Ontogeny of osmoregulatory structures and functions in the green crab Carcinus maenas (Crustacea, Decapoda)
1 Biologische Anstalt Helgoland/Stiftung Alfred-Wegener-Institut für
Polarund Meeresforschung, Meeresstation, D-27498 Helgoland, Germany
2 Equipe Adaptation Ecophysiologique et Ontogenèse, UMR 5000 GPIA,
Université Montpellier II, F-34095 Montpellier cedex 05,
France
* Author for correspondence (e-mail: ucieluch{at}awi-bremerhaven.de)
Accepted 16 October 2003
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Summary |
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Key words: osmoregulation, ontogeny, hemolymph osmolality, immunolocalization, Na+/K+-ATPase, gill, larva, ionocyte, Carcinus maenas
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Introduction |
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In the process of ionic regulation, Na+/K+-ATPase is
one of the most important enzymes (reviewed by Towle,
1981,
1984a
,b
;
Péqueux, 1995
;
Charmantier, 1998
;
Lucu and Towle, 2003
). By
using ATP as a source of energy, it enables an active ion-exchange across
epithelial membranes (Neufeld et al.,
1980
; De Renzis and Bornancin,
1984
). Immunolocalization of Na+/K+-ATPase
using monoclonal antibodies has recently been used as a tool to identify
transporting epithelia, e.g. in the terrestrial isopod Porcellio
scaber (Ziegler, 1997
),
lobster Homarus gammarus (Lignot
et al., 1999
; Lignot and
Charmantier, 2001
), and in crayfish Astacus leptodactylus
(Barradas et al., 1999
). By
investigating the development, location and functionality of transporting
epithelia, the precise cellular location of
Na+/K+-ATPase is of special interest
(Flik et al., 1994
;
Haond et al., 1998
;
Lignot et al., 1999
;
Lignot and Charmantier,
2001
).
Several studies have been conducted on the ontogeny of osmoregulation in
various species (reviewed by Charmantier,
1998). However, investigations on the ontogeny of osmoregulating
tissues and its potential variations throughout development are still very
limited (Hong, 1988
;
Bouaricha et al., 1994
;
Charmantier, 1998
;
Anger, 2001
;
Lignot and Charmantier, 2001
).
Among the few species in which the ontogeny of ion-transporting epithelia have
been investigated by histological and/or electron microscopical studies are
Farfantepenaeus aztecus (Talbot
et al., 1972
), Callianassa jamaicense
(Felder et al., 1986
),
Penaeus japonicus (Bouaricha et
al., 1994
) and Homarus gammarus
(Lignot and Charmantier,
2001
). From these studies it appears that organs other than gills
can also play a major role in ion-transport and that the location of epithelia
involved in ion-exchange can change during development (reviewed by
Charmantier, 1998
).
The adult green crab Carcinus maenas (L.) is a euryhaline species
that exhibits the ability of effective hyperosmoregulation in habitats of low
and/or fluctuating salinity (Theede,
1969; Siebers et al.,
1982
,
1985
). In European waters,
this ability has enabled the crab to cover a wide geographical area from the
Baltic Sea to the Azores, living in habitats where salinity ranges from
9
to 35
(Winkler et al.,
1988
). Its euryhalinity has also aided in it becoming an invasive
species in estuarine habitats of the east and west coasts of the USA and
Canada, as well as in West and South Africa and Australia
(Cohen et al., 1995
;
Grosholz and Ruiz, 1995
;
Lafferty and Kuris, 1996
).
The gills of adult C. maenas have received much attention as the
potential site of ionic exchange and much information, including the location
and fine structure of ionocytes, is known (e.g.
Compere et al., 1989;
Taylor and Taylor, 1992
;
Lawson et al., 1994
;
Hebel et al., 1999
). In
addition, an ultracytochemical approach conducted in gills of C.
maenas showed that the presence of Na+/K+-ATPase is
mainly restricted to basolateral infoldings of epithelial cells in posterior
gill lamellae (Towle and Kays,
1986
).
