Branchial expression of an aquaporin 3 (AQP-3) homologue is downregulated in the European eel Anguilla anguilla following seawater acclimation
School of Biology, Bute Medical Buildings, University of St Andrews, St Andrews, Fife KY16 9TS, Scotland
* Author for correspondence (e-mail: cpc{at}st-andrews.ac.uk)
Accepted 7 June 2002
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Summary |
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Key words: fish aquaporin, teleost, urea, ammonia, carbon dioxide, glycerol, messenger RNA, European eel, Anguilla anguilla
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Introduction |
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In teleost fish, water transport plays a crucial role in a number of tissues that work together to maintain body fluid homeostasis. In freshwater (FW) fish, water incursion occurs principally across the surface of the gill, which has a large surface area and significant water permeability. This net branchial water influx is counteracted by the relatively high production of dilute urine by the kidney. In marine teleosts, water loss across the gill or in the form of urine is balanced by drinking the seawater (SW). Water and salts are taken up across the gut to maintain body fluid volume with the excess salts excreted across the gills.
In order to study the mechanisms associated with body fluid homeostasis in
teleosts, the changes occurring during the FW or SW acclimation of euryhaline
species have been extensively investigated. In the European eel (Anguilla
anguilla) most life stages (including FW `yellow' and migratory `silver'
adult eels) are capable of immediate transfer between FW and SW environments
(Tsukamoto et al., 1998;
Birrell et al., 2000
). Mainly
due to this physiological plasticity, much of the information available
concerning epithelial water transport in fish has been determined in eels.
In teleost fish, a major fraction of water exchanges with the external
environment occurs across the gill (Rankin
and Bolis, 1984; Kirsch,
1972
), underlining the importance of this organ to water balance.
The results of a number of experiments have indicated that water transport can
take place across the gill epithelium independently of simple diffusion
through the lipid membranes or movement through the paracellular pathway.
While measurements of branchial diffusional water permeability are similar in
FW- and SW-acclimated eels, measurements made in the presence of an osmotic
gradient (osmotic water permeability) were between three- and 11-fold higher
(depending on temperature) in FW- compared to SW-acclimated fish
(Motais and Isaia, 1972
). In
addition, the osmotic water permeability of FW eels was two- to fourfold
higher than the diffusional permeability, which has been taken as evidence
that the branchial epithelium of FW eels contains (presumably osmotically
activated) `water-filled pores' (Motais et
al., 1969
; Motais and Isaia,
1972
; Isaia,
1972
). As the osmotic water permeability of the branchial
epithelium in SW eels was similar or lower (particularly at low temperatures)
than the diffusional permeability, these factors indicated not only the lack
of pores but also the presence of some unknown water re-absorption system
operating against the osmotic gradient
(Motais and Isaia, 1972
;
Motais and Garcia-Romeu,
1972
). This water transport pathway may or may not be associated
with some some unknown solute-uptake transport system. Further evidence of
regulated branchial water transport comes from the acute transfer of eels
between FW and SW. Under these conditions, net osmotic water fluxes across the
gills are non-symmetrical (rectified) when the osmotic gradient is reversed
(Isaia, 1984
). In FW fish,
gill epithelial cell apical membrane permeability was found to be eightfold
higher than that of basal membranes, suggesting that water `pores' in the
epithelium predominate in this membrane
(Isaia et al., 1978a
).
Adrenaline increases the permeability to water and ions of branchial and
opercular epithelia via a ß-adrenergic receptor pathway
(Haywood et al., 1977; Isaia
et al., 1978a
,
b
; Isaia,
1979
,
1984
;
Zadunaisky, 1984
). The effect
was greater in FW-than SW-acclimated fish (Isaia,
1979
,
1984
). Adrenaline was also
found to effect the permeability of both apical and basal membranes of the
gill epithelium of FW fish (Isaia et al.,
1978a
). Although the evidence is controversial
(Rankin and Bolis, 1984
), the
hormone prolactin probably reduces water influxes across FW eel gills (Ogawa,
1974
,
1975
;
Ogasawara and Hirano, 1984
),
whereas cortisol increases water fluxes
(Ogawa, 1975
;
Rankin and Bolis, 1984
) and
arginine vasotocin (AVT) is without effect
(Rankin and Bolis, 1984
).
The branchial transport/permeability of the gill to urea also parallels
changes in water transport. Depending on the diet, eels excrete between 23-42%
of their nitrogenous waste as urea (Engin
and Carter, 2001) and fish acclimated to FW have a threefold
higher branchial urea clearance rate than SW eels
(Masoni and Payan, 1974
).
