression
were not from cells that expressed Raldh2 themselves,
but from adjacent cells that migrated through the
Raldh2-expressing cells to a lateral location forming
the lateral LMCs (FIG. 8d). This indicates that there is a
paracrine inductive event, in which one cell generates
RA from RALDH2, and an adjacent cell is induced to
form a specific type of motor neuron (FIG. 8d).
Implications for neurological disease
Do these concepts have any relevance for postnatal life or
in understanding the causes of neurological disease? As
RA is a necessary component of the adult diet, it is highly
likely that RA signalling also occurs in the adult CNS.
Indeed, the signalling components — namely, RALDH1,
RALDH2, RA, the RARs, the RXRs, CRABP and CRBP
— are all present in the adult nervous system115–118. A
failure to function of any one of these components, such
as an enzyme malfunction or a receptor mutation, would
be expected to lead to the malfunction or degeneration
of the neuron or group of neurons concerned.
Do the expression domains of these signalling components give us any indication as to which groups of
neurons might be susceptible to such a malfunction?
There are at least four examples that can be put forward.
First, RALDH1 is expressed specifically by the neurons
of the SUBSTANTIA NIGRA119. The neurons with which they
synapse in the striatum have dopamine receptors, and
the levels of these receptors are controlled by RA120–122.
Double-null mutant mice for the RARβ and RXRβ or
RXRγ genes have reduced levels of dopamine receptors,
in addition to locomotor deficits123. It is easy to imagine,
therefore, that the malfunctioning of RALDH1 could
lead to decreased levels of RA, a failure of dopamine signalling and death of the nigral neurons, as could malfunctioning of RARβ or the RXRs in the striatum. In
other words, a failure of RA signalling might lead to a
movement disorder, such as Parkinson’s disease.
Second, as RALDH2 is required for LMC development at limb levels112, it is possible that, in the adult, the
failure of RA signalling might lead to degeneration of
limb motor neurons, leading to motor neuron disease.
Indeed, early studies in which adult pigs and chickens
were deprived of vitamin A resulted in animals with
degeneration of spinal cord motor neurons, loss of
coordination and spasms2,3. We have recently shown
that this is also the case in adult rats, and that there are
defects in the retinoid signalling pathway in human
cases of motor neuron disease124,125.
Third, RARβ- and RXRγ-null mutant mice have
severely impaired abilities in the MORRIS WATER MAZE, which
is a test of spatial learning and memory126. This is reflected
in the virtual elimination of long-term potentiation or
NATURE REVIEWS | NEUROSCIENCE
long-term depression in the hippocampus. So, a gradual
failure of RA signalling could be responsible for the
gradual decline in these abilities with age.
Finally, another neurological disease that might be
related to defects in retinoid metabolism is schizophrenia106,127,128. The evidence for this came from the
observation that genes that have been implicated in
schizophrenia are co-localized on the chromosomes
with the major retinoid genes. Also, dysfunction of the
dopaminergic system has been seen in patients with
schizophrenia, and as discussed above, dopamine receptors are regulated by RA. In addition, schizophrenia is
a disease that affects the forebrain, and certain aspects
of forebrain development are under the control of RA
synthesized by RALDH3 (REF. 129).
We do not know the causes of Parkinson’s disease,
motor neuron disease, schizophrenia or the decline in
mental abilities with age, but these considerations raise
the possibility of investigating a failure of retinoid signalling as one potential cause. If a failure to generate RA
itself underlies any of these diseases, then the potential
for a cure by administering this small lipophilic molecule
is an exciting prospect.
Summary and conjectures
Retinoid signalling is involved in several aspects of the
development of the CNS. In lower vertebrates, it is
required for generating the correct numbers of primary
neurons, and for their correct positioning. RA signalling is also responsible for aspects of AP patterning.
It was originally thought to be involved in the organization of patterning along the whole AP axis of the early
CNS, through the action of a gradient of RA with a
high point at the posterior end. Integration of more
recent data indicates that the hindbrain and anterior
spinal cord are the foci for the action of RA. Experiments in which RA signalling has been decreased have
revealed that RA seems to add posterior rhombomeres
(r4–r7), one by one, from a three-rhombomere ground
state (r1–r3) by generating rhombomere territories.
The presence of a gradient of RA across the developing
hindbrain, with a high point at the posterior end, is
implied by the distribution of Raldh2 and Cyp26a1
expression, but the proposed gradient has proved to be
difficult to measure and does not accord with the
Cyp26a1-knockout phenotype. A gradient might not
be necessary, as a constant source supplied over time
to a growing system could generate the same result.