In contrast to the ability of adult C. maenas to live over
extended periods in habitats with low salinity, the reproduction,
embryogenesis and larval development of this species require higher salt
concentrations (Green, 1968;
Kinne, 1971
;
Nagaraj, 1993
). A laboratory
study on the tolerance of C. maenas larvae from the North Sea facing
hypo-osmotic stress showed that a salinity of at least 25
is needed
for successful development (Anger et al.,
1998
). At reduced salinities (
20
), significant
decreases were found in the rates of early zoeal survival, development,
growth, respiration and assimilation (Anger
et al., 1998
). It is thus likely that the osmo-physiological
pattern changes during the course of development.
The present investigation was conducted (i) to study the ontogeny of osmoregulation by direct measurements of the hemolymph osmolality, (ii) to locate and follow the development of osmoregulatory epithelia and the expression of Na+/K+-ATPase using transmission electron microscopy (TEM) and immunofluorescence light microscopy (ILM), and (iii) to relate the ontogeny of osmoregulation to the development of transporting epithelia and to ecological traits.
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Materials and methods |
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Preparation of media
Experimental media were obtained by diluting 1 µm-filtered sea water
(32) with desalinated freshwater or by adding Tropic Marin® salt
(Wartenberg, Germany). Salinity was expressed as osmotic pressure (in mOsm
kg-1) and as salt content of the medium (in
); a value of
3.4
is equivalent to 100 mOsm kg-1 (29.41 mOsm
kg-1
1
). The osmotic pressure of the media was measured
with a micro-osmometer Model 3 MO plus (Advanced Instruments, Needham Heights,
MA, USA), requiring 20 µl per sample. The following media were prepared,
stored at 18°C and used in the osmoregulation experiment: 30 mOsm
kg-1 (1.0
), 155 mOsm kg-1 (5.3
), 300
mOsm kg-1 (10.2
), 500 mOsm kg-1 (17.0
),
749 mOsm kg-1 (25.5
), 947 mOsm kg-1
(32.2
, referred to as seawater) and 1302 mOsm kg-1
(44.3
).
Osmoregulation
The experiment was carried out at a constant temperature of 18°C,
representative of typical summer conditions in the area of origin of our
material, the North Sea, and known to be favourable for both larval and adult
C. maenas, both in the laboratory
(Dawirs, 1985;
Anger et al., 1998
) and in the
field (Harms et al.,
1994
).
Larvae and juveniles were transferred directly to the experimental media
and exposed for 24 h (72 h in large juveniles from the field) in covered Petri
dishes. Following their capture, large juvenile crabs from the field were kept
in seawater (32
) for 48 h at 18°C. The number of exposed
animals was kept to a minimum level of 9-11 individuals per condition. Dead
animals were counted at the end of the exposure time to obtain survival rates.
The surviving specimens were superficially dried on filter paper and quickly
immersed into mineral oil to prevent evaporation and dessication. Any
remaining adherent water was removed using a glass micropipette. A new
micropipette was then inserted into the heart for hemolymph sampling. For all
experimental stages, hemolymph osmolality was measured with reference to the
medium osmolality on a Kalber-Clifton nanoliter osmometer (Clifton Technical
Physics, Hartford, CT, USA) requiring about 30 nl. Results were expressed
either as hemolymph osmolality or as osmoregulatory capacity. The latter is
defined as the difference between the osmolality of the hemolymph and that of
the medium. Analysis of variance (ANOVA) and Student's t-tests were
used for multiple and pairwise statistical comparisons of mean values,
respectively, after appropriate checks for normal distribution and equality of
variance (Sokal and Rohlf,
1995
).
Immunofluorescence light microscopy
After removal of the carapace, anterior and posterior gills of adult C.
maenas were dissected from the inner body wall and fixed for 24 h in
Bouin's fixative. Zoeae, megalopae and crab I were fixed by direct immersion
in the same fixative. After rinsing in 70% ethanol, samples were fully
dehydrated in a graded ethanol series and embedded in Paraplast-extra (Sigma).
