1 Carnegie Institution of Washington, Department of Embryology, 115 W.
University Parkway, Baltimore, MD, 21210, USA
2 IGBMC, CNRS/INSERM/ULP, 1 rue Laurent Fries, BP10142, CU de Strasbourg, 67404
Illkirch cedex, France
* Author for correspondence (e-mail: halpern{at}ciwemb.edu)
Accepted 26 November 2002
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SUMMARY |
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Key words: Habenula, Epithalamus, Epiphysis, Pineal organ, Brain laterality
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INTRODUCTION |
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Left-right (LR) asymmetries have also been described in the forebrains of
lower vertebrates. Examples include the left hemisphere specialization of
rodents in the production and perception of vocalizations
(Ehret, 1987;
LeMay, 1999
), and the
preferential eye use in food-searching behavior and predator recognition
exhibited by amphibians, fish and birds
(Deng and Rogers, 1997
;
Miklosi et al., 2001
;
Miklosi et al., 1998
;
Rogers, 2000
;
Vallortigara et al., 1998
).
The wide phylogenetic distribution of asymmetric behaviors suggests that
functional lateralization of the brain is not a novel feature of the human
cerebral cortex, but arose early in vertebrate evolution. Thus, it has been
argued that asymmetry of the brain is a consequence of the cortex evolving
from an already asymmetric neural template
(Trevarthen, 1996
).
The dorsal diencephalon displays striking anatomical asymmetries in many
species (reviewed by Concha and Wilson,
2001), providing a promising entry point to study the
developmental origin of forebrain laterality. The pineal complex of fish,
reptiles and amphibians consists of the pineal organ (or epiphysis) and an
unpaired accessory organ (the parapineal, parietal eye or frontal organ,
respectively), which is frequently situated on the left side of the brain. In
several species of fish, including stickleback, lamprey and trout, the
parapineal preferentially innervates the left dorsal habenula
(van Veen et al., 1980
;
Yanez and Anadon, 1994
;
Yanez and Anadon, 1996
), one
of a bilateral pair of brain nuclei that directly neighbor the pineal complex.
The left and right dorsal (or designated medial for some species) habenular
nuclei of several amphibians and reptiles show notable differences in their
size, structure and molecular composition. In the frogs Rana
esculenta and Rana temporaria, for example, the medial habenula
consists of two compartments on the left side and only one on the right
(Kemali and Guglielmotti,
1977
; Morgan et al.,
1973
). Ultrastructural asymmetries have also been noted in the
habenular nuclei, including LR differences in cell morphology and packing
density, synaptic vesicle specializations and neuropil organization
(Kemali and Guglielmotti,
1977
). Accordingly, anti-acetylated tubulin labeling of the larval
zebrafish brain reveals a denser region of neuropil in the left dorsal
habenula compared with the right (Concha
et al., 2000
). The major efferent fiber tract projecting from the
dorsal/medial habenula, the fasciculus retroflexus, shows LR differences in
its cross-sectional diameter in the brains of some fish
(Braford and Northcutt, 1983
).
In addition, the habenulae can be distinguished by their biochemical
properties: the left habenula of dogfish expresses the Ca2+-binding
protein calbindin more abundantly
(Rodriguez-Moldes et al.,
1990
) and, in Rana esculenta, it exhibits higher nitric
oxide synthetase activity than the right
(Guglielmotti and Fiorino,
1999
). LR differences in levels of monoamine neurotransmitters
have also been observed (Ekstrom and
Ebbesson, 1988
).