Last, on the DV axis, RA signalling is required for
interneuron development, and it is also required
for the later development of a subset of motor neurons,
the LMCs.
What is now required is a greater understanding of
RA and RA-like molecules in the CNS: how and where
are they synthesized and what are their endogenous
distributions? For example, it will be interesting to find
out whether there are novel retinoids in the CNS that
are not found elsewhere in the body. In addition, it will
be important to identify the downstream targets of RA
signalling so that we can really get to grips with its
mechanisms of action. It is an increasingly common
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belief that developmental mechanisms are used in the
adult to maintain the differentiated state, and RA signalling mechanisms certainly seem to be a good example of this. There are several neurological diseases in
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4.
5.
6.
7.
8.
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852
Wolbach, S. B. & Howe, P. R. Tissue changes following
deprivation of fat soluble A vitamin. J. Exp. Med. 42,
753–777 (1925).
Hart, E. B., Miller, W. S. & McCollum, E. V. Further studies on
the nutritive deficiencies of wheat and grain mixtures and the
pathological conditions produced in swine by their use.
J. Biol. Chem. 25, 239–260 (1916).
Hughes, J. S., Lienhardt, H. F. & Aubel, C. E. Nerve
degeneration resulting from avitaminosis A. J. Nutr. 2,
183–186 (1929).
Hale, F. Pigs born without eye balls. J. Hered. 24, 105–106
(1933).
Cohlan, S. Q. Excessive intake of vitamin A as a cause of
congenital abnormalities in the rat. Science 117, 535–536
(1953).
Langman, J. & Welch, G. W. Effect of vitamin A on
development of the central nervous system. J. Comp.
Neurol. 128, 1–16 (1967).
Shenfelt, R. E. Morphogenesis of malformations in hamsters
caused by retinoic acid: relation to dose and stage at
treatment. Teratology 5, 103–118 (1972).
Shum, A. S. W. et al. Retinoic acid induces down-regulation of
Wnt-3a, apoptosis and diversion of tail bud cells to a neural
fate in the mouse embryo. Mech. Dev. 84, 17–30 (1999).
Tibbles, L. & Wiley, M. J. A comparative study of the effects
of retinoic acid given during the critical period for inducing
spina bifida in mice and hamsters. Teratology 37, 113–125
(1988).
Jones-Villeneuve, E. M. V., McBurney, M. W., Rogers, K. A.
& Kalnins, V. I. Retinoic acid induces embryonal carcinoma
cells to differentiate into neurons and glial cells. J. Cell Biol.
94, 253–262 (1982).
McBurney, M. W., Jones-Villeneuve, E. M. V., Edwards,
M. K. S. & Anderson, P. J. Control of muscle and neuronal
differentiation in a cultured embryonal carcinoma cell line.
Nature 299, 165–167 (1982).
Maden, M. Role and distribution of retinoic acid during CNS
development. Int. Rev. Cytol. 209, 1–77 (2001).
Duester, G. Families of retinoid dehydrogenases regulating
vitamin A function. Production of visual pigment and retinoic
acid. Eur. J. Biochem. 267, 4315–4324 (2000).
Fujii, H. et al. Metabolic inactivation of retinoic acid by a
novel P450 differentially expressed in developing mouse
embryos. EMBO J. 16, 4163–4173 (1997).
White, J. A. et al. Identification of the retinoic acid-inducible
all-trans-retinoic acid 4-hydroxylase. J. Biol. Chem. 271,
29922–29927 (1996).
White, J. A. et al. Identification of the human cytochrome
P450, P450RAI-2, which is predominantly expressed in the
adult cerebellum and is responsible for all-trans-retinoic acid
metabolism. Proc. Natl Acad. Sci. USA 97, 6403–6408 (2000).
Sonneveld, E., van den Brink, C. E., Tertoolen, L. G. J.,
van der Burg, B. & van der Saag, P. T. Retinoic acid
hydroxylase (CYP26) is a key enzyme in neuronal
differentiation of embryonal carcinoma cells. Dev. Biol. 213,
390–404 (1999).
Niederreither, K. et al. Genetic evidence that oxidative
derivatives of retinoic acid are not involved in retinoid signaling
during mouse development. Nature Genet. 31, 84–88 (2002).
Kastner, P., Chambon, P. & Leid, M. in Vitamin A in Health
and Disease (ed. Blomhoff, R.) 189–238 (Marcel Dekker,
New York, 1994).
Kliewer, S. A., Umesono, K., Evans, R. M. & Mangelsdorf,
D. J. in Vitamin A in Health and Disease (ed. Blomhoff, R.)
239–255 (Marcel Dekker, New York, 1994).