Sections (4 µm) were cut on a Leitz Wetzlar microtome, collected on
poly-L-lysine-coated slides and stored overnight at 38°C.
Sections were then pre-incubated for 10 min in 0.01 mmol l-1 Tween
20, 150 mmol l-1 NaCl in 10 mmol l-1 phosphate buffer,
pH 7.3. To remove the free aldehyde groups of the fixative, samples were
treated for 5 min with 50 mmol l-1 NH4Cl in
phosphate-buffered saline (PBS), pH 7.3. The sections were then washed in PBS
and incubated for 10 min with a blocking solution (BS) containing 1% bovine
serum albumin (BSA) and 0.1% gelatine in PBS. The primary antibody (monoclonal
antibody IgG5, raised against the avian
-subunit of
the Na+/K+-ATPase) was diluted in PBS to 20 µg
ml-1, placed in small droplets of 100 µl on the sections and
incubated for 2 h at room temperature in a wet chamber. Control sections were
incubated in BS without primary antibody. To remove unbound antibodies, the
sections were then washed (3x 5 min) in BS and incubated for 1 h with
small droplets (100 µl) of secondary antibody, fluorescein-isothiocyanate
(FITC)-labeled goat anti-mouse IgG (Jackson Immunoresearch, West Baltimore,
USA). After extensive washes in BS (4x 5 min), the sections were covered
with a mounting medium and examined using a fluorescence microscope (Leitz
Diaplan coupled to a Ploemopak 1-Lambda lamp) with an appropriate filter set
(450-490 nm band-pass excitation filter) and a phase-contrast device.
Transmission electron microscopy
Anterior and posterior gills of adult crabs were cut into small pieces and
fixed for 1.5 h in 5% glutaraldehyde solution buffered at pH 7.4 with 0.1 mol
l-1 cacodylate buffer. Zoeae, megalopae and early crab stages were
fixed for 1 h by direct immersion in the same fixative. For adjustment to the
osmotic pressure of the hemolymph, NaCl was added to the fixative and buffer
to give a final osmolality of 735 mOsm kg-1. Samples were then
rinsed in buffer and postfixed for 1.5 h at room temperature in buffered 1%
OsO4. After extensive washes in buffer, the samples were fully
dehydrated in graded acetone and embedded in Spurr low viscosity medium.
Semithin sections (1 µm) were prepared using glass knives with a LKB
microtome and stained with Methylene Blue for light microscopic observations.
Ultrathin sections were obtained using a diamond knife, contrasted with uranyl
acetate (Watson, 1958) and
lead citrate (Reynolds, 1963
)
and examined with a transmission electron microscope (EM 902, Zeiss, Germany)
operated at 80 kV.
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Results |
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Osmoregulation
The developmental stages were exposed to a wide range of salinities. The
experimental results are given as variations in hemolymph osmolality and as
osmoregulatory capacity in relation to the osmolality of the experimental
medium (Fig. 1A,B).
|
The pattern of osmoregulation changed during development. With the
exception of the first zoea, no significant differences were observed between
successive zoeal stages exposed to the same salinities. Only zoeae I larvae
were able of a slight hyperregulation at 500 mOsm kg-1
(17.0). All later zoeal stages (ZII-ZIV) osmoconformed over the entire
tested salinity range, 300-1302 mOsm kg-1 (10.2-44.3
). A
significant change in the pattern of osmoregulation was noted in the
megalopae. This stage osmoconformed at high salinities (947 mOsm
kg-1 or 32.2
; 1302 mOsm kg-1 or 44.3
).
At lower salinities (300-749 mOsm kg-1 or 10.2-25.5
), the
megalopae showed a strong ability for hyper-regulation. Later developmental
stages (crabs instars I, II, larger juveniles) maintained the osmoregulatory
pattern displayed by the megalopae, but with an increased osmoregulatory
capacity in media from 300 to 749 mOsm kg-1 (10.2-25.5
).
For instance, at 500 mOsm kg-1 (17
), the osmoregulatory
capacity in mOsm kg-1 was 33 in the zoea I, 1-5 in zoeal stages
II-IV, 89 in megalopa, and 188, 216 and 228, respectively, in crab I, crab II
and larger juveniles. All juveniles hyper-regulated at a low salinity of 155
mOsm kg-1 (5.3
).