The mechanisms that underlie the development of epithalamic asymmetry are
starting to be understood in the zebrafish system, where many molecular and
genetic tools are available. The earliest LR difference detected in this
region of the brain is a transient asymmetry in gene expression found during
embryonic development. Genes that encode components of the Nodal signaling
pathway, including the Nodal-related TGFß signal Cyclops (Cyc/Ndr2), the
Nodal antagonist Antivin/Lefty1 (Lft1) and a Nodal-activated transcription
factor Pitx2, are expressed on the left side of the presumptive pineal organ
during midsomitogenesis (Bisgrove et al.,
2000; Concha et al.,
2000
; Essneretal, 2000; Liang
et al., 2000
; Rebagliati et
al., 1998a
; Sampath et al.,
1998
; Thisse and Thisse,
1999
). Mutations in genes that act at gastrulation can affect the
later gene expression asymmetry of the epithalamus. For example, in no
tail (ntl) and floating head (flh) mutant
embryos, in which formation of the axial mesoderm is perturbed, gene
expression in the pineal anlage is later bilateral rather than left-sided
(Bisgrove et al., 2000
;
Concha et al., 2000
;
Liang et al., 2000
). The
mutation casanova (cas), which affects the initial
differentiation of endoderm and other early embryonic structures
(Alexander et al., 1999
),
causes cyc expression to become LR randomized or bilateral in the
brain (Liang et al., 2000
).
Although the results of such mutant analyses are indirect, they suggest that
LR specification of the zebrafish nervous system is already under way during
gastrulation.
As Nodal signaling is essential for early tissue patterning, and mutants
defective in this signaling pathway are lethal
(Feldman et al., 1998;
Rebagliati et al., 1998b
;
Sampath et al., 1998
;
Zhang et al., 1998
), it is
difficult to evaluate the specific function of the transient, asymmetric gene
expression in the brain. However, it is possible to rescue the early
requirement for Nodal activity using embryos mutated at the one eyed
pinhead (oep) locus, which encodes an essential co-factor for
the reception of Nodal signals. Injection of oep RNA at the one-cell
stage is sufficient to rescue Nodal signaling during gastrulation
(Yan et al., 1999
;
Zhang et al., 1998
), but does
not restore asymmetric gene expression (cyc, lft1 or pitx2)
in the developing brain (Concha et al.,
2000
; Liang et al.,
2000
). Using this approach, it was determined that the Nodal
pathway is required for normal positioning of the pineal organ stalk, which in
the adult zebrafish brain typically emanates from the roof of the diencephalon
with a subtle left bias (Liang et al.,
2000
). In rescued oep mutants, the position of the pineal
stalk becomes displaced along the LR axis
(Liang et al., 2000
), and
parapineal sidedness and the asymmetry in habenular neuropil are LR randomized
(Concha et al., 2000
;
Gamse et al., 2002
). These
findings suggest that transient activation of the Nodal signaling pathway is
involved in regulating the directionality of diencephalic asymmetry.
In the present study, we describe a molecular marker of LR identity of the dorsal habenular nuclei. The leftover (lov) gene is expressed at high levels in many cells of the left dorsal habenula, but at reduced levels and in a smaller number of cells in the right habenula. In mutants with disrupted midline development or defective Nodal signaling, laterality of the lov expression pattern is LR randomized. However, the habenula that shows stronger expression is always found on the same side of the brain as the parapineal. We show that laser-mediated ablation of the parapineal results in the loss of the lov expression asymmetry, with both habenulae adopting the weaker pattern characteristic of the right side. Our data indicate that the LR molecular identity of the habenulae is influenced by the unpaired parapineal organ, suggesting that lateralization of the diencephalon occurs through a step-wise accumulation of asymmetries and biases in cellular interactions.
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MATERIALS AND METHODS |
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To produce the flh:eGFP fish lines, 5.5 kb of genomic DNA sequence upstream of the flh transcriptional start site was fused in frame to DNA encoding enhanced GFP (eGFP; from the EGFP-1 plasmid, Clontech). Linearized DNA was injected into one-cell stage embryos, which were grown to adulthood and assayed for germline transmission of the transgene. Two stable transgenic founder fish were independently isolated. Fluorescent labeling of transgenic embryos resembles endogenous flh expression; however, eGFP is not detected in pineal precursors until the 18-20 somite stage. Labeling in Tg (flh:eGFP)c162 is similar to Tg (flh:eGFP)c161, except for an additional domain of fluorescence in the rostral hindbrain, probably caused by enhancer trapping at the site of transgene integration.