Sharpe, C. & Goldstone, K. The control of Xenopus
embryonic primary neurogenesis is mediated by retinoid
signalling in the neurectoderm. Mech. Dev. 91, 69–80 (2000).
Franco, P. G., Paganelli, A. R., Lopez, S. L. & Carrasco, A. E.
Functional association of retinoic acid and hedgehog
signaling in Xenopus primary neurogenesis. Development
126, 4257–4265 (1999).
This paper considers the involvement of RA in primary
neurogenesis, and how prepattern genes, proneural
genes and neural differentiation genes are linked
together by a cascade that starts with RA.
Papalopulu, N. & Kintner, C. A posteriorising factor, retinoic
acid, reveals that anteroposterior patterning controls the
timing of neuronal differentiation in Xenopus neuroectoderm.
Development 122, 3409–3418 (1996).
Sharpe, C. R. & Goldstone, K. Retinoid receptors promote
primary neurogenesis in Xenopus. Development 124,
515–523 (1997).
which a failure of RA signalling has been suggested to be
an underlying cause, and research over the next few
years should be directed towards investigating these
exciting possibilities.
25. Sharpe, C. & Goldstone, K. Retinoid signalling acts during
the gastrula stages to promote primary neurogenesis. Int. J.
Dev. Biol. 44, 463–470 (2000).
26. Blumberg, B. et al. An essential role for retinoid signalling in
anteroposterior neural patterning. Development 124,
373–379 (1997).
An analysis of the effects of dominant-negative and
constitutively active RARs on the AP patterning of
gene expression in the Xenopus CNS.
27. Bertrand, N., Castro, D. S. & Guillemot, F. Proneural genes
and the specification of neural cell types. Nature Rev.
Neurosci. 3, 517–530 (2002).
28. Gomez-Skarmeta, J., Glavic, A., de la Calle-Mustienes, E.,
Modolell, J. & Mayor, R. Xiro, a Xenopus homolog of the
Drosophila Iroquois complex genes, control development at
the neural plate. EMBO J. 17, 181–190 (1998).
29. Cho, K. W. Y. & De Robertis, E. M. Differential activation of
Xenopus homeobox genes by mesoderm-inducing growth
factors and retinoic acid. Genes Dev. 4, 1910–1916 (1990).
30. Dekker, E.-J. et al. Xenopus Hox-2 genes are expressed
sequentially after the onset of gastrulation and are differentially
inducible by retinoic acid. Dev. Suppl., 195–202 (1992).
31. Durston, A. J. et al. Retinoic acid causes an anteroposterior
transformation in the developing nervous system. Nature
340, 140–144 (1989).
A study of the effect of excess RA on Xenopus
embryos, showing a CNS action that reignited interest
in RA and CNS development.
32. Leroy, P. & De Robertis, E. M. Effects of lithium chloride and
retinoic acid on the expression of genes from the Xenopus
Hox 2 complex. Dev. Dyn. 194, 21–32 (1992).
33. Lopez, S. L. & Carrasco, A. E. Retinoic acid induces
changes in the localization of homeobox proteins in the
antero-posterior axis of Xenopus laevis embryos. Mech.
Dev. 36, 153–164 (1992).
34. Pannese, M. et al. The Xenopus homologue of Otx2 is a
maternal homeobox gene that demarcates and specifies
anterior body regions. Development 121, 707–720 (1995).
35. Sive, H. L., Draper, B. W., Harland, R. M. & Weintrub, H.
Identification of a retinoic acid-sensitive period during
primary axis formation in Xenopus laevis. Genes Dev. 4,
932–942 (1990).
36. Creech-Kraft, J., Schuh, T., Juchau, M. & Kimelman, D. The
retinoid X receptor ligand, 9-cis-retinoic acid, is a potential
regulator of early Xenopus development. Proc. Natl Acad.
Sci. USA 91, 3067–3071 (1994).
37. Drysdale, T. A. & Crawford, M. J. Effects of localized
application of retinoic acid on Xenopus laevis development.
Dev. Biol. 162, 394–401 (1994).
38. Kolm, P. J. & Sive, H. L. Regulation of the Xenopus labial
homeodomain genes, HoxA1 and HoxD1: activation by
retinoids and peptide growth factors. Dev. Biol. 167, 34–49
(1995).
39. Pijnappel, W. W. M. et al. The retinoid ligand 4-oxo-retinoic
acid is a highly active modulator of positional specification.
Nature 366, 340–344 (1993).
40. Saha, M. S., Michel, R. B., Gulding, K. M. & Grainger, R. M.
A Xenopus homeobox gene defines dorsal–ventral domains
in the developing brain. Development 118, 193–202 (1993).