Immunolocalization of Na+/K+-ATPase
The method of fixation and Paraplast-embedding procedures led to a good
tissue preservation and a good antigenic response, as observed by
phase-contrast microscopy (Figs
2B,D,F,
3B,D,F) and fluorescent
microscopy (Figs 2A,C,E,
3A,C,E). Control sections of
posterior gills without the primary antibody showed no specific immunolabeling
along the epithelial cells of the gill filaments or along the gill shaft
(Fig. 3E,F). A non-specific
auto-fluorescence was observed along the surrounding cuticle of anterior and
posterior gills (Fig.
3A,E).
|
|
In the zoea IV stage, gill buds were present within the branchial cavity. Only very weak traces of immunofluorescence staining were noted in gill buds (Fig. 2A,B). In the megalopa, the branchial cavity contained slightly lamellated anterior and posterior gills. Anterior gills lacked immunofluorescence whereas posterior gills showed specific binding of antibodies within the filaments and the central shaft of the gill (Fig. 2C,D). In the first crab instar, immunoreactivity was observed in the now well-formed filaments, in the marginal vessels at the tip of each filament and along the central shaft of the posterior gills (Fig. 2E). Immunofluorescence was mainly observed in the basal filaments of the gills, whereas apical gill parts appeared free of specific immunolabeling (Fig. 2E). In adults (Fig. 3A-D), no immunofluorescence was noted in the filaments, in the marginal vessels or along the gill shaft of anterior gills (Fig. 3A,B). A specific fluorescence was observed in the epithelial cells and pillar cells of proximal posterior gill filaments (Fig. 3C,D). The marginal tips and the central shaft of posterior gills showed no immunolabeling (not illustrated).
Ultrastructure of epithelial gill cells
Gills from adults
In the filaments of posterior gills, several principal cell types were
recognized, including chief cells, pillar cells, nephrocytes and glycocytes
(not illustrated). Ionocytes, or striated cells, which were mainly distributed
towards the proximal part of gill filaments, showed distinct features of
ion-transporting cells. These include apical microvilli in close contact to
the cuticle and numerous elongated mitochondria often in close contact to
basolateral infoldings of the cytoplasmic membrane
(Fig. 4F). A basal membrane
separates the epithelial cells from hemolymphatic spaces
(Fig. 4F).
|
Gills from larvae and juveniles
Epithelial cells in posterior gill buds of the zoea IV were rather
undifferentiated. The cells possessed a central nucleus surrounded by a few
mitochondria (Fig. 4A). In the
megalopa, epithelial ionocytes were found in basal parts of posterior gill
filaments. The cytoplasmic membrane showed basolateral infoldings and formed a
microvillious border at the apical part of the epithelial cells
(Fig. 4B). In the first crab
stage, ionocytes with typical features of ion-transporting cells could also be
recognized in basal filaments of posterior gills
(Fig. 4C-E).
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Discussion |
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Osmoregulation
Three alternative ontogenetic patterns can be recognized by comparing the
ontogeny of osmoregulation in decapod crustaceans
(Charmantier, 1998): (a)
osmoregulation is weak and varies only little during the course of
development; (b) the first postembryonic stage possesses the same
osmoregulatory pattern as the adults; (c) the osmoregulatory pattern changes
during development, usually at or after metamorphosis, from an osmoconforming
to an osmoregulating response. The shore crab C. maenas clearly
belongs to the third category, in which the pattern of osmoregulation changes
during the postembryonic development. Adult C. maenas are euryhaline
hyper-regulators in habitats with low and/or fluctuating salinity
(Theede, 1969
; Siebers et al.,
1982
,
1985
;
Winkler et al., 1988
). In
contrast to the euryhalinity in adults, successful larval development through
metamorphosis requires, at least in marine populations, salinities of at least
20
or 25
(Nagaraj,
1993
; Anger et al.,
1998
). In our experiments, the zoeal stages II-IV were stenohaline
osmoconformers, while the zoea I was a weak hyper-osmoregulator in dilute
medium (17
). Remarkably, this ability to hyper-regulate in brackish
water was already present in newly hatched zoea I, disappeared in the
subsequent zoeal stages and than reappeared in the megalopa. The ecological
implications will be discussed below. A similar osmoregulatory pattern has
also been noted in the larval development of the strongly hyper-regulating
grapsoid crab Chasmagnathus granulata
(Charmantier et al., 2002
).