Identification of the lov and cpd2 genes
Individual clones were selected from an adult zebrafish kidney cDNA library
that contained inserts between the EcoRI and XhoI sites of
pBK-CMV (Stratagene). Expression of cDNAs was assayed during embryonic stages
by a whole-mount in situ hybridization screen. Partial f-spondin2
(Higashijima et al., 1997),
lov and cpd2 cDNAs were isolated from this screen. The
theoretical translation of the partial zebrafish Cpd2 cDNA is 63% identical to
mouse Cpd2 (GenBank accession number NP_694803). The 5' end of the
lov transcript was identified by RNA-ligase mediated RT-PCR (RLM-RACE
kit, Ambion) using total RNA from 4 day-old larvae. RLM-RACE products were
subcloned into the pCRII vector (Invitrogen) and sequenced. All products were
identical in sequence, indicating that there is a single transcriptional start
site for lov.
RNA in situ hybridization and immunofluorescence
Whole-mount in situ hybridization was performed as described previously
(Gamse et al., 2002) with
reagents obtained from Roche Molecular Biochemicals. To synthesize antisense
RNA probes, pBK-CMV-lov was linearized with EcoRI and
transcribed with T7 RNA polymerase and pBK-CMV-f-spondin2 and
pBK-CMV-cpd2 were linearized with SalI and transcribed with
T7 RNA polymerase. Probes were labeled with UTP-digoxigenin or UTP-fluorescein
and incubated with embryos at 70°C in hybridization solution containing
50% formamide. Hybridized probes were detected using alkaline
phosphatase-conjugated antibodies and visualized by 4-nitro blue tetrazolium
(NBT) and 5-bromo-4-chloro-3-indolyl-phosphate (BCIP) staining. For
cross-sections, embryos were embedded, sectioned and counterstained as
described previously (Liang et al.,
2000
). For combined in situ hybridization and immunofluorescence,
the lov probe was visualized using the Fast Red (Roche) substrate,
followed by labeling with an anti-acetylated
-tubulin primary antibody
(Sigma) and an Alexa 488-conjugated secondary antibody. Images were captured
using a Kontron ProgRes 3012 or a Zeiss AxioCam HRm camera mounted on a Zeiss
Axiophot microscope, and with a Leica TCS-NT confocal microscope.
Laser ablation
Transgenic embryos were dechorionated, anesthetized with tricaine (Sigma),
and positioned dorsal side upwards in 1.2% molten agar layered on bridged
cover slips. GFP labeling at 28-32 h confirmed parapineal location. Ablation
was performed with the 440 nm beam from a Photonic Instruments MicroPoint
laser system mounted on a Zeiss Axiophot microscope using a 40x water
immersion objective. Laser output was calibrated to the same level for each
experiment. Approximately 15-20 cells were ablated using 5-10 pulses per cell.
Control embryos received a similar treatment, except that the ablated cells
were situated contralateral to the parapineal. Loss of cell integrity during
laser treatment was assessed by a change in contrast under Nomarski
optics.
Accession numbers
GenBank accession numbers are: lov, AY120891; zebrafish
cpd2, AY120892.
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RESULTS |
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LR differences are also found in the dorsal habenular nuclei of larval
zebrafish. The size difference is not as dramatic as in some amphibian brains
(see Harris et al., 1996);
however, the left habenula is typically larger in 4 d zebrafish larvae [an
approximately 18% greater area expressing cerebellum postnatal development
associated protein 2 (cpd2) and other markers; see
Fig. 7B,F and data not shown].