41. Taira, M., Otani, H., Jamrich, M. & Dawid, I. B. Expression of
the LIM class homeobox gene Xlim-1 in pronephros and CNS
cell lineages of Xenopus embryos is affected by retinoic acid
and exogastrulation. Development 120, 1525–1536 (1994).
42. von Bubnoff, A., Schmidt, J. E. & Kimelman, D. The
Xenopus laevis homeobox gene Xgbx-2 is an early marker
of anteroposterior patterning in the ectoderm. Mech. Dev.
54, 149–160 (1995).
43. Maden, M., Gale, E., Horton, C. & Smith, J. C. in Retinoids
in Normal Development and Teratogenesis (ed. Morriss-Kay,
G. M.) 119–134 (Oxford Univ. Press, Oxford, UK, 1992).
44. Zhang, Z., Balmer, J. E., Lovlie, A., Fromm, S. H. &
Blomhoff, R. Specific teratogenic effects of different retinoic
acid isomers and analogs in the developing anterior
central nervous system of zebrafish. Dev. Dyn. 206, 73–86
(1996).
45. Avantaggiato, V., Acampora, D., Tuorto, F. & Simeone, A.
Retinoic acid induces stage-specific repatterning of the rostral
central nervous system. Dev. Biol. 175, 347–357 (1996).
46. Cunningham, M. L., MacAuley, A. & Mirkes, P. E. From
gastrulation to neurulation: transition in retinoic acid
sensitivity identifies distinct stages of neural patterning in the
rat. Dev. Dyn. 200, 227–241 (1994).
47. Simeone, A. et al. Retinoic acid induces stage-specific
| NOVEMBER 2002 | VOLUME 3
48.
49.
50.
51.
52.
53.
54.
55.
56.
57.
58.
59.
60.
61.
62.
63.
64.
65.
66.
67.
68.
69.
antero-posterior transformation of rostral central nervous
system. Mech. Dev. 51, 83–98 (1995).
Agarwal, V. R. & Sato, S. M. Retinoic acid affects central
nervous system development of Xenopus by changing cell
fate. Mech. Dev. 44, 167–173 (1993).
Ruiz i Altaba, A. & Jessell, T. M. Retinoic acid modifies the
pattern of cell differentiation in the central nervous system of
neurula stage Xenopus embryos. Development 112,
945–958 (1991).
Ruiz i Altaba, A. & Jessell, T. Retinoic acid modifies
mesodermal patterning in early Xenopus embryos. Genes
Dev. 5, 175–187 (1991).
Sive, H. L. & Cheng, P. F. Retinoic acid perturbs the expression
of Xhox.lab genes and alters mesodermal determination in
Xenopus laevis. Genes Dev. 5, 1321–1332 (1991).
Taira, M., Saint-Jeannet, J.-P. & Dawid, I. B. Role of the Xlim-1
and Xbra genes in anteroposterior patterning of neural tissue
by the head and trunk organiser. Proc. Natl Acad. Sci. USA
94, 895–900 (1997).
Dekker, E.-J. et al. Overexpression of a cellular retinoic acid
binding protein (xCRABP) causes anteroposterior defects in
developing Xenopus embryos. Development 120, 973–985
(1994).
van der Wees, J. et al. Inhibition of retinoic acid receptormediated signalling alters positional identity in the
developing hindbrain. Development 125, 545–556 (1998).
Hollemann, T., Chen, Y., Grunz, H. & Pieler, T. Regionalized
metabolic activity establishes boundaries of retinoic acid
signalling. EMBO J. 17, 7361–7372 (1998).
Godsave, S. F. et al. Graded retinoid responses in the
developing hindbrain. Dev. Dyn. 213, 39–49 (1998).
Holder, N. & Hill, J. Retinoic acid modifies development of
the midbrain–hindbrain border and affects cranial ganglion
formation in zebrafish embryos. Development 113,
1159–1170 (1991).
Lopez, S. L., Dono, R., Zeller, R. & Carrasco, A. E.
Differential effects of retinoic acid and a retinoid antagonist
on the spatial distribution of the homeoprotein Hoxb-7 in
vertebrate embryos. Dev. Dyn. 204, 457–471 (1995).
Papalopulu, N. et al. Retinoic acid causes abnormal
development and segmental patterning of the anterior
hindbrain in Xenopus embryos. Development 113,
1145–1158 (1991).
Sundin, O. & Eichele, G. An early marker of axial pattern in
the chick embryo and its respecification by retinoic acid.
Development 114, 841–852 (1992).