Adrenaline at 10-6 mol 1-1 increased both urea and water
permeability by around 100 % (Haywood et
al., 1977
; Isaia et al.,
1978b
), with higher doses (10-5 mol 1-1)
causing much larger increases (400-440 %) in branchial urea efflux
(Bergman et al., 1974
;
Sorenson and Fromm, 1976
).
Studies measuring unidirectional urea fluxes suggest that gill urea transport
occurs via passive diffusion rather than by facilitated transport
processes such as those involving UT type, urea transporters
(Wright et al., 1995
;
Wright and Land, 1998
).
At the initiation of this study, the existence of aquaporins had not been
demonstrated in any fish species, despite having been identified in many other
organisms ranging from bacteria to mammals. However, it seemed possible that
these proteins, which are known water- and urea-transporter/channels, might
represent the branchial `pores' suggested to be present in previous reports.
Aspects of aquaporin structure, function and regulation have been reviewed
extensively (see Hamann et al.,
1998; Borgnia et al.,
1999
; Ma and Verkman,
1999
; Marples et al.,
1999
; van Os et al.,
2000
). In mammals the aquaporins are a large gene family,
currently with 12 members that are related by amino acid homology and genomic
structure, which have been grouped into three broad subfamilies. These include
the `water-selective' aquaporins, comprising AQPs 0-2 and AQPs 4-6 (although
AQP-0 is also permeable to glycerol;
Ishibashi et al., 2000
), the
glyceroaquaporin group, which contains AQPs 3, 7 and 9, which are permeable to
water and/or glycerol and urea (Echevarria
et al., 1996
; Yang and
Verkman, 1997
; Ishibashi et al.,
1997
,
1998
), and finally AQPs 8, X1
and X2, which are an anomalous group of channels with lower amino acid
homology where AQP 8 is permeable to water and/or urea, AQP X1 to water but
not glycerol, and the properties of AQP X2 are unknown
(Ishibashi et al., 2000
).
Many of the mammalian aquaporins such as AQP-3 are expressed in renal
tissues, where much research has been focussed. However, in addition, AQP-3
has also been shown to be expressed in other tissues such as the eye,
digestive, respiratory and urinary tracts as well as in the bladder, spleen,
skin, epidermis and in erythrocytes
(Hamann et al., 1998;
Borgnia et al., 1999
; Matsuzaki
et al., 1999
,
2000
). AQP-3 is expressed on
the basolateral surface of epithelial cells, particularly in tissues
interfacing with the external environment, where it may act to prevent cell
dehydration. In these tissues, expression may be controlled by extracellular
fluid osmolality (Ecelbarger et al.,
1995
; Terris et al.,
1996
; Ishibashi et al.,
1997
; Matsuzaki et al.,
1999
,
2000
). Several hormones have
also been implicated in AQP-3 regulation: both anti-diuretic hormone (ADH) and
corticosteroids may be involved in the long-term regulation of expression of
AQP-3 protein abundance (Terris et al.,
1996
; Tanaka et al.,
1997
). The characteristics of body fluid homeostasis in teleost
fish, and in particular water and urea transport, suggested the existence of a
water and/or urea transport pathway across the eel branchial epithelium. This
putative transport pathway, probably involving one or more transporters, is
likely to be of importance in the control of water balance and/or urea
excretion. This study set out to investigate whether members of the aquaporin
gene family are expressed in aquatic organisms such as the eel. The expression
of these putative water transporters was investigated in both indigenous
freshwater `yellow' and migratory `silver' eels to determine whether
developmental maturation prior to seawater exposure had any regulatory affect
on gene expression.
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Materials and methods |
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Total RNA extraction
RNA used for cloning experiments was extracted by a modified LiCl procedure
as described in Cutler et al.
(1995). RNA for northern
blotting experiments was extracted by a modification of the Chomczynski and
Sacchi method (Chomczynski and Sacchi,
1987
) as described in Cutler et al.
(2000
). Messenger RNA was
purified for reverse transcriptase-polymerase chain reaction (RT-PCR)
experiments as previously described (Cutler
et al., 1995
).