A significant molecular asymmetry was discovered in this region of the
diencephalon by screening the gene expression patterns of random cDNA clones
during embryonic and larval development. The lov gene shows a high
level of expression within a localized domain on the left side of the brain,
but only weak expression and in fewer cells in the corresponding region on the
right side (Fig. 2A). The vast
majority of wild-type embryos (97%, n=232) exhibited this pattern.
Double labeling (Fig. 2C,D,E)
confirmed that the left-sided domain of lov expression corresponded
to the previously described region of the left dorsal habenula associated with
denser neuropil (Concha et al.,
2000
). Expression of lov is restricted to the habenulae
at 4 d, although transient expression in the pituitary anlage and caudal blood
island is found between 1 and 2 d (not shown). In contrast to genes that
encode components of the Nodal signaling pathway, which are expressed
transiently, asymmetric expression of lov persists in the dorsal
habenulae throughout larval development and is retained in the adult
diencephalon (Fig. 2B).
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The lov cDNA encodes a 288 amino acid protein of unknown function
that possesses a conserved domain near the N terminus
(Fig. 2F,G) that is similar to
the oligomerization site of the Shaker voltage-gated K+ channel
(Papazian, 1999). This
protein-protein interaction domain is also found in members of the POZ/BTB
family of transcription factors (Collins
et al., 2001
). The Lov protein shares no other structural features
of ion channel proteins or transcription factors.
Correlation between the parapineal and habenular laterality
Several zebrafish mutations have been shown to perturb LR patterning of the
epithalamic region, including those that affect the development of specific
tissues at gastrulation (i.e. casanova, no tail and floating
head), and those that block the Nodal signaling pathway (i.e.,
one-eyed pinhead, cyclops and schmalspur). Asymmetric
expression of cyc, lft1 and pitx2 in the pineal anlage
becomes bilateral or is absent in the majority of mutant embryos and the
parapineal position is LR randomized
(Bisgrove et al., 2000;
Concha et al., 2000
;
Essner et al., 2000
;
Gamse et al., 2002
;
Liang et al., 2000
;
Rebagliati et al., 1998a
). To
determine whether molecular asymmetry of the dorsal habenulae is similarly
affected, lov expression was examined in cas, ntl, flh and
rescued MZoep mutant larvae. LR reversal of the asymmetric
lov expression pattern was found in approximately 50% of mutant
larvae examined (Table 1; Fig. 3A-H). Less frequently
(3-7% of mutants, Table 1),
equivalent levels of lov expression were observed on both sides of
the brain (Fig. 3I,J).
|
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The temporal relationship between parapineal formation and lov
expression in the diencephalon was examined in 26-48 h embryos at 2 hour
intervals. A distinct parapineal cell cluster could be recognized by 28 h
(Fig. 5). However, lov
transcripts were not detected in the habenular region until 38-40 h and were
asymmetrically distributed from the onset
(Fig. 5). Therefore, the
appearance of the parapineal (as assayed by otx5 expression) precedes
asymmetric lov expression in the dorsal habenular nuclei by 12
hours.
|
LR asymmetry of the dorsal habenulae requires the parapineal
On the basis of their spatial and temporal relationship, the parapineal
could play an instructive role in regulating the LR molecular identity of the
dorsal habenulae. To test this hypothesis, we selectively destroyed the
parapineal by laser-mediated cell ablation. As the parapineal is difficult to
visualize in living embryos, laser ablation was performed using two
independently isolated transgenic lines, which carry the flh promoter
driving expression of the green fluorescent protein reporter gene (GFP).
Transgenic flh:GFP embryos exhibit labeling in the developing midline
and pineal organ (Fig. 6A,B,E,F
and data not shown) similar to endogenous flh expression
(Masai et al., 1997). A
notable exception is the transient appearance of fluorescence in the
parapineal organ, a structure in which flh transcripts have never
been detected (Gamse et al.,
2002
). In 28-30 h transgenic embryos, the GFP-positive parapineal
organ is found anterior and to the left of the pineal anlage
(Fig. 6A,E), consistent with
the position of the emerging otx5-positive parapineal domain
(Fig. 5).