Lee, Y. M. et al. Retinoic acid stage-dependently alters the
migration pattern and identity of hindbrain neural crest cells.
Development 121, 825–837 (1995).
Leonard, L., Horton, C., Maden, M. & Pizzey, J. A.
Anteriorization of CRABP-I expression by retinoic acid in the
developing mouse central nervous system and its
relationship to teratogenesis. Dev. Biol. 168, 514–528 (1995).
Morriss-Kay, G. M., Murphy, P., Hill, R. E. & Davidson, D. R.
Effects of retinoic acid excess on expression of Hox-2.9 and
Krox-20 and on morphological segmentation in the hindbrain
of mouse embryos. EMBO J. 10, 2985–2995 (1991).
Morriss, G. M. Morphogenesis of the malformations induced
in rat embryos by maternal hypervitaminosis A. J. Anat. 113,
241–250 (1972).
Wood, H., Pall, G. & Morriss-Kay, G. Exposure to retinoic acid
before or after the onset of somitogenesis reveals separate
effects on rhombomeric segmentation and 3′ HoxB gene
expression domains. Development 120, 2279–2285 (1994).
Conlon, R. A. & Rossant, J. Exogenous retinoic acid rapidly
induces anterior ectopic expression of murine Hox-2 genes
in vivo. Development 116, 357–368 (1992).
A thorough analysis of the effect of RA on the
induction of ectopic Hox gene expression in mouse
embryos, relating it to the concentration, time and
position of the gene in the Hox cluster.
Mallo, M. & Brandlin, I. Segmental identity can change
independently in the hindbrain and rhombencephalic neural
crest. Dev. Dyn. 210, 146–156 (1997).
Hill, J., Clarke, J. D. W., Vargesson, N., Jowett, T. & Holder, N.
Exogenous retinoic acid causes specific alterations in the
development of the midbrain and hindbrain of the zebrafish
embryo including positional respecification of the Mauthner
neuron. Mech. Dev. 50, 3–16 (1995).
Marshall, H. et al. Retinoic acid alters hindbrain Hox code
and induces transformation of rhombomeres 2/3 into a 4/5
identity. Nature 360, 737–741 (1992).
This study shows the remarkable effect of RA on
www.nature.com/reviews/neuro
© 2002 Nature Publishing Group
REVIEWS
70.
71.
72.
73.
74.
75.
76.
77.
78.
79.
80.
81.
82.
83.
84.
85.
86.
87.
88.
89.
90.
mouse embryos — transforming rhombomeres into a
different fate (r2 and r3 into r4 and r5).
Manns, M. & Fritzsch, B. Retinoic acid affects the
organization of reticulospinal neurons in developing
Xenopus. Neurosci. Lett. 139, 253–256 (1992).
Kessel, M. Reversal of axonal pathways from rhombomere 3
correlates with extra Hox expression domains. Neuron 10,
379–393 (1993).
Gale, E., Zile, M. & Maden, M. Hindbrain respecification in
the retinoid-deficient quail. Mech. Dev. 89, 43–54 (1999).
Maden, M., Gale, E., Kostetskii, I. & Zile, M. Vitamin Adeficient quail embryos have half a hindbrain and other
neural defects. Curr. Biol. 6, 417–426 (1996).
This paper shows that a complete lack of RA in the
embryo results in loss of the posterior hindbrain
rhombomeres, as well as other neural defects.
White, J. C. et al. Defects in embryonic hindbrain
development and fetal resorption resulting from vitamin A
deficiency in the rat are prevented by feeding
pharmacological levels of all-trans-retinoic acid. Proc. Natl
Acad. Sci. USA 95, 13459–13464 (1998).
White, J. C., Highland, M., Kaiser, M. & Clagett-Dame, M.
Vitamin A deficiency results in the dose-dependent acquisition
of anterior character and shortening of the caudal hindbrain
of the rat embryo. Dev. Biol. 220, 263–284 (2000).
Niederreither, K., Subbarayan, V., Dolle, P. & Chambon, P.
Embryonic retinoic acid synthesis is essential for early
mouse post-implantation development. Nature Genet. 21,
444–448 (1999).
This study locates the source of RA for hindbrain
development in the paraxial mesoderm, showing that
the Raldh2 mutant mouse has the same hindbrain
defects as animals that are subject to complete
vitamin A deficiency.
Niederreither, K., Vermot, J., Schubaur, B., Chambon, P. &
Dolle, P. Retinoic acid synthesis and hindbrain patterning in
the mouse embryo. Development 127, 75–85 (2000).