The authors suggested that a limited hyper-osmoregulatory capability of the
freshly hatched zoea I larvae should allow for survival at low salinity after
hatching within the parental estuarine habitat, until the larvae are
transported to regions with higher salinities (for more detailed discussion of
ecological implications of our findings, see below).
The later zoeal stages (II-IV) of C. maenas were iso-osmotic over
the entire range of tolerable salinities and can thus be regarded as true
marine osmoconformers. The intolerance of dilute medium was limited to
25
, as increasing mortality levels occurred at lower salinities.
This limited osmotic tolerance of the zoeal stages supports the findings by
Anger et al. (1998
), in which
salinities below 25
led to decreasing early zoeal survival,
development, growth, respiration and assimilation.
A considerable shift in the osmoregulatory pattern occured after the first
metamorphic molt, from the last zoeal stage (IV) to the megalopa. The megalopa
was still osmoconforming in salinities 32
, but was able to
hyper-regulate in dilute media down to
10
. Although its capability
to hyper-regulate was still limited compared to the following instars (crabs I
and II, later juveniles), the osmoregulatory pattern (hyperisoregulation) of
adult C. maenas is in principle already established in the megalopa.
The next metamorphic molt, from the megalopa to the first juvenile crab stage,
showed considerably increased ability for hyper-regulation, allowing now for a
tolerance of salinities down to as low as
5
. The osmoregulatory
capacity of the crab I did not differ greatly from that in the following stage
(crab II), and it increased only slightly in later juveniles. However,
survival rates at salinities from 1.0
to 25.5
as well as
hemolymph osmolality at 5.3
observed in larger juveniles from the
field were below those of laboratory-reared crab I and II. Other factors such
as temperature, salinity, water and food quality, which can be controlled and
kept constant in the laboratory, are known to influence larval development and
overall fitness (reviewed by Anger,
2001
). Unknown natural variabilities in those factors in the field
might thus account for the slight but significant reduction in salinity
tolerance and osmoregulatory capability observed in later juvenile crabs.
As a preliminary conclusion, the establishment of the adult osmoregulatory
pattern in C. maenas is accomplished through two metamorphic steps:
(1) the zoea-megalopa transition with the appearance of limited
hyper-regulation, and (2) the megalopa-crab transition with a further
substantial increase in the osmoregulatory capacity and, in consequence, a
higher tolerance of lower salinities. A similar timing of
metamorphosis-related changes in osmoregulatory patterns has been reported for
other brachyuran crabs such as the strongly regulating grapsoids Armases
miersii (Charmantier et al.,
1998), Sesarma curacaoense
(Anger and Charmantier, 2000
),
and Chasmagnathus granulata
(Charmantier et al., 2002
), or
in the ocypodid Uca subcylindrica
(Rabalais and Cameron, 1985
).
An exception was found in the grapsoid Sesarma reticulatum, where the
megalopa maintained the initial zoeal osmoregulatory pattern and the
osmo-physiological shift only appeared after the megalopa-crab transition
(Foskett, 1977
). Sharing a
pattern of ontogenetic changes in osmoregulation similar to those of strongly
regulating grapsoids, but still limited by its osmotic tolerance, this study
confirms that C. maenas is a transitional species between true marine
osmoconformers like Cancer spp.
(Charmantier and Charmantier-Daures,
1991
), and very strongly regulating, freshwater-invading species
such as Eriocheir sinensis (G. Charmantier, unpublished data).