Cloning and sequencing
As no information was available on the possible existence of any aquaporins
present in fish, primers used for amplification were designed based on the
available information in mammalian species. As the presence of direct
counterparts to mammalian aquaporins could not be guaranteed, a more general
approach to primer design had to be taken for the amplification of fish
aquaporins. Unfortunately, the amino acid (aa) homology between mammalian
aquaporins is not very high, which further led to compromises in primer
design, and consequently the primers used could not encompass the complete
range of degeneracy found within the sequence data. Amino acids are referred
to by the single-letter nomenclature. Within aquaporins a major site of
relatively high conservation is around the NPA motifs thought to be associated
with the channel pore (Bill et al.,
2000). The synthetic degenerate primers (MWG Biotech AG,
Ebersberg, Germany), corresponded to approximately 6 aa upstream and up to 2
aa downstream of the NPA motif for the sense primer and 1 aa upstream and up
to 7 aa downstream of the NPA motif for the anti-sense primer. Primers were
designed to be relatively long (34-35mers) to allow a certain amount of
mismatching to be permitted whilst maintaining a reasonably high annealing
temperature (see Table 1).
|
RT-PCR using the degenerate primers
(Table 1) was performed on a
single strand cDNA template prepared using 5 µg of mRNA from the gill of
7-day SW-acclimated yellow eels, as previously described
(Cutler et al., 1995). PCR was
performed using a hot-start technique with an initial 2 min incubation at
92°C, followed by 40 cycles of 94 °C for 1 min, 53 °C for 30 s and
72 °C for 30 s, with a final incubation of 72 °C for 10 min. Reactions
of 20 µl were produced with separate additions of primers (4 µmol
l-1 final concentration) and template (0.5 µl undiluted reverse
transcriptase reaction mix), and initiated with the addition of the master mix
(to give final concentrations of 50 mmol l-1 KCl, 1.5 mmol
l-1 MgCl2 10 mmol l-1 Tris-HCl, pH 9.0, 200
µmol l-1 dNTPs and 1.5 units/20 µl Taq DNA polymerase). DNA
fragments within PCR reactions were then separated by Tris-acetate-EDTA
agarose gel electrophoresis (Sambrook et
al., 1989
) and bands of interest purified using Geneclean II (Bio
101, Carlsbad, California).
Further AQP-3 5'- and 3'-RACE (Rapid Amplification of cDNA
Ends) DNA fragments were produced from SW-acclimated yellow eel gill mRNA and
a Marathon cDNA amplification kit (Clontech, Basingstoke, UK) as described
previously (Cutler et al.,
2000). 5'-RACE products were produced in nested PCR
reactions using eel AQP-3-specific antisense primers 1 and then 2
(Table 1) in conjunction with
the Marathon kit nested primers. PCR fragments generated by the degenerate
primers or by 5'-RACE amplification were cloned into an Original TA
Cloning Kit (Invitrogen, Leek, The Netherlands) and were sequenced from colony
PCR-amplified fragments using a Big Dye Terminator sequencing kit (Perkin
Elmer Biosystems, Warrington, UK) as described previously
(Cutler et al., 2000
). The
3'-RACE nested amplifications generated a series of faint bands with
varying length. Cloning of the largest fragment proved unsuccessful. DNA from
pooled, multiple, nested 3'-RACE PCR amplifications (using Sense primer
3 then 4) was therefore purified directly using Geneclean II and partially
sequenced (Cutler et al.,
2000
) using a further nested specific primer (Sense primer 2). In
order to sequence the reverse strand a further specific primer (3'-RACE
Antisense 1) was synthesised using the initial sequence data obtained.
Northern blotting and analysis
Northern blots were performed as described previously
(Cutler et al., 2000). The
probe used for northern analysis was a colony PCR-amplified plasmid insert of
the original fragment of AQP-3 produced with the degenerate sense and
antisense primers. The amount of total RNA present in each lane of the gel was
determined before blotting using ethidium bromide staining of rRNA quantified
by a gel documentation and analysis system (Syngene, Cambridge, UK).
Quantified values were used to adjust the final radioactive signals obtained
per µg total RNA, from the hybridisation. Quantitative analysis of
radiolabelled AQP-3 DNA probes hybridising to blots was determined by
electronic autoradiography using an Instant Imager (Canberra Packard, Meriden,
CT). Statistical analysis was performed using StatView 4.01 software (Abacus
Concepts, Berkeley, CA, USA). To reduce potential problems with
heteroscedasticity, the data was log10-transformed to reduce the
heterogeneity of variances. The data were analysed using ANOVA with levels of
significance determined using Scheffe's F-test.