Begemann, G., Schilling, T. F., Rauch, G.-J., Geisler, R. &
Ingham, P. W. The zebrafish neckless mutation reveals a
requirement for raldh2 in mesodermal signals that pattern
the hindbrain. Development 128, 3081–3094 (2001).
Grandel, H. et al. Retinoic acid signalling is the zebrafish
embryo is necessary during pre-segmentation stages to
pattern the anterior–posterior axis of the CNS and to induce
a pectoral fin bud. Development 129, 2851–2865 (2002).
Dupe, V. & Lumsden, A. Hindbrain patterning involves
graded responses to retinoic acid signalling. Development
128, 2199–2208 (2001).
This paper shows that the hindbrain responds in a
graded way to loss of RA by gradually losing
rhombomeres, rather than in an all-or-nothing manner.
Dupe, V., Ghyselinck, N., Wendling, O., Chambon, P. &
Mark, M. Key roles of retinoic acid receptors α and β in the
patterning of the caudal hindbrain, pharyngeal arches and
otocyst in the mouse. Development 126, 5051–5059 (1999).
Wendling, O., Ghyselinck, N., Chambon, P. & Mark, M.
Roles of retinoic acid receptors in early embryonic
morphogenesis and hindbrain patterning. Development
128, 2031–2038 (2001).
Different combinations of RAR knockouts result in
different types of hindbrain defect in mouse embryos.
Gavalas, A. ArRAnging the hindbrain. Trends Neurosci. 25,
61–64 (2002).
Chen, Y.-P., Huang, L. & Solursh, M. A concentration
gradient of retinoids in the early Xenopus laevis embryo.
Dev. Biol. 161, 70–76 (1994).
Horton, C. & Maden, M. Endogenous distribution of
retinoids during normal development and teratogenesis in
the mouse embryo. Dev. Dyn. 202, 312–323 (1995).
Maden, M., Sonneveld, E., van der Saag, P. T. & Gale, E.
The distribution of endogenous retinoic acid in the chick
embryo: implications for developmental mechanisms.
Development 125, 4133–4144 (1998).
A detailed analysis of endogenous retinoids in the
chick embryo, showing the anteroposterior boundary
of RA at very early stages, as well as data on later
stages.
Ang, H. L., Deltour, L., Hayamizu, T. F., Zgombic-Knight, M.
& Duester, G. Retinoic acid synthesis in mouse embryos
during gastrulation and craniofacial development linked to
class IV alcohol dehydrogenase gene expression. J. Biol.
Chem. 271, 9526–9534 (1996).
LaMantia, A. S., Colbert, M. C. & Linney, E. Retinoic acid
induction and regional differentiation prefigure olfactory
pathway formation in the mammalian forebrain. Neuron 10,
1035–1048 (1993).
Wagner, M., Han, B. & Jessell, T. M. Regional differences in
retinoid release from embryonic neural tissue detected by an
in vitro reporter assay. Development 116, 55–66 (1992).
Balkan, W., Colbert, M., Bock, C. & Linney, E. Transgenic
indicator mice for studying activated retinoic acid receptors
91.
92.
93.
94.
95.
96.
97.
98.
99.
100.
101.
102.
103.
104.
105.
106.
107.
108.
109.
110.
111.
112.
during development. Proc. Natl Acad. Sci. USA 89,
3347–3351 (1992).
Mendelsohn, C., Ruberte, E., LeMeur, M., Morriss-Kay, G. &
Chambon, P. Developmental analysis of the retinoic acidinducible RAR-β2 promoter in transgenic animals.
Development 113, 723–734 (1991).
Reynolds, K., Mezey, E. & Zimmer, A. Activity of the
β-retinoic acid receptor promoter in transgenic mice.
Mech. Dev. 36, 15–29 (1991).
Rossant, J., Zirngibl, R., Cado, D., Shago, M. & Giguere, V.
Expression of a retinoic acid response element–hsplacZ
transgene defines specific domains of transcriptional activity
during mouse embryogenesis. Genes Dev. 5, 1333–1344
(1991).
Shen, S., van den Brink, C. E., Kruijer, W. & van der Saag, P. T.
Embryonic stem cells stably transfected with mRARb2–lacZ
exhibit specific expression in chimeric embryos. Int. J. Dev.
Biol. 36, 465–476 (1992).
Niederreither, K., McCaffery, P., Drager, U. C., Chambon, P.
& Dolle, P. Restricted expression and retinoic acid-induced
downregulation of the retinaldehyde dehydrogenase type 2
(RALDH-2) gene during mouse development. Mech. Dev.
62, 67–78 (1997).
Swindell, E. C. et al. Complementary domains of retinoic
acid production and degradation in the early chick embryo.