Fluorescent labeling of the parapineal in flh:GFP transgenic larvae served as a useful guide for targeted laser-mediated ablation of this structure. Approximately 15-20 cells corresponding to the labeled parapineal were destroyed in 28-32 h transgenic embryos (Fig. 6C,D) using five to ten pulses of a focused laser beam. The embryos were subsequently allowed to develop to 4 d, at which time ablation of the parapineal was confirmed by the absence of the parapineal-specific otx5 expression domain (refer to Fig. 7J-M). In these larvae, the otx5-expressing pineal anlage appeared normal, indicating that only the parapineal was affected. In control larvae, either a comparable region on the right side of the pineal anlage (n=31; Fig. 6G,H) or cells adjacent to the parapineal were ablated (n=9; data not shown).
The denser neuropil characteristic of the left dorsal habenula was present in control-ablated larvae (Fig. 7A,E). However, this neuropil density was absent in the left habenula of parapineal-ablated larvae (n=12/12, Fig. 7I). To confirm that the habenular nuclei were intact after laser-mediated destruction of the parapineal, we examined expression of two markers. The cpd2 and f-spondin2 genes are transcribed at equal levels in the L and R habenular nuclei. Expression of these genes was unaffected in all parapineal-ablated larvae (n=18/18 for cpd2, 8/8 for f-spondin2, Fig. 7J,K), and indistinguishable from untreated or control ablated larvae (Fig. 7B,C,F,G), indicating that the habenulae were not directly damaged by the laser treatment.
The habenulae of control-ablated and untreated sibling larvae displayed the typical asymmetric pattern of lov expression (n=18/18 for control ablated, Fig. 7D,H). Destruction of the pineal organ, with the parapineal intact, also had no effect on lov expression (n=4/4, Fig. 7N). By contrast, both the left and right habenulae of larvae in which the parapineal organ was selectively destroyed prior to the onset of lov transcription (Fig. 7L,M) showed the pattern of lov expression normally found on the right side (weaker expression in a subset of cells, n=24/24). Thus, elimination of the parapineal specifically abolished the neuropil and molecular LR asymmetry of the adjacent brain nuclei.
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DISCUSSION |
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Progression of LR asymmetry in the zebrafish epithalamus
The epithalamus had previously been identified as a site of molecular and
morphological LR differences in the zebrafish brain. These differences appear
in a progressive manner during embryonic and larval development. The earliest
manifestation of brain asymmetry is the transient expression of components of
the Nodal-related Cyc signaling pathway on the left side of the presumptive
pineal organ during somitogenesis (Bisgrove
et al., 2000; Concha et al.,
2000
; Essner et al.,
2000
; Liang et al.,
2000
; Rebagliati et al.,
1998a
; Sampath et al.,
1998
; Thisse and Thisse,
1999
). Shortly afterwards, the parapineal appears to the left of
the pineal anlage (Gamse et al.,
2002
). Fluorescent labeling of the parapineal in flh:GFP
larvae suggests that it derives from the same precursor population as the
pineal, and results from the persistence of GFP in lineally related cells.
Laser ablation of the pineal anlage prior to parapineal formation supports
this idea (K. Cygnar and J. T. G., unpublished); however, direct fate-mapping
analysis is required to confirm the precise cellular origin of the parapineal.
Adjacent to the pineal complex lie the paired habenular nuclei, which exhibit
LR differences in their size, neuropil density
(Concha et al., 2000
) and (as
shown in this study) in expression of the lov gene as early as 2 d.