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Results |
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Although vertebrate AQP-3 aa sequences are on average reasonably well
conserved, the level of conservation shows considerable heterogeneity along
the aa chain. The most highly conserved regions of the eel AQP-3 homologue in
comparison to AQP-3 sequences from other species occur around the first
putative membrane spanning domain (aa 20-50) and a broad region encompassing
the end of the extracellular loop C
(Borgnia et al., 1999) through
the fifth putative membrane spanning domain and through most of (conserved
second NPA motif containing) loop E (aa 160-230). A number of small conserved
motifs are also distributed throughout the protein. Notable regions of low
conservation include a region from aa 6-16 within the cytoplasmic N-terminal
tail, a region in the middle of the extracellular loop C (aa 127-142), and the
C-terminal tail including the putative sixth transmembrane domain (from aa 237
to the end). The C-terminal tail also contains an insertion of 4 aa in
comparison to other AQP-3 sequences.
Tissue distribution of mRNA expression
Northern blots (Fig. 2)
indicated that high levels of expression were present in the gill of FW eels,
where two distinct mRNA transcripts were evident: a major 2.4 kb band and a
minor 7 kb band, which probably represents an immature precursor or extended
3'-spliceoform of the mature mRNA. Much lower levels of expression were
present in SW-acclimated eel gill, in the intestine of both FW and
SW-acclimated eels, and in the oesophagus and eye of a SW-acclimated eel.
Expression was notably absent in the kidney (SW-acclimated eel) as well as in
other tissues.
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Quantitative mRNA expression in yellow and silver eel gill and
intestine
Quantitative northern blots were used to investigate possible changes in
expression in the three major osmoregulatory organs, namely the gill,
intestine and kidney of both FW or 21-day SW-acclimated yellow and silver eels
(Fig. 3). In the kidney, long
autoradiographic exposures (32 h) were required to visualise bands in FW
yellow and silver eel RNA samples (data not shown). However, the presence of
signals was not consistent between samples. In the intestinal RNA samples,
there was no significant difference in expression between FW-or SW-acclimated
yellow or silver eels (Fig.
4).
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In the gill, significant differences were found in AQP-3 mRNA expression both between yellow and silver eels and following transfer from FW to SW environments. FW silver eels exhibited a 60 % increase in AQP-3 mRNA expression over the FW yellow eels. 3 weeks after transfer to SW there was a marked reduction in AQP-3 expression in both groups of fish, with a much larger decrease in silver (97 %) compared to yellow (76 %) eels. As a result, there was still a sixfold higher level of AQP-3 expression in the yellow compared to the silver SW groups at this time point.
Time course of changes in mRNA expression in yellow eels
The acclimation of yellow eels to SW resulted in a decrease in AQP-3 mRNA
expression in the gill that occurred with a half time of approx. 10 h (Figs
5,
6). AQP-3 expression levels
became significantly reduced 1 day after transfer, compared to the FW to FW
transferred control group. AQP-3 mRNA abundance reached a minimum 2 days post
SW transfer, where the level of expression was reduced by 94 % (compared to
the FW time-matched control). After 7 days, the level of AQP-3 mRNA expression
in SW-transferred fish partially recovered, with transcript abundance
increasing significantly compared to the 2- or 4-day (SW) expression levels.
Consequently, by the end of the time course (21 days), the level of expression
in SW-transferred fish was only reduced by 84 % (compared to FW controls).
There was also an indication that AQP-3 mRNA expression levels in the FW
control fish increased slightly over the time course, although only the 6 hour
time point was significantly different to those at 1 and 21 days.
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Discussion |
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One of the more interesting amino acid sequence features is the loss of the
single putative N-linked glycosylation site in AQP-3s from other species,
where this motif is NGT in other species (at position 141-143), but is KAT in
eel AQP-3. This suggests that the eel AQP-3 may not be glycosylated. The eel
KAT motif is also identical to that found in mouse AQP-7 at the same alignment
position (see EMBL accession no. AB010100). Despite the loss of this site, eel
AQP-3 does have another putative glycosylation site, although this is the
normally conserved first NPA amino acid motif in loop B, which is thought to
form part of the water pore of aquaporin within the membrane
(Borgnia et al., 1999). In eel
AQP-3, the normally conserved alanine residue (in mammalian AQPs) within this
NPA site is changed to a threonine, creating the potential glycosylation site,
although the central role of this motif in aquaporin channel function suggests
that this site is unlikely to be glycosylated. In mammals the only similar
non-conservation of NPA motifs occurs within the second NPA motif region of
AQP-7, where the normally conserved alanine is replaced by a serine (an aa
chemically similar to threonine). The yeast Fps1p channel also has a similar
alanineserine replacement within the first NPA motif site
(Bill et al., 2000
) as in eel
AQP-3. Replacement of the alanine of the first NPA motif of the E.