Dev. Biol. 216, 282–296 (1999).
This study shows that CYP26A1 and RALDH2 form
complementary domains to potentially generate a
source/sink diffusion gradient of RA across a field of
cells that constitute the presumptive hindbrain.
Maden, M. Heads or tails? Retinoic acid will decide.
Bioessays 21, 809–812 (1999).
Grapin-Botton, A., Bonnin, M.-A., Sieweke, M. & Le Douarin,
N. M. Defined concentrations of a posteriorizing signal are
critical for MafB/Kreisler segmental expression in the
hindbrain. Development 125, 1173–1181 (1998).
MacLean, G. et al. Cloning of a novel retinoic-acid
metabolizing cytochrome P450, Cyp26B1, and comparative
expression analysis with Cyp26A1 during early murine
development. Mech. Dev. 107, 195–201 (2001).
Berggren, K., McCaffery, P., Drager, U. & Forehand, C. J.
Differential distribution of retinoic acid synthesis in the
chicken embryo as determined by immunolocalization of the
retinoic acid synthetic enzyme, RALDH-2. Dev. Biol. 210,
288–304 (1999).
Mic, F. A., Haselbeck, R. J., Cuenca, A. E. & Duester, G.
Novel retinoic acid generating activities in the neural tube
and heart identified by conditional rescue of Raldh2 null
mutant mice. Development 129, 2271–2282 (2002).
Solomin, L. et al. Retinoid-X receptor signalling in the
developing spinal cord. Nature 395, 398–402 (1998).
Grapin-Botton, A., Bonnin, M.-A. & Le Douarin, N. M. Hox
gene induction in the neural tube depends on three
parameters: competence, signal supply and paralogue
group. Development 124, 849–859 (1997).
Itasaki, N., Sharpe, J., Morrison, A. & Krumlauf, R.
Reprogramming Hox expression in the vertebrate hindbrain:
influence of paraxial mesoderm and rhombomere
transposition. Neuron 16, 487–500 (1996).
Gould, A., Itasaki, N. & Krumlauf, R. Initiation of
rhombomeric Hoxb4 expression requires induction by
somites and a retinoid pathway. Neuron 21, 39–51 (1998).
La Mantia, A.-S. Forebrain induction, retinoic acid, and
vulnerability to schizophrenia: insights from molecular and
genetic analysis in developing mice. Biol. Psychiatry 46,
19–30 (1999).
Abu-Abed, S. S. et al. The retinoic acid-metabolising
enzyme, CYP26A1, is essential for normal hindbrain
patterning, vertebral identity, and development of posterior
structures. Genes Dev. 15, 226–240 (2001).
Sakai, Y. et al. The retinoic acid-inactivating enzyme CYP26
is essential for establishing an uneven distribution of retinoic
acid along the anterio-posterior axis within the mouse
embryo. Genes Dev. 15, 213–225 (2001).
Pierani, A., Brenner-Morton, S., Chiang, C. & Jessell, T. M.
A sonic hedgehog-independent, retinoid-activated pathway
of neurogenesis in the ventral spinal cord. Cell 97, 903–915
(1999).
Liu, J.-P., Laufer, E. & Jessell, T. M. Assigning the positional
identity of spinal motor neurons: rostrocaudal patterning of
Hox-c expression by FGFs, Gdf11, and retinoids. Neuron
32, 997–1012 (2001).
Zhao, D. et al. Molecular identification of a major retinoic
acid-synthesising enzyme, a retinaldehyde-specific
dehydrogenase. Eur. J. Biochem. 240, 15–22 (1996).
Sockanathan, S. & Jessell, T. M. Motor neuron-derived
retinoid signalling specifies the subtype identity of spinal
motor neurons. Cell 94, 503–514 (1998).
This study reveals the role of RA in generating a
subset of motor neurons by the paracrine action of
RALDH2 in the ventral horn.
NATURE REVIEWS | NEUROSCIENCE
113. McCaffery, P. & Drager, U. C. Hot spots of retinoic acid
synthesis in the developing spinal cord. Proc. Natl Acad.
Sci. USA 91, 7194–7197 (1994).
This paper reveals the presence of a RA-synthesizing
enzyme (RALDH1) in a specific set of neurons in the
adult brain, and shows that these neurons are involved
in Parkinson’s disease. Could loss of RALDH1 be an
aetiological factor in Parkinson’s disease?
114. Ensini, M., Tsuchida, T., Betling, H.-G. & Jessell, T. The
control of rostrocaudal pattern in the developing spinal cord:
specification of motor neuron subtype identity is initiated by
signals from paraxial mesoderm. Development 125,
969–982 (1998).