Lateralized features of the epithalamus persist throughout larval stages and
are found in the adult brain. The mature pineal organ consists of the
photoreceptive pineal end vesicle and the pineal stalk, with the stalk
emanating from the diencephalic roof at a left-of-medial position
(Liang et al., 2000
). The
otx5-expressing parapineal remains a rudimentary structure at the
base of the pineal stalk adjacent to the left habenular nucleus. In the adult,
the left habenula continues to express lov at high levels relative to
the right side.
Analyses of zebrafish mutants have shed some light on how the LR
differences of the diencephalon are regulated. Although the initial
symmetry-breaking event is unknown, early patterning processes, including the
formation of midline tissues at gastrulation, influence the later laterality
of the epithalamic region. Activation of the Nodal signaling pathway in the
left pineal anlage during segmentation stages appears to be an essential cue
for setting the position of both the parapineal
(Concha et al., 2000) and
pineal outgrowth along the LR axis (Liang
et al., 2000
). In mutants defective in Nodal signaling (or those
that show bilateral activation of the pathway), we presume that this cue is
lost in the developing epithalamus. As a consequence, placement of the pineal
stalk and parapineal is determined by a stochastic mechanism, and at the
populational level, sidedness of the pineal complex becomes LR randomized.
Our data reveal that the lateralized features of the habenular nuclei are
associated with the parapineal. A discrete parapineal cellular cluster is
detected in the left dorsal diencephalon almost 12 hours before the onset of
lov expression in the dorsal habenulae. Notably, lov
expression is asymmetric from the outset, with the stronger left-sided domain
closely apposed to the parapineal. In all mutant and wild-type larvae, the
direction of parapineal sidedness (L versus R) corresponds with the neuropil
density of the dorsal habenulae (left-biased versus right-biased; this study)
(see also Concha et al., 2000)
and the sidedness of the lov expression pattern. Furthermore, in the
few larvae that develop bilateral parapineal organs, both the left and right
habenulae show high levels of lov expression. The findings indicate
that parapineal formation precedes and strictly correlates with habenular
asymmetry.
The parapineal is required for habenular laterality
One interpretation of the data is that the parapineal plays an instructive
role in the generation of habenular asymmetry. An alternative hypothesis is
that habenular asymmetry is not dependent on the parapineal; rather, the
laterality of both structures is coordinately regulated through a common
signal. The latter hypothesis, however, is not supported by the results of
parapineal ablation experiments. In all larvae in which the parapineal was
selectively destroyed before the onset of lov transcription,
equivalent low levels of lov expression were later detected on both
sides of the brain. The loss of lov expression and dense neuropil
characteristic of the left habenula was specific to parapineal ablation.
Moreover, only these asymmetric features were lacking, as other habenular
markers that are normally expressed at equal levels on the right and left were
unaffected. Removal of the pineal anlage had no effect on habenular asymmetry
as long as the left-sided parapineal organ was preserved. The analysis of
flh mutant larvae, which show a greatly reduced pineal anlage
(Masai et al., 1997) and an
intact parapineal (Gamse et al.,
2002
), further suggests that the parapineal is not only necessary
but is sufficient to influence habenular laterality. Whether an earlier
event(s) establishes the LR asymmetry of the habenulae, and the parapineal
serves to maintain or reinforce this lateralization cue(s), remains to be
determined. It is also possible that loss of the parapineal might delay the
appearance of rather than inhibit habenular asymmetry. This is currently under
investigation by examining the persistence of bilateral symmetry in the
habenulae of juvenile and adult fish that have been reared after parapineal
ablation (J. T. G and M. E. H., unpublished).
At present, the nature of the interaction between the parapineal and left
habenula is unknown. Preferential innervation of the left habenula by the
parapineal has been described for some fish species, including trout
(Yanez and Anadon, 1996),
lamprey (Yanez and Anadon,
1994
) and stickleback (van
Veen et al., 1980
). Connectivity between the parapineal and
diencephalon has not yet been rigorously demonstrated for zebrafish, although
axons from opsin-immunoreactive parapineal neurons do selectively project
towards the left habenula (Concha et al.,
2000
). Direct cell-cell signaling is another potential mechanism,
as the close apposition of the developing parapineal and left habenula
cellular domains appears to be essential for generating habenular asymmetry.