coli glycero-aquaporin homologue (GlpF) with a serine has been shown to
reduce the glycerol transport of this channel
(Bill et al., 2000
). While the
range of molecules transported by eel AQP-3 and their inherent permeabilities
are currently unknown, this information suggests that the channel
characteristics may well be somewhat different from the AQP-3 homologues of
other species.
The cysteine residue in mammalian AQP-1 (equivalent to the position of aa
211 in eel AQP-3) is thought to be responsible for the mercury sensitivity of
aquaporins, although a cysteine at this position is absent in mammalian AQP-0
(MIP), AQP-3, AQP-4 (MIWC or mercury-insensitive water channel) and AQP-7
(Borgnia et al., 1999). In eel
AQP-3, in common with mammalian AQP-3s and AQP-7s, a tyrosine is substituted
at this site, suggesting that if the eel AQP-3 (and others) is sensitive to
mercurial reagents, this probably is a result of interactions at some other
site.
The tissue distribution of eel AQP-3 was very similar to that found for
AQP-3 in mammals, where it is primarily expressed in tissues exposed to the
external environment such as the eye, respiratory tract (equivalent to the
teleost gill) and digestive tract (Borgnia
et al., 1999; Hamann et al.,
1998
; Matsuzaki et al.,
1999
,
2000
). However, there are one
or two notable exceptions between the tissue distribution of eel and mammalian
AQP-3. Firstly, eel AQP-3 was only expressed in the kidney at a low level and
in an inconsistent fashion between individual fish, whereas in mammals the
kidney is a major site of AQP-3 expression, where it is found in abundance in
renal collecting duct principal cells
(Borgnia et al., 1999
). The
lack of a consistent or high level of renal AQP-3 expression increases the
likelihood of the presence of other aquaporins in eel kidney, and this may
include the presence of a duplicate copy of AQP-3, as has been demonstrated
for other ion transporters in the eel
(Cutler et al., 2000
).
Although there was no evidence for the presence of AQP-3 mRNA in the eel
stomach, a tissue that exhibits significant levels of expression in mammals
(Matsuzaki et al., 2000
),
AQP-3 mRNA was easily detected in other parts of the eel gastrointestinal
tract.
The presence of AQP-3 mRNA in the oesophagus is of special interest as
there is thought to be little net water flux across this epithelium in
SW-acclimated eels that are drinking the hyperosmotic seawater environment
(Hirano and Mayer-Gostan,
1976). Despite this report, a high unidirectional water flux has
been reported to occur across the flounder oesophagus when in SW
(Parmelee and Renfro, 1983
).
Taken together, this suggests that while there may be no significant net flow
of water across this epithelium, there may be water transport mechanisms
present that counteract the osmotic loss of water to the hyperosmotic SW in
the lumen. Since it is unlikely that there is a fundamental difference in the
physiological function of eel and flounder oesophagus, AQP-3 could play an
active role in both species by providing a conduit for serosal (basolateral)
transport of water into the epithelia, thus preventing cellular dehydration,
as has been previously suggested (Matsuzaki et al.,
1999
,
2000
).
In the quantitative expression studies, FW to SW transfer induced no change
in the AQP-3 mRNA levels found in intestinal samples taken from either yellow
or silver eels. This suggested that AQP-3 was unlikely to play any role in the
absorption of water that occurred following the drinking response in marine
teleosts. Subsequent to these experiments more recent immunohistochemical
studies have indicated that AQP-3 is not expressed in intestinal enterocytes
but, is in fact localised to discrete structures, possibly macrophages, within
the intestinal epithelium of both FW and SW eels
(Lignot et al., 2002). These
results further support our contention that AQP-3 has little role in water
absorption across the intestinal epithelium.
Quantitative northern blots indicated that there was a major downregulation
in AQP-3 mRNA expression in the branchial epithelium after SW-transfer. 3
weeks after SW transfer, AQP-3 mRNA abundance had decreased by 76 % in yellow
eels and 97 % in silver eels compared to FW control fish. These differences in
branchial AQP-3 expression between FW- and SW-acclimated eels correlates well
with the three- to 11-fold higher levels of osmotic water permeability found
in the gills of FW fish (Motais and Isaia,
1972). There is also an additional correlation between AQP-3
expression and the threefold higher levels of branchial urea clearance rates
found in FW compared to SW eels (Masoni
and Payan, 1974
), indicating that AQP-3 may be involved in either
water and/or urea transport.