Immunolocalization of Na+/K+-ATPase and gill
ultrastructure
Osmoregulation is based on efficient ionic regulation (mainly of
Na+ and Cl-), accomplished by specialized transporting
epithelia where the enzyme Na+/K+-ATPase is abundantly
located (Thuet et al., 1988;
Lignot et al., 1999
;
Lignot and Charmantier, 2001
;
reviewed by Lucu and Towle,
2003
). The ontogeny of osmoregulation of C. maenas is
correlated with the expression of Na+/K+-ATPase (present
study) and the development of gills (for detailed discription of gill
development in C. maenas, see
Hong, 1988
).
In the last zoeal stage (zoea IV), which is an osmoconformer,
undifferentiated gill buds are formed within the branchial chamber.
Na+/K+-ATPase was almost absent within these organs,
suggesting that these simple branchial extensions are not yet involved in
effective ionic exchange. This suggestion is supported by the ultrastructure
of epithelial cells found within these organs (see
Fig. 4A). They lack typical
features of ionocytes such as apical microvilli and basolateral infoldings.
Gill morphology begins to differentiate after metamorphosis to the megalopa,
which has limited ability to hyper-osmoregulate. The arthobranchs and
pleurobranchs arising from the thoracal appendages then become differentiated
into a central gill shaft and partially lamellated filaments. Epithelial cells
with typical ion-tansporting features can now be found within the gill
filaments. This morphological and ultrastructural change coincides with a
possible involvement of gills in osmoregulation, also supported by the
presence of Na+/K+-ATPase in epithelia of the gill shaft
and of the filaments of the posterior two pleurobranchs. The presence of
Na+/K+-ATPase can be related to an involvement of the
epithelial cells in ionic exchange (Lignot
et al., 1999; Lignot and
Charmantier, 2001
). Different immunoreactivity between anterior
and posterior gills was observed in the megalopa and in the following juvenile
stages. In the crab I, a strong hyper-osmoregulator, the three posterior gills
(1 arthobranch and 2 pleurobranchs) are well developed within the branchial
chamber, and Na+/K+-ATPase is distributed mainly in the
basal filaments. Ionocytes found in the filaments are similar to those
observed in adults, including typical features such as a microvillous border
and numerous mitochondria in close contact to basolateral infoldings of the
cytoplasmic membrane. Thus, most posterior gills are involved in
osmoregulation, whereas the anterior gills with thin epithelial cells and lack
of Na+/K+-ATPase seem to attain respiratory functions.
These findings agree with previous studies conducted on the crabs
Callinectes sapidus and Carcinus maenas
(Towle and Kays, 1986
). Our
study supports also the observation that, in adult C. maenas,
Na+/K+-ATPase is mainly restricted to basolateral
infoldings of thick epithelial gill cells in posterior gills
(Towle and Kays, 1986
).
In other decapod crustaceans species, organs like branchiostegites and
epipodites play, at least at certain points during development, an important
role in ionic exchange. For instance, an immunohistochemical approach in
juvenile Homarus gammarus showed that
Na+/K+-ATPase is mainly restricted to epithelia of the
epipodite and the inner epithelium of the branchiostegite
(Lignot et al., 1999).
Following the ontogeny of osmoregulatory functions in H. gammarus,
the presence of Na+/K+-ATPase in epipodites has been
already established in embryos, and the branchiostegite appears as an
additional osmoregulatory organ after metamorphosis
(Haond et al., 1998
;
Flik and Haond, 2000
;
Lignot and Charmantier, 2001
).
In C. maenas, no shift in location or function of ion-transporting
epithelia was observed in this study. The temporary and low
hyper-osmoregulatory ability that we report in zoea I might originate from the
temporary occurrence of ionocytes along the branchiostegites, but this remains
to be studied. The posterior gills appear as the dominant organs involved in
the process of ionic regulation. An increase in
Na+/K+-ATPase after abrupt transfer to low salinity has
been observed in anterior gills of C. maenas
(Lucu and Flik, 1999
), but the
present study confirms the major role of posterior gills of C. maenas
in the process of ionic exchange (Towle
and Kays, 1986
; Goodman and
Cavey, 1990
; Taylor and
Taylor, 1992
; Lawson et al.,
1994
; Hebel et al.,
1999
). The development of gills and the expression of
Na+/K+-ATPase can therefore be considered as one of the
main processes enabling the effective hyper-osmoregulatory abilities of C.
maenas.