Known mutations of candidate signaling molecules, including Ace/Fgf8
(Reifers et al., 1998
),
Syu/Shh (Schauerte et al.,
1998
) and Snh/Bmp7 (Dick et
al., 2000
; Schmid et al.,
2000
), do not disrupt the lov expression pattern in a
specific manner (J. T. G. and M. E. H., unpublished). To determine how the
parapineal exerts its influence on the brain, an unbiased genetic screen is
under way to identify zebrafish mutations that block the parapineal/left
habenula interaction.
Implications of epithalamic asymmetry
On the basis of their work on amphibia, Braitenberg and Kemali
(Braitenberg and Kemali, 1970)
proposed that asymmetry of the parapineal complex imposes a LR bias on the
brain that could extend beyond the habenular region. The major efferent
projection of the medial/dorsal habenular nuclei, the fasciculus retroflexus
(FR), terminates on the unpaired interpeduncular nucleus as part of an
evolutionarily conserved conduction pathway that extends from telencephalic
nuclei to the midbrain (Butler and Hodos,
1996
). The number of axons can differ between the left and right
FR (Braford and Northcutt,
1983
), but this is not a consistent feature of vertebrate brains
(Kemali and Guglielmotti,
1982
). Thus, it will be important to determine whether the
asymmetry of the zebrafish habenulae extends to the axonal tracts of the FR
and influences the laterality of other brain regions.
Epithalamic asymmetry has been described in fishes, amphibians, reptiles
and birds, and in a few mammalian species (see
Concha and Wilson, 2001);
however, the significance of LR differences in this region of the brain is
unknown. Despite a high degree of anatomical conservation
(Sutherland, 1982
), the
habenular region and habenulo-interpeduncular circuit are poorly understood at
the functional level. LR differences in the habenular nuclei have been linked
to seasonal variation, reproductive behavior and sexual dimorphism in some
vertebrates (Harris et al.,
1996
; Bisazza et al.,
1998
). Characterization of the Lov protein, its subcellular
localization, multimeric structure and binding partners may provide insight
into potential asymmetric functions of the habenular nuclei. Preliminary
experiments to deplete Lov protein using morpholino antisense oligonucleotides
have so far been uninformative, but genetic redundancy could be a complicating
factor. The lov gene is a member of a multigene family with
homologues identified so far in frog, mouse, rat and humans (J. T. G., C.
Brösamle and M. E. H., unpublished). Examination of gene expression in
other species will reveal to what extent the lov molecular asymmetry
is preserved in the vertebrate diencephalon.
Many species of fish display lateralized behaviors at both the individual
and populational level (reviewed by
Vallortigara and Bisazza,
2002). Zebrafish, for example, prefer to use the right eye when
fixating on foreign objects and the left eye for viewing familiar ones
(Miklosi and Andrew, 1999
;
Miklosi et al., 1998
),
properties presumed to play a role in feeding, schooling and escaping
behaviors (see Andrew, 2002
).
Adult zebrafish have also been found to show biases in their turning behavior
while swimming (Heuts, 1999
).
Whether there is a connection between epithalamic asymmetry and the
lateralized motor behaviors elicited by the LR discrimination of visual
stimuli remains unclear. Intriguingly, activity of the pineal gland has been
found to modulate the swimming behavior of Xenopus tadpoles under
some conditions (Jamieson and Roberts,
1999
; Jamieson and Roberts,
2000
), although it is not known whether this behavior has a
lateralized component. The molecular genetic approaches afforded by the
zebrafish system will permit the perturbation of epithalamic asymmetry so that
the relevance of this asymmetry can be explored in a behavioral context.
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ACKNOWLEDGMENTS |
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