It is still not clear why water (and urea) transport across the gills is
much higher in FW- than SW-acclimated fish, and what role an aquaporin channel
such as AQP-3 might play. There is no reason to believe that the respiratory
requirements of FW or SW eels should be markedly different, so changes in
water permeability are unlikely to relate to respiratory gas transfer. One
possible explanation for the higher osmotic water permeability of FW eels is
that, as a result of differences in expression of one or more apically located
solute transporters, there is an increase in water uptake across the apical
membrane into the branchial epithelial cells. As AQP-3 has universally been
shown to have a basolateral localisation in mammalian epithelia, it is
possible that the eel AQP-3 may act as a conduit in the basolateral membrane
for the release of water to the serosal fluid and the prevention of cell
swelling (Cutler and Cramb,
2000; Matsuzaki et al.,
2000
).
One pathway that may exist in branchial surface epithelial cells is the
putative `bulk fluid flow' ion transport mechanism thought to operate through
the tubular system of chloride cells
(Isaia, 1984). Here, AQP-3
would be most likely to act as a conduit for water into or out of the tubular
system. Links between ion and water transport have already been established in
mammals, where AQP-3 is thought to be regulated by the cystic fibrosis
transmembrane conductance regulator (CFTR;
Schreiber et al., 2000
).
However, as the `bulk flow' mechanism is associated with ion transport in
marine teleosts, it is likely to be less, rather than more, active in FW
eels.
Finally, the possibility also exists that water transport is just the
consequence of some primary role of AQP-3 such as the transport of some other
solute(s) molecule. One obvious possibility is that AQP-3 is essential for
urea excretion across the gills. This would correlate well with the parallels
between water and urea permeability/clearance across the gill
(Motais and Isaia, 1972;
Masoni and Payan, 1974
) as
well as their regulation by adrenaline
(Bergman et al., 1974
;
Sorenson and Fromm, 1976
;
Haywood et al., 1977
; Isaia et
al., 1978; Isaia, 1979
,
1984
;
Zadunaisky, 1984
). Another
potential substrate for the AQP-3 transporter is ammonia. Ammonia, a more
abundant nitrogenous waste product than urea in many fish, is also excreted
across the gills, and certain plant (Cooper
et al., 2000
) and animal
(Nakhoul et al., 2001
)
aquaporins have been shown to transport this compound. FW fish are known to
excrete a higher percentage of total body ammonia across the gills than marine
teleosts (Wilkie, 1997
). This
may be because SW fish can excrete much of their nitrogen waste as ammonium
ions through ion transport mechanisms in the gill
(Wilkie, 1997
). Another
molecule that might additionally be transported by eel AQP-3 is
CO2. Several studies have shown that the transport of
CO2 can be facilitated by mammalian AQP-1
(Cooper and Boron, 1998
;
Nakhoul et al., 1998
;
Prasad et al., 1998
;
Yang et al., 2000
). As is the
case for oxygen uptake, it would seem unlikely from a respiratory standpoint
that a significantly greater branchial permeability for CO2 would
exist in FW than SW fish. However, it could be argued that there is a need for
additional controls of body fluid pH while in FW environments. If the latter
is indeed the case, changes in environmental pH should have profound effects
on the expression of branchial AQP-3 in the eel. Recently it has been
suggested that mammalian AQP-3 but not AQPs 0, 1, 2, 4 or 5 may be gated by
H+ ions (Zeuthen and Klaerke,
1999
). Whether this mechanism operates in teleosts for the
regulation of water permeability or body fluid pH awaits further
investigation.
Possible clues to the principle role of AQP-3 in the eel gill may reside in
studies that reveal the cellular location of this transporter in the branchial
and intestinal epithelium. The accompanying paper
(Lignot et al., 2002) presents
some initial immunohistochemical studies which go some way to answering these
questions.
Although this paper has taken a significant first step in improving our understanding of the processes that underlie water transport in fish, much remains to be determined. Future studies will need to determine the range of solutes that can be transported by eel AQP-3 and also to focus on the hormonal regulation that underpins the changes in AQP-3 expression and action during salinity acclimation.
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Acknowledgments |
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References |
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