115. Cullingford, T. E. et al. Distribution of mRNAs encoding the
peroxisome proliferator-activated receptor α, β, and γ and
the retinoid X receptor α, β, and γ in rat central nervous
system. J. Neurochem. 70, 1366–1375 (1998).
116. Krezel, W., Kastner, P. & Chambon, P. Differential expression
of retinoid receptors in the adult mouse central nervous
system. Neuroscience 89, 1291–1300 (1999).
117. Zetterstrom, R. H., Simon, A., Giacobini, M. M. J., Eriksson,
U. & Olson, L. Localization of cellular retinoid-binding
proteins suggests specific roles for retinoids in the adult
central nervous system. Neuroscience 62, 899–918 (1994).
118. Zetterstrom, R. H. et al. Role of retinoids in the CNS:
differential expression of retinoid binding protein and
receptors and evidence for presence of retinoic acid. Eur. J.
Neurosci. 11, 407–416 (1999).
119. McCaffery, P. & Drager, U. C. High levels of a retinoic acidgenerating dehydrogenase in the meso-telencephalic
dopamine system. Proc. Natl Acad. Sci. USA 91,
7772–7776 (1994).
120. Farooqui, S. M. Induction of adenylyl cyclase sensitive
dopamine D2-receptors in retinoic acid induced
differentiated human neuroblastoma SHSY-5Y cells. Life Sci.
55, 1887–1893 (1994).
121. Samad, T. A., Krezel, W., Chambon, P. & Borrelli, E.
Regulation of dopaminergic pathways by retinoids: activation
of the D2 receptor promoter by members of the retinoic acid
receptor-retinoid X receptor family. Proc. Natl Acad. Sci. USA
94, 14349–14354 (1997).
122. Valdenaire, O., Maus-Moatti, M., Vincent, J. D., Mallet, J. &
Vernier, P. Retinoic acid regulates the developmental
expression of dopamine D2 receptors in rat striatal primary
cultures. J. Neurochem. 71, 929–936 (1998).
123. Krezel, W. et al. Impaired locomotion and dopamine
signalling in retinoid receptor mutant mice. Science 279,
863–867 (1998).
124. Malaspina, A., Kaushik, N. & De Belleroche, J. Differential
expression of 14 genes in amyotrophic lateral sclerosis
spinal cord detected using gridded cDNA arrays.
J. Neurochem. 77, 132–145 (2001).
125. Corcoran, J., So, P.-L. & Maden, M. Absence of retinoids
can induce motoneuron disease in the adult rat and a retinoid
defect is present in motoneuron disease patients. J. Cell Sci.
(in the press).
126. Chiang, M.-Y. et al. An essential role for retinoid receptors
RARβ and RXRγ in long-term potentiation and depression.
Neuron 21, 1353–1361 (1998).
127. Goodman, A. B. Three independent lines of evidence
suggest retinoids as causal to schizophrenia. Proc. Natl
Acad. Sci. USA 95, 7240–7244 (1998).
128. Goodman, A. B. Chromosomal locations and modes of
action of genes of the retinoid (vitamin A) system support
their involvement in the etiology of schizophrenia. Am. J.
Med. Genet. 60, 335–348 (1995).
129. Schneider, R. A., Hu, D., Rubenstein, J. L. R., Maden, M. &
Helms, J. A. Local retinoid signalling coordinates forebrain
and facial morphogenesis by maintaining FGF8 and SHH.
Development 128, 2755–2767 (2001).
Online links
DATABASES
The following terms in this article are linked online to:
LocusLink: http://www.ncbi.nlm.nih.gov/LocusLink/
ADHs | CRABP1 | CRABP2 | CRBP1 | CRBP2 | CYP26A1 |
CYP26B1 | Eph | MafB | Meis2 | RALDH1 | RALDH2 | RALDH3 |
RARα | RARβ | RARγ | RoDHs | RXRα | RXRβ | RXRγ
OMIM: http://www.ncbi.nlm.nih.gov/Omim/
Parkinson’s disease | schizophrenia
Swiss-Prot: http://ca.expasy.org/sprot/
CEK8 | chordin | En2 | follistatin | Gli3 | Hoxb3 | Hoxb4 | Hoxb9 |
Krox20 | noggin | Otx2 | Pax2 | Pax6 | XASH-3 | X-delta-1 |
X-MyT1 | X-ngnr-1 | X-shh | Zic2
FURTHER INFORMATION
Encyclopedia of Life Sciences: http://www.els.net/
vertebrate central nervous system: pattern formation
Access to this interactive links box is free online.
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