Ecological implications
In the natural environments of the shore crab C. maenas, low
and/or fluctuating salinities are common, but osmotic stress is initiated or
compensated by effective hyper-regulation of internal ionic concentration
(Theede, 1969; Siebers et al.,
1982
,
1985
). This mechanism was
observed, although only weakly developed, as early as in the zoea I stage,
which hatches in the same habitat where adults live. In contrast, later zoeal
stages of C. maenas from the North Sea are true marine
osmoconformers, which suffer osmotic stress when they are exposed to
constantly low or varying salinities (Anger
et al., 1998
). Behavioural mechanisms such as tide-related release
of larvae and endogenous vertical migration rhythms are known from estuarine
crab populations living adjacent to the sea
(Queiroga et al., 1994
; Zeng
and Naylor,
1996a
,b
,c
).
These mechanisms provide a rapid off-shore export of larvae to regions with
higher salt concentrations (Anger,
2001
). In the case of a retention in areas with lower salinity,
zoea larvae must face hypo-osmotic stress. During a short-term exposure to
such conditions, mechanisms of isosmotic intracellular regulation may allow
for survival, as observed in the lobster Homarus gammarus
(Haond et al., 1999
).
Typical of a brachyuran crab, the morphological metamorphosis of C.
maenas is accomplished over two molts
(Rice and Ingle, 1975). After
the first metamorphic molt, the megalopa resembles an intermediate stage
between the planktonic zoeae and the benthic crabs. Towards the end of this
instar, the megalopa settles and molts to the first juvenile crab instar
(Crothers, 1967
). The megalopa
can be also regarded as an intermediate stage in terms of osmoregulation,
differing from the zoeae by its hyper-osmoregulatory ability in salinities
<25
, yet limited in its osmotic tolerance and ion-regulating
capacity compared to the subsequent juvenile crab instars. However, the
osmoregulatory ability in the megalopa allows for a reinvasion of areas with
low salt concentrations. The second shift, with another substantial increase
in the hyperregulating ability and, consequently, an enhanced tolerance of low
salinities, takes place at the transition from the megalopa to the first crab
stage, in which the morphological, anatomical and osmo-physiological
metamorphosis of C. maenas can be considered as complete. The young
crab is able to cope with low and/or fluctuating salinities, which extends its
habitat to areas with brackish water conditions, e.g. estuaries (Siebers et
al., 1982
,
1985
).
Populations of C. maenas that live in coastal areas and estuaries
of the North Sea are influenced by tidally fluctuating salinities, while their
counterparts living in the Baltic Sea are exposed to rather constant
conditions of low osmotic pressure. It is still unknown whether the population
of C. maenas in the western Baltic Sea is capable of reproduction.
Although freshly hatched zoeae of C. maenas are seasonally abundant
in the surface plankton at average salinities of 15 in the Kiel Fjord,
Baltic Sea (Kändler,
1961
), advanced developmental stages have not so far been
observed. Comparative studies have shown that adult C. maenas from
the Baltic Sea have a higher capacity of hyper-regulation than crabs from the
North Sea, and a cross-wise adaptation to higher or lower salinities was only
partially reversible (Theede,
1969
). Anger et al.
(1998
) suggested that there
might be genetic differences between these populations. Although physiological
variations have been studied in adult crabs from geographically separated
populations (Theede, 1969
) and
in different colour morphs (McGaw and Naylor,
1992a
,b
),
no information is available about the larval response of crabs from the Baltic
Sea to salinity variations. A comparative study on the ontogeny of
osmoregulation and of reproductive traits in C. maenas from the North
Sea and the Baltic Sea might thus provide valuable information on the
processes required for a succesful establishment of decapod crustacean species
in habitats with constantly low salinity.
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Acknowledgments |
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References |
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